ADVIA Centaur ®

ADVIA Centaur ® XP
Immunoassay Systems

AFP

Assay Summary
Sample Type Serum, amniotic fluid
Sample Volume 10 µL
Calibrator D
Sensitivity and Assay Range 1.3 – 1000 ng/mL (1.08 – 830 IU/mL)

Contents
REF Contents Number of Tests
03305838 5 ReadyPack® primary reagent packs containing ADVIA Centaur® 500
(110764) AFP Lite Reagent and Solid Phase
ADVIA Centaur AFP Master Curve card
or
03974780 1 ReadyPack primary reagent pack containing ADVIA Centaur 100
(110763) AFP Lite Reagent and Solid Phase
ADVIA Centaur AFP Master Curve card

Intended Use
For in vitro diagnostic use in the quantitative determination of alpha-fetoprotein (AFP) in the
following:
• human serum and in amniotic fluid from specimens obtained at 15 to 20 weeks gestation,
as an aid in detecting open neural tube defects (NTDs) when used in conjunction with
ultrasonography and amniography testing,
• human serum, as an aid in managing non-seminomatous testicular cancer when used in
conjunction with physical examination, histology/pathology, and other clinical evaluation
procedures, using the ADVIA Centaur and ADVIA Centaur XP systems.

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AFP ADVIA Centaur and ADVIA Centaur XP Systems

WARNING: The concentration of AFP in a given specimen, as determined by assays from different
manufacturers, can vary due to differences in assay methods and reagent specificity. The results
reported by the laboratory to the physician must include the identity of the AFP assay used.
Values obtained with different AFP assay methods cannot be used interchangeably. Before
changing assay methods, the laboratory must do the following:
• For prenatal testing, the laboratory must establish a range of normal values for the new
assay based on normal serum and amniotic fluid from pregnant women with a confirmed
gestational age.
• For cancer management, the laboratory must perform additional testing to confirm baseline
values for patients being serially monitored.
United States federal law restricts this device to sale and distribution by or on the order of a
physician, or to a clinical laboratory; and use is restricted to, by, or on the order of a physician.
Use AFP results only as part of the overall clinical evaluation of a patient. Do not use AFP
results as the only criterion for diagnosis. (Refer to Warnings, Summary and Explanation of the
Test, and Limitations.)

Warnings
Elevated MSAFP (maternal serum AFP) levels may indicate open NTD, but are not used to
diagnose the defect without additional testing. In addition, elevated MSAFP levels may
indicate other forms of fetal distress or malformation, which include placental malformations,
ventral wall defects, fetal kidney dysfunction, and fetal death. MSAFP levels may also be
elevated in certain benign and malignant conditions not related to pregnancy. These conditions
include hepatitis, cirrhosis, ataxia telangiectasia, primary hepatocellular carcinoma, and
certain germ cell cancers. Furthermore, incorrect estimation of gestational age can result in
either under- or over-estimation of open NTD risk. Therefore, AFP testing requires accurate
gestational dating for reliable risk assessment for open NTDs. Confirmatory procedures such
as ultrasonography, amniography, amniotic fluid acetylcholinesterase, and amniotic fluid AFP
must be used in conjunction with MSAFP testing for accurate NTD risk assessment.
When using AFP in the evaluation of fetal defects, laboratories must establish their own
median values for each gestational week. Absolute AFP values may vary for each lab
depending on the demographics of its population including race and maternal weight.
Collect maternal serum specimens for NTD testing before amniocentesis. Refer to Special
Precautions for detailed information.
The ADVIA Centaur AFP assay is not a screening test for cancer and must never be used as
such. AFP testing is a safe and effective supplement to patient care when used as part of the
overall management strategy for patients undergoing treatment for non-seminomatous
testicular cancer or for patients being monitored after therapy is complete.
Do not interpret serum AFP as absolute evidence of the presence of malignant disease. At time
of presentation, patients with confirmed non-seminomatous testicular cancer may have serum
AFP concentrations within the range observed in healthy individuals. Since elevated AFP
levels are often found in patients with other malignant and non-malignant conditions, the
physician should rule out all other conditions associated with elevated AFP levels prior to the
use of the ADVIA Centaur AFP values in non-seminomatous testicular cancer management.
Conversely, low concentrations of AFP are not necessarily indicative of absence of disease,
particularly post-surgery or after chemotherapy. Testicular tumors that are histologically
categorized as pure seminoma do not synthesize AFP. The ADVIA Centaur AFP assay, as a
useful adjunct in cancer management, is intended for the evaluation of non-seminomatous
testicular cancer, or mixed tumors with non-seminomatous elements, but not for pure
seminoma. Additionally, several histologic subtypes of non-seminoma either do not synthesize
AFP (choriocarcinoma) or do so unpredictably (teratoma). Therefore, AFP levels should be
used concurrently with other diagnostic and clinical patient information.

