Gel Doc XR+ and

ChemiDoc XRS+
Systems with Image Lab
Software
User Guide
Version 5.1

Notice
No part of this publication may be reproduced or transmitted in any form or by any
means, electronic or mechanical, including photocopy, recording, or any
information storage or retrieval system, without permission in writing from
Bio-Rad Laboratories, Inc.
Bio-Rad reserves the right to modify its products and services at any time. This
user guide is subject to change without notice. Although prepared to ensure
accuracy, Bio-Rad assumes no liability for errors or omissions, or for any damage
resulting from the application or use of this information.

Credits
1. Image Lab software is based in part on the work of the Qwt project
(http://qwt.sf.net).
2. Image Lab software is based in part on the work of the CImg project
(http://cimg.sourceforge.net/).
See license for details at
www.cecill.info/licences/Licence_CeCILL-C_V1-en.html
3. Image Lab software is based in part on the work of the Independent JPEG
Group (www.ijg.org/)
Alexa Fluor, Coomassie Fluor, Qdot, and SYPRO are trademarks of Invitrogen
Corporation. Coomassie is a trademark of BASF Aktiengesellschaft. Cy2 and Cy3
are trademarks of GE Healthcare. DyLight and Krypton are trademarks of Thermo
Fisher Scientific Inc. Excel, PowerPoint, and Windows XP, and Windows 7 are
trademarks of Microsoft Corporation. iWork, Macintosh, Mac OS, and Numbers
are trademarks of Apple Inc. GelGreen and GelRed are trademarks of Biotium, Inc.
Intel Core and Pentium are trademarks of Intel Corporation. Mitsubishi is a
trademark of Mitsubishi Companies. PulseNet International is a trademark of
Centers for Disease Control and Prevention. Slo-Blo is a trademark of Littelfuse.
CHEF (U.S. Patent Number 5,549,796, issued to Stanford University) is exclusively
licensed to Bio-Rad Laboratories, Inc.
Precision Plus Protein standards are sold under license from Life Technologies
Corporation, Carlsbad, CA, for use only by the buyer of the product. The buyer is
not authorized to sell or resell this product or its components.
SYBR is a trademark of Life Technologies, Inc. Bio-Rad Laboratories, Inc. is
licensed by Life Technologies, Inc. to sell reagents containing SYBR Green I for
use in real-time PCR, for research purposes only.
Bio-Rad Laboratories, Inc. is licensed by Invitrogen Corporation to sell SYPRO
products for research use only under U.S. Patent Number 5,616,502.
Copyright 2014 Bio-Rad Laboratories, Inc.

Table of Contents
Safety and Regulatory Compliance . . . . . . . . . . . . . . . . . . . . . . . . . .11
Important Safety Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Warranty . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12
General Precautions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12
Regulatory Notices . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
Instrument Safety Warnings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
Notice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Power Safety Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15
Voltage Setting Information. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15
Fuses. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15

Chapter 1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .17

Gel Doc XR+ Imaging System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
ChemiDoc XRS+ Imaging System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18
System Components . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18
CCD Camera and Lenses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18
Universal Hood . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19
Image Lab Software . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19
Emission Filters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
Optional Accessories . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
Printer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
Conversion Screens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20

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Chapter 2 Gel Doc XR+ Imager . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21

Gel Doc XR+ Imager Workflow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
Gel Doc XR+ Applications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22
Gel Doc XR+ Technical Specifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

Chapter 3 ChemiDoc XRS+ Imager. . . . . . . . . . . . . . . . . . . . . . . . . . . 25

ChemiDoc XRS+ Imager Workflow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25
ChemiDoc XRS+ Applications. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26
ChemiDoc XRS+ Technical Specifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27

Chapter 4 Setting Up the Instrument with Image Lab Software . . . 29

System Requirements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29
Image Lab Security Edition . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29
Computer Specifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29
Installing Image Lab Software . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30
Installing the Drivers on a Windows XP Computer . . . . . . . . . . . . . . . . . . . . . . . . 36
Installing the Drivers on a Windows 7 Computer . . . . . . . . . . . . . . . . . . . . . . . . . 39
Setting Up Image Lab Security Edition . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39
Activating and Deactivating Security Edition. . . . . . . . . . . . . . . . . . . . . . . . . . . 39
Enabling and Disabling Image Lab Secure Mode . . . . . . . . . . . . . . . . . . . . . . . 46
Setting Security Preferences . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50
Renaming Security Groups . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53
Using Groups on a Local Domain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53
Changing Security Preferences . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54

Chapter 5 Image Lab Software Overview . . . . . . . . . . . . . . . . . . . . . . 55

Interface Overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 57
Main Window. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 57
Main Toolbar . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58
Results Data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58
Display Toolbox . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58
Start Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59
Analysis Tool Box . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59
Status Bar . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61
Menu Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62
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Chapter 6 Acquiring Images . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69

The Protocol Setup Window . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70
Creating a Protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
Setting Up a New Protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72
Step 1. Gel Imaging for Single-Channel Protocols . . . . . . . . . . . . . . . . . . . . . . 73
Step 2. Detect Lanes and Bands. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78
Step 3. Analyze Molecular Weight. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79
Step 4. Specify Output . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 82
Review Protocol Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 83
Creating a Multichannel Image from Single Images . . . . . . . . . . . . . . . . . . . . . . . 84
Editing a Saved Protocol. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85
Positioning the Gel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 87
Running a Protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 88
Running Signal Accumulation Mode (SAM) Protocols . . . . . . . . . . . . . . . . . . . . . 89
Saving Signal Accumulation Mode (SAM) Images . . . . . . . . . . . . . . . . . . . . . . 90
Setting Up a Custom Application . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 91
Regression Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 93
Application Tables . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 94

Chapter 7 Viewing Images . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 97

Displaying Gel Images . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 98
Display Gel Options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 98
Zoom Tools . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99
Fit in Window . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 100
Image Transform . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 100
Image Colors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 102
3-D Projection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 103
Image Info . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 104
Displaying Data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 104
Analysis Table Options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 105
Lane and Band Table Measurement Definitions . . . . . . . . . . . . . . . . . . . . . . . 107
Volume Measurement Definitions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 108
Lane Profile . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 109
Standard Curve . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 112
Report . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 113
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Chapter 8 Analyzing Images . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 115

Using Auto Analysis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 115
Analysis Tool Box Tools . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 117
Image Tools . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 118
Correcting a Slanted Gel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 119
Cropping a Gel Image . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 119
Inverting Data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120
Merging Images. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120
Lane and Bands Tools . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121
Detecting Lanes. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
Using the All Lanes and Single Lane Tools . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
Copying Lanes. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 125
Detecting Bands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 126
Editing the Detected Bands. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 129
Molecular Weight (MW) Analysis Tools . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 131
Quantity Tools . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135
Annotation Tools . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 141
Add Annotations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 142
Alignment. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 142
Text Properties . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 143
Color . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 143
Rotate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 143
Volume Tools . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 144

Chapter 9 Generating Reports. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 151

Report . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 151
Report Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 152
Print Report . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 155
Print Report to a PDF File . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 155
Adjust the Printer Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 156

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Chapter 10 Exporting Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 157

Exporting Gel Images . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 157
Exporting Gel Images for Publication . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 158
Exporting Gel Images for Analysis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 160
Exporting Gel Images to PulseNet International . . . . . . . . . . . . . . . . . . . . . . . 160
Exporting Lane and Band Tables to Excel . . . . . . . . . . . . . . . . . . . . . . . . . . . 161
Exporting Volume Tables to File . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161
Screenshot Tool Export. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161
Analysis Table Export . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161

Chapter 11 System Calibration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 163

Recalibrating Your Imager. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 164

Chapter 12 Image Lab Logs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 167

Image Lab Logs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 167
Viewing the Instrument Log. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 168
Viewing the System Log . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 169
Viewing the Document Log . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 170
Displaying Log Data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 171
Displaying Data Columns in Logs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 171
Filtering Data in Logs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 172
Collapsing or Expanding Data Rows . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 174
Exporting Logs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 175
Printing Logs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 176

Chapter 13 Using the Security Edition . . . . . . . . . . . . . . . . . . . . . . . 179

21 CFR Part 11 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 179
Standard Mode versus Secure Mode . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 180
User Names, Groups, and Roles. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 180
Role Restrictions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 182
Starting Image Lab Security Edition . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 183
Electronic Records . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 183
Unsecured Documents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 184
Secure Documents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 184

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Modifying Secure Documents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 185

Signing Documents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 186
Document Logs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 187
Viewing the Document Log . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 188

Appendix A Maintenance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 189

UV Transilluminator Lamp and Starter Replacement . . . . . . . . . . . . . . . . . . . . . 189
Epi-Illumination Lamp Replacement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 191
Fuse Replacement. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 192

Appendix B Troubleshooting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193

Appendix C Setting Up Users and Groups . . . . . . . . . . . . . . . . . . 195
Setting Up Image Lab Users and Groups . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 195
User Accounts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 195
User Authentication and Group Membership . . . . . . . . . . . . . . . . . . . . . . . . . 195
Finding the name of your authentication domain . . . . . . . . . . . . . . . . . . . . . . 197
Configuring Users and Groups on a Local Computer . . . . . . . . . . . . . . . . . . . 198
Configuring Users and Groups on a Network Domain . . . . . . . . . . . . . . . . . . 202
Password Security. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 206
Password Policy Setting Examples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 208
Account Lockout Policy Setting Examples . . . . . . . . . . . . . . . . . . . . . . . . . . . 208
Auditing Windows Event Logs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 208
Miscellaneous Security Measures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 209

Appendix D Accessories . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 211

Calibrating Accessories. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 211
Installing Optional Accessories . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 211
UV/White Light Conversion Screen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 211
XcitaBlue Conversion Screen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 212
Gel Alignment Template Kit. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 214
White Light Transilluminator . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 215
Orange Fluorescence Reference Plate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 216
Ordering Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 218
viii

| Gel Doc XR+ and ChemiDoc XRS+ with Image Lab Software

| Table of Contents

Appendix E Using Bio-Rads Stain-Free Technology . . . . . . . . . 223

Stain-Free Workflow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 224
Electrophoresis with Stain-Free Gels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 225
Imaging Gels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 225
Imaging Blots. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 226

Appendix F Mitsubishi P93/P95 Thermal Printer . . . . . . . . . . . . . 227

Setting up a Thermal Printer on Windows. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 227
Setting up a Thermal Printer on a Mac . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 228

Appendix G Regression Calculation Methods . . . . . . . . . . . . . . . 229

Glossary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 231

User Guide

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x |

Table of Contents

Gel Doc XR+ and ChemiDoc XRS+ with Image Lab Software

Safety and Regulatory

Compliance
Important Safety Information
Please read these instructions before attempting to operate the
Gel Doc XR+ or ChemiDoc XRS+ imaging system.
This instrument is suitable for research use only. It must be used, therefore, only by
specialized personnel who know the health risks associated with the reagents that
are normally used with this instrument.
Use of the Gel Doc XR+ or ChemiDoc XRS+ imaging system involves UV
illumination. Proper precautions must be taken to avoid eye and skin exposure to
the UV radiation. This instrument is meant for use only by trained personnel who
know the health risks associated with the UV radiation normally used with this
instrument. The acrylic shield provides some UV protection. However, it does not
guarantee complete protection, and it is designed to shield only the person working
in front of the imager.
WARNING! Use of the acrylic screen does not guarantee the user protection
from UV radiation. The use of protective eyeglasses, mask, and/or gloves is
strongly recommended.

User Guide

| 11

| Safety and Regulatory Compliance

Warranty
The Gel Doc XR+ or ChemiDoc XRS+ imaging system is warranted against defects
in materials and workmanship for one year. If any defect occurs in the instrument
during this warranty period, Bio-Rad Laboratories, Inc. will repair or replace the
defective parts at its discretion without charge. The following defects, however, are
specifically excluded:

Defects caused by improper operation

Repair or modification done by anyone other than Bio-Rad Laboratories,

Inc. or the companys authorized agent

Use of spare parts supplied by anyone other than Bio-Rad Laboratories,

Inc.

Damage caused by accident or misuse

Damage caused by disaster

Corrosion caused by improper solvents or samples

General Precautions

12

Read the user guide carefully.

The instrument must be used only for the intended purpose of gel
documentation in research laboratories.

The instrument must be connected to a grounded power source line and

protected by a circuit breaker.

Do not pour liquids directly on or inside the instrument.

Switch off all lights on the instrument immediately after use.

Clean the transilluminator sample area after use.

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Regulatory Notices

Regulatory Notices
The Gel Doc XR+ or ChemiDoc XRS+ imaging system is designed and certified to
meet EN 61010, the internationally accepted electrical safety standard, EMC
regulations, and TUV requirements. Certified products are safe to use when
operated in accordance with this user guide. Do not modify or alter this instrument
in any way. Modification or alteration of this instrument will:

Void the manufacturers warranty

Void the regulatory certifications

Create a potential safety hazard

WARNING! Bio-Rad Laboratories, Inc. is not responsible for any injury or

damage caused by use of this instrument for purposes other than those for
which it is intended or by modifications of the instrument not performed by
Bio-Rad Laboratories, Inc., or an authorized agent.

Instrument Safety Warnings

Notes, cautions, and warnings are used to highlight certain operating procedures
and recommendations. The table below describes how each is used in this
document.
Table 1. Notes, Cautions, and Warnings
Icon

Meaning

Note:

Note: A note indicates a special procedure, an exception to normal

operation, or something else of specific interest to the reader. Notes are
preceded by the word Note.
Caution: A caution precedes an operational step that could damage the
instrument or destroy data unless the operator takes certain precautions.
Cautions located in the main text are preceded by the word Caution and are
accompanied by the caution symbol in the left margin.

User Guide |

13

| Safety and Regulatory Compliance

Table 1. Notes, Cautions, and Warnings, continued

Icon

Meaning
Caution: With the exception of cleaning or replacing light bulbs, refer all
servicing to qualified Bio-Rad personnel or their agents. If you experience
technical difficulties with the instrument, contact Bio-Rad to schedule
service. The instrument should not be modified or altered in any way.
Alteration voids the manufacturers warranty and might create a potential
safety hazard for the user.
Caution: If the case interlock is defeated, there is a possibility of UV-B
radiation hazard due to UV-B light exposure. Exercise caution when
servicing the instrument.
Caution: Bio-Rad is not responsible for any injury or damage caused by the
use of this instrument for purposes other than that for which it is intended, or
by the modification of this instrument when not performed by qualified
Bio-Rad personnel or their agents.
Caution: Disconnect the AC power cord before removing the instrument

cover.
Warning: A warning precedes an operating procedure that could cause
injury to the operator if not followed correctly. Warnings located in the main
text are preceded by the word Warning and are accompanied by the
warning symbol in the left margin.
Warning: This instrument must be connected to an appropriate AC voltage
outlet that is properly grounded.

Notice
TheGel Doc XR+ or ChemiDoc XRS+ imager is intended for laboratory use only. This
device is meant for use by specialized personnel who know the health risks
associated with reagents normally used in electrophoresis. The UV light source is
computer controlled, and proper interlocks are implemented to avoid users
accidental exposure to UV radiation. Bio-Rad Laboratories, Inc. is not responsible
for any injury or damage caused by use of this instrument for purposes other than
those for which it is intended, or for instrument modifications not performed by
Bio-Rad Laboratories, Inc. or an authorized agent.

14

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Power Safety Information

Power Safety Information

Voltage Setting Information
The universal hood of the Gel Doc XR+ or ChemiDoc XRS+ imager has a power
supply that automatically chooses the correct voltage for your country or region.

Fuses
The universal hood of the Gel Doc XR+ or ChemiDoc XRS+ imager has two userserviceable fuses, F1 and F2, which are located on the bottom rear panel and are a
part of the power entry module. See Fuse Replacement on page 192 for information
about replacing the fuses.

User Guide |

15

| Safety and Regulatory Compliance

16

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

1 Introduction
Gel Doc XR+ and ChemiDoc XRS+ gel documentation systems offer high
performance and ease of use. They both contain a charge coupled device (CCD)
camera to capture images in real time and allow you to accurately position your
sample and generate optimized image data.
The Gel Doc XR+ and ChemiDoc XRS+ systems use the same lighttight enclosure
(the universal hood), which contains built-in UV and white light illumination, but
different CCD cameras. Both systems feature dynamic flat fielding technology for
superior image uniformity and accurate quantitation.
Bio-Rad Image Lab software controls image capture and optimization for your
selected applications, analyzes results, and produces reports based on your
specified output, all in a single workflow.

Gel Doc XR+ Imaging System

Gel Doc XR+ imaging system is a high-resolution gel documentation system that
allows fast, easy quantitation of gels and blots. Position your sample inside the
imager and follow the onscreen steps to run a protocol with only one click. You can
customize your applications within an existing protocol or create a new protocol
using many options presented by the Image Lab software.
Features include:

Detection of dynamic ranges that cover >3 orders of magnitude

Dynamic flat fielding specific to fluorescent and colorimetric applications

User Guide

| 17

1 |

Introduction

Smart, application-based protocol setup using Image Lab software, which

assists you by presenting appropriate filter and illumination sources for
each application

Ability to upgrade to the ChemiDoc XRS+ system

ChemiDoc XRS+ Imaging System

In addition to the gel documentation applications of Gel Doc XR+ system, the
ChemiDoc XRS+ imaging system offers sensitive chemiluminescent detection. The
system includes a supersensitive 16-bit CCD camera that is deeply cooled for the
long imaging times required for faint sample detection and for accurate
quantification of image data.
Features include:

Suitability for imaging applications that require excellent sensitivity

(chemiluminescent western blots)

Smart, application-based protocol setup using Image Lab software, which

assists you by presenting appropriate filter and illumination sources for
each application

Dynamic range >4 orders of magnitude

Flexibility to image chemiluminescent, fluorescent, and colorimetric

samples with dynamic flat fielding specific to each application

System Components
CCD Camera and Lenses
Either the Gel Doc XR+ or the ChemiDoc XRS+ camera is placed on top of a
lighttight enclosure (the universal hood) for capturing images. The camera comes
with a motorized zoom lens (MZL) that allows remote adjustment of the lens control
functions (zoom, focus, and iris).

18

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

System Components

A patent-pending software algorithm controls the MZL, giving the user automatic
image focus once an initial calibration is performed during system installation. See
the Technical Specifications table, included in the chapter which discusses your
camera, for complete specifications of each system.
A +1 diopter lens is factory installed to allow the entire sample stage to be visible.
This lens should always remain on the MZL assembly.

Universal Hood
The universal hood is designed to capture fluorescent and chemiluminescent
images without using a photographic darkroom. The enclosure has built-in white
light epi-illumination and UV transillumination. For easy sample loading, the UV
transilluminator is located in the drawer of the universal hood and can be accessed
from the front of the enclosure. When not imaging, the lights in the darkroom
enclosure turn off automatically.
The universal hood has touchpad buttons to perform various functions; however,
Image Lab software controls all of these functions remotely, removing any
requirement for manual control of the lens and lights. Running a protocol overrides
any touchpad input.

Image Lab Software

Each system ships with a full version of Image Lab software. In addition to
controlling the imager systems, image capture, and optimization, Image Lab
software can be used to annotate and document images, analyze molecular weights
(or base pairs, when imaging nucleic acid gels), and to determine accurate
quantitation and purity of samples.
You can print all or a subset of your data in a report. Alternatively, you can export
your data to other software, such as Microsoft Office programs, for further analysis
or presentation options. See Chapters 5 to 11 for detailed instructions on the
software.

User Guide |

19

1 |

Introduction

Emission Filters
The universal hood can hold two different emission filters for fluorescent
applications. No filter is required to image chemiluminescent samples.
A standard filter (catalog #170-8081) is used for colorimetric (white light)
applications and is included in the installation kit.

Optional Accessories
Bio-Rad Laboratories, Inc. offers a selection of optional filters and illumination
sources. See Appendix D, Accessories, for ordering information and a complete
listing of accessory filters, UV light sources, optional parts, and replacement parts.

Printer
For your convenience, Bio-Rad offers an optional USB printer for use with the
Gel Doc XR+ and ChemiDoc XRS+ systems: the Mitsubishi thermal printer, (catalog
#170-8089).

Conversion Screens
White Light Conversion Screen
The white light conversion screen (catalog #170-8001) is a phosphor screen that
produces white light transillumination when placed on top of the UV transilluminator.

White Light Transilluminator

An optional white light transilluminator (catalog #170-7950) can also be used with
either system. The white light transilluminator plugs inside the universal hood and is
controllable by the touchpad or the computer software.

