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Batch Record

The document describes the manufacture of a HEK293 Master Cell Bank including thawing cells from frozen storage, seeding cells into flasks at a target density, and incubating. Materials used include cell banks, media, flasks and equipment for thawing, counting, and incubating cells. Detailed steps are provided for preparing media, thawing and counting cells, calculating seeding densities and volumes, and labeling and incubating seeded flasks to establish the Master Cell Bank.

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Ines Concepcion Tupas
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2K views11 pages

Batch Record

The document describes the manufacture of a HEK293 Master Cell Bank including thawing cells from frozen storage, seeding cells into flasks at a target density, and incubating. Materials used include cell banks, media, flasks and equipment for thawing, counting, and incubating cells. Detailed steps are provided for preparing media, thawing and counting cells, calculating seeding densities and volumes, and labeling and incubating seeded flasks to establish the Master Cell Bank.

Uploaded by

Ines Concepcion Tupas
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
Available Formats
Download as PDF, TXT or read online on Scribd
You are on page 1/ 11

Document Title: Manufacture of HEK Master Cell Bank

Company: Cells R Us
Reference Project Code: MATC-HEK-MCB
Lot Number:
Document Approval Date:

EXAMPLE BATCH RECORD


Bill of Materials
Item Description

Vendor/Cat#

RM #

Lot #

Expiration
Date

DMEM
Fetal Bovine Serum
100X Glutamax
HEK293 Cell Bank (2x106
cells/vial, P20)
50ml Conical Tubes
T75 Flasks
T175 Flasks

Non-RM Materials
Item Description

Vendor/Cat #

Pipets
Trypan Blue

Ancillary Equipment
Item Description

Equipment ID #

Page 1 of 11

Lot#

Quantity

Document Title: Manufacture of HEK Master Cell Bank


Company: Cells R Us
Reference Project Code: MATC-HEK-MCB
Lot Number:
Document Approval Date:

BSC
Waterbath
Centrifuge
Incubator
Refridgerator

Estimated Quantities of Materials Required:


Item Description

Quantity

DMEM
Fetal Bovine Serum
100X Glutamax
Pipets

Manufacturing Room:

Mfg Rm Release Ref: SOP-XXX-XXX Log # _______________

Date/Time Begin:

Signature:

Step 1: Date/Time _________________ Room ____________


Step #

Step Description

1.0

Preparation of Media

Operator
Initials

1.1 Thaw 1 (one) x 100ml bottle of Fetal Bovine Serum


(RM-XXX, lot ____) .
Page 2 of 11

Verifier
Initials

Document Title: Manufacture of HEK Master Cell Bank


Company: Cells R Us
Reference Project Code: MATC-HEK-MCB
Lot Number:
Document Approval Date:

Thaw bottle in a water bath set at 37 3 C


Waterbath ID# __________ Temp ______ C
Time in water bath ________________
Time out of water bath _____________
1.2 Sanitize the BSC with Cavicide followed by 70% IPA.
Sanitize and transfer required materials into the
BSC per SOP-XXX-XXX. Follow aseptic technique for
all steps performed within BSC.

____________ ____________

____________

1.3 Prepare three (3) individual bottles of Complete


Medium (DMEM, 10% FBS, 1X Glutamax)
Into each 500 ml of DMEM Medium (RM-XXX, lot
____ ) add:

____________ ____________

55 ml FBS (RM-XXX, lot ____ )


5.6 ml 100x Glutamax (RM-XXX, lot ____ )
Volume FBS added _______ ml/bottle
Volume 100X Glutamax added _______ ml/bottle
# of bottles of Complete Medium prepared
_______

Step #

Operator
Initials

Step Description

1.4 Label bottles: Complete Medium per SOP-XXX-XXX.


Include 45 day expiration date. Place example of
label below. If not immediately using, store
Complete Medium in refrigerator at 4C 2C.

Page 3 of 11

Verifier
Initials

____________ ____________

Document Title: Manufacture of HEK Master Cell Bank


Company: Cells R Us
Reference Project Code: MATC-HEK-MCB
Lot Number:
Document Approval Date:

Fridge ID # _______________ Temp ________


Date/time to fridge _____________________

1.5 End Date/Time ___________________________


____________ ____________

Step 2: Date/Time _________________ Room _______________


Estimated Materials Required:
Item Description

Quantity

HEK293 Cell Bank (2x10e6


cells/vial, P20)
Complete Medium (step 1.4)
50ml Conical Tubes
T75 Flasks
T175 Flasks
Pipets
Trypan Blue

Step #

Operator
Initials

Step Description

Page 4 of 11

Verifier
Initials

Document Title: Manufacture of HEK Master Cell Bank


Company: Cells R Us
Reference Project Code: MATC-HEK-MCB
Lot Number:
Document Approval Date:

2.0

Thaw/Seed of HEK Cell Bank (Passage 20)


2.1 Obtain two (2) vials of HEK Cell Bank @ 2x106
cells/vial (RM-XXX, lot _____ ) from the liquid N2
freezer and place on dry ice.
2.2 Sanitize the BSC with Cavicide followed by 70% IPA.
Sanitize and transfer required materials into the
BSC per SOP-XXX-XXX. Follow aseptic technique for
all steps performed within BSC.
2.3 Pipet 18 ml of Complete Medium (prepared in step
1.4) into a 50ml conical tube (RM-XXX, lot ____)
2.4 Thaw two (2) vials of cells in a water bath set at 37
+ 3 C with constant swirling until a small amount
of ice remains. Remove vial from water bath and
continue thaw at room temperature until
completely thawed.