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Materials Required But Not Provided
REF Description Contents
00133564 Calibrator D 6 vials of low calibrator
(672183) 6 vials of high calibrator
or
04956492 Calibrator D 2 vials of low calibrator
(672173) 2 vials of high calibrator

Optional Reagents
REF Description Contents
07948423 ADVIA Centaur Multi-Diluent 2 2 ReadyPack ancillary reagent packs
(110314) containing 10 mL/pack
04855629 Multi-Diluent 2 50 mL/vial
(672260)
672428 AFP Master Curve Material 7 x 1 mL

Summary and Explanation of the Test

AFP is a single chain glycoprotein with a molecular weight of approximately 70,000 daltons.1
AFP was first described as a fetal protein by Bergstrand and Czar in 1956.2 AFP and albumin
share considerable sequence homology and some physiological functions.3,4 Fetal AFP
synthesis occurs in the liver, yolk sac, and gastrointestinal tract.5 AFP produced by the fetus is
secreted into fetal serum, reaches a peak at 13 weeks gestation, and then gradually declines
during gestation. Shortly after birth, the newborn’s AFP level reaches the normal adult level.
In adults, serum AFP concentrations remain low except during pregnancy, benign liver
diseases (hepatitis, cirrhosis), primary hepatocellular carcinoma, and certain germ cell tumors.

Prenatal Testing
During pregnancy, maternal serum AFP (MSAFP) levels rise through the third trimester.
Elevated or depressed AFP levels may indicate fetal problems. Elevated MSAFP levels during
the second trimester of pregnancy are often associated with one of the most common types of
birth defects, open neural tube defects (NTDs).6-8 A number of studies9-13 have confirmed the
utility of AFP testing to detect NTDs during the second trimester of pregnancy. In addition to
AFP testing, maternal factors such as race, weight, age, diabetes, and family history must be
considered when assessing the open NTD risk.14,15 Final determination of open NTD depends
on information provided by confirmatory testing since conditions other than open NTDs, such
as cirrhosis, hepatitis, certain types of cancer, and other fetal malformations (ventral wall
defects,16 defective kidneys,17 and others), may also cause elevated MSAFP levels.14,15
Such testing includes amniotic fluid AFP (AFAFP), acetylcholinesterase, amniography,
and ultrasonography. Depressed MSAFP levels have been reported in other conditions.

Cancer Management
Interest in AFP as a tumor marker originated with a report by Abelev in 1963.18 Tatarinov
provided the first evidence linking elevated serum AFP concentrations to primary cancer of
the liver.19 Since then, investigators have demonstrated elevated serum AFP levels in
hepatocellular cancer,20-22 malignant germ cell tumors of the ovary and testis,23,24 and
teratocarcinoma of the testis.25 Although at a very low rate of incidence, increased circulating
AFP concentrations may also occur in serum specimens from patients with gastrointestinal,
pancreatic, and pulmonary cancers.26

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The most important application of AFP testing in cancer management is for testicular cancer.
Although not present in pure seminoma,27 elevated serum AFP is closely associated with
non-seminomatous testicular cancer.28-30 The measurement of AFP in serum, in conjunction
with serum hCG, is an established regimen for monitoring patients with non-seminomatous
testicular cancer.31-34 In addition, monitoring the rate of AFP clearance from serum after
treatment is an indicator of the effectiveness of therapy.35,36 Conversely, the growth rate
of progressive cancer can be monitored by serially measuring serum AFP concentration
over time.37
Serial serum AFP testing is a useful adjunctive test for managing non-seminomatous
testicular cancer.

Assay Principle
The ADVIA Centaur AFP assay is a two-site sandwich immunoassay using direct
chemiluminometric technology, which uses constant amounts of two antibodies. The first
antibody, in the Lite Reagent, is an affinity purified polyclonal rabbit anti-AFP antibody
labeled with acridinium ester. The second antibody, in the Solid Phase, is a monoclonal mouse
anti-AFP antibody covalently coupled to paramagnetic particles.
The system automatically performs the following steps:
• dispenses 10 μL of sample into a cuvette
• dispenses 50 μL of Lite Reagent and 250 μL of Solid Phase and incubates for
7.5 minutes at 37°C
• separates, aspirates, and washes the cuvettes with reagent water38
• dispenses 300 μL each of Acid Reagent and Base Reagent to initiate the
chemiluminescent reaction
• reports results according to the selected option, as described in the system operating
instructions or in the online help system
A direct relationship exists between the amount of AFP present in the patient sample and the
amount of relative light units (RLUs) detected by the system.

Specimen Collection and Handling

Serum and amniotic fluid are the recommended sample types for this assay. The following
recommendations for handling and storing blood samples are furnished by the Clinical and
Laboratory Standards Institute (CLSI, formerly NCCLS):39
• Collect all samples observing universal precautions.
• Allow blood samples to clot adequately before centrifugation.
• Keep tubes stoppered and upright at all times.
• Do not use samples that have been stored at room temperature for longer than 8 hours.
• Tightly cap and refrigerate specimens at 2° to 8°C if the assay is not completed within
8 hours.
• Freeze samples at or below -20°C if the assay is not completed within 48 hours.
• Freeze samples only once and mix thoroughly after thawing.
Before placing samples on the system ensure that:
• Samples are free of fibrin or other particulate matter. Remove particulates by
centrifugation at 1000 x g for 15 to 20 minutes.
• Samples are free of bubbles.

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Special Precautions
• Collect maternal serum specimens for open NTD testing before amniocentesis. Significant
quantities of AFP may pass into the maternal circulation during amniocentesis causing
MSAFP levels to increase. Since the estimated half-life of AFP in serum is 4 to 6 days,40,41
the MSAFP levels may be falsely elevated.42 Therefore, if serum specimens are not
collected before amniocentesis, postponement of MSAFP testing for at least 2 weeks after
amniocentesis is recommended.
• Treat amniotic fluid specimens contaminated with red blood cells with caution because
fetal blood drawn with amniotic fluid may artificially elevate the AFP result. If red blood
cell contamination is suspected, then evaluate the amniotic fluid for the presence of fetal
hemoglobin.
• Centrifuge amniotic fluid specimens to clarify them before testing or freezing.
• All amniotic fluid samples can be automatically diluted 1:100 by the system or prepared
manually using Multi-Diluent 2.