XcitaBlue Conversion Screen

The optional XcitaBlue screen kit (catalog #170-8182) is a UV to blue light
conversion screen, which allows you to visualize DNA samples while protecting
them from UV damage.

20

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Gel Doc XR+ Imager

Refer to the Gel Doc XR+ Installation Guide found in your Gel Doc XR+ Installation
Kit for information about installing and calibrating your system. Refer to Chapter 4,
Setting Up the Instrument with Image Lab Software for information about installing
Image Lab software.
To recalibrate your system because you have acquired new accessories, refer to
Chapter 11, System Calibration.

Gel Doc XR+ Imager Workflow

Following are the basic steps to acquiring, analyzing, and archiving an image using
the Gel Doc XR+ system and Image Lab software:
1.

Select an existing protocol or customize a new one.

2.

Position the sample to be imaged.

3.

Run a selected protocol.

4.

View the displayed results.

5.

Optimize the analysis.

6.

Generate a report.

7.

Save or export the results.

User Guide

| 21

2 |

Gel Doc XR+ Imager

Gel Doc XR+ Applications

The Gel Doc XR+ system is capable of running a wide variety of protocols. Below
are examples of the detection reagents, listed by application, that can be used to
image your samples. Contact Bio-Rad technical support to determine whether your
application can be imaged on this instrument.
See Chapter 6, Acquiring Images, for information about designing protocols.

Nucleic Acid Gels

Ethidium bromide

SYBR Gold

Fast Blast

SYBR Green

GelGreen

SYBR Safe

GelRed

Protein Gels

Coomassie Blue

Oriole fluorescent gel stain

Coomassie Fluor Orange

Silver stain

Copper stain

SYPRO Ruby

Flamingo fluorescent gel

stain

Zinc stain

Krypton

Blots

22

Alexa Fluor 488

Fluorescein

Colorimetric reagent

Qdot 525

Cy2

Qdot 565

DyLight 488

Qdot 625

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Gel Doc XR+ Technical Specifications

Gel Doc XR+ Technical Specifications

Applications
Chemiluminescence

No

Fluorescence*

Yes

Colorimetry/densitometry

Yes

Gel documentation

Yes

Hardware Specifications
Maximum sample size

Maximum image area

Excitation source

Illumination control

Length: 28 cm

Width: 36 cm

Length: 19.4 cm

Width: 26 cm

Trans-UV and epi-white are standard (302 nm

included, with 254 and 365 nm available as
options).

Optional trans white, self-powered or conversion

screen.

Optional XcitaBlue UV/blue conversion screen.

3 modes:

Trans-UV

Trans white

Epi-white

Detector

CCD

Pixel size

4.65 x 4.65 (H x V in microns)

Cooling system

Not applicable

Camera cooling temperature

Not available

Filter holder

3 positions:

2 for emission filters

1 for using no filter

User Guide |

23

2 |

Gel Doc XR+ Imager

Emission filters

1 included (standard)

3 optional

Dynamic range

>3.0 orders of magnitude

Pixel density (gray levels)

4,096

Dynamic flat fielding

Application-specific, for all applications

Instrument size

Length: 36 cm

Width: 60 cm

Height: 96 cm

Instrument weight

32 kg

Operating Ranges
Operating voltage

110/115/230 V AC nominal

Operating temperature

1028C (21C recommended)

Operating humidity

<70% noncondensing

Automation Capabilities
Workflow automated selection

Application driven, user selected or recalled by a

protocol

Workflow automated execution

Controlled by a protocol via application-specific

setup for image area, illumination source, filter,
analysis, and reporting

Workflow reproducibility

100% repeatability via recallable protocols; from

image capture to quantitative analysis and reports

Autofocus (patent pending)

Precalibrated focus for any zoom setting

Image flat fielding (patent

pending)

Dynamic; precalibrated and optimized per

application

Autoexposure

2 user-defined modes (intense or faint bands)

* Using the optional XcitaBlue kit (catalog #170-8182) is highly recommended for SYBR Safe
DNA applications because the UV-to-blue conversion screen allows you to visualize DNA
samples while protecting them from UV damage.

24

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

3 ChemiDoc XRS+ Imager

Refer to the ChemiDoc XRS+ Installation Guide found in your Image Lab
upgrade for the ChemiDoc XRS+ system for information about assembling and
calibrating the ChemiDoc XRS+ system. Refer to Chapter 4, Setting Up the
Instrument with Image Lab Software for information about installing Image Lab
software.
To recalibrate your system because you have acquired new accessories, refer to
Chapter 11, System Calibration.

ChemiDoc XRS+ Imager Workflow

Following are the basic steps to acquiring, analyzing, and archiving an image using
the ChemiDoc XRS+ system and Image Lab software:
1.

Select an existing protocol or customize a new one.

2.

Position the sample to be imaged.

3.

Run the selected protocol.

4.

View the displayed results.

5.

Optimize the analysis.

6.

Generate a report.

7.

Save or export the results.

When running chemiluminescent blots, you can use a signal accumulation mode
(SAM), which takes a series of exposures within a set period of time. This enables
you to choose an image with optimal signal. Refer to Chapter 6, Acquiring Images
for more information.
User Guide

| 25

3 |

ChemiDoc XRS+ Imager

ChemiDoc XRS+ Applications

The ChemiDoc XRS+ system is capable of running protocols to image all of the
nucleic acid gels, protein gels, and blots listed in Gel Doc XR+ Applications on
page 22 plus the others added to this list. Those listed are examples of the
detection reagents that can be used to image your samples and is not considered a
complete list. Contact Bio-Rad technical support to determine whether your
application can be imaged on this instrument.
See Chapter 6, Acquiring Images, for information about designing protocols.

Nucleic Acid Gels

Ethidium bromide

SYBR Gold

Fast Blast stain

SYBR Green

GelGreen

SYBR Safe

GelRed

Protein Gels

Coomassie Blue

Oriole

Coomassie Fluor Orange

Silver stain

Copper stain

SYPRO Ruby

Flamingo

Zinc stain

Krypton

Blots

26

Alexa Fluor 488

Fluorescein

Chemiluminescent substrate

Qdot 525

Colorimetric substrate

Qdot 565

Cy2

Qdot 625

DyLight 488

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

ChemiDoc XRS+ Technical Specifications

ChemiDoc XRS+ Technical Specifications

Applications
Chemiluminescence

Yes

Fluorescence*

Yes

Colorimetry/densitometry

Yes

Gel documentation

Yes

Hardware Specifications
Maximum sample size

Maximum image area

Excitation source

Illumination control

Length: 28 cm

Width: 36 cm

Length: 26 cm

Width: 35 cm

Trans-UV and epi-white are standard (302 nm included, with

254 and 365 nm available as options)

Optional trans white, self-powered or conversion screen

Optional XcitaBlue UV/blue conversion screen

5 modes:

Trans-UV (standard)

Epi-white (standard)

No illumination for chemiluminescence (standard)

Trans white (optional)

XcitaBlue (optional)

Detector

Supercooled CCD

Pixel size

6.45 x 6.45 (H x V in microns)

Cooling system

Peltier cooled

Camera cooling
temperature

30C controlled

Filter selector

3 positions:

2 for filters

1 without filter (for chemiluminescence)

User Guide |

27

3 |

ChemiDoc XRS+ Imager

Emission filters

1 included (standard)

3 optional

Dynamic range

>4.0 orders of magnitude

Pixel density (gray levels)

65,535

Dynamic flat fielding

Application-specific, for all applications

Instrument size

Length: 36 cm

Width: 60 cm

Height: 96 cm

Instrument weight

32 kg

Operating Ranges
Operating voltage

110/115/230 V AC nominal

Operating temperature

1028C (21C recommended)

Operating humidity

<70% noncondensing

Automation Capabilities
Workflow automated
selection

Application driven, user selected or recalled by a protocol

Workflow automated
execution

Controlled by a protocol via application-specific setup for image

area, illumination source, filter, analysis, and reporting

Workflow reproducibility

100% repeatability via recallable protocols; from image capture

to quantitative analysis and reports

Autofocus (patent
pending)

Precalibrated focus for any zoom setting or sample height

Image flat fielding (patent

pending)

Dynamic; precalibrated and optimized per application

Autoexposure

2 user-defined modes (intense or faint bands)

* Using the optional XcitaBlue Conversion Screen kit (catalog #170-8182) is highly recommended if
performing SYBR Safe DNA applications, because the UV-to-blue conversion screen allows you to
visualize DNA samples while protecting against UV damage.

28

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Setting Up the Instrument

with Image Lab Software

System Requirements
Image Lab software runs on Microsoft Windows XP Professional, Microsoft
Windows 7, and Mac OS X. Images scanned at high resolution can be quite large.
The amount of memory required for using the program is determined by the size of
the images you scan and analyze.
For this reason, we recommend that you archive images on a network file server or
on removable storage media. Bio-Rad can also provide an appropriate computer to
use with this system. Contact your local Bio-Rad representative for more details.

Image Lab Security Edition

Note: The system requirements for Image Lab Security Edition are the same as
those for the Standard Edition. The software must be installed on a computer
running the Windows XP Professional or Windows 7 operating system to take
advantage of the secure mode features.

Computer Specifications
Specifications

Minimum

Recommended

Operating system

Windows XP SP3

Windows XP SP3 Professional

Windows 7, 32- and 64-bit

Windows 7 Professional, 64-bit

Mac OS X 10.6

Mac OS X 10.6

Pentium 4 or equivalent (Windows)

at 2.0 GHz

Intel Core 2 Duo 2.0 GHz or

higher

Processor

User Guide

| 29

4 |

Setting Up the Instrument with Image Lab Software

Specifications

Minimum

Recommended

Hard disk space

20 GB

>100 GB

Memory (RAM)

1,024 MB

>1,024 MB

Ports for connecting

instrument

1 free USB 2.0 port

1 free USB 2.0 port

Other software

Microsoft Excel 2003 or later

(Windows)

Microsoft Excel 2003 or later

(Windows)

Office 2008 or iWork software

(Mac)

Office 2008 or iWork software

(Mac)

(optional)

Installing Image Lab Software

The process of installing Image Lab differs depending on your operating system.
This section explains how to install Image Lab software on a Windows PC and on a
Mac.
Note: During the Windows installation process, you are prompted to install
Image Lab Standard Edition or Security Edition. In order to enable secure
mode, you must have Image Lab Administrator role privileges. Bio-Rad
recommends that you create the required groups and assign the Image Lab
roles to users within those groups before installing Image Lab software.
See User Names, Groups, and Roles on page 180 for additional information about
the required Image Lab roles, groups, and users.
See Setting Up Users and Groups on page 195 for information about setting up
groups, user names, and passwords.

To install Image Lab on a Windows PC

1.

30

Insert the Image Lab software CD in your CD-ROM drive.

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Installing Image Lab Software

The Image Lab installer automatically launches.

2.

On the Welcome screen, click Next.

3.

Accept the license agreement and click Next.

4.

On the Edition Selection screen, choose the edition to install.

User Guide |

31

4 |

Setting Up the Instrument with Image Lab Software

Note the following.

If you are licensed to install the Security Edition but choose to install the
Standard Edition, you will need to uninstall the Standard Edition and install
the Security Edition before you can use it.

If you are licensed to install the Security Edition and choose to install it,
only a user assigned the Image Lab Administrator role (or group) privileges
can enable and disable secure mode.
Note: A user assigned the Image Lab Administrator role will not
necessarily also be the network or IT administrator. You must be
assigned the correct role within Image Lab to enable or disable secure
mode.
For more information, see Setting Up Image Lab Security Edition on
page 39.

5.

32

If you are not licensed to install the Security Edition but choose to install it,
you will be prompted for a license key when you start Image Lab.

Click Next.

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Installing Image Lab Software

6.

A screen appears requesting your license code.

Note: Your 18-digit license code can be found in the Image Lab product
folder pocket.

Enter the 18digit code in the three text boxes. The software verifies the code.
Tip: If you do not know or do not have access to the code, contact your
Bio-Rad customer service representative.
Note: Until you provide a license code, Image Lab will function only in
standard mode.
7.

Click Next.

8.

On the Install Location screen, accept the default location or click Change and
browse to another folder.

9.

Click Next.

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10. On the Ready to Install the Program screen, click Install.

The wizard installs Image Lab.

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Installing Image Lab Software

11. When the installation is complete, you are prompted to display the Release
Notes and/or the Windows Installer log.

The following screen shot is an example of the Windows Installer log.

12. Select or clear the checkboxes in the Install Wizard Completed dialog.
13. Click Finish to exit the wizard.

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Setting Up the Instrument with Image Lab Software

The Image Lab icon appears on your desktop. Follow the instructions in the next
section to connect your system.

To install Image Lab on a Mac

1.

Insert the Image Lab software CD in your CD-ROM drive.

2.

Double-click the CD icon on your desktop to see the folder contents.

3.

Double-click the file Image Lab.dmg.

4.

Drag the Image Lab application icon into the Applications folder.

Installing the Drivers on a Windows XP Computer

After the instrument is connected and the power is turned on, the PC automatically
recognizes the instrument, and the hardware installation wizard starts, prompting
you to install the drivers.
The initial Welcome dialog box appears.

Installing the drivers on a Windows XP computer

1.

36

On the Welcome dialog box, click No, not this time, and then click Next.

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Installing the Drivers on a Windows XP Computer

The dialog box listing the driver that will be installed appears.

2.

By default, the radio button for Install the software automatically

(Recommended) is selected.
Keep the default selection and click Next.
The following warning dialog box appears.

3.

Click Continue Anyway.

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Setting Up the Instrument with Image Lab Software

Windows installs the driver.

When the installation is complete, the following dialog box appears.

4.

Click Finish.
A message appears briefly: Your new hardware is ready to use.

Note: Windows XP might prompt you to go through the driver installation a

second time. If this happens, repeat this section until you receive the message
Your new hardware is ready to use.

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Installing the Drivers on a Windows 7 Computer

Installing the Drivers on a Windows 7 Computer

If you are running Windows 7, the device driver is installed during the Image Lab
installation process. After successful installation, you see a message similar to the
following.

Note: During the installation process you might see a warning similar to the
following. You can ignore this warning because it appears even when the driver
has been installed correctly.

Setting Up Image Lab Security Edition

Activating and Deactivating Security Edition

Before you activate or deactivate Image Lab Security Edition, close any
open document files.

After you change the active status of the Security Edition, you must restart
Image Lab.

To activate the Security Edition via the Internet

1.

Double-click the Image Lab icon on your desktop to open Image Lab.

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Setting Up the Instrument with Image Lab Software

The License Code field is populated with the 18-digit license code that you
entered when you installed Image Lab.
The Security Edition Activation dialog box appears.

40

2.

Select Activate Via Internet.

3.

Click Activate.

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Setting Up Image Lab Security Edition

Within about 30 seconds you will receive a confirmation that your Image Lab
Security Edition has been activated.

To activate the Security Edition via email

1.

Double-click the Image Lab icon on your desktop to open Image Lab.
The Security Edition Activation dialog box appears.
The License Code field is populated with the 18-digit license code that you
entered when you installed Image Lab.

2.

Select Activate Via Create Activation Email.

3.

Click Create Email.

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Setting Up the Instrument with Image Lab Software

A Save File window appears with the File name field already filled in. Do not
change this file name.

4.

Click Browse Folders to choose a location for the file, and click Save.

5.

In your email program, create an email addressed to

[email protected], with the subject line: Request to Activate
Image Lab software Security Edition.

6.

Attach the ActivationEmail.txt file to the email and click Send.

The Bio-Rad Technical Support Department will process your request and reply
with an email containing an attachment with your activation code.

42

7.

When you receive your reply email, open it and save the attached
UnlockCode.txt file to the folder in which you saved the ActivationEmail.txt file.

8.

On the Security menu, click Activate Security Edition to display the Security
Edition Activation dialog box.

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Setting Up Image Lab Security Edition

9.

Select Activate Via Receive Activation Email.

10. Click Receive Email.

An Open File window appears.
11. Navigate to the location where you saved the UnlockCode.txt file, select the
file, and click Open at the bottom of the window.

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The Security Edition activation complete window appears.

12. Click OK to close the window.

Deactivate Image Lab Security Edition

Image Lab can be installed on more than one computer. Bio-Rad Laboratories
recommends that you install the software on only one desktop computer and one
laptop. To load the Image Lab Security Edition on a second computer, the software
must be deactivated on the first computer before it can be activated on another. You
can deactivate the Security Edition automatically via the Internet, or you can
deactivate it manually by sending a deactivation email.

To deactivate the Security Edition automatically via the Internet

44

1.

On the Security menu, click Deactivate Security Edition to display the

Deactivate Security Edition dialog box.

2.

Select Deactivate Via Internet.

3.

Click Deactivate.

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Setting Up Image Lab Security Edition

The system sends a message to the Bio-Rad Technical Support Department, a

window stating that your deactivation was successful appears, and Image Lab
Security Edition is immediately deactivated.

To deactivate the Security Edition via email

1.

From the Security menu, click Deactivate Security Edition to display the
Deactivate Security Edition dialog box.

2.

Select Deactivate Via Create Deactivation Email.

3.

Click Deactivate.

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Setting Up the Instrument with Image Lab Software

A Save File window appears.

4.

Navigate to the folder in which you want to save the deactivation email and
click Save.

5.

Create an email addressed to [email protected], with the

subject line: Request to Deactivate Image Lab software Security Edition.

6.

Attach the DeactivationEmail.txt file to the email and click Send in your email
program.
The Bio-Rad Technical Support Department processes your request and
deactivates Image Lab Security Edition.

Enabling and Disabling Image Lab Secure Mode

You must have Image Lab Administrator role privileges to switch between
Image Lab secure mode and standard mode.

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Setting Up Image Lab Security Edition

To enable secure mode

1.

From the Security menu, click Security Preferences to display the Security
Preferences dialog box.

2.

Select Enable secure mode.

3.

Click OK to display the Enable Secure Mode authentication dialog box.

4.

Enter your Image Lab Administrator user name and password.

5.

If you are set up on a Windows network server, type the name of the Windows
domain in the Domain field.
Note: The default is the domain on which the current Windows user is
located, and this name will appear in the field

6.

Click OK to save your changes.

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Setting Up the Instrument with Image Lab Software

A message appears stating that you must restart Image Lab for the new secure
mode enable setting to take effect.

7.

Click OK.

The application exits.

Important: For full details on why and how to set your security preferences, see
Setting Security Preferences on page 50.

To disable secure mode

48

1.

From the Security menu, click Security Preferences to display the Security
Preferences dialog box.

2.

Clear the Enable secure mode checkbox.

3.

Click OK.

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Setting Up Image Lab Security Edition

The Admin Authentication dialog appears.

4.

Enter your Image Lab Administrator user name and password.

5.

If you are set up on a Windows network server, type the name of the Windows
domain in the Domain field.
Note: The default is the domain on which the current Windows user is
located, and this name will appear in the field.

6.

Click OK to save your changes.

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A message appears stating that you must restart Image Lab for the new
security settings to take effect.

7.

Click OK.

The application exits.

Setting Security Preferences

Here are three useful definitions used in these instructions.

Network domain a remote domain-controlling computer or system

which ensures that only authorized users with valid credentials can access
and run Image Lab

Local domain (or local computer) the computer on which Image Lab is
running, and which ensures that only authorized users with valid user
credentials can access and run Image Lab

Credentials the valid user name and password that allows or prohibits
specific user actions

There are three possible combinations of settings in the Security Preferences dialog
box.

To choose settings in the Security Preferences dialog box

1.

50

To set preferences so that only users who are set up on a network domain can
use Image Lab:

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Setting Up Image Lab Security Edition

In the Domain to be used in authentication field, enter the name of your

network domain. See To find the name of your network domain on
page 198 for instructions on how to determine this name.
Note: The default domain name that will appear in this field is the
domain on which the current user is logged in.

2.

Do not select the Use local groups for establishing user security levels
checkbox.

To set preferences so that only users who are domain users and who are also
valid members of specific local groups can run Image Lab:

In the Domain to be used in authentication field, enter the name of your

network domain. See To find the name of your network domain on
page 198 for instructions on how to determine this name.
Note: The default domain name that will appear in this field is the
domain on which the current user is logged in.

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Setting Up the Instrument with Image Lab Software

3.

Select the Use local groups for establishing user security levels checkbox.

To set preferences so that only local users can run Image Lab:

In the Domain used in authentication field, enter your local computer name.
See To find the name of your local domain on page 197 for instructions on
how to determine this name.
Note: The default domain name that will appear in this field is the
domain on which the current user is logged in.