____________ ____________

____________

____________ ____________

____________ ____________

Water Bath ID# __________ Temp ____ oC


2.5 Remove label from vial of cells and place
representative label below.

2.6 Using a pipet, transfer thawed cells to 50 ml conical


(step 2.3) adding drop wise while carefully swirling
to distribute cells in medium.

Page 5 of 11

____________

____________

Document Title: Manufacture of HEK Master Cell Bank


Company: Cells R Us
Reference Project Code: MATC-HEK-MCB
Lot Number:
Document Approval Date:

2.7 Centrifuge at 200g x 5 minutes at 18 3oC.


Centrifuge ID# __________ Speed _____ g _______
rpm

____________ ____________

Time ________ Temp ______ C


2.8 Aspirate supernatant and re-suspend pellet in 2 ml
Complete Medium (prepared step 1.4).

____________ ____________

Volume Complete Medium added ______ ml


2.9 Prepare dilution for cell count (suggestion 1:4
dilution):

____________ ____________

_____L cells + _____ L trypan blue =____ L


total
_______ L total _______ L cells = _______
dilution factor
Step #

Operator
Initials

Step Description

2.10 Record counts from four diagonal 1 mm2 squares


(#of viable cells / # of dead cells.)
A) _____/b _____ C) _____/d)_____
E)_____/f_____ G)_____/h_____
a). Average Live Cells = A+C+E+G 4 = ________
b.) Average Dead Cells = b+d+f+h 4 = ________
c). Avg. Live _______ + Avg. Dead _______ = Total
Cells _________
d). Avg. Live Cells ________ Total Cells ________

Page 6 of 11

Verifier
Initials

____________ ____________

Document Title: Manufacture of HEK Master Cell Bank


Company: Cells R Us
Reference Project Code: MATC-HEK-MCB
Lot Number:
Document Approval Date:

= ____% Viable Cells


2.11 Calculate total number of viable cells/ml as follows:
____________ ____________
______ Avg live cells (step 2.10.a) X ____ Dilution
factor (step 2.9) X 104 = __________ cells/ml
2.12 Calculate total number of viable cells as follows:

Step #

_________ cells/ml (step 2.11) X _______resuspension volume (step 2.8) = _________ total
viable cells

____________ ____________

Step Description

Operator
Initials

Verifier
Initials

2.13 Intention is to seed 1x T175 flasks at 2x10e4


0.5x10e3cells/cm2. Calculate size and number of
flasks to seed in this seeding density range.
3.5x10e6 cells/T175 flask __________ cells/ml
(step 2.11) = _______ml cell suspension per flask
Based on total re-suspension volume (step 2.8)
_______ml, determine size and number of flasks to
seed. Record information below.
Flask
Type

# Flasks

Seed
Density

Volume Cell
Suspension

T75
T175
Page 7 of 11

____________ ____________

Document Title: Manufacture of HEK Master Cell Bank


Company: Cells R Us
Reference Project Code: MATC-HEK-MCB
Lot Number:
Document Approval Date:

2.14 Pipet Complete Medium (step 1.4) into each flask


required (T75 use 20ml, T175 use 40ml).
T75 (RM-XXX, lot _____ )
T175 (RM-XXX, lot ______ )
Record volume Complete Medium added per flask.
T75 _________ ml/flask

____________ ____________

T175 ________ ml/flask


Step #

Operator
Initials

Step Description

2.15 Add calculated volume of cell suspension (step


2.13) to each flask.

Verifier
Initials

____________ ____________

T75 _________ ml/flask


T175 ________ ml/flask
2.16 Label and date seeded flasks per SOP-XXX-XXX.
Place representative label below.
2.17 Place all flasks into incubator at 37 2 C, 5 1%
CO2

____________ ____________

____________ ____________

Incubator ID# _______ Temp _________ %CO2


______
Date/time ______________________
2.18 End Date/Time _______________________
____________ ____________

Page 8 of 11

Document Title: Manufacture of HEK Master Cell Bank


Company: Cells R Us
Reference Project Code: MATC-HEK-MCB
Lot Number:
Document Approval Date:

Operator Signature

Date

Verifier Signature

Date

Operator Comments:

Deviations Notes:
Step #

Description

Page 9 of 11

Document Title: Manufacture of HEK Master Cell Bank


Company: Cells R Us
Reference Project Code: MATC-HEK-MCB
Lot Number:
Document Approval Date:

Page 10 of 11

Document Title: Manufacture of HEK Master Cell Bank


Company: Cells R Us
Reference Project Code: MATC-HEK-MCB
Lot Number:
Document Approval Date:

QA Review/Approval Notes:

Manufacturing Management Approval

Date

Quality Assurance Management Approval

Date

Page 11 of 11

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