Reagents
Store the reagents upright at 2° to 8°C.
Mix all primary reagent packs by hand before loading them onto the system. Visually inspect the bottom of the
reagent pack to ensure that all particles are dispersed and resuspended. For detailed information about
preparing the reagents for use, see the system operator’s guide.

Reagent Pack Reagent Volume Ingredients Storage Stability

ADVIA Centaur Lite Reagent 5.0 mL/ polyclonal rabbit anti-AFP 2–8°C until the expiration date on
AFP ReadyPack reagent antibody (~0.16 μg/mL) the pack label.
primary reagent pack labeled with acridinium ester For onboard stability, refer
pack in buffered saline with sodium to Onboard Stability and
azide (0.13%) and Calibration Interval.
preservatives
Solid Phase 25.0 mL/ monoclonal mouse anti-AFP 2–8°C until the expiration date on
reagent antibody (~0.064 mg/mL) the pack label.
pack covalently coupled to For onboard stability, refer
paramagnetic particles in to Onboard Stability and
buffered saline with sodium Calibration Interval.
azide (0.11%) and
preservatives
ADVIA Centaur Multi-Diluent 2 10.0 mL/ goat serum with sodium azide 2–8°C until the expiration date on
reagent (0.1%) and preservatives the pack label
ReadyPack pack or
ancillary reagent 28 consecutive days after
pack accessing the ancillary
reagent pack
Safety data sheets (MSDS/SDS) available on www.siemens.com/diagnostics.
NOTE: Sodium azide can react with copper and lead plumbing to form explosive metal azides.
On disposal, flush reagents with a large volume of water to prevent the buildup of azides, if
disposal into a drain is in compliance with federal, state, and local requirements.
R22 Harmful! Harmful if swallowed. After contact with skin, wash immediately with plenty of soap
S28 and water. Contains: sodium azide; Lite Reagent, Solid Phase

CAUTION: This device contains material of animal origin and should be handled as a potential
carrier and transmitter of disease.
For In Vitro Diagnostic Use.

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Loading Reagents
Ensure that the system has sufficient primary and ancillary reagent packs. For detailed
information about preparing the system, refer to the system operating instructions or to the
online help system.
Mix all primary reagent packs by hand before loading them onto the system. Visually inspect
the bottom of the reagent pack to ensure that all particles are dispersed and resuspended. For
detailed information about preparing the reagents for use, see the system operator’s guide.
Load the ReadyPack reagent packs in the primary reagent area using the arrows as a placement
guide. The system automatically mixes the primary reagent packs to maintain homogeneous
suspension of the reagents. For detailed information about loading reagents, refer to the
system operating instructions or to the online help system.
If automatic dilution of a sample is required, load ADVIA Centaur Multi-Diluent 2 in the
ancillary reagent entry.

Onboard Stability and Calibration Interval

The ADVIA Centaur AFP assay can be run using either the high or low volume usage option.
If you use the low volume usage option, you will need to modify the test definition parameters
for the assay. For detailed information about modifying test definitions, refer to the system
operating instructions or to the online help system.
• High volume usage = use of more than one primary reagent pack within the Onboard
Stability period indicated in the table for High Volume Usage.
• Low volume usage = use of less than one primary reagent pack within the Onboard
Stability period indicated in the table for High Volume Usage.

High Volume Usage

Onboard Stability Calibration Interval
14 days 28 days

Additionally, the ADVIA Centaur AFP assay requires a two-point calibration:

• when changing lot numbers of primary reagent packs
• when replacing system components
• when quality control results are repeatedly out of range
NOTE:
• Discard the primary reagent packs at the end of the High Volume Usage onboard stability
interval.
• Do not use reagents beyond the expiration date.

Low Volume Usage

NOTE: For low volume usage, load only one primary reagent pack on the system at a time.
Perform a calibration every time you load a fresh primary reagent pack.
Onboard Stability Calibration Interval
28 days 14 days

Additionally, the ADVIA Centaur AFP assay requires a two-point calibration:

• when changing lot numbers of primary reagent packs
• when replacing system components
• when quality control results are repeatedly out of range

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NOTE:
• Discard the primary reagent pack at the end of the Low Volume Usage onboard stability
interval. When the Primary Reagent Stability option is set to USE BEYOND, the system does
not display a flag when the reagent exceeds the onboard stability.
• Do not use reagents beyond the expiration date.

Master Curve Calibration

The ADVIA Centaur AFP assay requires a Master Curve calibration when using a new lot
number of Lite Reagent and Solid Phase. For each new lot number of Lite Reagent and Solid
Phase, use the barcode reader or keyboard to enter the Master Curve values on the system. The
Master Curve card contains the Master Curve values. For detailed information about entering
Master Curve values, refer to the system operating instructions or to the online help system.