52

The Use local groups for establishing user security levels checkbox is
grayed out (not accessible).

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Setting Up Image Lab Security Edition

Renaming Security Groups

To rename any of the four default Security Groups
1.

From the main menu, select Security > Rename Security Groups.

Note: This menu option is visible only if the person logged on to the local
computer is logged on as a member of the Windows Administrators group.

2.

Click in any of the four Group Name fields.

3.

Enter a new name.

4.

Click OK to save your changes.

Note: The new user group name must comply with standard Windows Local
Users and Groups user names rules.
For more information on setting up security groups, see Setting Up Users and
Groups on page 195.

Using Groups on a Local Domain

If you choose not to create or use groups on the network domain, set up local
groups. Add the authorized users to the groups on the local domain. In the Security
Preferences dialog box, select Use local groups for establishing user security levels.
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For information about setting up users and groups for Image Lab Security Edition,
see Appendix C, Setting Up Users and Groups on page 195.

Changing Security Preferences

Changing the domain that is used to authenticate users is a two-step process. You
first authenticate on the first domain, then authenticate on the second domain. This
change in domains can be performed in either of two ways. It can be performed by
one individual assigned the Image Lab Administrator role on both domains, or it can
be performed by two individuals, one with the administrator role on the first domain,
and the other with the administrator role on the second domain. See User
Authentication and Group Membership on page 195 for more information about
using this dialog box.

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5 Image Lab Software

Overview
Image Lab image acquisition and analysis software works with the
Gel Doc XR+ and ChemiDoc XRS+ imaging systems to create a reproducible,
automated, and time-saving workflow for imaging and analyzing gels.
In Image Lab, a protocol is any combination of imaging, analysis, and report settings
that has been saved to run as a single workflow. Researchers can run one protocol
repeatedly or easily design a wide range of protocols.
With Image Lab you can view analyzed data, edit the analysis, and produce
customized reports that show precisely the settings applied in order to ensure
repeatable results.
Image Lab generates two types of files:

Protocol files describe the parameters for imaging and analyzing your gel
images.

Image files contain the imaged gel, annotations, and analysis performed on
the gel. An imaged gel, run according to a protocol file, generates an image
file.

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Table 2 lists the extensions and icons for the type of files that Image Lab generates.
Table 2. Image Lab file extensions and icons
File type

56

File Extension

Icon

Unsigned

Signed

Protocols

.ptl

.sptl

Images

.scn

.sscn

Unsigned

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Signed

Interface Overview

Interface Overview
The following illustration shows the Image Lab main window. This section explains
the main software elements.

Main Window
Image Lab displays a single main window. All image and protocol dialog boxes that
present choices open in the workspace, which is the gray area of the main window.
If many screens are open in the workspace, you can make one active by clicking the
title bar at the top of the selected screen. A list of open protocols and image files
also appears in the main window menu. Select one to make it active.
You can view complete analyses for images or protocols one at a time or compare
image results by arranging screens in the workspace.

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Main Toolbar
Many Image Lab tools can be selected by clicking toolbar buttons. The Screenshot
tool enables you to send a screen capture of your image to the clipboard or to save
it as a file. You can view demonstrations of various functions by clicking Tutorials.
The unlimited Undo and Redo buttons enable you to correct missteps easily.

File management

View results data

Results Data
Results data associated with gel images can be viewed as an analysis table, a lane
profile, a standard curve, or in a report. Different tools for viewing the results data
are easily accessible from the main toolbar. These tools are described in Chapter 7,
Viewing Images.
The views display the analysis for the selected image. All of the views can be
displayed at the same time. See Displaying Data on page 104 for details.

Display Toolbox
The display toolbox at the top of every image enables you to display images in the
most useful ways. See Chapter 7, Viewing Images for a description of each option.

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Interface Overview

Start Page
The Start Page guides you through creating, opening, and viewing protocols and
images.

Analysis Tool Box

The Auto-Analysis

button quickly analyzes images. The remaining

tools customize the analyzed data.

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Image Lab Software Overview

An image file must be selected to make analysis tools available.

Image Tools enables you to flip, rotate, and crop images and to transform the
image files.
Lane and Bands enables you to resize, adjust, and bend lanes and to detect,
adjust, add, or delete bands.
Normalization enables you to normalize volume data in multichannel images, so
you can correct for sample loading errors in your gels. For more on multichannel
images, see the System with Image Lab Software User Guide.

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Interface Overview

MW Analysis Tools (Molecular Weight Analysis) enables you to choose standard

samples, assign standard lanes, and choose a regression method.
Quantity Tools enables you to automatically quantify bands, using either relative or
absolute values.
Annotation Tools enables you to add formatted text and arrows to any area of a
gel.
Volume Tools enables you to manually quantify an object inside a boundary that
you define.
These tools are described in Analysis Tool Box Tools on page 117.

Status Bar
The status bar at the bottom of the main window shows the imager in use and the X
and Y values for the cursor position on the image file.
Note: The status bar also displays the intensity (Int) values for the image at the
cursor position. The maximum data range is 065,535. However, the actual
range varies depending on the values contained within each image.

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Menu Commands
The following section describes all menu commands in the File, Edit, View, Window,
and Help menus. Many commands are also available on the toolbar or the Start
Page.

File Menu Commands

New Protocol enables you to create a new protocol that contains the necessary
steps and choices. See Chapter 6, Acquiring Images for detailed instructions.
Protocols can also be altered and stored for reuse.
Open browses the file system to retrieve a previously saved protocol file or image
file.
Recent Images enables you to open a recent image file.
Recent Protocols enables you to open a recent protocol.
Save enables you to save a protocol or image file after it is named.
Save As enables you to name and store a protocol or image. Protocols are stored
with a .ptl or .sptl extension. Image files are stored with an .scn or .sscn extension.
Close closes the active window.
Close All closes all the windows.
Export enables you to export gel images or analysis tables with the following
options:

Export for Publication exports a displayed image to a file. You can

select from .bmp, .png, .jpg, and .tif formats. The gel displays with any
lanes, bands, and annotations that appear on the screen. See Exporting
Gel Images for Publication on page 158 for more information.

Export for Analysis creates a .tif file that retains all gel image data.
Analysis data are not included. Use this option to analyze the image in
other software such as Quantity One,FPQuest, or InfoQuestFP. See
Exporting Gel Images for Publication on page 158 for more information.

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Interface Overview

Export for PulseNet reduces the image to an 8-bit .tif file. Resolution is
limited and file size is restricted to 300 dots per inch (dpi).

Lane and Band Table to Excel exports your lane and band table data
to an Excel (or Numbers on a Mac) spreadsheet.
Note: Excel or Numbers must be installed on your computer.

Lane and Band Table to File exports as a comma-separated values

(CSV) file so that your lane and band table can be opened in a database
application.

Volume Table to Excel exports your volume table data to an Excel (or
Numbers on a Mac) spreadsheet.
Note: Excel or Numbers must be installed on your computer.

Volume Table to File exports as a CSV file so your volume table can be
opened in a database application. See Exporting Volume Tables to File on
page 161 for detailed information about exporting files.
See Chapter 10, Exporting Results for more information about exporting files.

Image Info displays information about individual gel and blot images, such as
acquisition date and data range, and image capture detail, such as exposure time
and illumination source used. Click the Image Details, Analysis Settings, and Notes
tabs to display these properties. See Image Info on page 104 for more information.
Page Setup contains print controls such as orientation (landscape or portrait),
margins, printer used, and paper size.
Print displays a print preview of the gel and the header information, which includes
the filename of the image, the users name, and the date and time it was printed.
The usual Windows Print screen is available as well, enabling you to select a printer
and the number of copies to print.
Exit closes Image Lab (after prompting you to save changes to your protocols or
images).

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Edit Menu Commands

Undo undoes the last action.
Redo restores the last action after an Undo.
Screenshot enables you to take a screen shot of the Lane Profile Window, the
Standard Curve Window, or the default choice, Current Image View. The screen shot
can include the name of the image, and it can be placed on the clipboard or saved
in a file.
Default Imager enables users who own two or more imagers to switch between
them.
Instrument Setup displays information about the instrument, including its name,
serial number, camera serial number, illumination options, and last calibration. If you
add accessories to the instrument, you can reset the system calibration in this
dialog box.
Report Settings enables you to configure reports. This dialog box has three tabs.
All of the checkboxes are selected by default. Clear the boxes to exclude
information from reports. Your selections apply to all reports until you change them.

The General tab has options for excluding or reporting information about
your gel image.

The Lane and Band Table tab enables the researcher to choose whether to
include all lanes or selected lanes, with appropriate identifiers. Lane
profiles can also be included.

The Volume Table tab enables the researcher to choose appropriate

identifiers for the volume table and provides the option of excluding the
table from reports.

Preferences enables you to set naming and color preferences for your image files.
This dialog box has two tabs.

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Interface Overview

The Protocol tab shows presets for naming image files. You can choose to
include a designated Prefix, User Name, Date, and/or Time in the name of
your image files.

The Colors tab enables you to choose colors for the graphic elements in
your gels, such as Lane Frame, Lane, Band, Band Attribute, and MW
Legend. This functionality ensures that these elements are visible,
regardless of the color of the gels.

View Menu Commands

Image Overview displays the gel image with a red rectangle outlining the area
visible in the larger main window. This is useful when you zoom in to a small section
of an image.
Image Transform displays a histogram that enables you to adjust the light and dark
values of a gel image. This adjustment does not change your data, only the way the
data display on your monitor.
Operations History displays the sequence of actions performed by both the user
and the software.
View System Log displays events related to running Image Lab software, including
enabling or disabling secure mode, and the users who log on to or log off of the
software.
View (Instrument) Log displays events related to the instrument, including
calibrating the instrument and the success or failure of the calibration. This log file is
visible only if Image Lab is connected to an instrument.
View (Document) Log displays events related to creating and modifying secure
protocol and image files. This log file is created only when Security Edition is
enabled.

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Security Menu Commands

Note: This menu is only visible in Image Lab Security Edition. For more
information about Security Edition, see Chapter 13, Using the Security Edition.
Security Preferences allows the Image Lab Administrator to enable and disable
secure mode. In this dialog box, the administrator chooses the domain to be used
for authentication and whether to use local groups for security levels.
Rename Security Groups allows a person logged on as a member the Windows
Administrators group to change any of the four default Image Lab security group
names (TDS_Administrator, TDS_User, TDS_Tech, and TDS_Guest).
Note: This menu option is visible only if the person logged on to the local
computer is logged on as a member of the Windows Administrators group.
Only users who are logged on as a member of that group maintain the authority
to change any of the four default Image Lab security group names. Any
changes made to these Security Group names must match the names your
Windows system administrator has given those groups.
Sign Document enables users to sign images and protocols. Users enter their user
name and password and provide a reason for signing. When the document is
signed, the reason is saved in the System Log file.

Window Menu Commands

The Window controls enable you to show and hide multiple open image files in your
workspace. A list of all currently open images and protocols appears in this menu.
Tile aligns all open image files so they are visible at the same time.
Tile Horizontal aligns all open image files from top to bottom.
Tile Vertical aligns all open image files from left to right.
Cascade stacks all open image files and protocols with overlapping title bars, so
each one can be easily chosen for viewing.
Imitate Zoom changes the zoom setting of all open images to the same zoom
setting as the current image file.

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Interface Overview

Imitate Transform changes the brightness and contrast of all open images to the
same transform settings as the current image file.
Next cycles through all open image files from oldest to newest.
Previous cycles through all open image files from newest to oldest.

Help Menu Commands

Image Lab Help displays the help system.
User Guide displays the instruction manual in .pdf form.
About displays Image Lab software version and release date.

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Image Lab Software Overview

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Acquiring Images

Image Lab software runs specific applications with repeatable workflows using
configurable protocols that have a wide variety of settings. These protocols can be
retrieved, revised, and reused.
In Image Lab, a protocol is any combination of settings for imaging, analyzing, and
reporting that runs as a single workflow.
A protocol allows you to choose one application for acquisition of a single image
from a sample, with the exception of signal accumulation mode (SAM) for
chemiluminescence.

To create a new protocol

Click New in the Protocol box on the Start Page, or choose New Protocol on
the toolbar or menu bar.

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Acquiring Images

The Protocol Setup Window

The left pane displays headings. Under the headings are numbered protocol steps.
You can enable or disable a step by selecting or clearing its checkbox. When you
select a step, the right pane of the window displays the detailed settings for that
step.

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Creating a Protocol

You can review protocol settings by selecting Protocol Summary, which lists the
settings for each step in the right pane of the Protocol Setup window.

Creating a Protocol
There are three categories of settings when setting up a protocol:

Acquisition Settings settings to acquire the image

Analyze Image settings to detect lanes and bands and to analyze the
molecular weight

Generate Output settings to generate the output

The three categories are listed in the left pane of the Protocol Setup window.
Numbered steps in each category appear under these headings. To select an option
under a protocol step, select the accompanying checkbox. Options for that step
appear in the right pane of the window. To disable any step, clear its checkbox.

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Setting Up a New Protocol

The following steps set up (or create) a protocol:

72

Step 1. Gel Imaging for Single-Channel Protocols

Step 2. Detect Lanes and Bands

Step 3. Analyze Molecular Weight

Step 4. Specify Output

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Creating a Protocol

Step 1. Gel Imaging for Single-Channel Protocols

To take an image of the gel or blot, you need to configure the acquisition settings for
the protocol.

To configure the acquisition settings

1.

In the right pane, click Select and choose an application from the menu. The
detection reagents appear in submenus under each application type.

When you choose an application and detection reagent, any required filter or
illumination source displays in the Protocol Setup window.

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Choose Custom to save and run an existing application with a new name or to
create and run a new application. Previously saved custom applications display
here. To create a custom application, see Setting Up a Custom Application on
page 91.
Tip: For a list of applications with all required detection reagents, light
sources, and any conversion screens or filters noted, see Application
Tables on page 94.
Note: If you select the Stain Free option, you may select the gel activation
time. See Appendix E, Using Bio-Rads Stain-Free Technology, on
page 223 for more information.
2.

In Imaging Area, select from the list of Bio-Rad gels or enter the image area
dimensions. The red box represents the imaging area for the selected gel, and
the gray rectangle represents the imager sample stage.

3.

In Image Exposure, select from one of the following options:

74

Auto Exposure this setting estimates an optimal exposure time and

ensures the best use of the dynamic range. Intense Bands optimizes for all
bands and Faint Bands makes faint bands more visible but might
overexpose more prominent bands.

Intense Bands optimizes exposure for all bands

Faint Bands a longer exposure time is used making faint bands

more visible, but more prominent bands might be overexposed

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Creating a Protocol

Manual Exposure use this setting to manually override automated

imaging. Exposure time can range from 0.001 to 7,200 seconds with the
ChemiDoc MP imager. Exposure time can range from 0.001 to 99 seconds
using the Gel Doc XR+ system and from 0.001 to 7,200 seconds for the
ChemiDoc XRS+ system. After imaging a gel with automatic exposure
optimization, the display provides a reference point from which to set your
manual exposure time.

Note: You can view the exposure time of the image in the Image Info
window (see Image Info on page 104).

Signal Accumulation Mode if you are running a chemiluminescence

application on your system, you can also use signal accumulation mode
(SAM).

SAM allows you to get a cumulative sequence of exposures from a

chemiluminescent sample to increase the likelihood that you will get the
correct image.

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Rather than manually acquiring a series of independent images with

different imaging times, SAM presents a cumulative series of images with
progressively greater signal in each image. SAM is a partially automated
method for image acquisition because it requires that you estimate the
shortest and longest times expected to generate an image with the
appropriate signal intensity. You then decide how many total images to
acquire in this time frame.
Note: For any given exposure time, you will get more background
noise with a SAM acquisition than you would with a single image taken
at the specified exposure time.
For example, if the expected minimum time to image the sample is 1
minute and the maximum is 5 minutes, these values are entered (in
seconds) in the setup window. If you enter 5 in the Total number of images
field (as shown below), three images will be acquired between the first and
last images.
To use SAM, click Signal Accumulation Mode and select Setup to display
the Signal Accumulation Setup dialog box.

In this example, the bar in the Signal Accumulation Setup dialog box
shows that images will be acquired at 1 minute intervals, beginning at
1 minute and ending at 5 minutes. The second 1 minute image is added to
the first 1 minute image, resulting in a 2 minute total integration time image.
The third 1 minute image is added to the previous image, and so on, until
the last image is presented.
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Creating a Protocol

Although SAM is useful for determining the optimum imaging time for a
chemiluminescent sample, it results in data that are not as accurate as
data from a single image. Signal that is near the intensity of background
noise becomes increasingly masked as the number of cumulative images
increases. To identify extremely faint signals in an image, reacquire it as a
single image, using the time the SAM tool found to be appropriate.
4.

In Display Options, you can set:

Highlight saturated pixels select this checkbox to see any saturated

pixels in red. This shows how much of the gel image is saturated. This
option can be changed later by selecting View > Image Transform.

Image color select color choice to display the sample image. Viewing
the image with a different color scheme can make it easier to see all of the
elements. See Image Colors on page 102 to view the color choices dialog
box.

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Step 2. Detect Lanes and Bands

To analyze the gel or blot, Image Lab must detect lanes and bands on the image.
Lanes are detected automatically, and then the background is subtracted
automatically. Refer to Using the All Lanes and Single Lane Tools on page 122 for
details. Customize band detection with the following options.

To configure the lane and band detection settings

78

1.

Select the Lane and Band Detection checkbox in the left pane of the Protocol
Setup window.

2.

In the right pane, select one of the following lane and band detection options:

Low Band Detection Sensitivity this option sets detection at a low

level (25) for images with more prominent bands. Faint bands are not
detected with this setting.

High Band Detection Sensitivity this option sets detection at a higher

level (75) for images that are fainter. Extraneous bands can be removed
later, using the Band tools in the Analysis Tool Box. See Lane and Bands
Tools on page 121.

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Creating a Protocol

Custom select a numeric value from 1 to 100 to choose the best

detection sensitivity for your sample.

Step 3. Analyze Molecular Weight

Determining molecular weight depends on selecting the proper protein standards.

Many protein standards are available from Bio-Rad and many different DNA
standards are also available. See Appendix D, Accessories for all Bio-Rad standards
and their catalog numbers.

To specify how molecular weight is analyzed

1.

Select the Analyze Molecular Weight checkbox in the left pane of the Protocol
Setup window.
The software calculates the molecular weight for each band based on the
specified standard.

2.

To estimate the size of the molecules in the bands of your gel, enter the
standard you are using.
Alternatively, you can choose from available standards:

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3.

80

a.

Click Change to display the Manage Standards dialog box.

b.

Use the Show dropdown list to show all, only the protein, or only the DNA
standards.

c.

Choose the standard that you need and click OK to exit the dialog box.

To create a new standard, click New in the Manage Standards dialog box and
complete the fields in the Edit Standard dialog.

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Creating a Protocol

a.

Enter a name for the new standard.

b.

Choose a standard type: protein or nucleic acid

A protein standard can be expressed in either isoelectric point (pI) or
molecular weight (kDa) units. A nucleic acid standard can be expressed in
base pairs (bp), kilo base pairs (kb), or mega base pairs (Mb) units.

c.

Click Add to display the Edit dialog box.

Note: At least two values must be added to create a new standard.

4.

d.

Enter the value and description of your new standard and click OK to
return to the Edit Standard dialog box.

e.

Click Add to display the Edit dialog box again, and enter a second value
and description for your new standard.

f.

Click OK to save your changes and close the Edit dialog box.

g.

Click OK in the Edit Standard dialog box to save your new standard.

h.

Click OK in the Manage Standards dialog box to save your changes.

Choose the lane(s) that contain your standards by typing lane numbers or the
words First and Last in the Standard Lanes field. The format is xx, xx, xx, and
so on, where xx is the lane number. For example, if you run an 18-well gel and
want your standards in lanes 1, 10, and 18, enter First, 10, Last.
Note: Lane detection works best when standards are placed in the first
and last lanes. For nucleic acid samples, use this step to determine the
size of the bands in base pairs.
For more information, see Molecular Weight (MW) Analysis Tools on page 131.

5.

Select the appropriate regression method for the gel type:

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Gradient gels the linear (semilog) regression method works well for
these gels because the mobility of the bands is linear to the log of their
molecular weight. As an alternative, the point-to-point (semilog) method
can be used if the R2 value is not sufficient.

