ADVIA® Chemistry Systems

Methods Introduction
V 2.00.00
This document contains general information for all methods performed on
ADVIA® Chemistry systems and explanations of important sections contained in the
method sheets (IFUs).
Bayer HealthCare provides the procedures necessary for the preparation, use, and
storage of the ADVIA Chemistry reagents in the specific method sheet. Calibrator
information is provided in their respective package inserts.
IMPORTANT NOTICE
Any modification of computer disks and/or the programs contained on such disks can
adversely affect the control of the instrument’s performance and thereby invalidate the
results obtained and the claims that have been made regarding system performance.
Bayer HealthCare cannot be responsible for errors that are introduced by, or results
from, any modifications or alteration of the computer disks and/or programs contained
on such disks by the user, or for any direct or indirect consequences resulting from such
modification or alteration.

Sample Collection and Preparation

WARNING - POTENTIAL BIOHAZARDOUS MATERIAL
Any samples of human blood, plasma, serum, urine, or cerebrospinal fluid
should be handled cautiously as a biohazardous material according to good
laboratory practices.
The blood sample should be collected in a commercially available collection
tube using routine venipuncture techniques. A full discussion of sample
collection and preparation is beyond the scope of this document; however this
subject is covered in detail in recent texts by Kaplan and Pesce and Tietz. In
addition, the Clinical and Laboratory Standards Institute (formerly NCCLS) has
published two documents on sample collection.

During sample collection and preparation, avoid hemolysis of erythrocytes. When the
effect of hemolysis on a method is significant, this effect is described in the Limitations
of the Procedure and Interferences sections of the specific clinical method. Process
samples in a manner that prevents the introduction of clots, fibrin strands, or other solid
materials into the system.
Unless otherwise specified in the clinical method IFU, collect plasma samples using
lithium heparin as an anticoagulant. The individual laboratory should qualify the use of
other anticoagulants.
The use of urine preservatives is beyond the scope of this document.1,2 Special analyte-
specific storage conditions such as freezing or protection from direct exposure to light
are noted in the specific clinical method. For example, timely separation of serum from
cells is especially important to prevent elevated results for aspartate aminotransferase,
lactate dehydrogenase, magnesium, potassium, and total bilirubin. Erythrocytes
normally contain high concentrations of these analytes, which may increase the serum

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 or plasma concentration upon standing. Also, glucose test values may decrease upon
prolonged contact with erythrocytes due to the presence of glycolytic enzymes.
In general, separate plasma and serum from cells within two hours of collection. If you
cannot assay samples immediately after separation, store them in stoppered containers
at 2–8°C.
1. After drawing the specimen from the patient, perform identification procedures
consistent with your laboratory protocol. For serum samples, place the collection
tube in an upright position and allow the specimen to stand undisturbed thirty
minutes to allow clot to form.
2. If you have not used gel separation tubes, carefully separate the clot from the glass
by rimming the collection tube with a wooden stick.
Do not allow the wooden stick to stand (remain) in the tube. Perform this procedure
carefully so the cells are not hemolyzed.
3. If plasma is an acceptable sample type for the particular method, draw the patient
specimen into a tube containing acceptable anticoagulant (lithium heparin or as
indicated in the specific clinical method).
4. Centrifuge the collection tube according to the tube manufacturer’s
recommendation.
Consult centrifuge manufacturer’s instructions for further information on
centrifuge performance.
5. If the clot is attached to the side of the tube and has not dropped to the bottom,
repeat rimming the tube and centrifugation. After centrifugation, examine the
primary collection tubes to ensure that there is a level horizontal separation for
serum and clot. If using a gel tube, you can place the primary collection tube
directly into the sample rack on the system.
6. Alternatively, you can carefully decant the serum from the primary collection tube
into a clean sample cup or secondary tube.
NOTE: Make certain that no clots are decanted into the clean cup or tube. In the
case of plasma, make certain that no fibrin or strands appear in the plasma.
When determining the sample volume requirements for the assays, include the
applicable dead volume:
• for collection tubes: 200 µL minimum
• for sample cups: 50 µL minimum
IMPORTANT
To avoid having insufficient sample, when using sample cups, carefully dispense
the specimen so no air bubbles are entrapped in bottom of the sample cup.

English - 2 Methods Introduction

Calibrator and Control Products
This section contains information about product part numbers and frequency of
calibrations and quality control procedures.