Quality Control
Follow government regulations or accreditation requirements for quality control frequency.
For detailed information about entering quality control values, refer to the system operating
instructions or to the online help system.
To monitor system performance and chart trends, as a minimum requirement, two levels of
quality control material should be assayed on each day that samples are analyzed. Quality
control samples should also be assayed when performing a two-point calibration. Treat all
quality control samples the same as patient samples.
Siemens Healthcare Diagnostics recommends the use of commercially available quality
control materials with at least 2 levels (low and high). A satisfactory level of performance is
achieved when the analyte values obtained are within the Acceptable Control Range for the
system or within your range, as determined by an appropriate internal laboratory quality
control scheme.
If the quality control results do not fall within the Expected Values or within the laboratory’s
established values, do not report results. Take the following actions:
• Verify that the materials are not expired.
• Verify that required maintenance was performed.
• Verify that the assay was performed according to the instructions for use.
• Rerun the assay with fresh quality control samples.
• If necessary, contact your local technical support provider or distributor for assistance.

Sample Volume
This assay requires 10 μL of sample for a single determination. This volume does not include
the unusable volume in the sample container or the additional volume required when
performing duplicates or other tests on the same sample. For detailed information about
determining the minimum required volume, refer to Sample Volume Requirements in the
ADVIA Centaur Reference Manual.
NOTE: The sample volume required to perform onboard dilution differs from the sample
volume required to perform a single determination. Refer to the following information for the
sample volume required to perform onboard dilutions:
Dilution Sample Volume (µL)
1:10, 1:20, 1:100, 1:200 20

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Assay Procedure
For detailed procedural information, refer to the system operating instructions or to the online
help system.

Procedural Notes
Calculations
For detailed information about how the system calculates results, refer to the system operating
instructions or to the online help system.
The system reports AFP results in ng/mL (common units) or IU/mL (SI units), depending
on the units defined when setting up the assay. The conversion formula is
1 ng/mL = 0.83 IU/mL.43
Based on a molecular weight of 70,000 daltons, 1 ng = 0.0143 nmol.

Dilutions
• Serum samples and amniotic fluid samples with AFP levels greater than 1000 ng/mL
(830 IU/mL) must be diluted and retested to obtain accurate results.
• For an automatic or manual dilution, the final AFP concentration in the diluted sample
must be ≥ 15 ng/mL (12.5 IU/mL).
• Patient samples can be automatically diluted by the system or prepared manually.
• For automatic dilutions, ensure that ADVIA Centaur Multi-Diluent 2 is loaded and set the
system parameters as follows:
Dilution point: ≤ 1000 ng/mL (830 IU/mL)
Dilution factor, serum samples: 10, 20, 100, 200
Dilution factor, amniotic fluid samples: 100
For detailed information about automatic dilutions, refer to the system operating
instructions or to the online help system.
• Manually dilute the patient samples when patient results exceed the linearity of the assay
using automatic dilution, or when laboratory protocol requires manual dilution.
• Use Multi-Diluent 2 to manually dilute patient samples, and then load the diluted sample
in the sample rack, replacing the undiluted sample.
• Ensure that results are mathematically corrected for dilution. If a dilution factor is entered
when scheduling the test, the system automatically calculates the result.

High Dose Hook Effect

Patient samples with high AFP levels can cause a paradoxical decrease in the RLUs (high dose
hook effect). In this assay, patient samples with AFP levels as high as 1,000,000 ng/mL
(830,000 IU/mL) will assay greater than 1000 ng/mL (830 IU/mL).

Disposal
Dispose of hazardous and biologically contaminated materials according to the practices of
your institution. Discard all materials in a safe and acceptable manner and in compliance with
all federal, state, and local requirements.

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Limitations
Heterophilic antibodies in human serum can react with reagent immunoglobulins, interfering
with in vitro immunoassays.44 Patients routinely exposed to animals or to animal serum
products can be prone to this interference and anomalous values may be observed. Additional
information may be required for diagnosis. The ADVIA Centaur AFP assay uses antibodies
from two animal species and routinely adds animal sera to its assay components to minimize
the interfering response.
Serum specimens that are . . . Have an insignificant effect on the assay up to . . .
hemolyzed 500 mg/dL of hemoglobin
lipemic 1000 mg/dL of triglycerides
icteric 20 mg/dL of bilirubin

Expected Results
AFP Values in Benign and Malignant Disease
The expected results for the ACS:180® AFP assay were previously established. Data was
obtained on 1858 serum samples as shown in the following table:
Distribution of AFP (ng/mL)
Sample Category N 0−8.0 8.1−20.0 20.1−500.0 500.1−1000.0 > 1000.0
Apparently Healthy Subjects 793 780 12 1 0 0
males 397 389 7 1 0 0
females 396 391 5 0 0 0
Malignant Diseases 717 513 64 88 11 41
Testicular Cancer
seminoma 41 37 3 1 0 0
non-seminoma 204 105 19 56 5 19
Liver Cancer
primary 80 29 11 20 4 16
secondary 93 79 8 5 0 1
Other Cancer
gastrointestinal 64 54 8 2 0 0
genitourinary 40 37 3 0 0 0
ovarian 78 73 5 0 0 0
pancreatic 18 16 1 1 0 0
other 99 83 6 3 2 5
Benign Diseases 348 316 18 8 1 5
cirrhosis 60 48 4 2 1 5
hepatitis 64 51 8 5 0 0
other 224 217 6 1 0 0

In this study, 98.4% of the apparently healthy subjects had AFP values less than 8.1 ng/mL.
These results were confirmed for the ADVIA Centaur AFP assay by analyzing serum samples
in the range of 1.3 to 943.6 ng/mL (1.1 to 783.2 IU/mL). Refer to Method Comparison.