Fixed percentage gels these gels have a nonlinear relationship

between the gels mobility and their molecular weight. For these gels,
choose the logistic or cubic spline regression method.

For more information about regression methods, see Regression Methods on

page 93.

Step 4. Specify Output

To specify the output of the protocol

1.

Select Specify Output in the left pane of the Protocol Setup window to display
output options.

2.

In the right pane, you can choose to

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Automatically print the image

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Creating a Protocol

Automatically print a report

Display the report

Image Lab prints to the default printer unless you select otherwise. For
information about customizing reporting options, see Report on page 151.

Review Protocol Settings

Click Protocol Summary in the left pane of any Protocol Setup window to see a
quick review of all protocol settings.

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Creating a Multichannel Image from Single Images

You can create a multichannel image from existing single images or from single
channels in other multichannel images. Only images with the same aspect ratio can
be combined in a multichannel image. These images are not linked to one another.
When you make a change to one image, it is not propagated to the other images.

To create a multichannel image from single images

1.

Open the single images from which you want to create a multichannel image.

2.

In the File menu, click Create Multichannel Image. The list of open images
appears in the left pane of the Create Multichannel Image dialog box.

3.

Drag each image into one of the channel boxes in the right pane. After you
select the first file, the Available Open Images list displays only files with the
same aspect ratio.

4.

(Optional) Specify a color for each channel in the accompanying dropdown

lists. The resulting multichannel image appears in the Image Preview section of
the dialog box.

5.

Click OK to save the multichannel image.

To replace a channel in a multichannel image

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1.

Open the multichannel image and the new image you want to use.

2.

In the File menu, select Create Multichannel Image. The open image files are
listed in the Compatible Open Images list of the Create Multichannel Image
dialog box.

3.

Drag the images you want to keep from the Available Open Images list (left
pane) into the channels in the New Multichannel Image pane (right pane).

4.

Drag the new image you want to use into one of the available channel boxes.

5.

Click OK to save the new multichannel image.

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Editing a Saved Protocol

Editing a Saved Protocol

You can change the protocol settings and rename the protocol using Image Lab
tools.

To edit a saved protocol

1.

To open a saved protocol, you can:

Click Open on the Start Page

Click File > Open on the menu bar

You can also choose a recently used protocol from the lists on the Start Page.

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You see the same set of menus and choices described in Creating a Protocol
on page 71.

86

2.

To update an existing protocol, edit and then save your changes without
renaming the protocol.

3.

To create a new protocol:

a.

Edit the protocol.

b.

Choose File > Save As and type a different name.

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Positioning the Gel

Positioning the Gel

To position a gel
1.

Click the yellow Position Your Gel button in the Protocol Setup window.

2.

Place a gel on the imager stage and view the gel in Image Lab.

3.

Use the slider below the image to zoom the image into place. You can also
move the gel manually until it is centered properly on the stage.
Note: The Bio-Rad gel alignment template kit allows four sizes of standard
agarose gels to be centered quickly and easily and ensures the consistent
placement of each gel. See Appendix D, Accessories, on page 211 for
more information.

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Running a Protocol
To run a protocol

Click Run Protocol in the left pane.

The software runs through the steps in imaging mode and a progress indicator
tracks the progress.

Note: If you run a chemiluminescence application using autoexposure, you

might see the following error message:
The sample is too faint to use the autoexposure setting. Change the Image
Exposure setting to use manual exposure or Signal Accumulation Mode (SAM), then
run the protocol again.
This error occurs if Image Lab, using the initial autoexposure algorithm, determines
that it would take too long (approximately 30 minutes or more) to acquire an image.
You can always use the manual Image Exposure setting and specify a longer
exposure time.
After the images are taken, Image Lab continues with the detection, analysis, and
reporting steps if these steps were selected in the Protocol Setup pane. When
Image Lab has completed running the protocol, it displays the images that were
acquired. You can then edit and save these images or do further analysis on them.

To end the protocol

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Click Cancel Run.

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Running Signal Accumulation Mode (SAM) Protocols

Running Signal Accumulation Mode (SAM) Protocols

You can interrupt the acquisition of images for a SAM-enabled protocol at any time
by clicking Stop Acquire and Continue with Selected. The acquisition process stops
and continues the protocol with the selected image. Any other images that were
acquired are discarded.
At the end of the acquisition process, a thumbnail of each image appears at the
bottom of the window. The last image acquired appears, by default, in the main
window. You can view any of the SAM images in the main window by clicking on the
thumbnail image.

Review the images, select the image you want to use in your analysis, and click
Select Image and Continue. Image Lab continues to the next step in the protocol
using the selected image.
Note: After you click Select Image and Continue, only the selected image is
retained. All other images are deleted. Therefore, save any images you want to
keep before continuing with the protocol. See Saving Signal Accumulation
Mode (SAM) Images on page 90 for more information.

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Saving Signal Accumulation Mode (SAM) Images

You can save any individual image or all images at once by right-clicking on the
image and selecting an option from the shortcut menu.

To save a single SAM image

1.

Right-click the thumbnail and click Save.

2.

In the Save File dialog box, accept the default name for the file or enter another
name. Click Save.

To save all SAM images

1.

Right-click any of the images and click Save All.

2.

In the Select Directory dialog, enter the name of the folder and click Select
Folder.
The images are saved in the specified folder. The name of the file includes the
user name, timestamp, and exposure time. For example: John Doe 2012-05-01
15 hr 44 min_Exposure_5.0sec.

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Setting Up a Custom Application

Setting Up a Custom Application

Use the Manage Custom Application dialog box to save an existing application with
a new name or to create an application unlike existing applications.

To create a custom application

1.

Select Custom on the Applications menu to display the Manage Custom

Applications dialog box. If you have stored custom applications, they display
here.

2.

Click New.

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The Create Custom Application dialog box appears.

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3.

Enter a unique application name.

4.

Select a light source, filter, and image color from the lists.

5.

Select a binning setting. Choosing a higher binning setting combines pixels to

increase the amount of signal without increasing noise. While a higher setting
provides optimal sensitivity for low-light applications such as
chemiluminescence, it also reduces image resolution.

6.

Click OK to create the custom application.

| Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Regression Methods

Regression Methods
A regression method is used to calculate the molecular weight of the unknown
bands. The software uses the relative front and molecular weight values of the
standard bands to calculate the standard curve. This standard curve is then used to
calculate the values of the unknown bands. The shape of the standard curve is
based on the selected regression method. Choose one of the four regression
methods listed in Table 3.
Table 3. Methods of generating a standard curve
Regression Method

Minimum number
of standard bands

Linear (semilog)

Point-to-point (semilog)

Logistic

Cubic spline

If you do not have enough data points for the selected method, the molecular
weight of the unknown bands is not calculated.
You can check how well each regression method fits the data in the standard curve
window (see Standard Curve on page 112 for more information). The linear (semilog)
regression method provides a measurement that describes how well the standard
curve fits the data R2 value. The closer the R2 value is to 1.0, the better the data fit
the standard curve.
The molecular weight of each band is displayed in the Mol. Wt./Base Pair column in
the analysis table. See Molecular Weight (MW) Analysis Tools on page 131 for more
information about molecular weight.
For information about the calculations behind the regression methods, see
Appendix G, Regression Calculation Methods.

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Application Tables
The following tables specify the light sources and conversion screens or filters you
need for each type of gel or blot to be imaged.
Table 4. Nucleic Acids
Detection
Reagent

Preferred
Light
Source

Alternate
Light
Source

Preferred
Filter

Alternate
Filter

Preferred
Flat Field

Alternate
FlatField

Ethidium
bromide

UV

None

Standard

630/30

Orange

Lens

SYBR
Green

XcitaBlue

UV

560/50

Standard

Lens

None

SYBR Safe

XcitaBlue

UV

560/50

Standard

Lens

None

SYBR Gold

XcitaBlue

UV

560/50

Standard

Lens

None

GelGreen

XcitaBlue

UV

560/50

Standard

Orange

Lens

GelRed

UV

XcitaBlue

630/30

Standard

Lens

None

Fast Blast

White trans

None

Standard

None

White
conv

Lens

Table 5. Protein Gels

Detection
Reagent

Preferred
Light
Source

Alternate
Light
Source

Preferred
Filter

Alternate
Filter

Preferred
Flat Field

Alternate
FlatField

Coomassie
Blue

White trans

None

Standard

None

White
conv

Lens

Copper stain

White trans

None

Standard

None

White
conv

Lens

Zinc stain

White trans

None

Standard

None

White
conv

Lens

Flamingo

UV

None

Standard

Standard

Orange

Lens

Oriole

UV

None

Standard

None

Orange

Lens

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Table 5. Protein Gels, continued

Detection
Reagent

Preferred
Light
Source

Alternate
Light
Source

Preferred
Filter

Alternate
Filter

Preferred
Flat Field

Alternate
FlatField

Silver stain

White trans

None

Standard

None

White
conv

Lens

Coomassie
Fluor Orange

XcitaBlue

UV

560/50

None

Orange

Lens

SYPRO Ruby

UV

None

Standard

None

Orange

Lens

Krypton

XcitaBlue

UV

560/50

Standard

Orange

Lens

Detection
Reagent

Preferred
Light
Source

Alternate
Light
Source

Preferred
Filter

Alternate
Filter

Preferred
Flat Field

Alternate
FlatField

Colorimetric

White epi

None

Standard

None

Lens

None

Qdot 525

UV

None

520/30

Standard

Lens

None

Qdot 565

UV

None

560/50

Standard

Lens

None

Qdot 625

UV

None

630/30

Standard

Lens

None

Cy2

UV

None

520/30

Standard

Lens

None

Alexa Fluor
488

UV

UV

520/30

Standard

Lens

None

DyLight 488

UV

UV

520/30

Standard

Lens

None

Fluorescein

UV

None

520/30

Standard

Lens

None

Table 6. Blots

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Viewing Images

After a gel is imaged, the image appears in the workspace. Many controls are
available to optimize viewing and to analyze the image.
The following screen shot shows a gel image with band and lane detection as well
as annotations. The labels are overlays that you can display or hide.
There are many ways to view the data associated with the results. You can view
data as an analysis table, a standard curve, and a report.

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Displaying Gel Images

The display toolbar is located above the gel image. Each tool is described in the
following sections.

Display Gel Options

This section describes the settings in the Display Gel Options dialog box.

Annotations
You can choose whether to show text and arrow annotations that have been drawn
on the image.

Lanes and Bands

You can turn on or off any image overlays, such as lane frames, lanes, bands, lane
labels, molecular weight legends, and band edges.

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Band Attributes
You can show the following attributes for selected lanes or for all lanes.

Band number

Band label

Molecular weight

Relative front

Volume

Absolute Quantity

Relative Quantity

Band %

Lane %

Volumes
If you drew volume boundaries on the gel, you can display the boundaries and their
volume labels.

Zoom Tools
The zoom tools resize the gel image. Click the magnifying glass with the plus sign to
make the image larger. Click the magnifying glass with the minus sign to make the
image smaller.
You can also zoom in on an area using the right mouse button. Right-click and drag
to select the area you want to magnify. You can also resize the image by rightclicking and using the scroll wheel on your mouse.
Tip: You can return to the original view by right-clicking anywhere on the
image.

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Fit in Window
If you zoomed in on an area of an image, this button brings the entire image back
into view.

Image Transform
The Image Transform dialog box adjusts image brightness and contrast, optimizing
the image display so faint details can be seen.

The minimum to maximum range varies depending on the light and dark values
present in the image. These adjustments do not change the data. They change only
the way the data are displayed. The human eye cannot see as great a range as the
image contains.
The frequency distribution histogram shows the total data range in the image and
the amount of data at each point in the range.
Auto Scale determines an optimal setting for the image automatically. The lightest
part of the image is set to the minimum intensity, and the darkest is set to the
maximum.

100

The High progress indicator determines which intensity value is shown at

the maximum gray scale (or other color) in the gel image.

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The Low progress indicator determines which intensity value is shown at

the minimum gray scale (or other color) in the gel image.

The Gamma progress indicator changes the gray scale curve. A value of 1
is linear. A value <1 redistributes a greater proportion of the gray scale to
the first half of the intensity values. A value >1 redistributes a greater
proportion of the gray scale to the second half of the intensity values.

You can also type numerical values in the boxes next to the progress indicators.
Clicking anywhere on the progress indicator bars moves the progress indicator
incrementally.

Options:

Invert image display inverts dark bands on a light background to light

bands on a dark background. Light bands on a dark background are
inverted to dark bands on a light background.

Highlight saturated pixels when this checkbox is selected, areas of the

image with saturated signal intensity (higher than a measurable range) are
highlighted in red.

Linear or logarithmic histogram this adjustment changes the y-axis on

the histogram to display the number of pixels at each intensity value using
either a linear or a logarithmic scale.

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Viewing Images

Image Colors

You can choose a color map for your image results file. Viewing the image with a
different color scheme can make it easier to see all of the elements in the image, but
it does not change your data.
The first eight color choices imitate the colors of stained gels. The remaining
choices supply enough color variation to highlight small differences in the image
data. The available colors include:

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Gray

EtBr (ethidium bromide)

Coomassie

Stain-free

SYBR Green

SYPRO Ruby

Flamingo

Silver

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Displaying Gel Images

False color

Spectrum

Gold-Silver

Pseudo

3-D Projection

The 3-D view transforms the gel image into a solid three-dimensional model
spinning in space with x, y, and z dimensions. Accentuate or diminish the relative
heights of data points by pulling the slider at the bottom of the window to the right
or left.

To view the intensity of various bands

1.

Select the 3-D button in the display toolbar.

2.

Click and drag the model to rotate it into your preferred view.

3.

Bring the window into focus by clicking the image.

4.

Press C to display an inverted green cone, which can be dragged around to

evaluate the intensity of various bands.

5.

Press C again to hide the tool.

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Viewing Images

Image Info
The Image Info dialog box provides information about the active image.
The dialog box has three tabs.

Image Details acquisition and image information appear in this tab.

Analysis Settings settings that were used when the gel was analyzed
are displayed here. For example, Band Detection and Molecular Weight
Analysis will appear if they were performed.

Notes on this tab you can add notes, point out the types of samples
used, and add any other information about the results. You can create
custom labels for the lanes in your image.

Displaying Data
You can view results from analyzed data associated with a gel image using an
analysis table, a lane profile, a standard curve, or a report.
The buttons in the main toolbar turn these views on or off. You can view your data
with all views simultaneously.

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Analysis Table Options

Numerical data associated with an analysis can be viewed in an analysis table. Data
from the Lane and Band analysis can be viewed in the Lane and Band Table tab. If
volume analysis was performed, these data can be viewed in the Volume Table tab.
The buttons above the table provide options for displaying and exporting analysis
table data.

To change the size of the Analysis Table window

Move your cursor to the top of the window until it changes into a
double-headed arrow. Click and drag the edge of the window until you can see
all of the data.

Note: Resizing the Analysis Table window is restricted when a protocol window
is open.

Display Data Options

The Display Data Options dialog box has three tabs.

Measurements enables you to choose the measurements to display in

the table. Use the arrows to move the columns between the Not Displayed
pane and the Displayed pane. By default, all measurements are displayed
in the Analysis table.
Note: For a description of each Lane and Band measurement type, see
Lane and Band Table Measurement Definitions on page 107. For a
description of each Volume measurement type, see Volume Measurement

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Definitions on page 108.

Display enables you to set the display for the analysis table. The
following settings appear on the Display tab:

Default display settings The Move selected lane to top checkbox is

selected by default. When you click a lane on the image, the Analysis table
scrolls so that the data for the selected lane appears first in either the
vertical or horizontal view of the table.

Per Measurement Precision set the precision (decimal places) for the
measurements in the Lane and Band table and the Volume table.

Example shows an example of how measurements will display with the

selected measurement and precision settings.

Export enables you to choose how to export the analysis data. The
following settings appear on the Export tab:

Export formatting select checkboxes to include lane headers (Lane

and Band table tab only) and/or column headers in the exported file.

Export delimiter select a delimiter option for the exported file.

Comma delimited

Tab delimited

Use other delimiter (user defined)

Change Analysis Table Orientation

This button toggles between two table orientations.
Horizontal displays the lanes/volumes beside each other, so you can scroll
through the table from left to right.
Vertical displays the lanes/volumes on top of each other, so you can scroll
through the table from top to bottom.

Copy Analysis Table to the Clipboard

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Displaying Data

Copies the table data to the clipboard so that you can paste the data into word
processing or presentation applications.
Tip: It is best to use the vertical table orientation when copying to an
8.5 x 11-inch page, to give the columns enough room to display.

Export Analysis Table to a File

Exports the table data as a CSV file so you can open it in a database application.

Export Analysis Table to Excel

Exports the table data to a spreadsheet so you can use the sorting and formula
functions to manipulate your data. If you have Excel (PC or Mac) or Numbers (Mac)
installed on your computer, the program opens with your spreadsheet displayed.

Lane and Band Table Measurement Definitions

This section defines the measurements that display in the Lane and Band tab in the
Analysis table. Use the Data display options button to choose the columns to
display.
Band Number each band in a lane has a unique number, sorted from top to
bottom.
Band Label you can assign a custom label to each band by clicking the Band
Label field of the Lane and Band table.
Molecular Weight the molecular weight of the band is calculated based on the
user-defined standard and regression method. Italic values indicate extrapolated
values. When using nucleic acid gels, the size of the band is displayed in base pairs.
Relative Front values between 01 indicate the relative movement of the band
from top to bottom.
Volume the sum of all intensities within the band boundaries.
Abs. Quant. absolute quantification of the band.
Rel. Quant. relative quantification of the band compared to the reference band.

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Viewing Images

Band % percentage of the bands volume compared to all band volumes in the
lane.
Lane % percentage of the bands volume compared to the entire volume of the
lane.

Volume Measurement Definitions

This section defines the measurements that display in the Volume tab in the Analysis
table. Use the Display Data Options button to choose the columns to display.
Volume Number a unique number is assigned to each volume.
Volume Label software-generated labels for different types of volumes
(U unknown, B background, S standard). Label can be changed in Volume
Properties.
Volume the sum of all the intensities within the band boundaries.
Adjusted Volume the background-adjusted volume.
Mean Background the mean value of the background.
Absolute Quantity Volume the quantity of the volume based on the standard
volumes and the regression method.
Relative Quantity Volume the ratio of the adjusted volume and the adjusted
volume of the reference volume.
# Pixels number of pixels inside the volume boundary.
Minimum Value intensity of the pixel with the minimum intensity inside the
volume.
Maximum Value intensity of the pixel with the maximum intensity inside the
volume.
Mean Value mean value of all pixels inside the volume boundary.
Standard Deviation standard deviation of all pixels inside the volume boundary.

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Area area of the volume in mm2.

Lane Profile

Background

Band
boundaries

The Lane Profile option shows a cross-section view of a single lane rotated 90.

To explore the Lane Profile window

Use the Next and Previous buttons at the top of the window to page
through profiles of the lanes in your image.

Click on a lane in your image to see its profile.

The title bar identifies which lane profile is in view (Lane 1, Lane 2, and so on).
There are several settings in the title bar.

Scale to fit graph

Include Background

Identify Bands by

These settings, as well as the zoom tools, are global. They apply to all the profiles.

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The zoom tools work the same way they work elsewhere in the software. For more
information, see Zoom Tools on page 99.
In addition to the graph of the lane intensities, the Lane Profile tool also shows an
image of the selected lane below the graph. The transform and color map are
applied to the gray-scale image.
As you move your cursor over the profile, the current relative front (Rf) value and the
average intensity (Int) value at the Rf value display in the lower-right corner of the
Lane Profile window.

Scale to Fit Graph

You can choose the highest point of the display to define the range of the graph.
This provides the best view of the lane profile.
You can clear the Scale to Fit Graph checkbox to display the entire range of
possible intensity values in the graph. Doing so allows valid comparisons between
different lanes.

Include Background
When the Include Background checkbox is selected, the Lane Profile window
shows the subtracted background under the blue line. The area used for band
quantification appears in green under the red line.
When you clear the Include Background checkbox, the area of the lane profile that
represents the background of the image does not display.

Identify Bands by
You can change how the bands are labeled by choosing from the options in the
Identify Bands by dropdown list. By default, the bands are labeled by band number.
You can display one of the following attributes:

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Band Number (Band No.)

Band Label

Molecular Weight (Mol. Wt.)

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Relative Front

Volume

Absolute Quantity (Abs. Quant.)

Relative Quantity (Rel. Quant.)