WARNING - POTENTIAL BIOHAZARDOUS MATERIAL

CAUTION! Contains human source material. While each human serum or
plasma donor unit used in the manufacture of this product was tested by FDA-
approved methods and found nonreactive for hepatitis B surface antigen
(HBsAg), antibody to hepatitis C (HCV), and antibody to HIV-1/2, all products
manufactured using human source material should be handled as potentially
infectious. Because no test method can offer complete assurance that hepatitis B
or C viruses, HIV, or other infectious agents are absent, these products should
be handled according to established good laboratory practices at the Biosafety
Level II as recommended for any potentially infectious human blood specimens
in Protection of Laboratory Workers from Infectious Disease Transmitted by
Blood, Body Fluids, and Tissue - Second Edition; Approved Guideline (1997),
Document M29-A, promulgated by the Clinical and Laboratory Standards
Institute (formerly NCCLS).

Calculation of Results
The system automatically performs all calculations necessary for obtaining final
results. The system reports the results for each sample qualitatively or as
concentration/activity (quantitatively).
The system questions questionable results. One of the following automatic system
responses is assigned for each flag: no action, rerun with no dilution, or rerun with
dilution. Samples designated for rerun can be repeated automatically from the sample
tray or the dilution tray.
Review results using the Review/Edit window or at the host computer. You can screen
sample results transmitted to the host computer using the Data Clean feature.

Interpretation of Results
System operators and laboratory supervisors are responsible for operating and
maintaining Bayer HealthCare products in accordance with the procedures described in
the applicable product labeling (such as Instructions for Use, package inserts, bulletins,
product labels, and online documentation) and for determining that product
performance conforms to the applicable claims.
All in vitro assays can generate erroneous results. There are many possible causes for
discordant results. Erroneous results may occur due to interferences from identifiable
serum constituents or patient-specific serum constituents.
If, under these prescribed conditions of operation and maintenance, an aberrant or
abnormal result, as defined by the laboratory protocol, occurs, laboratory personnel
should first make certain that the system is performing and is being operated in
accordance with the product labeling. Laboratory personnel should then follow a
protocol consistent with applicable legal requirements for advising the clinician of a
result that appears to deviate from the norms established by the laboratory.
Test results alone are not diagnoses of medical conditions. A physician’s diagnosis
involves evaluation of the test result in conjunction with, and in the context of, the

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 patient’s medical history, physical examination, and other test results. Consultation
with other medical experts is sometimes required.
Laboratory personnel should address any product issues (such as, failure to meet a
performance specification or to perform as intended) and report to Bayer HealthCare
and/or the appropriate regulatory agency (such as the FDA) to the extent required by
legal or regulatory requirements. Various areas of the product labeling previously noted
address malfunctions and their possible effect on results.

Waste Disposal Requirements

Laws and regulations enacted to protect the environment and to encourage resource
conservation, require the disposal of hazardous and biohazardous wastes in a specified
manner.
Some of the wastes from the ADVIA Chemistry systems can be classified as hazardous
or biohazardous wastes. It is essential that the laboratory take appropriate steps to
determine the laws and regulations applicable to their disposal and to effect
compliance.
You must sample instrument effluent to evaluate compliance with applicable
regulations. The laboratory should contact a local licensed biohazardous waste disposal
firm for assistance.
The principal wastes in the ADVIA Chemistry systems come from the following items:
reaction cuvettes, dilution cuvettes, method reagent containers, sample containers,
ancillary reagent containers, and the system wastes.
Handle and dispose of sample containers with human specimens, control material, and
all reagents in accordance with all applicable federal, state, and local requirements.
Refer to the product labeling and to the Material Safety Data Sheets for details
concerning any special precautions related to reagents handling.

Clinical Methods (Instructions for Use)

Each clinical method provides the information relevant to an individual analyte. The
clinical method contains the information and data described in CLSI publication GP2-
A2 and by FDA regulations.
A laboratory procedure manual is mandatory for good laboratory practice and
regulatory compliance. To aid in the preparation of a laboratory procedure manual,
refer to the cross-reference of the information contained in our clinical methods with
the guidelines for Clinical Laboratory Procedure Manual published by CLSI
(Approved Guidelines GP2-A2).
Bayer HealthCare clinical methods are presented in a consistent format with the
headings listed below. The following is a brief description of the information included
under each heading:
Method Summary
This table summarizes method information such as the method principle, specimen
type, on-board stability, calibration frequency, and analytical range.