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AFP values in Maternal Serum

The expected results for the ACS:180 AFP assay were previously established. Data were
obtained from 1713 serum samples at three sites as shown in the following table. MSAFP
values are reported in ng/mL.
Median* Multiples of Median (MoM, ng/mL)
Gestational Week N (ng/mL) 2.0 MoM 2.5 MoM 3.0 MoM
15 347 31.3 62.6 78.3 93.9
16 412 36.3 72.6 90.8 108.9
17 320 42.0 84.0 105.0 126.0
18 330 48.7 97.4 121.8 146.1
19 201 56.5 113.0 141.3 169.5
20 103 65.4 130.8 163.5 196.2
* Medians are determined based on a weighted linear regression model.14
These results were confirmed for the ADVIA Centaur AFP assay by analyzing serum samples
in the range of 1.3 to 943.6 ng/mL (1.1 to 783.2 IU/mL). Refer to Method Comparison.

AFP Values in Amniotic Fluid

The expected results for the ACS:180 AFP assay were previously established. Data was
obtained from 714 amniotic fluid samples at two sites as shown in the following table. AFAFP
values are reported in μg/mL.
Median* Multiples of Median (MoM, µg/mL)
Gestational Week N (µg/mL) 2.0 MoM 2.5 MoM 3.0 MoM
15 92 17.3 34.6 43.3 51.9
16 138 14.3 28.6 35.8 42.9
17 152 11.9 23.8 29.8 35.7
18 134 9.8 19.6 24.5 29.4
19 104 8.1 16.2 20.3 24.3
20 94 6.7 13.4 16.8 20.1
* Medians are determined based on a weighted linear regression model.14
These results were confirmed for the ADVIA Centaur AFP assay by analyzing amniotic fluid
samples in the range of 0.3 to 86.9 μg/mL. Refer to Method Comparison.
Due to potential variability in AFP values attributable to differences in regional populations
and assay methods, each laboratory should establish its own gestational age-specific median
values. Various options for obtaining a reliable set of medians appropriate for your screened
population have been described.14 Once medians are available, it is customary to report AFP
test results as a multiple of the median (MoM) to normalize for gestational age. Each
laboratory must select a MoM screening cut-off that meets its needs.9-11
As with all diagnostic assays, each laboratory should determine its own reference range(s)
for the diagnostic evaluation of patient results.45

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Performance Characteristics
Specificity
The potential interference of various endogenous proteins found in increased concentrations
during pregnancy was tested by adding these proteins to serum pools containing AFP at three
different concentrations. The AFP levels were then determined and compared to a serum
control containing no added protein. Human proteins tested were α1-glycoprotein,
α1-antitrypsin, α-globulin, ceruloplasmin, chorionic gonadotropin, γ-globulin, placental
lactogen, transferrin, luteotropic hormone, fetal hemoglobin, and pregnancy-associated
glycoprotein. There was no interference or cross-reactivity by any of these compounds.
Common drugs such as aspirin and acetaminophen and vitamins commonly prescribed during
pregnancy also demonstrated no interference in the measurement of AFP.

Interference by Chemotherapeutic Agents

The potential interference of chemotherapeutic agents was tested by adding these agents to
serum pools containing AFP at three different concentrations. The level of AFP in each of
these pools was then determined and normalized to the level without the respective drugs.
Amount Added Mean % Recovery
Substance (µg/mL) (Spike/control x 100)
Bleomycin 1300 101
Cisplatin 1500 97
Cyclophosphamide 330 101
Doxorubicin 10 99
5-fluorouracil 360 101
Methotrexate 13 106
Mitomycin-C 60 99
Vinblastine 1200 100
Vincristine 700 99

Interference testing was determined according to CLSI Document EP7-A2.46

Sensitivity and Assay Range

The ADVIA Centaur AFP assay measures AFP concentrations up to 1000 ng/mL (830 IU/mL)
with a minimum detectable concentration (sensitivity) of 1.3 ng/mL (1.08 IU/mL) based on
eight patient profiles. Sensitivity was determined by diluting low patient specimens with the
zero standard. The sensitivity of the assay with a particular patient specimen was taken as that
concentration which was statistically different from both the zero standard and the next lowest
dilution of patient sample.

Method Comparison
For 498 serum samples in the range of 1.3 to 943.6 ng/mL (1.1 to 783.2 IU/mL), the
relationship of the ADVIA Centaur AFP assay to the ACS:180 AFP assay is described
by the following equation:
ADVIA Centaur AFP = 1.05 (ACS:180 AFP) – 0.3 ng/mL
Correlation coefficient (r) = 0.99
For 355 amniotic fluid samples in the range of 0.3 to 86.9 μg/mL, the relationship of the
ADVIA Centaur AFP assay to to the ACS:180 AFP assay is described by the following
equation:
ADVIA Centaur AFP = 0.94 (ACS:180 AFP) + 0.1 μg/mL
Correlation coefficient (r) = 0.99