Band %

Lane %

Adjusting Band Boundaries

Below the profile of each lane there is a strip that displays the bands. Each band is
surrounded by a pair of vertical lines delimiting its boundaries. You can move the
vertical lines and change these boundaries.

To change the boundaries of a band

1.

Hold your cursor over one of the boundary lines until a double arrow appears.

2.

Click and drag the cursor to the new position.

Note: Boundary lines cannot overlap one another. Therefore, you will not
be able to move any boundary line beyond the next boundary line.

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Standard Curve

The Standard Curve dialog box displays the best curve fit for the defined standards
and the bands relative to this curve for the lane selected in the image. The tabs at
the bottom of the dialog box display the standard curves for three different
analyses.
Standards appear in green. Unknown bands appear in red. You can toggle the
molecular weight display on the y-axis between linear and log scale by clicking the
Log y-axis box at the upper left. The regression method you chose in Molecular
Weight Analysis Tools appears, as well as the formula (if applicable) and the R2 value
of the regression method.
Tabs in this window enable you to view the molecular weight standard curve, the
absolute quantity standard curve, or the volume standard curve.

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Report
See Generating Reports on page 151, for information about reports.

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Analysis Tool Box tools are enabled when an image file is opened and in focus. An
active or in focus window has a darker blue menu bar on a Windows PC. On a
Macintosh, the window control icons display more brightly when a window is active.
This distinction helps you to identify the active window among many open image
files in your workspace.

Using Auto Analysis

Auto analysis detects the lanes and bands and, optionally, calculates the molecular
weight of the bands in your image. Click Auto-Analysis in the Analysis Tool Box to
do the following:

Analyze images obtained with protocols that did not include steps for auto
detection and analysis

Change your analysis parameters to reanalyze your images

Note: If you change any settings for an analyzed gel, the initial analysis is
overwritten. To preserve both analyses, save each image file with a different
name.

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Auto Detection Settings

The band detection sensitivity options are as follows:

Low Band Detection Sensitivity sets detection at a low level for images with
prominent bands. Faint bands are not detected with this setting.
High Band Detection Sensitivity sets detection at a higher level for images that
are faint. Extraneous bands can be removed using the Band Tools in the Analysis
Tool Box. See Lane and Bands Tools on page 121.
Custom enables you to set a value between 1 and 100 to select the best
detection sensitivity for your sample. You can also drag the sliding bar left or right to
set the value.
When Low Band Detection Sensitivity is used, the numerical value is set at 25; when
High Band Detection Sensitivity is used, the value is set at 75.

Molecular Weight Analysis Settings

Molecular Weight Standard choose any of the many Bio-Rad standards or
another standard you added to your standards list.
Standard Lanes choose or change the lanes in which the standards are placed.

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Regression Method four regression methods are available. For more

information, see Regression Calculation Methods on page 229.

Analysis Tool Box Tools

All Analysis Tool Box tools customize the analyzed data in image files. These tools
are available only when an image file is open. Click a specific image to select among
the many windows that might be open in your workspace.
Note: Some tools delete the existing analysis.

To access a tool

Click any of the toolbox buttons.

To return to the Analysis Tool Box menu

Click the green Up arrow to the right of the tool name.

Up arrow

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Image Tools
The image tools enable you to manipulate your images.

To display the image tools menu

Click Image Tools.

Flip you can flip the gel image horizontally or vertically.

Rotate you can rotate the gel image 90 using the Left or Right buttons.
Or you can set a custom rotation using the Custom button.

Crop you can trim the outer edges of your image to any shape or area.

Invert Data you can toggle the image data from positive to negative.

Merge you can merge a chemiluminescent blot image with a

colorimetric image of the same blot.

The sections that follow describe how to use these tools in greater detail.

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Correcting a Slanted Gel

To correct a slanted gel
1.

In the Rotate section, click Custom.

Red arrows appear over the gel image.

2.

Rotate the red arrows to any degree between 0 and 360 by dragging them.

3.

Right-click the gel image and click Rotate to set your gel in the new position.
Click Cancel if you do not want to set the rotation.

Cropping a Gel Image

You can save crop settings and use them to crop other images. This feature is useful
when you want to crop the same area in several images.

To crop a gel image

1.

Click Crop. A red box outlines the image area.

2.

Drag the red box to surround the image area you want to keep.

3.

(Optional) Right-click the image to open the Crop menu and click Save Crop
Settings.
The Save Crop Settings dialog box appears.

4.

(Optional) Type a name for the crop settings and click OK.

5.

Right-click and select Crop or Cancel. Selecting Crop crops the image to the
area inside the red box.

To crop an image using saved crop settings

1.

Click Crop. A red box outlines the image area.

2.

Right-click the image to open the Crop menu and click Load Crop Settings.

3.

Select the saved crop settings that you want to use and click Load. The red box
resizes and the crop specifications appear on the image.

4.

Right-click and select Crop. The image is cropped to the area specified in the
crop settings you selected.

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To delete crop settings

1.

With an image open, click Crop.

2.

Right-click the image inside the red box to open the Crop menu and click
Delete Crop Settings.

3.

Select the crop settings in the dialog box that appears and click Delete.

Inverting Data
Use this button to change the image data from positive to negative. Invert Data is
used for negative stains and zymograms. Intensity values of bands must be greater
than background to perform analysis on the gel. View the gel as a 3-D projection to
determine if the data must be inverted.

Merging Images
Use this button to merge a chemiluminescent blot image with a colorimetric image
of the same blot. If you have used colorimetric prestained standards for a
chemiluminescent blot, you can acquire an epi-white light image of the blot to show
the standards and a chemiluminescent image to show immunodetection. These two
images can then be merged into a combined image with both signals.
Note: Merging images can have an adverse effect on quantification. If accurate
quantification is required, perform analysis on the original, separate images.
Only images of the same size can be merged.

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Lane and Bands Tools

The Lane and Bands tools enable you to identify the lanes and bands in your
images.

To open the Lane and Bands tool

In the Analysis Tool Box, click Lane and Bands, then select the tab for the
Lanes or the Bands tool.

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Detecting Lanes
To detect the lanes in the image, do one of the following:

Click Automatic if the gel image is fairly typical.

Click Manual to detect a specific number of lanes or if automatic lane

detection did not find all the lanes.

Using the All Lanes and Single Lane Tools

To use the All Lanes and Single Lane tools

Click the lane tool first, then click the lane to which you want to apply the
change.

All Lanes Tools

You can modify all the lanes in your image using these tools.

Resize resizes the lanes in your image. Click Resize and then drag the
handles of the red squares to resize all lanes.

Adjust adjusts the orientation of all lanes. Click Adjust and then drag a
single corner of the lane frame. The Adjust All Lanes tool does not resize
lane width.
You can add anchor points to the top or bottom borders of the rectangle by
clicking the lane frame. Remove any unneeded anchor point by right-clicking it.
By dragging these additional anchor points, you can adjust for smiling gels.
Note: You can move the entire lane frame when resizing or adjusting the
lanes. Click Resize or Adjust and make the changes to your lanes. Then
click anywhere in the frame and move it to the desired location.
Tip: On the PC, you can press the Shift key or the Ctrl key and use the
arrow keys on your keypad to move the lane frame. On a Mac, press the
Shift or the Command key and use the arrow keys.

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Delete deletes all lanes.

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Width changes the width of all lanes at the same time. Click Width and
then drag one of the anchor points on any lane to change the width of all
lanes.
Tip: You can use the plus key (+) on your keyboard to increase the lane
width, and the minus key (-) to decrease the width.
You can also change the lane width by specifying the size of the lanes. Click
Width, enter a number (in mm) in the box, and click Apply.
Note: You might see that the number in the box changes after you click
Apply. For example, you enter 5.75 and the number changes to 5.71. The
image is made up of pixels and the Image Lab software can only draw a
boundary for the lane between pixels. If the number you specify would
cause the boundary to fall anywhere on the pixels, Image Lab moves the
boundary so that it falls between two pixels and updates the number
accordingly.
Note: The All Lanes Width tool makes the width of all lanes uniform.
Therefore, if you used the Single Lane tool to change the width of an
individual lane, this change is overridden by the All Lanes Width tool.

Single Lane Tools

Specific information about how to use each lane tool is described in the following
section. You can modify an individual lane in your image using these tools.

Add adds a lane to a gel image. Click Add, then click within the lane
frame where you want to place the new lane. The lanes are automatically
renumbered.
Note: To add a lane outside the frame, add a lane inside the frame and
click Move to expand the lane outside the frames boundaries.

Delete deletes a lane. Click Delete, then click either the lane or its lane
number. The lanes are automatically renumbered.

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Bend bends a lane to better fit the gel image. Click Bend, then drag
square anchor points to fit the lane to the image.
Note: To add anchor points, left-click within the lane. Drag these anchor
points to adjust the lane to fit the gel image. To remove an anchor point,
right-click on it.

Move moves a lane to a new position on a gel image. Click Move, then
click the lane you want to move. Drag it to a new location. The lanes are
renumbered according to their new positions.

Width changes the width of a lane. Click Width, then click within the
lane. Click on the anchor points to adjust the lane width.
You can also change the lane width by specifying the size of the lanes. Click the
Width button, enter a number (in mm) in the box, and click Apply.
Note: You might notice that the number in the box changes after you click
Apply. For example, you enter 5.75 and the number changes to 5.71. The
image is composed of pixels and the software can draw a boundary only in
the lane between pixels. If the number you specify would cause the
boundary to fall anywhere on the pixels, Image Lab moves the boundary so
that it falls between two pixels and updates the number accordingly.

Lane Background Subtraction

To perform lane-based background subtraction

Select Enable Subtraction in the Background Subtraction field. Use the Lane
Profile view to see the subtracted lane background.

Disk Size specifies the size of a hypothetical rolling disk (between 0.5 and
99.5 mm in 0.5 mm increments) that removes background levels along the length of
the lane. The size of the disk determines how closely the background level follows
the intensity profile.
A large disk follows the profile trace less closely, touching fewer points along the
trace and removing less background. A smaller disk more closely follows the profile
trace, removing more background.

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A disk radius that is too large will result in poor background removal. A disk radius
that is too small might subtract actual data. For most samples, a size of <10 mm is
usually appropriate. You can perform this task several times until you are satisfied
with the amount of background removed. Use the Lane Profile tool to evaluate the
appropriate disk size for background subtraction.
Apply to selected Lane applies the specified level of background subtraction
only to the selected lane. This option enables you to set different background
subtraction levels for each lane.

Copying Lanes
You can copy the lanes from one image into any other image. The entire frame and
all the lanes are copied. Individual lanes cannot be copied.

To copy lanes between channels

1.

Open the Lane and Bands tool.

2.

Select the channel that contains the lanes you want to copy.

3.

From the Edit menu, select Copy.

The message Copying All Lanes appears.

4.

Select the channel that you want to copy the lanes into.

5.

From the Edit menu, select Paste.

A channel can contain only one lane frame at a time. Therefore, if you paste
lanes into a channel that already contains a lane frame, you are prompted to
confirm the deletion of the existing lanes. If you click Yes, the existing lanes are
deleted and replaced with the copied lanes.

After the lanes are copied into the channel, you can manipulate individual lanes
using the lane tools so that they are correctly positioned.

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Detecting Bands
Bands are detected for individual images.

To detect bands in the image

126

Click Detect Bands to open the Band Detection dialog box. Select band
detection sensitivity and the lanes to which it applies.

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Displaying Advanced Detection Options

You can also select Display Advanced Options to set specific parameters for band
detection. When initially expanded, the values in Advanced Options are set in
relation to the band detection sensitivity that you select in Detection Settings. You
can set specific parameters for the sensitivity level and apply the parameters to all
lanes or to a specific lane.
If later you change the band detection sensitivity in Detection Settings, the values in
Advanced Options change in relation to the new sensitivity levels. You might need to
reset these parameters to the original values.

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Advanced Detection Options

Sensitivity determines the minimum optical density that will be defined

as a band. The higher the sensitivity value, the lower the minimum signal
intensity, therefore the more bands will be detected.
If the sensitivity is set too high, background staining might be detected as
bands. If the setting is too low, bands of interest might not be detected.
The default sensitivity setting is 10.0. If the gel has faint bands (for example, if
the optical density is less than 0.05, and counts are less than 2,000), you may
want to increase this value to 20.0.

Size Scale distinguishes between trends in signal intensity and random

intensity fluctuations. It is the number of pixels in a vertical column that are
taken together to determine whether a band is present.
The Size Scale parameter uses the size of objects in the image to determine the
nature of those objects. If a gel image has high levels of background noise, a
larger size scale is appropriate. At low noise levels, a smaller value is preferable.
You can also increase the size scale if the gel has only a small number of thick
bands scanned at high resolution.

Noise Filter minimizes the number of small fluctuations (or noise) in the
image that are called bands while still recognizing larger features (the real
bands). This filter becomes especially important at higher sensitivity levels.
The noise filter value refers to the size of the filter in pixels (for example, a value
of 2.50 equals a filter size of 2.50 x 2.50 pixels). Features smaller than the filter
size will not be recognized as bands. Entering a noise filter size of zero turns it
off completely.
If band detection identifies doublets as single bands, decrease the noise filter
setting and/or increase the sensitivity level.
Tip: You can also decrease the Size Scale parameter instead of the noise
filter to improve the detection of closely spaced bands. However, if you
decrease both the noise filter and the size scale, the fuzziness around
bands might be mistakenly detected as separate bands.

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Shoulder band detection tries to distinguish shoulders as separate

bands. When looking at a lane trace, these bands appear as flat or gently
sloping abutments to darker, better-defined bands (that is, there is no dip
on the trace between the two bands). Increasing the shoulder sensitivity
results in more shoulders being detected as bands. Changing this setting
to zero results in no shoulders being recognized as separate bands.
If band detection calls a doublet a single band, check the lane trace to see if
there is a dip between the peaks of the two bands. If there is no dip, increasing
the shoulder sensitivity value will help resolve the two bands.

Normalize Sensitivity compensates for differences in intensity between

lanes.
The intensity of each lane is determined by the darkest band in that lane. For
example, suppose that in all but one of the lanes the darkest band has an
intensity of 50,000 counts. In the light lane, the darkest band is only 25,000
counts. With normalization, band detection will be twice as sensitive when
processing the light lane, improving the detection of faint bands.
Note: It does not normalize for band quantitation.

Band Limit enables you to limit the number of bands that will be
detected in each lane, thus reducing the need for later editing.

Editing the Detected Bands

You can optimize the bands in your images using these tools.

Add manually add a band to a lane. Click Add, then click at the desired
location in a lane. Image Lab then locates a faint band close to where you
clicked.
Tip: You can darken your entire image to view faint bands more easily
using the sliders in the Image Transform dialog box. For instructions, see
Image Transform on page 100.

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Delete deletes a band from a lane. Use this feature to remove bands
that are not relevant to your analysis. Click Delete, and then click on the
band you want to remove.

Adjust adjusts the height of a band. Click Adjust. Boundary lines appear
on either side of each band. Move the cursor over a boundary line until you
see a double-headed arrow. Move the boundary line up or down. The
center is recalculated and the band appears there.
Note: You can also adjust band boundaries in the Lane Profile view.

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Molecular Weight (MW) Analysis Tools

Molecular Weight Analysis tools enable you to determine molecular weight (or base
pairs, if using nucleic acid gels) by comparing a test sample with known standards.
Note: Before you can use the Molecular Weight Analysis tools, you must detect
the lanes and bands in your image.
You can view each bands molecular weight in the molecular weight column of the
Lane and Band tab in the Analysis Table.

You can also display the molecular weight of the bands on the gel image by opening
the Display Gel Options window and selecting Mol. Wt. from the dropdown list in the
Band Attributes section. (See Displaying Gel Images on page 98 for information
about displaying band attributes.)

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Molecular Weight Standard

You can change the standards to ones that are relevant to your samples.

To change standards

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Click Change to access the Manage Standards dialog box. Choose another
standard or add your own standards.

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Standard Lanes
Standard samples are placed in the first and last lanes by default. You can specify
other standard lanes by selecting the box below each lane or by entering the
standard lane numbers separated by commas in the MW Analysis Tools dialog box.
In the Lane and Bands view, Std appears below the lanes, identifying them as
standard lanes. In the Molecular Weight Analysis view, these lanes are indicated
with a check mark below the lane.
There are three standard lanes in the screen shot below (lanes 1, 16, and 17).
Standard lanes are identified with a check mark in the boxes below the lane. The
molecular weights of the bands in the standard lanes appear on either side of the
lane frame. The red lines running from one end of the lane frame to the other identify
the location of the bands in the standard lanes. You can use these lines to see where
the bands in the other lanes fall relative to the bands in the standard lanes.
Note: You can use the lane labels to identify your standard lanes.

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The values for the molecular weight in any standard lane appear in bold. In this
example, the values for the first lane are in bold. (Lanes 16 and 17 are not included
in the screen shot, but their values would also be in bold.) The molecular weight of
the bands in the remaining lanes is calculated relative to these standards.

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Regression Methods
There are four regression methods.

Linear (semilog)

Point-to-point (semilog)

Logistic

Cubic spline

See Regression Calculation Methods on page 229.

Quantity Tools
You can quantify bands in test samples automatically using either the Relative or
Absolute tab under Quantity Tools.

Relative Quantity Tab

To compare the relative quantities of bands
1.

Select the Relative tab.

2.

Click Select.

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3.

Click the band you want to use as a reference. A small R appears near the band
you selected.

To review the relative quantities of bands

Go to the Rel. Quant. column of the Analysis table (Lane and Band tab). The
relative quantity is the ratio of the band volume divided by the reference band
volume.

All other bands now display numerical values that are relative to the reference band.
If the reference band value is 1.00, values higher than 1.00 indicate that the band
quantity is greater than the reference band. Values lower than 1.00 indicate the band
quantity is less than that of the reference band.

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Absolute Quantity Tab

Absolute quantification is used to quantify bands based on known standard bands
using a calibration curve.

To calculate the absolute quantities of the bands

1.

Select the Absolute tab.

2.

Click Select.

3.

Select at least two standard (known) bands and assign quantity values. Small
As appear (shown below circled in red) near the bands you selected.

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The values display in the Standard Bands table. The greater the number of
known bands and the wider the range of their values, the more accurate the
absolute quantity calculation of the unknown bands will be.

Note: Any standard band selection can be deleted. To do so, select the
entry in the Standard Bands field and then click Delete.
4.

Select a unit of measure from the Units dropdown list.

5.

Select a regression method from the dropdown list.

Consider the following guidelines when making your choice.

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Linear generates a straight line that is the best fit of the values you provided
and is preferred in most cases.
Point-to-point generates a curve in which each data point is connected
directly to the next, regardless of the shape of the resulting curve.
Cubic spline generates a smooth curve that connects each data point. At
least four standard points are required to use this method of least-squares
polynomial fits.
Table 10. Regression Methods
Regression
Method

Minimum Number
of Standard Bands

Minimum Number with Force

Through Origin Option

Linear

Point-to-point

Cubic spline

6.

Click Standard Curve on the toolbar and select the Absolute Quantity Standard
Curve tab.

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The calibration curve displays. Standards are represented by green triangles.

Unknown values are represented by red triangles.
Note: Selecting the Force Through Origin checkbox always starts the
standard curve graph at 0,0, regardless of the best curve fit.

Note: Clicking the Standard Curve table generates a crosshair tool that
displays the numerical values associated with the placement of the cursor
in the graph.

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Annotation Tools
You can annotate results with text and arrows to emphasize areas of interest.

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Add Annotations

Text enables you to add text annotations to images to emphasize

important details. Click Text, then click an area on the image where you
want to insert your comment. A box appears with a dotted-line border.
Type your comment in the box.
Tip: To add a new line to the box, place your cursor where you want the
break and press Shift+Enter.

To move the box

Click and drag the box to change its position.

Arrow enables you to add arrows to images to emphasize important

details. Click where you want the arrow to start and drag to stretch the
arrow point to the location you want to emphasize. To move the arrow on
the image, click the middle of the arrow and drag it to the new position.

To change where the arrow points

Click either end of the arrow. Square boxes appear. Drag a box to change the
length or orientation of the arrow.

Alignment
The alignment buttons enable you to align multiple annotations, such as lane
numbers, which you have manually added.

To select multiple annotations

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Press the Ctrl key (Command key on a Mac) and click each item or drag a
selection box around them.

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Text Properties
You can change the type size and font of your text annotations.

Font click the box you want to change. Open the dropdown Font menu
to show all fonts installed on your system. Select a new font for the text
annotation.