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 Intended Use
This section contains a brief statement of intended use for the method as an in vitro
diagnostic procedure for quantitating or detecting analytes in human serum, plasma,
urine, or other human fluids.
Summary and Explanation
This section contains a short history of the method with pertinent references to
previous related methods.
Principles of the Procedure
This section provides a physical, chemical, and biological explanation of the method
and a description of the chemical reactions, where applicable.
Reagents
This section describes the reagents required to perform the method including reagent
packaging information, product descriptions, materials required but not provided, and
reagent preparation instructions. Analyte-specific calibrator information also may be
included.
It also contains the appropriate hazard warnings pertaining to the reagents specific to
the method. You must maintain a constant awareness of the hazards and warnings
associated with all reagents. Where necessary, use appropriate safety equipment, such
as eye protection, gloves, and lab coats. You should follow the handling and safety
precautions specified in the Material Safety Data Sheets provided by Bayer
HealthCare.
Components and Concentrations
The reagent components and the percentage concentration of each component are listed
in this section.
Reagent Preparation and Use
The preparation instructions for reagents are provided in the specific clinical method.
On-board Reagent Stability (OBS)
Reagent and calibrator storage and expiration dates are printed on the product label.
When stored unopened, these products are stable until the printed expiration date or, if
a specific day is not provided, through the last day of the month.
The on-system stability stated in the clinical method represents the minimum stable
period for the working reagent. If the laboratory uses reagents for longer than the
recommended time, because the laboratory’s Quality Control Protocol indicates
acceptable performance, then the laboratory is responsible for completing a risk
analysis and verifying/validating the extended use.
When stored unopened at the appropriate temperature, the system solutions are stable
through the last day of the month stated in the expiration date.
Sample Handling
For a detailed explanation of sample handling, refer to Sample Collection and
Preparation.
Refer to the specific clinical method for details about any special sample handling
requirements or dilutions.

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 Materials Required but Not Provided
This topic lists the additional materials other than the reagents and calibrators that are
required to perform the methods.
Calibration
This topic specifies the calibration type, frequency, and procedure for each method and
identifies the calibration material to be used, where appropriate.
Refer to the Calibration Overview in the online Operator’s Guide for information about
how to setup and run calibration.
Calibration Frequency
This section lists the minimum duration of the calibration stability.
Reagent Blank (RBL) Frequency
This section describes when the RBL is measured.
Quality Control
This topic specifies recommended quality control material and quality control
frequency for the individual method.
In general, Bayer HealthCare recommends monitoring the ADVIA Chemistry methods
using the Bayer Assayed Chemistry Control 1 and 2 or at least two levels of other
commercially available control materials.
Run controls at the beginning of each shift or at some interval chosen by the laboratory,
whenever a new reagent is used, and following the performance of any detection
system maintenance or cleaning.
Bayer HealthCare recommends that the laboratory evaluate all control results prior to
reporting patient results.
If control results fail to meet the laboratory’s established criteria for acceptability,
evaluate all patient test results obtained in the unacceptable test run to determine if
patient test results were adversely affected. The laboratory should take and document
appropriate corrective actions, which may include recalibration, before reporting
patient results.
If commercially available controls are used, the laboratory must verify that the controls
respond properly on the system.
Refer to the Quality Control Overview in the online Operator’s Guide for information
about how to setup and run controls.
Limitations of the Procedure
The section describes substances, conditions, and patient states that have been reported
to cause actual or apparent changes in the analyte concentration.
The interference effects of Bilirubin, Hemoglobin, and Lipemia were studied by
spiking pooled serum at various levels. Unless otherwise noted, the serum interferences
were evaluated at two analyte concentrations; if no significant effect is observed, only
the lowest analyte level is presented. Interference is judged to be significant if it causes
a ≥ 10% deviation from the analyte mean concentration.
Bayer HealthCare makes every effort to identify in the Troubleshooting, Maintenance,
and Bibliography sections of the product labeling, the potential effects caused by
instruments, reagents, or endogenous substances, including known interferences on