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Dilution Recovery
Six serum samples in the range of 336.3 to 594.7 ng/mL (279.1 to 493.6 IU/mL) were diluted
1:2, 1:4, 1:8, 1:16, and 1:32 with Multi-Diluent 2 and assayed for recovery and parallelism.
The recoveries ranged from 78.1% to 117.2% with a mean of 100.5%.
Observed Expected Observed Expected
Sample Dilution (ng/mL) (ng/mL) (IU/mL) (IU/mL) Recovery %
1 — 594.7 493.6
1:2 286.6 297.4 237.9 246.8 96.4
1:4 147.7 148.7 122.6 123.4 99.3
1:8 78.3 74.3 65.0 61.7 105.4
1:16 39.3 37.2 32.6 30.9 105.6
1:32 20.0 18.6 16.6 15.4 107.5
Mean 102.8
2 — 472.1 391.9
1:2 239.0 236.1 198.4 195.9 101.2
1:4 123.9 118.0 102.8 98.0 105.0
1:8 63.7 59.0 52.9 49.0 108.0
1:16 33.1 29.5 27.5 24.5 112.2
1:32 16.9 14.8 14.1 12.2 114.2
Mean 108.1
3 — 405.3 336.4
1:2 203.2 202.7 168.7 168.2 100.2
1:4 103.3 101.3 85.7 84.1 102.0
1:8 53.3 50.7 44.2 42.1 105.0
1:16 28.1 25.3 23.3 21.0 111.1
1:32 14.5 12.7 12.0 10.5 114.2
Mean 106.5
4 — 388.1 322.1
1:2 191.4 194.1 158.8 161.1 98.6
1:4 99.2 97.0 82.3 80.5 102.3
1:8 52.0 48.5 43.2 40.3 107.2
1:16 27.9 24.3 23.1 20.1 114.8
1:32 14.2 12.1 11.8 10.1 117.4
Mean 108.1
5 — 346.3 287.4
1:2 161.0 173.2 133.7 143.7 93.0
1:4 78.1 86.6 64.8 71.9 90.2
1:8 38.7 43.3 32.1 35.9 89.4
1:16 20.7 21.6 17.2 18.0 95.8
1:32 11.1 10.8 9.2 9.0 102.8
Mean 94.2
6 — 336.3 279.2
1:2 146.5 168.2 121.6 139.6 87.1
1:4 67.8 84.1 56.3 69.8 80.6
1:8 32.8 42.0 27.2 34.9 78.1
1:16 17.3 21.0 14.4 17.4 82.4
1:32 9.1 10.5 7.5 8.7 86.7
Mean 83.0
Mean 100.5

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ADVIA Centaur and ADVIA Centaur XP Systems AFP
Five amniotic fluid samples in the range of 8.6 to 11.5 μg/mL of AFP were diluted 1:20, 1:40,
1:80, 1:160, and 1:320 with Multi-Diluent 2 following an initial dilution to bring the sample
within the range of the ADVIA Centaur AFP assay. The recoveries ranged from 96.8% to
115.5% with a mean of 104.0%. The first measurable dilution was assigned a value of 100%.
Observed Expected
Sample Dilution (µg/mL) (µg/mL) Recovery %
1 —
1:20 11.2 11.5 97.4
1:40 11.3 11.5 98.3
1:80 11.5 11.5 100.0
1:160 11.9 11.5 103.5
1:320 12.0 11.5 104.3
Mean 100.7
2 —
1:20 8.6 8.6 100.0
1:40 8.7 8.6 101.2
1:80 9.2 8.6 107.0
1:160 9.5 8.6 110.5
1:320 9.6 8.6 111.6
Mean 106.1
3 —
1:20 9.5 9.7 97.9
1:40 10.0 9.7 103.1
1:80 10.2 9.7 105.2
1:160 10.7 9.7 110.3
1:320 11.2 9.7 115.5
Mean 106.4
4 —
1:20 9.8 9.9 99.0
1:40 9.9 9.9 100.0
1:80 10.2 9.9 103.0
1:160 10.3 9.9 104.0
1:320 10.8 9.9 109.1
Mean 103.0
5 —
1:20 9.2 9.5 96.8
1:40 9.5 9.5 100.0
1:80 9.8 9.5 103.2
1:160 10.1 9.5 106.3
1:320 10.8 9.5 113.7
Mean 104.0
Mean 104.0

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AFP ADVIA Centaur and ADVIA Centaur XP Systems

Spiking Recovery
Known amounts of AFP ranging from 21.2 to 342.4 ng/mL (17.6 to 284.2 IU/mL) were added
to five patient samples with endogenous AFP levels between 35.6 and 47.4 ng/mL (29.6 to
39.3 IU/mL). When compared to expected values, the measured (recovered) levels of AFP
averaged 99.0% with a range of 91.9% to 109.2%.
Amount Added Observed Amount Added Observed
Sample (ng/mL) (ng/mL) (IU/mL) (IU/mL) Recovery %
1 — 35.6 — 29.5
21.2 56.4 17.6 46.8 98.1
89.7 118.3 74.5 98.2 92.2
155.1 197.6 128.7 164.0 104.4
342.4 387.7 284.2 321.8 102.8
Mean 99.4
2 — 39.2 — 32.5
21.2 58.8 17.6 48.8 92.5
89.7 122.0 74.5 101.3 92.3
155.1 199.6 128.7 165.7 103.4
342.4 377.5 284.2 313.3 98.8
Mean 96.7
3 — 39.9 — 33.1
21.2 61.4 17.6 51.0 101.4
89.7 125.0 74.5 103.8 94.9
155.1 209.3 128.7 173.7 109.2
342.4 386.1 284.2 320.5 101.1
Mean 101.7
4 — 47.4 — 39.3
21.2 69.3 17.6 57.5 103.3
89.7 136.3 74.5 113.1 99.1
155.1 205.1 128.7 170.2 101.7
342.4 387.8 284.2 321.9 99.4
Mean 100.9
5 — 41.5 — 34.4
21.2 62.2 17.6 51.6 97.6
89.7 133.3 74.5 110.6 102.3
155.1 185.1 128.7 153.6 92.6
342.4 356.2 284.2 295.6 91.9
Mean 96.1
Mean 99.0