Size click the box whose font you want to resize. Open the dropdown
Size list to increase or decrease the size of the text. You can set the font
size from 6 to 72 points using the dropdown list.

Color
You can change the color of text annotations to make them visible with any color
scheme and emphasize them further by adding a color to the annotations
background, which is invisible by default.

To change the color of multiple items

Press the Ctrl key and click each item.

Foreground click a text annotation or arrow. This activates the

Foreground field, so you can select a foreground color from the dropdown
list.

Background click a text annotation. This also activates the Background

field so you can select a background color from the dropdown list.

Rotate
You can rotate text annotations 90 to the left or right by clicking the Rotate buttons.

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Volume Tools
Volume tools enable you to manually quantify features on a sample image when
automated lane and band analysis is not appropriate or possible, such as in dot
blots.

You can use Volume tools to quantify the signal intensity of bands, spots, arrays,
and other image data. Define an area of interest by surrounding it with a shape. You
can choose a rectangle, circle, freehand, or lane shape by clicking the appropriate
button under the Volumes field.

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A default label appears within the shape drawn. The volume label is assigned a
sequential number and can be one of three types:

U unknown

Std standard

B background

Each new volume you create initially has a red border, which indicates that the
volume is selected. When you click elsewhere on the image, the border changes to
blue, indicating that the volume is no longer selected.
Note: Double-click a volume area to change its properties.

To review data for the volumes

Open the analysis table and select the Volume tab. Volumes are listed based on
their number and/or the associated information per volume. See Volume
Measurement Definitions on page 108.

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Volume Types
You can define the volume type (unknown, standard, or background), the quantity of
standard volumes, or enter a custom name to replace the default label.

Unknown volumes are volumes you want to quantify.

Standard volumes are used for absolute quantities. See Absolute Volume Quantity
on page 148.
Background volumes are used to remove the background from the calculation. The
result of volume background subtraction appears in the Adjusted Volume column of
the analysis table (Volume Table tab).
Note: This volume type needs to be assigned only when using Global
Background subtraction.

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Volume Tools

Volume Background Subtraction

When you draw a volume, some nondata background pixels might be included
inside the volume. These background pixels usually have an intensity value that you
do not want to include in your volume quantification. There are two methods of
calculating this background intensity for your volumes: local and global.

Local local background subtraction calculates a separate background

intensity for each unknown and standard volume you create. For each
volume, the intensities of the pixels in a 1-pixel border around the volume
are added together and divided by the total number of border pixels. This
gives an average intensity for the background around each, which is then
subtracted from the intensity of each pixel inside the volume. If the
background value is greater than the pixel value inside the volume, the
background-adjusted quantity of the volume could be <0. In this case,
redraw the border for this volume.

Global global background subtraction calculates a single background

intensity for the entire gel. This average background intensity is then
subtracted from all the volumes in the gel. The average intensity of the
pixels in the background volume is calculated and subtracted from each
pixel in all standard and unknown volumes. Therefore, it is not necessary
for the background volume area to be the same size as your unknown.

To calculate global background subtraction

1.

Use one of the Volume Tools to create a volume in a representative background

region of your image (that is, a nondata region similar to the background
surrounding your data).

2.

Double-click the volume. This opens the Volume Properties dialog box.

3.

Select the Background option button.

Note:

If you select Global in the Volume Tools toolbox but do not define a
background volume as described, no background subtraction is
performed.

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If you create more than one background volume, all the pixels in those
background volumes are used to calculate the average background. Your
background volume(s) will have default names B1, B2, and so on based on
the sequence in which they were created.

If the region you defined as background has a higher average intensity

value than your data object, you obtain a negative value for your adjusted
volume in the analysis table. If this happens, select a new background
region with less intensity than your data object.

Relative Volume Quantity

You can choose any one volume as a reference volume by selecting the Reference
Volume checkbox in the Volume Properties dialog box. The reference volume is
indicated by an asterisk on the volume label, for example, U1*.
Relative quantities are displayed in the Relative Quantity column in the analysis
table (Volume Table tab). The relative quantity is the ratio of the backgroundadjusted volume divided by the background-adjusted reference volume.
All other volumes now display numerical values relative to your reference volume.
Values higher than 1.00 indicate that the volume is greater than the reference
volume. Values lower than 1.00 indicate the volume is less than the reference
volume.

Absolute Volume Quantity

If you have drawn your volume around an object of known quantity, you can use it to
calculate the quantity of your unknown volumes. The quantities of your unknown
volumes are calculated based on the standard volumes and the selected regression
method.

To classify a particular volume as a standard

148

1.

Double-click the volume. This opens the Volume Properties dialog box.

2.

Select the Standard option button and enter the quantity in the Quantity box.

3.

Click OK to close the dialog box.

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Standard volumes will have the default names S1, S2, and so on, based on the
sequence of their creation.

To review the regression curve

Open the Standard Curve window and select the Volume Standard Curve tab.

Regression Methods
Three regression methods are available to generate the volume quantification curve
used for absolute quantity: linear, point-to-point, and cubic spline. To display the
standard curve, click the Standard Curve button on the toolbar and select the
Volume Standard Curve tab in the Standard Curve dialog box. See Regression
Calculation Methods on page 229 to learn how each of these methods is calculated.
The data for volume standards are found in the Absolute Quantity column of the
Volume Table.
Note: Selecting the Force Through Origin checkbox always starts the standard
curve graph at 0,0, regardless of the best curve fit.

Alignment
Align volumes by using the appropriate alignment button. To select several volumes,
Ctrl-click each one, then select one of the alignment buttons. Hover over any of the
six alignment buttons to display its function (Align Left, Align Right, etc.)
Copy and paste selected volumes by pressing Ctrl+C to copy. Press Ctrl+V to
paste.
When you click the Standard Curve button on the toolbar, a chart displays all
unknown and standard quantities.

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Generating Reports

After viewing results, you can generate a report that displays the analyzed gel
images, all of the settings used in the protocol, and as much information about the
data as you want to include.
You can choose print settings within the Report Settings dialog box in the Edit menu
or by clicking Report in the main toolbar.

Report
To produce a preview of your report

Click Report on the toolbar.

Use the following dialog boxes to customize the content in your reports. Doing so
does not modify the data from the analysis.

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Report Settings
Use this dialog box to customize the content of your report.

General Tab

The General tab includes the following settings:

Include Gel Image specify whether the image is included in the report.
If the image is included, the following options determine which overlays are
displayed on the gel image:

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Show Lanes and Bands

Show Volumes

Show Annotations

Include Unannotated Image

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Report

When you select this checkbox, both the annotated image (if available) and
unannotated images are printed in the report. The unannotated image
precedes the annotated image in the report.
Note: If you disable the other checkboxes in the Include Gel Image
group, the Include Unannotated Image checkbox is automatically
disabled.

Image Info specify what information is included in the report.

Acquisition Information

Analysis Settings

Image Information

Notes

Signature History details on when and why a secure document was

signed
If a secure document has been signed, the user name, the date and time of the
signature, and the reason for signing are all included in the Signature History
section. If a document has not been signed, this section is omitted from the
image report.

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Lane and Band Table Tab

The Lane and Band Table tab includes the following settings:

154

Include Lane and Band Table specify whether to include the Lane and
Band table in the report.

Lanes to show specify which lanes to display in the report.

Show Lane Profile specify whether to display the Lane Profile view.

Print one lane per page specify whether all lanes are printed on one
page or whether each lane is printed on a separate page (adds a page
break after each lane).

Show Lane Profile include the lane profile for each lane.

Not Displayed/Displayed remove columns that you do not want to

display in the report. By default, the report displays all columns from the
Lane and Bands table.

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Report

Volume Table Tab

The Volume Table tab includes the following settings:

Include Volume Table clear to exclude this information from your

report.

Not Displayed/Displayed remove columns that you do not want to

display in the report. By default, the report displays all columns from the
Volume table.

Print Report
Click the Print Report button to print your report.

Print Report to a PDF File

The Print Report to .pdf File button opens a Save dialog box so the PDF file can be
saved on your system.

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Generating Reports

Adjust the Printer Settings

The Printer Settings button accesses options for paper size, orientation, and page
margins.

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10 Exporting Results
The most convenient way to archive complete information about experiments is to
produce reports. However, you might want to export only gel images or analysis
table data for analysis in different programs, such as Quantity One, FPQuest, or
InfoQuestFP software. Or you might need exported files for presentation or
publication.

Exporting Gel Images

Image Lab software includes features for exporting gel images several ways.

You can export displayed image data to a publication (choose Export for
Publication).

You can export raw image data as a 16-bit .tif file (choose Export for
Analysis).

You can export image data to PulseNet. Doing so reduces the image to an
8-bit .tif file, limits its resolution, and restricts its file size to 300 Kb.

You can export lane and band tables as well as volume tables to a
spreadsheet program or to a file.

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Exporting Results

The options to export gel images are available using the Export option in the File
Menu.

Exporting Gel Images for Publication

Use this format only to export visual information to presentation or word processing
software, such as PowerPoint or Word.

To export a displayed image to a file

Select File > Export > Export for Publication.

Note: You can select from .bmp, .png, .jpg, and .tif formats. The gel displays with
any lanes, bands, and annotations that appear on the screen.

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Exporting Gel Images

In this dialog box you can:

Select the entire image or the current view

Select the resolution or specify a custom resolution

Specify the publishing dimensions

View the resulting published image size and dimensions

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Exporting Results

You can zoom in on an area in a current view and export only that area, or you can
export the entire image. You can exclude annotations or overlays by clicking Display
Gel Options on the toolbar to access the appropriate settings.

Exporting Gel Images for Analysis

To export an image for analysis
Select File > Export > Export for Analysis.
This exports the raw data only as a 16-bit .tif file.
Note: 16-bit .tif images are not compatible with all image viewers.
The image might require contrast adjustment when it is imported into analysis
software. This option creates a file that can be analyzed in other programs such as
Quantity One, FPQuest, or InfoQuestFP software.

Exporting Gel Images to PulseNet International

To export an image to PulseNet International

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Select File > Export > Export for PulseNet.

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Exporting Gel Images

Image Lab reduces the image to an 8-bit .tif image file. Resolution is limited and file
size is restricted to 300 Kb.

Exporting Lane and Band Tables to Excel

If you have Excel (or Numbers on a Mac) installed on your computer, you can export
the data to the spreadsheet application.

To export the data to Excel (or Numbers)

Select File > Export > Lane and Band Table to Excel.

This opens a table directly in the spreadsheet program. You can then use the Save
As option to produce other formats.

Exporting Volume Tables to File

To export an image as a CSV file

Select Export > Volume Table to File.

Image Lab exports the image as a comma-separated values (CSV) file so the data
file can be opened in a database application.

Screenshot Tool Export

Use the Screenshot tool, available on the toolbar, to capture a displayed image to
the clipboard or to save it to a file (.bmp, .gif, .jpg, or .png).

Analysis Table Export

You can export table analysis data from the File menu or by using the export buttons
at the top of the Analysis Table window.
The Analysis Table window has several buttons to export data to different formats,
depending on how the data are to be presented.

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Exporting Results

Copy Analysis Table to the Clipboard

Copies the analysis table to the clipboard and then pastes the analysis table into
word processing or presentation applications. It is best to use the vertical table
orientation when copying to an 8.5 x 11-inch page to accommodate the columns.

Export Analysis Table to a File

Exports an analysis table as a CSV file, so your data file can be opened in a
database application.

Export Analysis Table to a Spreadsheet

Enables you to use sort and formula functions of a spreadsheet program with your
data. If you have Excel (or Numbers on a Mac) installed on your computer, the data
open in the spreadsheet program.

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11 System Calibration
When your imager is installed, instrument calibration is performed with a calibration
wizard. See the installation guide for your imager in your Image Lab upgrade kit
for detailed instructions.
The instrument calibration wizard performs several procedures required to automate
the system and prevent focus problems. Each of these calibrations affects your
system as follows:

Focus Calibration this calibration allows automated focus settings at

any zoom point, using a (patent pending) software algorithm. Therefore,
your focus remains correct whether you view an entire sample or an area of
interest.

Focus Calibration with Height Offset this calibration takes the tallest
of the available conversion screens into account, and extrapolates values
for the others, so that focus remains optimal, for whichever screen is used.

Dark Reference Image this calibration determines and corrects any

background signal present in your imager (Gel Doc XR+ system only).

UV Flat Field Calibration this calibration generates the flat field correction
profiles needed for the UV light source. Because of this calibration, your
images have more accurate quantity reporting and backgrounds of even
intensity.

Lens Flat Field Calibration this calibration corrects for the intensity rolloff inherent in any lens.

White Conversion Screen Calibration this calibration generates a flat

field correction profile needed for the white light conversion screen.

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System Calibration

Recalibrating Your Imager

When you add light sources or filters to a Gel Doc XR+ or ChemiDoc XRS+
imager, recalibrate your imager using the Instrument Setup dialog box.

To recalibrate your imager

1.

In Image Lab, click Edit > Instrument Setup to open the Instrument Setup dialog
box.

If you are adding a new light conversion screen (refer to UV/White Light
Conversion Screen on page 211), select the appropriate box in the
Illumination Options field.

If you are adding additional filters, select the appropriate box in the Filter
Options field.

For any other changes to the optical pathway, perform a flat field calibration by
clicking Reset in the Instrument Calibration > Flat Field panel and following the
on screen instructions.

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Recalibrating Your Imager

2.

The software prompts you to restart the calibrations needed for the new
illumination sources.

3.

Click OK to exit the dialog box. Your settings persist until you make further
changes.

Note: If the Gel Doc XR+ and ChemiDoc XRS+ systems are used to conduct
quantitation with Bio-Rads stain-free gel technology, Bio-Rad recommends
performing the flat field calibration with the orange fluorescence reference plate
(catalog # 170-8008) after installing Image Lab 5.1. For instructions on
performing this calibration, see Calibrating the Imager to Use the Orange
Fluorescence Reference Plate on page 218.

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12 Image Lab Logs

Image Lab Logs
Image Lab software provides three types of logs.

Instrument log records events related to the instrument, including

calibrating the instrument and the success or failure of the calibration. This
log file is visible only if the computer running Image Lab is connected to an
instrument.

System log records events related to running Image Lab, including

enabling or disabling of secure mode and the users who log on to or log off
of Image Lab.

Document log (Security Edition only) records events related to the

creating and modifying of secure protocol and image files.

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Image Lab Logs

Viewing the Instrument Log

Instrument operations that you perform with Image Lab are recorded in the
instrument log.

To open the Instrument Log Viewer

From the View menu, click the log for your instrument.

The menu displays the specific instrument to which you are connected. The
GS-900 densitometer is shown as an example, above.
Note: You can view an instrument log only if Image Lab is connected to the
specified instrument.
The instrument log includes the following information.

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date and time of the action

the user name of the user logged into the computer at the time of the
action

the users security level

the domain where the current user is logged in

the type of action performed

the result of the action taken, including complete records of the calibration

the users reason for the action, if provided

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Image Lab Logs

Viewing the System Log

Events related to running Image Lab, including enabling or disabling of secure mode
and users logging on to or logging off from Image Lab, are recorded in the system
log.

To open the System Log Viewer

From the View menu, click View System Log.

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Image Lab Logs

Viewing the Document Log

The View menu displays a list of logs for each open document. The document log
captures information about creating and editing the Image Lab protocol and image
files.
Note: Document logs are viewable only in Image Lab Security Edition, and only
on Windows-based computers. Document logs are not viewable on the Mac.
The document log captures changes from:

Image tools

Lane and bands tools

Normalization tools

Molecular weight tools

Quantity tools

Volume tools

You can view the document log for any open file. This file can be a previously saved
file, or it can be a newly created protocol or image file that is open on your desktop
but which has not yet been saved.
If the document was previously saved, the log is identified with this name. If the
document has not been saved, the log is identified with the time stamp, showing
when the document was created.

Log file for saved image files

Log file for open but unsaved image file

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Image Lab Logs

To open the Document Log Viewer

From the View menu, click the document log that you want to view. A typical
document log is shown below.

Displaying Log Data

The display toolbar is located above the log.

Displaying Data Columns in Logs

By default, the logs display the following columns:

Date and time

User

Level the security role of the user

Domain domain where the current user is logged in

Type the type of event

Description the actual event captured

Reason (For Secure Mode only) the reason for signing a document

You can change your view of any log by displaying or hiding data columns.

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Image Lab Logs

To display or hide columns

1.

From the View menu, open the log file.

2.

Click the Display log viewer options icon.

3.

In the Display Column Options dialog box, use the arrow keys to move columns
between the Not Displayed and Displayed lists.

4.

Click OK.

Filtering Data in Logs

For all Image Lab logs, you can filter the entries in the following columns:

Date and time

User

Type

For example, you can set the filter in the Type column to File and Image Lab will
display only the rows where Type is equal to File.

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Image Lab Logs

Setting filters in logs

To filter the entries of a column
1.

In the log, click the heading of the column that you want to filter. The Display
Filter options icon in the Actions tool bar is enabled. (By default, the log opens
with all event types displayed.)
Note: The Display Filter options icon is enabled only when you click on a
column that can be filtered. The Display Filter options icon remains
disabled if you click a column that cannot be filtered.

2.

Click the Display Filter options icon to display the Select filter values dialog box.
Tip: Alternatively, right-click the heading of a filterable column to display
the Select filter values dialog box.

3.

Select a value and click OK.

Tip: You can select multiple values to filter.

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Image Lab Logs

Removing filters in logs

The Remove all filters icon is enabled only after you set a filter on a column. This
icon clears all filters on all columns.

Each filterable column has a Remove filters icon as well. This icon removes the filter
for that column only.

Collapsing or Expanding Data Rows

You can expand the size of rows in any log to display the full content of the row, or
you can collapse the row size to display more rows in the table. The icon toggles
between collapse and expand.

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Image Lab Logs

To display the full content of a row

Click the Collapse or expand rows icon to expand the row height. The row
adjusts to fit the text of the longest entry.
This action expands all rows in the log where an entry is longer than the width
of the column. Rows that do not need to wrap are not affected.

To display more rows in the table

Click the Collapse or expand rows icon to collapse the row height to the default
height setting.
This action collapses all rows in the log to display data on a single line.

Exporting Logs
From the log viewer, you can:

Copy log entries to the clipboard copies the log entries to the
clipboard, enabling you to paste them into a word processing or
presentation application.
Tip: You can also open a log in the log viewer and press Ctrl+C to copy
the log entries to the clipboard.

Export log entries to a file exports the log entries as a CSV file that can
be opened in a database application.

Export log entries to Excel exports the log entries to an Excel file
where you can use Excels sorting and formula functions to manipulate
your data. If Excel is not installed on your computer, this feature is
disabled.

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Image Lab Logs

To export a log file

1.

From the View menu, click the log you want to view and export.

2.

Click one of the icons to export the data in one of the supported formats.
Copy log entries to clipboard
Export log entries to File
Export log entries to Excel

Printing Logs
Log data can be sent to a printer or saved to a PDF file.

To print a log

176

1.

From the View menu, click the log you want to view and print.

2.

In the log viewer, click Print log.

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Image Lab Logs

The Log Print Preview window displays the contents of the log file.

From the Log Print Preview window, you can:

Click Print log to print the log to a printer.

Click Print log to PDF to save the log to a PDF file.

Click Adjust printer settings to prepare the file for printing.

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Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

13 Using the Security Edition

21 CFR Part 11
Image Lab Security Edition is a module within the Bio-Rad Image Lab software
that assists users in meeting the Food and Drug Administrations regulations on
good lab practices in the pharmaceutical and biotechnology industries. The Security
Edition enables system administrators to ensure that Image Lab operates in
compliance with Title 21 of the Code of Federal Regulations (CFR) Part 11 within a
closed system. A closed system is defined as an environment in which system
access is controlled by the persons who are responsible for the content of electronic
records that are on the system (Section 11.3 (b) (4).
Note:

Image Lab Security Edition is not supported by the Mac.

The security controls built into Image Lab Security Edition must be properly
configured and administered by the system administrator(s) in your
organization in order to be secure and in compliance with 21 CFR Part 11.

Bio-Rad makes no claim that Image Lab Security Edition software is

CFR-compliant in and of itself, nor does the company guarantee
compliance for the user. Your organization must establish policies and
standard operating procedures that work in conjunction with the tools
provided by Bio-Rad to ensure compliance with 21 CFR Part 11.

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Using the Security Edition

Standard Mode versus Secure Mode

Image Lab Security Edition can be run in either of the following modes:

Standard Mode in this mode, there are no restrictions on controlling the

instrument, operating the software, or changing the documents.