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 patient results. However, it is impossible to provide information that would be
applicable to every clinical situation. This information is provided in lieu of attempting
to define the need for additional testing, which involves medical judgment in individual
cases. The decision whether or not to report a diagnostic result rests with the
laboratory.
Results
The system flags questionable results. One of the following automatic system
responses is assigned for each flag: no action, rerun with no dilution, or rerun with
dilution. Samples designated for rerun can be repeated automatically from the sample
tray or the dilution tray.
You can review results using the Review/Edit window or at the host computer. You can
screen sample results transmitted to the host computer using the Data Clean feature.
When appropriate, this section contains information concerning the dilution of samples
that exceeded the analytical range.
Interferences
This section lists potential interferents, if any, and their affect on various sample
concentrations.
Performance Characteristics
This section provides technical information related to method performance
characteristics, including precision, correlation, linearity, and sensitivity.
A description of each statistical parameter and the protocol followed to compute that
parameter are contained in the paragraphs that follow.
Precision
Precision is defined as the degree of variability among repeated independent
measurements of the same sample using external and internal evaluation data.
Within run precision refers to the reproducibility of the method when samples are
assayed within a single run.
Total precision extends the within-run estimates to include between run and between
day components of variation.
Precision estimates are computed at specified analyte levels using human serum pools,
urine pools, or control materials in a protocol similar to that recommended in the CLSI
document EP5-A, Evaluation of Precision Performance of Clinical Chemistry Devices;
Approved Guideline.
Unless otherwise noted, all precision data was obtained over a single calibration period
using a single lot of individual reagents and calibrator. Precision was evaluated using
two systems per method, at least ten days per system, one to two runs per day, and two
or more samples per run.
The precision values for each method, described on the specific clinical method, are
point estimates of average precision you can expect on the system. Since this is a point
estimate, any single system tested with a similar protocol could yield a different
estimate of precision and yet not be significantly different from the published average
precision.
The CLSI document, EP5-A, describes a statistical test (Chi Square) for comparing an
instrument’s performance and the expected performance. You should be able to obtain
precision data that pass the Chi Square test at the 95% confidence level.

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 Analytical Range
This section describes the concentration range across which you can obtain acceptable
results from the method.
For specimens that produce flags, you can dilute and reassay them according to
directions available in the specific clinical method.
For the IgA, IgG, and IgM methods, the low end of the analytical range is extended
automatically by having the system rerun patient samples when the assay absorbance is
less than the first non-zero calibrator assay absorbance. To ensure that this feature
operates properly, confirm the Re-absorb (d) and Re-absorb (u) parameter values
whenever the IgA, IgG, and IgM methods are recalibrated.
Prozone Effect
This section describes whether a prozone effect was demonstrated for various
concentrations.
Expected Values
The expected values and therapeutic ranges were taken from the literature cited in each
clinical method. For some methods, the serum (and plasma, if appropriate) expected
range was determined in a study conducted by Bayer HealthCare using the Bayer
method on an ADVIA Chemistry systems.
Correlation Data
Correlation data is determined using human serum, plasma, and urine samples. These
studies are conducted internally or externally at evaluation sites by comparing the
performance of the applicable ADVIA Chemistry system with the performance of a
reference method or a comparative system.
Regression statistics are computed using a protocol similar to that recommended in the
CLSI document EP9-A, Method Comparison and Bias Estimation Using Patient
Samples; Approved Guideline (1995).
Unless noted otherwise, regression data in each method indicates the linear least
squares fit between the appropriate ADVIA Chemistry System (y) and the comparative
system/method (x).

Bibliography
1. Kaplan LA, Pesce AG. Clinical Chemistry – Theory, Analysis, and Correlation. St.
Louis: CV Mosby Company; 4th Edition, 1996: 69–81.
2. Tietz Textbook of Clinical Chemistry and Molecular Diagnostics. 4th ed.
Philadelphia, PA: WB Saunders Company; 2006: 41–57.
3. CLSI Standard Procedure for the Collection of Diagnostic Blood Specimens by
Venipuncture. CLSI Standard H3-A3. 1991. Available from CLSI, 940 West Valley
Road, Suite 1400, Wayne, PA, 19087-1898.
4. CLSI Standard Procedures for the Collection of Diagnostic Blood Specimens by
Skin Puncture, CLSI Standard H4-A3. 1991. Available from CLSI, 940 West
Valley Road, Suite 1400, Wayne, PA, 19087-1898.
5. Refer to the Information Bulletin titled Sample Handling Guidelines (Publication
No. TN9-5729-31).

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Trademarks
Bayer, the Bayer cross, ADVIA, ADVIA Centaur, ADVIA IMS, and Technicon DAX
are trademarks of Bayer.
Beckman is a trademark of Beckman Coulter, Inc.
Dade Behring is a trademark of Dade Behring, Inc.
Dimension is a trademark of Dade Behring, Inc.
Emit is a trademark of Syva Company.
Hitachi is a trademark of Hitachi Medical Systems America, Inc.
Synchron is a trademark of Beckman Coulter, Inc.
Roche, Cobas Fara, and Cobas Integra are trademarks of Roche Diagnostics, Inc.

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