Precision
Seven samples were assayed 3 times, in ≥ 4 runs, on ≥ 2 systems, (n = 113 for each sample),
over a period of 2 days. The following results were obtained:
Mean (ng/mL) Mean (IU/mL) Within-run % CV Run-to-run % CV Total % CV
16.5 13.7 3.6 4.4 5.7
21.9 18.2 4.0 3.8 6.1
37.3 31.0 3.3 3.9 5.7
67.1 55.7 2.7 3.4 4.9
173.9 144.3 2.8 3.7 5.0
499.9 414.9 3.1 2.0 5.0
732.0 607.6 3.4 2.0 5.5

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ADVIA Centaur and ADVIA Centaur XP Systems AFP
Standardization
The ADVIA Centaur AFP assay standardization is traceable to World Health Organization
(WHO) Reference Preparation for human AFP (72/225)47-48 using highly purified AFP.
A comparison over the full assay range gave the following correlation:
Siemens AFP = 0.99 (WHO) – 2.8 ng/mL
r = 0.99
Assigned values for calibrators are traceable to this standardization.

Evaluating Results
The following is recommended when you observe poor reproducibility of AFP values at low
levels or if you are not satisfied with assay performance:
• Ensure that the assay reagent and calibrator lot numbers and expiration dates match those
entered in the system.
• Ensure that the calibrators, quality control materials, and assay reagents were prepared
according to the recommended procedures.
• Ensure that the recommended sample collection and handling procedures were followed.
• Ensure that the recommended system cleaning procedures were followed.
• Ensure that Type II reagent water was used when operating the system.38
• Visually check the probe and tubing for obstructions, leaks, and deformities such as
pinched or crimped tubing.
• Take further corrective action following established laboratory procedures.
• Calibrate the system using new assay reagents, calibrators, and quality control samples.
• Contact Siemens for technical assistance.

Technical Assistance
For customer support, please contact your local technical support provider or distributor.
www.siemens.com/diagnostics

References
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fetoprotein. Int J Cancer 1971;7:218–25.
2. Bergstrand CG, Czar B. Demonstration of a new protein fraction in serum from the human fetus. Scand J Clin
& Lab Invest 1956;8:174.
3. Ruoslahti E, Engvall E, Kessler MJ. Chemical properties of alpha-fetoprotein. In: Herberman RB, McIntire
KR, editors. Immunodiagnosis of cancer. NY: Marcel Dekker, 1979. p.101–17.
4. Morinaga T, Sakai M, Wegmann T, Tamaoki T. Primary structures of human α fetoprotein and its mRNA. Proc
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5. Gitlin D, Perricelli A, Gitlin G. Synthesis of α-fetoprotein by liver, yolk sac and gastrointestinal tract of the
human conceptus. Cancer Res 1972;32:979–82.
6. Harris R, Jennison RF, Barson AJ, Laurence KM, Ruoslahti E, Seppala M. Comparison of amniotic-fluid and
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Lancet 1974;i(855):428.
7. Brock DJH, Bolton AE, Monaghan JM. Prenatal diagnosis of anencephaly through maternal serum alpha
fetoprotein measurement. Lancet 1973;ii(835):923–4.
8. Wald NJ, Brock DJH, Bonnar J. Prenatal diagnosis of spina bifida and anencephaly by maternal serum alpha-
fetoprotein measurement, a controlled study. Lancet 1974;i:765–7.

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AFP ADVIA Centaur and ADVIA Centaur XP Systems