Secure Mode in this mode, in addition to the features available in

standard mode, extensive security features are enabled, including
document signing and log creation.
When Image Lab is in secure mode, a padlock symbol appears in the left corner
of the status bar. If no padlock is present, the software is running in standard
mode.

Lock indicates secure

mode is enabled

No lock indicates standard

mode is enabled

When Image Lab is installed, by default it is set to run in standard mode. It continues
to run in this mode until a user with Image Lab Administrator privileges enables
secure mode.
This chapter assumes you are running the application in secure mode unless
otherwise noted.

User Names, Groups, and Roles

When Image Lab Security Edition is run in secure mode, you must log in with a user
name and password in order to run the application. The Microsoft Windows system
administrator generally creates the user names and passwords. The Image Lab
administrator defines the groups to which the Security Edition roles will be
associated.
Note: See Setting Up Users and Groups on page 195 for additional information
about setting up groups, user names, and passwords.

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User Names, Groups, and Roles

There are four default roles in Image Lab Security Edition. Each role is associated
with one of the four default Image Lab user groups, and each user is assigned a role
that provides the user access to specific features in the software. Table 11 lists the
default Image Lab Security Edition user groups and the corresponding roles. It also
provides a brief description of the permissions for each role.
Note: System administrators can change the user group names, if necessary,
to meet their local company standards.
Table 11. Image Lab Security Edition groups and roles
User Group

Image Lab Role

Description

TDS_Administrator

Administrator

Users with this role can enable or

disable secure mode.
Administrator users can also
view log files, but they do not
have access to any other
features.

TDS_User

Supervisor

Users with this role have full

access to all features and
functions of the application. They
can also sign files. Supervisors
cannot enable or disable secure
mode.

TDS_Tech

Clinician

Users with this role can perform

instrument operations, run
existing protocols, and view
protocol files, results files, and
log files. They can also sign files.
All other access is restricted. For
example, they cannot create or
edit protocols.

TDS_Guest

Reviewer

Users with this role can open

and view protocol files, results
files, and log files. They can also
sign files. Users with this role
cannot create new protocols and
cannot change protocols. All
other access is restricted.

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Using the Security Edition

Role Restrictions
Your role determines which features of the security edition you have permission to
use. If you attempt to perform an action that is not permitted for a user in your role,
you will see an error message. In some instances the users role determines which
Security Edition features are visible and/or enabled. Therefore, you might not see all
of the features described in this chapter.
Table 12 lists the Image Lab Security Edition functions that each role has permission
to perform.
Table 12. User access to features by role
Function

Administrator

Enable/disable secure mode

View log files

Supervisor

Clinician

Reviewer

Set up and recalibrate the

instrument

Create new protocols

Open existing protocols

Run protocols

Edit protocols

Create new images

Open existing images

Edit images

Analyze images

Sign documents

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Starting Image Lab Security Edition

Starting Image Lab Security Edition

When secure mode is enabled, you are prompted to log on when you start the
Image Lab application.

To start Image Lab in secure mode

1.

Click the Image Lab icon to start the application.

2.

In the Log on to Image Lab dialog box, enter your user name and password.

3.

Click OK.

Note: If you have any questions or problems logging on, see your system
administrator.

Electronic Records
Image Lab Security Edition enables you to create secure electronic records as
defined by 21 CFR Part 11. In Image Lab, the following are electronic records:

Protocol files

Image files

Document log files

Instrument log files

System log files

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Using the Security Edition

Unsecured Documents
Protocol and image files created in standard mode or created with previous versions
of Image Lab are unsigned, unsecure documents. Unsecure documents remain
unsecure. When you open these files in secure mode, you can make changes to
these files and save them without restrictions. A document log is generated for
changes to these files, but the log is not viewable in standard mode. You must log in
to secure mode to see the log.
You can also open unsecure documents in secure mode and sign them. You can
make changes to these files and save them as secure documents. The original
unsecure document remains unsigned and unsecure. The new document is saved
as read-only with an incremental revision number and Image Lab generates an audit
log. Image Lab will not be able to overwrite the secure file, but it can overwrite the
original unsecure file.

Secure Documents
Protocol and image files created in secure mode are known as secure documents
and generate a log. However, until they are signed, such documents save as
standard files. Only after they are signed are they saved with the secure
extension(.sptl or .sscn). Image files generated from signed protocols automatically
open the signing dialog box when the scan completes.
If you create a default protocol based on an existing signed protocol, the resulting
new default protocol cannot be edited.
Note: A secure document can be saved unsigned. Likewise, a signed
document can be saved as a new, unsigned file. In this case, the saved file is
still a secure document. All changes to the document are captured in the
document log.

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Electronic Records

Documents created in secure mode can be signed at any time. When a secure
document is signed, it is saved as read-only. Image Lab cannot overwrite the
document. You can open signed documents and sign them again. The newly signed
file is saved as a second revision, and the new signatures are captured in the
document log.
Note: Image Lab can never overwrite any signed file.
You can open secure documents in standard mode of Image Lab. Selecting Save As
from the File menu creates an unsecure file. The original file is preserved. No log is
generated for the new, unsecure document. However, an entry is added to the
original log that the file has been reverted to unsecure.

Secure Documents in SAM mode

Images saved in signal accumulation mode (SAM) follow the same rules as the main
image. The rules apply when the image is saved from within a protocol. If you create
an image from a signed protocol, all resultant images must also be signed. If you try
to save a file more than once, a warning message appears asking you to confirm the
re-save. And if you confirm this re-save and click OK, Image Lab creates a new file
with an incremental revision number.

Modifying Secure Documents

You can open and change a signed document in Image Lab. The original document
is read-only and cannot be overwritten. Saving the revisions opens the Save As
dialog box. The changes are saved into a new revision with an incremental revision
number.
Each time a secure document is modified, you must provide a reason for each
change before you can sign the document. The modifications are logged in the
document log. The new signed document takes with it the entire history of the
original document in its log.

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Using the Security Edition

Signing Documents
To sign a document
1.

Select the protocol or results file.

2.

From the Security menu, click Sign Document. The Signing Document dialog
box appears.

3.

Enter the user name and password of a user authorized to sign documents.
Note: The user name and password can be for a user other than the
current user.

4.

Enter a reason for signing the document. Typical reasons include review,
approval, responsibility, or authorship.
Note: You must provide a reason in order to sign the document.
The user name, date and time of the signature, and reason for signing are
always included in the Signature History section of the image report (see
page 153).

5.

Click OK.
A Save File dialog box appears.

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6.

Enter a new name for the file and click Save.

Note: Signed protocols are saved with an .sptl extension, signed images
are saved with an .sscn extension.

Document Logs
Any changes made or actions performed on a protocol or image file generate a
document log documenting each change or action. This log is created as soon as
the protocol or scan file is created. Protocols are updated when the protocol is
saved or run. If you make changes to a file and save it as a new file, regardless of
whether you are in secure mode, the document log is preserved in the new file. If
you are in secure mode, the signature of the previous file is noted as part of the
document log.
Note: When a protocol is run, its log entries are copied to the resulting image
file.
All major actions and changes are audited (generate a document log). Examples of
auditable actions include:

Signing a file

Changing protocol settings

Changing the image, for example, by cropping or rotating it

Modifying the analysis, for example, changing the lanes and bands, or
adding or editing annotation to the images

Minor changes that affect only the display are not audited, such as:

Selecting different columns in the Analysis Table

Changing the display options in Lane Profile

Annotating or labeling the image using Annotation Tools

Each change you make to a signed protocol or image file must be documented in
the Reason for Change dialog box.

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Viewing the Document Log

The View menu displays a list of logs for each open document. The document log
captures information about creating and editing Image Lab protocol and image files.
You can view the document log for any open file. This file can be a previously saved
file, or it can be a newly created protocol or image file that is open on your desktop
but which has not yet been saved.
If the document was previously saved, the log is identified with this name. If the
document has not been saved, the log is identified with the time stamp of when the
document was created.

Log file for saved image file

Log file for open but unsaved image file

The document log includes:

Date and time

User

Level the security role of the user

Type the type of event

Description the actual event captured

Reason the reason for signing a document

For more information about logs, see Chapter 12, Image Lab Logs on page 167.

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A Maintenance
This chapter includes instructions for maintaining the universal hood in proper
working condition by replacing parts.

UV Transilluminator Lamp and Starter Replacement

Note: Keep the UV filter surface clean from the chemical agents used as gel
dyes. Use protective gloves when touching the UV transilluminator cover.
Depending on usage, the UV bulbs and starters last for many years. Replace bulbs
when you notice them flickering. If a bulb does not turn on when it is new or moved,
replace the bulb starter and test the bulb again.
There are three types of bulbs available. The catalog numbers are listed in Appendix
D, Accessories. The standard bulb is 302 nm. Optionally, the 254 nm bulb is used
for cross-linking of proteins and nucleic acids, and the 365 nm bulb is used to
minimize denaturing of DNA.

To replace the lamps

1.

Turn off the power.

2.

Disconnect the power cord from the universal hood.

3.

Remove the four screws located on the left/right sides of the transilluminator
cover.

4.

Remove the cover with the UV glass by sliding it forward, then lifting up.

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Maintenance

5.

Place it on a nonabrasive surface so that the glass does not get scratched or
damaged.
Note: Do not put the UV cover directly on the bench. Wear gloves when
touching the lamps.

6.

Rotate the lamp until it becomes loose and the pins come to a vertical position.

7.

Remove the lamp. Install the new lamp by rotating it so that the pins are
horizontal and the lamp is tight.

8.

Remove the starter by rotating it counterclockwise and then pulling it out.

9.

Insert the new starter into the holder and rotate clockwise.

10. .Reassemble the cover and retighten the screws on both sides.

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Epi-Illumination Lamp Replacement

The lamps are located behind two cover panels on the left and right sides of the
universal hoods interior.

To replace the lamps

1.

Turn system power off and remove the power cord.

2.

Open the enclosure door.

3.

Locate the epi-white light housing inside the hood and the screw on the center
of the housing cover.

4.

Use a socket wrench to remove the screw.

5.

Pull the cover to remove it from the housing and the lamp will become visible.

6.

To remove the lamp, hold it at the plastic receptacle and pull it from the
receptacle.

7.

Insert the new lamp into the lamp receptacle and push until it is firmly seated in
the receptacle.

8.

Reassemble the epi-white light cover.

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Maintenance

Fuse Replacement
Caution: Always unplug the instrument before changing or checking the fuses.

Power outlet

Fuse holder

This unit is protected by two fuses (5 x 20 mm, 2A Slo-Blo). The fuses are located in
the left side rear of the universal hood, in a fuse holder housed in the power entry
module.

To replace the fuses

192 |

1.

Unplug the main power cable from the power outlet.

2.

Use a screwdriver as a lever on the fuse holder to extract the fuse.

3.

Remove the blown fuses and replace them with two new fuses
(catalog #900-8935).

4.

Slide the fuse holder into the power entry module until it snaps into place.

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

B Troubleshooting
Follow these suggestions to troubleshoot your system.
Problem

Possible Cause

Solution

Camera does not

respond/camera not
found

Power to the camera

may be turned off.

Turn on the power to the camera.

The camera cables may

not be seated properly.

Make sure that all cables are

connected as shown in the
Installation Guide.

The software driver for

the camera is missing.

If the camera driver is not present,

reload the camera driver from the
Image Lab software CD.

Computer power-saving
modes may be
interfering with the
camera driver.

Disable the power-saving modes on

the computer.

The cables may be

defective.

Replace the cables.

The camera may be

defective.

Replace the camera.

Horizontal stripes in
image when using
the UV mode

The emission filter may

not be positioned
properly.

Move the filter lever so that the filter

slider positions the filter under the
camera lens.

Image is not visible

on the monitor

The monitor settings are

incorrect.

See your computer manual for the

proper settings.

The lens cap is

attached.

Remove the lens cap.

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| Troubleshooting

Problem

Possible Cause

Solution

Printout does not

look like the monitor
image

The monitor settings are

wrong.

See your monitor manual for the

appropriate settings.

The printer settings are

wrong.

See your printer manual for the

appropriate settings.

Light leakage into the

darkroom

The lens body is not

seated properly against
the gasket on the hoods
adapter plate.

Loosen the thumbscrew and seat the

lens properly against the gasket on
the hoods adapter plate.

Unable to focus on
the sample using
white light
transilluminator or
conversion screen

Focus is not calibrated

for samples using this
light source.

Select Edit > Instrument Setup to

recalibrate the focus for use with this
accessory.

Lens limits seem

artificially restricted

The camera lens is not

seated properly on the
lens mounting plate.

Re-seat the camera on the lens

mounting plate.

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C Setting Up Users and

Groups
Setting Up Image Lab Users and Groups
This appendix explains how to set up users and groups to run Image Lab Security
Edition software in secure mode.
Note: This task requires System Administrator privileges on the client
computers (and possibly their domain) on which Image Lab is installed.

User Accounts
To give users access to Image Lab Security Edition, you can create new Windows
user accounts, add existing user accounts to the four default user groups specified
in Table 11 on page 181, or rename any of the four default user groups.

A user account can have any name, but must have a password defined for
the account. See the section on Password Security, on page 206 for
information about setting passwords for maximum security.

Each user can belong to the Image Lab Administrator group and one other
Image Lab user group.
For example, a user can belong to the Administrator group and the Supervisor
group, but a user cannot belong to both the Clinician group and the Supervisor
group.

User Authentication and Group Membership

Authentication in Image Lab is made up of two processes: user authentication and
group membership evaluation.

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User Authentication
In the user authentication process, Image Lab matches (authenticates) a user name
with permissions assigned to that user name on the authentication domain. That
domain can reside on your local computer (a local domain) or on a network server (a
network domain). You set the location of this domain in the Domain to be used for
authentication field of the Security Preferences dialog box. This name must be the
exact name of your local computer or network server. For details on how to find this
name see Finding the name of your authentication domain, on page 197.

Enter the name of your local

computer, or the name of the
network server used for your
authentication domain.

In the Security Preferences dialog box, if you (or your network administrator) choose
a local domain to be used for authentication, you are considered a local user. If you
or your network administrator choose a network domain, you are considered a
domain user.

Group Membership Evaluation

In the group membership evaluation process, Image Lab verifies that a user is a
member of one or more of the four default Image Lab user groups
(TDS_Administrator, TDS_User, TDS_Tech, or TDS_Guest). The valid members for
each of these groups can be specified in one of two places, as defined by the Use
local groups for establishing user security levels checkbox on the Security
Preferences dialog box.

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This checkbox specifies whether

valid users and groups are defined
on (1) the local Image Lab
computer, or (2) on the network
domain.

If this checkbox is selected, only users and groups that are defined on the local
computer (on which Image Lab is installed) are recognized. If this checkbox is not
selected, only users and groups that are defined on the network domain are
recognized.

Finding the name of your authentication domain

Your authentication domain can be hosted on your local computer (a local domain)
or on a network server (a network domain).

To find the name of your local domain

1.

From your Start menu, open the Control Panel.

2.

Click System.

The System window appears. In the Computer name, domain, and workgroup
settings section, the name of your local computer is shown as Computer name.

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Setting Up Users and Groups

To find the name of your network domain

1.

From your Start menu, open the Control Panel.

2.

Click System.

The System window appears. In the Computer name, domain, and workgroup
settings section, the name of your network domain is shown as Domain.

Configuring Users and Groups on a Local Computer

To set up users and groups on a local computer

198

1.

In the Windows Control Panel, select Administrative Tools.

2.

Select Computer Management.

3.

In the Computer Management window, expand the System Tools folder, and
then expand the Local Users and Groups folder.

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Setting Up Image Lab Users and Groups

To create a new user on a local computer

1.

Click on the Users folder to open it and select Action > New User. Alternatively,
use the right-click context menu.
The New User dialog box appears.

2.

Fill out all the fields:

User name the user name must be unique.

Full name the full name must be unique.

Bio-Rad recommends using the users actual full name, as this name will
be shown in the document log and all the log reports. This is a requirement
of 21 CFR 11.50a.

Description this field must also be filled out.

Bio-Rad recommends entering the users title as the description.

Password enter and confirm a password for the user.

Tip: Select the User must change password at next logon checkbox.
This prevents the Windows system administrator from knowing the
passwords of the users.
Note: If you select the User must change password at next logon
checkbox, the user must actually log on to Windows and change the
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Setting Up Users and Groups

password before using Image Lab Security Edition. Otherwise,

Security Edition will not recognize the user.

To create a new group on a local computer

1.

In the Computer Management window, click on the Groups folder to open it and
select Action > New Group. Alternatively, use the right-click context menu.
The New Group dialog box appears.

2.

In the Group name field, enter the name of one of the groups specified in
Table 11 on page 181 (TDS_Administrator, TDS_User, TDS_Tech, TDS_Guest).
You can also enter a description in the Description field.
The group does not need any special operating-system level privileges.

3.

Click Create to create and save the new group.

4.

Repeat this task for the remaining groups specified in Table 11 on page 181.

To add a user to a group on a local computer

1.

In the New Group dialog box, click Add. Alternatively, double-click an existing
group in the Groups folder to open its Properties dialog box, and click Add.
You see the Select Users dialog box.

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2.

Click Advanced to expand the dialog box.

3.

In the expanded dialog box, click Find Now to populate the bottom field with all
the users on the local computer.

4.

Click a user name in the list to select it, or press the Ctrl key and click multiple
users to select them.

5.

When you have selected all the users to add to the group, click OK, and click
OK again to close the Select Users dialog box.

6.

Click Create to close the New Group dialog box and create the group.
Alternatively, you can click OK to close the existing groups Properties dialog
box and accept the changes.

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Setting Up Users and Groups

Configuring Users and Groups on a Network Domain

Note: The network administrator must know how users and groups are set up
using the Windows server software at your site. The following example is used
to illustrate the choices.

To locate the users and groups on a Windows server

Open Administrative Tools and select Active Directory.

Note that in the Active Directory window, the Users folder lists groups as well.

To create a new user on a Windows server

1.

With the Users folder open, select Action > New User. Alternatively, use the
right-click context menu.
You see the New User dialog box.

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Setting Up Image Lab Users and Groups

2.

Fill out all the fields:

User name The user name must be unique.

Full name The full name must be unique.

Bio-Rad recommends using the users actual full name, as this name will be
shown in the document log and all the log reports. This is a requirement of
21 CFR 11.50a.

Description This field must also be filled out.

Bio-Rad recommends that you enter the users title as the description.

Password Enter and confirm a password for the user.

Tip: Select the User must change password at next logon checkbox. This
prevents the Windows system administrator from knowing the users
passwords.
Note: If you select the User must change password at next logon
checkbox, the user must actually log on to Windows and change their
password before using Image Lab Security Edition. Otherwise, Security
Edition will not recognize the user.

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Setting Up Users and Groups

To create a new group on a Windows server

1.

With the Users folder open, select Action > New Group. Alternatively, you can
use the right-click menu.
You see the New Object - Group dialog box.

2.

In the Group name field, enter one of the group names specified in Table 11 on
page 181 (TDS_Administrator, TDS_User, TDS_Tech, TDS_Guest).
Type the name exactly as specified. You can also enter a description for the
group in the Description field.
Note: The group does not need to have any special operating-system
level privileges.

To add a user to a group on a Windows server

1.

In the New Group dialog box, click Add. Alternatively, double-click an existing
group in the User Manager folder to open its Properties dialog box and click
Add.
You see the Select Users dialog box.

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Setting Up Image Lab Users and Groups

2.

Click Advanced to expand the dialog box.

3.

In the expanded dialog box, click Find Now to populate the bottom field with all
the users.

4.

Click a user name in the list to select it, or press the Ctrl key and click multiple
users to select them.

5.

When you have selected all the users to add to the group, click OK, and click
OK again to close the Select Users dialog box.

6.

Click Create to close the New Group dialog box and create the group, or click
OK to close the existing groups Properties dialog box and accept the changes.

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Setting Up Users and Groups

Password Security
21 CFR 11.300 (b) requires that passwords be periodically checked, recalled, or
revised. Password policies are therefore recommended, although the password
duration and rules are up to the system administrator and the organization. For
instance, the exact duration between password changes is flexible.

To set password policies on a local computer

1.

Open the Control Panel and select Administrative Tools > Local Security Policy.

2.

In the left pane of the Local Security Policy window, expand Account Policies
and then select Password Policy.
You see the password policies listed in the right pane of the window.

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Password Security

3.

4.

To change password policy settings:

a.

Right-click the policy and select Properties to open its properties dialog
box.

b.

Modify the default setting to meet your company policy.

c.

Click Apply.

d.

Click OK to close the Properties dialog box.

Close the Local Security Policy window.

To set password policies in Active Directory

1.

Open the Control Panel and select Administrative Tools > Domain Controller
Security Policy.

2.

In the left pane of the Domain Controller Security Policy window, expand
Security Settings > Account Policies and select Password Policy.

3.

To change password policy settings:

4.

a.

Right-click the policy and select Properties to open its properties dialog
box.

b.

Modify the default setting to meet your company policy.

c.

Click Apply.

d.

Click OK to close the Properties dialog box.

Close the Domain Controller Security Policy window.

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Password Policy Setting Examples

The following examples are only suggestions. Your organization needs to establish
its own password policy.

Enforce password history: 12 passwords remembered

Minimum password age: 5 days

Maximum password age: 30 days

Minimum password length: 8 characters

Password must meet complexity requirements: Enabled

Store passwords using reverse encryption: Enabled

Account Lockout Policy Setting Examples

Account lockout duration: 0 (The account is locked out until the

administrator unlocks it.)

Account lockout threshold: 3 logon attempts

Reset account lockout counter after: 30 minutes

Auditing Windows Event Logs

Some global auditing information is stored in the Windows Event logs. It is a
requirement of 21 CFR Part 11 that these logs be archived. However, by default,
Windows systems automatically remove these data without warning.
Note: It is therefore critical that the event log is reconfigured to generate and
preserve all necessary log data. Regular manual intervention is also required to
preserve these data.

To open the Event Properties Log

208

1.

Open Administrative Tools and click Event Viewer.

2.

Right-click on each log and select Properties.

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Miscellaneous Security Measures

3.

In the section When maximum event log size is reached: select Do not overwrite
events (Clear logs manually).

4.

Increase the maximum size of the event log to cover any possible messages.
The smaller the maximum size of the event log, the more often the system
administrator must manually view, archive, and clear the system log.

Auditing information generated by the operating system is recorded in the Security

Log. Logon failures in Image Lab Security Edition are recorded in this log.
During the review process, the log should be examined for attempted breaches of
security, such as a series of failed logon attempts. To avoid the risk of losing data,
the size should be very large and this inspection/archive process should occur daily.
The Audit Policy should be set as follows:

Audit account logon events Failure should be checked at a minimum

Audit account management both Success and Failure should be

checked

Audit logon events Failure should be checked at a minimum

Audit policy change both Success and Failure should be checked

Miscellaneous Security Measures

Bio-Rad recommends that you use the built-in protections that Windows Server
offers in order to protect the computer while the user is absent.
Note: Microsoft is continually updating its operating systems in response to
security issues. It is critical to keep all components of the Windows operating
system, especially any domain controllers, up to date.

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Setting Up Users and Groups

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

D Accessories
Calibrating Accessories
If you are installing accessories along with your original system installation, calibrate
your system with a one-time Instrument Calibration wizard. Complete instructions
are in the installation guide that arrives with your imager. If you acquire new
conversion screens, light sources, or filters after your original system installation,
you will have to recalibrate your imager to use them.

Installing Optional Accessories

UV/White Light Conversion Screen
This optional white light conversion screen (catalog #170-8289) converts the UV
light generated in the universal hood to white light. Your imager must be calibrated
to use the white light conversion screen.

To use the white light conversion screen

1.

Center the conversion screen on the imager stage.

2.

Center your samples on top of the conversion screen.

3.

Image the gel using your preferred application

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| Accessories

XcitaBlue Conversion Screen

The optional XcitaBlue conversion screen kit (catalog #170-8182) converts UV to
blue light, which enables you to visualize DNA samples while protecting them
against UV damage.

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The XcitaBlue conversion screen is held in place by adhesive-backed edge guides.

After the edge guides are installed, the conversion screen remains centered and will
not slide, even if you close the drawer rapidly.

Adhesive-backed
edge guide
Transilluminator
drawer front

Transilluminator
border edge

To install the XcitaBlue conversion screen

1.

Perform a trial placement first, without removing the paper tape. Place the edge
guides in each corner of your transilluminator, as shown. The edge guides
should touch the inside of the drawer front and fit over the edge of the metal
transilluminator border (shown in red).

2.

Remove the paper tape from the bottom surface of each edge guide.

3.

Press each edge guide into position carefully. After the adhesive surfaces
touch, it is difficult to reposition the guide.

4.

Calibrate your imager to use this accessory by going to Edit > Instrument
Setup. Select the XcitaBlue Conversion Screen checkbox under Illumination
Options. The software prompts you to calibrate the focus with height offset.

5.

To visualize a sample using the XcitaBlue conversion screen, place the screen
between the edge guides.

6.

Center the gel on top of the XcitaBlue conversion screen, and proceed with
normal image capture. Use the gel alignment template kit to center your gels
easily and consistently.
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| Accessories

Gel Alignment Template Kit

The Bio-Rad gel alignment template kit (catalog #170-8184) allows four sizes of
standard agarose gels to be centered quickly and easily and ensures the consistent
placement of each gel.

Magnetic
locator
frame

7 x 7 cm

15 x 7 cm

7 x 10 cm
15 x 10 cm

The kit contains:

Magnetic locator frame

Instruction sheet

Alignment guides for the following gel trays:

Sub-Cell GT UV-transparent mini-gel tray, 7 x 7 cm

Sub-Cell GT UV-transparent wide mini-gel tray, 15 x 7 cm

Sub-Cell GT UV-transparent mini-gel tray, 7 x 10 cm

Sub-Cell GT UV-transparent gel tray, 15 x 10 cm

The gel alignment templates fit exactly into the XcitaBlue conversion screen frame
(catalog #170-8182).

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Installing Optional Accessories

To install and use the gel alignment template kit

1.

Place the locator frame over the transilluminator with the magnetic side down.
Match the corners of the magnetic locator frame with the edges of the
transilluminator. The UV symbol on the frame will be in the same orientation as
the UV symbol on the imager.

2.

Place the gel alignment template that matches the size of your sample tray or
agarose gel into the magnetic locator frame.

3.

Place your gel or gel tray into the open area of the template.

Note: No recalibration is necessary to use the gel alignment template kit.

White Light Transilluminator

The white light transilluminator provides a white light that is independent of the UV
bulbs. Between uses, this transilluminator can be stored in the rear of the universal
hood.

To install the white light transilluminator

1.

Open the enclosure door and pull the drawer toward you.

2.

Locate the power cable positioned on the inside left side of the enclosure,
behind the drawer slide.

3.

Remove the black rubber boot that covers the connector plug and insert it into
the white light transilluminator. Make sure that the main power switch on the
transilluminator is in the ON position.
Note: Power to the white light transilluminator is controlled from either the
software or the membrane touch pad on the universal hood.

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D | Accessories

4.

Place the white light transilluminator on top of the UV transilluminator.

Your system must be calibrated to use the white light transilluminator.

To calibrate the system to use the white light transilluminator

Choose Edit > Instrument Setup and select the appropriate checkbox in the
dialog box. The Image Lab software will guide you through the calibration.

Orange Fluorescence Reference Plate

The orange fluorescence reference plate enables you to apply UV flat fielding
corrections to your imager. Corrections are made for all UV illumination sources,
filters, and the camera lens.
The orange fluorescence reference plate is recommended for quantitation with
Bio-Rads stain-free gels, and can be used to correct for image nonuniformities for
red and orange gels, including:

216

Ethidium bromide

GelRed

Flamingo

Coomassie Fluor Orange

SYPRO Ruby

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Installing Optional Accessories

Krypton

Qdot 625

The orange fluorescence reference plate (catalog #170-8008) can be used with
several of the Bio-Rad Molecular Imager series of products, including:

Gel Doc XR+ imaging system with Image Lab software

ChemiDoc XRS+ imaging system with Image Lab software

ChemiDoc XRS+ imaging system with Quantity One software

When you order the reference plate, you receive:

Orange fluorescence reference plate, overall dimensions: 29.5 x 29.5 cm;

viewing surface: 27 x 27 cm

Instruction sheet (10017296)

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D | Accessories

Calibrating the Imager to Use the Orange Fluorescence Reference

Plate
Your imager must be calibrated to use the orange fluorescence reference plate.

To calibrate the imager to use the orange fluorescence reference plate

Choose Edit > Instrument Setup and select the appropriate checkbox in the
dialog box. Image Lab software guides you through the calibration.

Ordering Information
The following table contains catalog numbers and descriptions for all parts available
for the Gel Doc XR+ or ChemiDoc XRS+ systems, plus all optional accessories and
replacement parts. For complete information, see the Bio-Rad catalog.
Catalog #

Description

Molecular Imager System (includes universal hood, camera, cables, and

accessories)
170-8195

218

Gel Doc XR+ with Image Lab software system for PC/Mac

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Ordering Information

Catalog #

Description

170-8265

ChemiDoc XRS+ with Image Lab software system for PC/Mac

Installation Kits
170-8199

Image Lab Upgrade for Gel Doc XR+ System

170-8299

Image Lab Upgrade for ChemiDoc XRS+System

Image Lab Software

170-9690

Image Lab software, Windows/Mac

Optional Filters
170-8074

Filter, 520DF30, GFP

170-8075

Filter, 560DF50, Cy3/rhodamine

170-8076

Filter, 630BP30, SYPRO Ruby

Optional Accessories
170-8289

UV/White light conversion screen (UV to white light)

170-7950

White light transilluminator (universal hood plug-in)

170-8182

XcitaBlue (UV to blue light) conversion screen kit, without standard

detection filter

170-8008

Orange fluorescence reference plate

170-8089

Mitsubishi thermal printer, 100/240 V, USB

170-3759

Bio-Rad fluorescent ruler

170-3760

Gel cutter ruler

170-8184

Gel alignment template kit

Replacement Parts
170-8026

Focus calibration target

170-8027

Image Lab flat fielding disc

170-8185

XcitaBlue viewing goggles

170-7581

Mitsubishi thermal printer paper, 4 rolls

100-2784

UV transilluminator lid (includes UV filter glass)

1001-4106

Thumbscrew for camera

User Guide |

219

D | Accessories

Catalog #

Description

170-8081

Standard emission filter replacement, 62 mm

100-1370

UV bulb starter, quantity 3

800-0142

Multicolor target

Lamps
100-2827

Lamp, epi-illumination, 5 W

170-8097

302 nm lamp kit, (6 lamps)

170-6887

365 nm lamp kit, (6 lamps)

Fuses
900-8935

Fuse T 2 A, 250 V, quantity 10

190-0234

Fuse T 4 A, 250 V, quantity 2

Optional Analysis Software

170-9600

Quantity One software

Protein Standards

220

161-0363

Precision Plus Protein Unstained Standards (single vial)

161-0396

Precision Plus Protein Unstained Standards (5 pack)

161-0373

Precision Plus Protein All Blue Standards (single vial)

161-0393

Precision Plus Protein All Blue Standards (5 pack)

161-0374

Precision Plus Protein Dual Color Standards (single vial)

161-0394

Precision Plus Protein Dual Color Standards (5 pack)

161-0377

Precision Plus Protein Dual Xtra Standards (single vial)

161-0397

Precision Plus Protein Dual Xtra Standards (5 pack)

161-0375

Precision Plus Protein Kaleidoscope Standards (single vial)

161-0395

Precision Plus Protein Kaleidoscope Standards (5 pack)

161-0385

Precision Plus Protein WesternC pack (single vial)

161-0398

Precision Plus Protein WesternC pack (5 pack)

161-0318

Prestained SDS-PAGE standards, broad range

161-0317

Unstained SDS-PAGE standards, broad range

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Ordering Information

Catalog #

Description

Nucleic Acid Standards

170-8351

EZ Load 20 base pairs molecular ruler

170-8352

EZ Load 100 base pairs molecular ruler

170-8353

EZ Load 100 base pairs PCR molecular ruler

170-8354

EZ Load 500 base pairs molecular ruler

170-8355

EZ Load 1 kb molecular ruler

170-8205

2.5 kb molecular ruler

170-8200

AmpliSize molecular ruler

170-8356

EZ Load precision molecular mass ruler (base pairs/ng of sample)

Pulsed Field Standards and Markers

170-3624

CHEF DNA size standard, 5 kb ladder

170-3707

CHEF DNA size standard, 8.348.5 kb

170-3635

CHEF DNA size standard, lambda ladder

170-3605

CHEF DNA size marker, 0.22.2 Mb

170-3667

CHEF DNA size marker, 13.1 Mb

170-3633

CHEF DNA size marker, 3.55.7 Mb

User Guide |

221

D | Accessories

222

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

E Using Bio-Rads
Stain-Free Technology
Bio-Rads stain-free gels eliminate the time-consuming staining and destaining
steps required by other protein detection methods. Stain-free gels include
unique trihalo compounds that allow rapid fluorescent detection of proteins with
the Gel Doc XR+ and ChemiDoc XRS+ imagers without staining.
When using Image Lab software version 5.1 or higher, the Gel Doc XR+ and
ChemiDoc XRS+ imagers are stain-free enabled to image these gels:

Criterion TGX Stain-Free precast gels

Criterion Stain Free precast gels

Mini-PROTEAN TGX Stain-Free precast gels

TGX Stain-Free FastCast acrylamide solutions for handcast gels

When trihalo compounds in the gels encounter tryptophan residues, a UV

lightinduced reaction produces fluorescence, which can be easily detected by
the Gel Doc XR+ and ChemiDoc XRS+ imagers within gels or on low
fluorescence polyvinyl difluoride (PVDF) membranes. Activation of the trihalo
compounds in the gels adds 58 Da moieties to available tryptophan residues and
is required for protein visualization. Proteins that do not contain tryptophan
residues cannot be detected using this technology. The sensitivity of stain-free
gels is comparable to staining with Coomassie Brilliant Blue for proteins with a
tryptophan content >1.5%; sensitivity superior to Coomassie staining is possible
for proteins with a tryptophan content >3%.

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E |

Using Bio-Rads Stain-Free Technology

The benefits of stain-free technology include:

Elimination of staining and destaining steps for faster time to results

No background variability within a gel or between gels (as is often seen

with standard Coomassie staining)

Elimination of the need for acetic acid and methanol in staining and
destaining, which reduces organic waste

Visualization of transferred or blotted proteins on low fluorescence PVDF

membranes

Stain-Free Workflow
For detailed information about the Activate/image gels step, refer to Acquiring
Images on page 69. For all other workflow steps, refer to the Criterion Precast
Gels Instruction Manual and Application Guide (bulletin #4110001) or to the
Mini-PROTEAN Precast Gels Instruction Manual and Application Guide
(bulletin #1658100).

Prepare buffers
Prepare gels and assemble
electrophoresis cell

Prepare and load samples

Perform electrophoresis

Activate/image gels

Analyze the separation

224

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Electrophoresis with Stain-Free Gels

Electrophoresis with Stain-Free Gels

Stain-free gels are made and packaged without sodium dodecyl sulfate (SDS),
allowing them to be used for both SDS and native polyacrylamide gel
electrophoresis (PAGE) applications.

To perform electrophoresis with stain-free gels

1.

Prepare the sample and running buffers.

2.

Set up the electrophoresis cell.

3.

Perform the run.

Imaging Gels
Use unstained standards with stain-free gels, as some prestained standards are not
compatible with stain-free technology. To monitor electrophoresis, use a 1:1 mixture
of unstained and prestained standards.
Setting up a protocol for stain-free gels is very similar to setting up protocols for
other applications. Follow the instructions in Setting Up a New Protocol on page 72.
Choose one of the following activation times based on your sample and the purpose
of your experiment:

Gels used in blotting use 1 min activation for optimal results when
performing western blotting followed by immunodetection.

Good sensitivity use 2.5 min activation when samples are abundant
and when a fully optimized signal-to-noise ratio is not necessary.

Best sensitivity use 5.0 min activation for detection of proteins that are
in low concentration and for the best quantification of the maximum
number of bands. Because the reaction is near completion after 5 min, this
method offers an optimal signal-to-noise ratio.

Note: If the gel has already been activated for 2.5 min, activating it for another
2.5 min might improve it. But activating an image for more than 5 min will not.

User Guide |

225

E |

Using Bio-Rads Stain-Free Technology

Imaging Blots
To blot stain-free gels, use standard blotting procedures as described in the
instruction manual you are using. Use only PVDF membranes with low background
fluorescence, as membranes other than low fluorescence PVDF can result in high
background or low sensitivity with the ChemiDoc XRS+ and Gel Doc XR+ imagers.
To assess transfer efficiency, be sure to activate and visualize the gel using the
imager before transfer.

226

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

F Mitsubishi P93/P95
Thermal Printer
Setting up a Thermal Printer on Windows
The printer driver is on the Image Lab software installation CD in the Misc folder.

To set up a thermal printer on a Windows system

1.

Install the printer driver.

2.

Open the printer section in Control Panel.

3.

Click the thermal printer icon and select Printing Preferences.

4.

Configure the correct paper size. Select 1280 x 1280 from the dropdown list.

5.

Click OK to apply your changes.

User Guide

| 227

| Mitsubishi P93/P95 Thermal Printer

Setting up a Thermal Printer on a Mac

The printer driver can be found on the Image Lab software installation CD in the
Misc directory.

To set up a thermal printer on a Mac system

1.

Install the printer driver.

2.

Connect the printer to the computer.

To configure the correct paper size

228

1.

Start Image Lab software.

2.

Select File > Page Setup.

3.

In the Settings dropdown list, select Page Attributes.

4.

In the Format For dropdown list, select the Mitsubishi printer.

5.

In the Paper Size dropdown list, select 1280 x 1280.

6.

In the Settings dropdown list, select Save as Default.

7.

Click OK to save the settings.

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

G Regression Calculation
Methods
Each regression method calculates a standard curve. Some of the methods provide
the formula for the standard curve. In this case, the molecular weight can be
calculated by:
x = relative front of the band of interest
y = molecular weight of the band of interest
Linear (semilog): The linear equation is y = a + bx, where a is the intercept and b is
the slope of the line.
Note: The linear equation is calculated on the log of the molecular weight
values.
The R2 value can be used to determine the overall quality of the linear fit. A linear
regression with an R2 value of >0.99 is considered a very good fit. The primary
advantage of this method is that it is extremely simple. The primary disadvantage is
that it will deliver incorrect results if the data are not very linear.
Point-to-point (semilog): No single equation is available for the point-to-point
method. The slope of each segment of the curve between data points is calculated
independently.
Note: The log of the molecular weight values is used to calculate the slope for
each segment of the curve.
Logistic: The logistic-4PL equation is

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G | Regression Calculation Methods

where:
x = mobility
y = molecular weight
a = estimated molecular weight at infinity
b = slope of the tangent at midpoint
c = midpoint
d = estimated molecular weight at zero mobility
Since the curve generated by the logistic-4PL regression method represents a
perfectly shaped S, it might not fit the data very well in all cases.
Cubic spline: Cubic spline curves are smooth curves that go through every data
point. The model is a cubic polynomial on each interval between data points. In
some cases, a spline curve can work well as a standard curve for interpolation.
However, because the curve is calculated individually for every pair of points, it does
not correspond to any single equation.

230

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Glossary

Aspect ratio

The ratio of the width to the height of an image.

CCD

(Charge-coupled device) A light-sensitive silicon chip used as a

photodetector in ChemiDoc MP and ChemiDoc XRS+ camera
systems.

Colormaps

Different color representations of a gel image.

Electrophoresis

A technique for separating molecules based on the differential

movement of charged particles through a matrix when subjected to an
electric field.

Example precision The number of decimal places chosen for displaying a measurement.
Flat fielding

An average intensity computation that compensates for

nonuniformities generated by an instrument.

Histogram

A graphed representation of the brightness, or gray value, of an image.

Native charge
density

The inherent electrical charge of a protein without the addition of SDS.

pl

Isoelectric point; the pH at which a protein molecule carries no net

charge.

Rf

Relative front value of the band. In Image Lab software, Rf has a

value between 0 and 1 and indicates the relative movement of the
band from top to bottom.

Quantitative
imaging

Determines the quantity of a proteins components through analysis of

the pixel values in a digital image of the sample.

UV-B

The range of ultraviolet light used by the system.

UV
transilluminator

The part of the imager that transmits UV light through a sample.

User Guide

231

| Glossary

232

Gel Doc XR+ and ChemiDoc XRS+ Systems with Image Lab Software

Bio-Rad
Laboratories, Inc.
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Life Science
Group

10017218 Rev G

US/EG

13-2297

1213

Sig 1213