9. Maternal serum alpha-fetoprotein measurement in antenatal screening for anencephaly and spina bifida in
early pregnancy. Report of the UK collaborative study on alpha-fetoprotein in relation to neural tube defects.
Lancet 1977;i:1323–32.
10. Second report of the UK collaborative study on alpha-fetoprotein in relation to neural-tube defects. Lancet
1979;ii:652–62.
11. Fourth report of the UK collaborative study on alpha-fetoprotein in relation to neural tube defects. J Epidemiol
Community Health 1982;36:87–95.
12. Johnson AM, Palomaki GE, Haddow JE. Maternal serum α fetoprotein levels in pregnancies among black and
white women with fetal open spina bifida; a US collaborative study. J Obstet Gynecol 1990;162:328–31.
13. Brock DJH. The prenatal diagnosis of neural tube defects. Obstet Gynocol Surv 1976;31(1):32–40.
14. Knight GJ. Maternal serum α-fetoprotein screening. In: Hommes FA, editor. Techniques in diagnostic human
biochemical genetics; a laboratory manual. NY: Wiley-Liss, Inc, 1991. p.491–518.
15. Burton BK. Elevated maternal serum alpha-fetoprotein (MSAFP); interpretation and follow-up. Clin Obstet
Gynecol 1988;31(1):293–305.
16. Palomaki GE, Hill LE, Knight GJ, et al. Second trimester maternal serum alpha-fetoprotein levels in
pregnancies associated with gastroschisis and omphalocele. Obstet & Gynecol 1988;71(6 Pt i):906–09.
17. Seppala M, Rapola J, Huttunen NP, et al. Congenital nephrotic syndrome: prenatal diagnosis and genetic
counseling by estimation of amniotic fluid and maternal serum alpha-fetoprotein. Lancet 1976;ii:123–24.
18. Abelev GI. Study of the antigenic structure of tumors. Acta Unio Internationalis Contra Cancrum 1963;
19:80–2.
19. Tatarinov YS. Detection of an embryo-specific alpha-globulin in the blood sera of patients with primary liver
tumor. Vopr Med Khim 1964;10:90–1.
20. Abelev GI. Alpha-fetoprotein in ontogenesis and its association with malignant tumors. Adv Cancer Res
1971;14:295–358.
21. Chen Ding-Shinn, Sung, Juei-Low. Serum alpha fetoprotein in hepatocellular carcinoma. Cancer
1977;40:779–83.
22. McIntire KR, Vogel CL, Princeler GL, et al. Serum α-fetoprotein as a biochemical marker for hepatocellular
carcinoma. Cancer Res 1972;32:1941–6.
23. Kawai M, Furuhashi Y, Kano T, et al. α fetoprotein in malignant germ cell tumors of the ovary. Gynecol Oncol
1990;39:160–6.
24. Javadpour N. Serum and cellular biologic tumor markers in patients with urologic cancer. Hum Pathol
1979;10(5):557–68.
25. Masopust J, et al. Occurrence of fetoprotein in patients with neoplasms and non-neoplastic diseases. Int J
Cancer 1968;3:364–73.
26. Waldmann TA, McIntire KR. The use of a radio-immunoassay for alpha-fetoprotein in the diagnosis of
malignancy. Cancer 1974;34(4 Sup):1510–5.
27. Javadpour N, McIntire KR, Waldmann TA. Human chorionic gonadotropin and alpha-fetoprotein in sera and
tumor cells of patients with testicular seminoma. Cancer 1978;42:2768–72.
28. Lange PH, McIntire KR, Waldmann TA, et al. Serum alpha-fetoprotein and human chorionic gonadotropin in
the diagnosis and management of nonseminomatous germ-cell testicular cancer. N Engl J Med
1976;295(22):1237–40.
29. Javadpour N, McIntire KR, Waldmann TA, et al. The role of the radioimmunoassay of serum alpha-fetoprotein
and human chorionic gonadotropin in the intensive chemotherapy and surgery of metastatic testicular tumors.
J Urol 1978;119:759–62.
30. Kohn J, Orr AH, McElwain TJ, et al. Serum alpha1-fetoprotein in patients with testicular tumors. Lancet
1976;ii:433–6.
31. Perlin E, Engeler JE, Edson M, et al. The value of serial measurement of both human chorionic gonadotropin
and alpha-fetoprotein for monitoring germinal cell tumors. Cancer 1976;37:215–9.
32. Scardino PT, Cox HD, Waldmann TA, et al. The value of serum tumor markers in the staging and prognosis of
germ cell tumors of the testis. J Urol 1977;118:994–9.
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34. Mason MD. Tumour markers. In:Horwich A, editor. Testicular cancer investigation and management.
Baltimore: Williams and Wilkins, 1991. p.35–50.
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of complete response and survival in non-seminomatous germ cell tumors. Proc Am Soc Clin Oncol
1990;9:133.
36. Kirkpatrick AM, Kirkpatrick KA. Clearance-corrected differencing and other analytic techniques useful in the
interpretation of serum AFP values. In: Kirkpatrick AM, et al, editors. Alpha-fetoprotein: laboratory
procedures and clinical applications. NY: Masson, 1981. p.135–48.
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38. Reagent Water Technical Bulletin. Siemens Healthcare Diagnostics, 107060.
39. Clinical and Laboratory Standards Institute (formerly NCCLS). Procedures for the Handling and Processing
of Blood Specimens; Approved Guideline - Third Edition. Wayne, PA: Clinical and Laboratory Standards
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delivery. Am J Obstet Gynecol 1972;112(2):208–12.
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43. Sizaret P. Equivalence between international units and mass units of alpha-fetoprotein. Clin Chimica Acta
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Approved Guideline - Second Edition. Wayne, PA: Clinical and Laboratory Standards Institute; 2005. NCCLS
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ADVIA Centaur, ReadyPack, and ACS:180 are trademarks of Siemens Healthcare Diagnostics.
© 2008 Siemens Healthcare Diagnostics Inc. All rights reserved.
US Pats 4,745,181; 4,918,192; 5,110,932; 5,609,822; 5,788,928

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AFP ADVIA Centaur and ADVIA Centaur XP Systems

Understanding the Symbols

The following symbols may appear on the product labeling:
Symbol Definition Symbol Definition

In vitro diagnostic medical device Catalog number

Authorized Representative in the European

Manufacturer
Community

CE Mark with identification number of notified

CE Mark
body

Consult instructions for use Caution! Potential Biohazard

Do not freeze (> 0°C) Temperature limitation (2–8°C)

Lower limit of temperature (≥ 2°C) Upper limit of temperature (≤ -10°C)

Keep away from sunlight Use by

Shake the reagent pack vigorously.

Refer to Loading Reagents in the assay-specific
Store upright ADVIA Centaur product instructions for detailed
information.

Batch code Contains sufficient for (n) tests

2010-01 Date format (year-month) Printed with soy ink

Green dot Recycle

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