11899040001V18.

Anti-HBs
Antibody to hepatitis B surface antigen (anti-HBs)

Elecsys 2010
MODULAR ANALYTICS E170
11820524 122 100 cobas e 411
cobas e 601
cobas e 602

English Precautions and warnings

Intended use For in vitro diagnostic use.
Exercise the normal precautions required for handling all laboratory
Immunoassay for the in vitro quantitative determination of human antibodies reagents.
to the hepatitis B surface antigen (HBsAg) in human serum and plasma. Disposal of all waste material should be in accordance with local guidelines.
The electrochemiluminescence immunoassay “ECLIA” is intended for use Safety data sheet available for professional user on request.
on Elecsys and cobas e immunoassay analyzers. All human material should be considered potentially infectious. The
Summary calibrators (A‑HBS Cal1 and A‑HBS Cal2) have been prepared exclusively
Anti‑HBs is a specific (generally IgG) antibody that is directed against the from the blood of donors tested individually and shown to be free from
hepatitis B surface antigen.1 Anti‑HBs can be formed following a hepatitis B HBsAg and antibodies to HCV and HIV. The testing methods applied were
infection or after hepatitis B vaccination. Antibodies are formed against the FDA‑approved or cleared in compliance with the European Directive
HBsAg determinant a, which is common to all subtypes, and against 98/79/EC, Annex II, List A.
subtype‑specific determinants.2 The HBsAg starting material used was inactivated prior to labeling with
Anti‑HBs tests are used within the scope of hepatitis B vaccination to check biotin or ruthenium by heating to 60 °C for 15 hours. In addition, any virus
the necessity and success of vaccination.3,4,5 In addition, anti‑HBs tests are particles remaining were removed by ultracentrifugation.
used to monitor the course of disease following acute hepatitis B infection.3 However, as no testing method can rule out the potential risk of infection
The Elecsys Anti‑HBs assay uses a mixture of purified antigens of the with absolute certainty, the material should be handled with the same level
HBsAg subtypes ad and ay from human serum. of care as a patient specimen. In the event of exposure, the directives of the
responsible health authorities should be followed.6,7
Test principle Avoid foam formation in all reagents and sample types (specimens,
Sandwich principle. Total duration of assay: 18 minutes. calibrators and controls).
▪ 1st incubation: Anti‑HBs in the sample (40 µL), biotinylated HBsAg Reagent handling
(ad/ay), and HBsAg (ad/ay) labeled with a ruthenium complexa) react to The reagents in the kit are ready for use and are supplied in bottles
form a sandwich complex. compatible with the system.
▪ 2nd incubation: After addition of streptavidin-coated microparticles, the Elecsys 2010 and cobas e 411 analyzers: The calibrators should only be
complex becomes bound to the solid phase via interaction of biotin and left on the analyzers during calibration at 20‑25 °C. After use, close the
streptavidin. bottles as soon as possible and store at 2‑8 °C.
▪ The reaction mixture is aspirated into the measuring cell where the Due to possible evaporation effects, not more than 5 calibration procedures
microparticles are magnetically captured onto the surface of the per bottle set should be performed.
electrode. Unbound substances are then removed with
ProCell/ProCell M. Application of a voltage to the electrode then induces MODULAR ANALYTICS E170, cobas e 601 and cobas e 602 analyzers:
chemiluminescent emission which is measured by a photomultiplier. Unless the entire volume is necessary for calibration on the analyzer,
transfer aliquots of the ready‑for‑use calibrators into empty snap‑cap bottles
▪ Results are determined via a calibration curve which is instrument- (CalSet Vials). Attach the supplied labels to these additional bottles. Store
specifically generated by 2‑point calibration and a master curve provided the aliquots at 2‑8 °C for later use.
via the reagent barcode. Perform only one calibration procedure per aliquot.
a) Tris(2,2'-bipyridyl)ruthenium(II)-complex (Ru(bpy) )
All information required for correct operation is read in from the respective
Reagents - working solutions reagent barcodes.
The reagent rackpack (M, R1, R2) is labeled as A‑HBS. Please note: Both the vial labels, and the additional labels (if available)
contain 2 different barcodes. The barcode between the yellow markers is
M Streptavidin-coated microparticles (transparent cap), 1 bottle, 6.5 mL: for cobas 8000 systems only. If using a cobas 8000 system, please turn
Streptavidin-coated microparticles 0.72 mg/mL; preservative. the vial cap 180° into the correct position so the barcode can be read by the
system. Place the vial on the instrument as usual.
R1 HBsAg~biotin (gray cap), 1 bottle, 10 mL:
Storage and stability
Biotinylated HBsAg (ad/ay) human, > 0.5 mg/L; MESb) buffer Store at 2‑8 °C.
85 mmol/L, pH 6.5; preservative.
Do not freeze.
R2 HBsAg~Ru(bpy) (black cap), 1 bottle, 8 mL: Store the Elecsys reagent kit upright in order to ensure complete
HBsAg (ad/ay) human, labeled with ruthenium complex > 0.3 mg/L; availability of the microparticles during automatic mixing prior to use.
MES buffer 85 mmol/L, pH 6.5; preservative. Stability of the reagent rackpack
b) MES = 2-morpholino-ethane sulfonic acid unopened at 2‑8 °C up to the stated expiration date
A-HBS Cal1 Calibrator 1 (white cap), 2 bottles of 1.3 mL each: after opening at 2‑8 °C 8 weeks
Anti‑HBs (human) in human serum; preservative. on MODULAR ANALYTICS E170, 8 weeks
cobas e 601 and cobas e 602
A-HBS Cal2 Calibrator 2 (black cap), 2 bottles of 1.3 mL each:
on Elecsys 2010 and cobas e 411 4 weeks
Anti‑HBs (human) in human serum; preservative.
Stability of the calibrators
unopened at 2‑8 °C up to the stated expiration date

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Anti-HBs
Antibody to hepatitis B surface antigen (anti-HBs)

Stability of the calibrators ▪  03023141001, PC/CC‑Cups, 12 cups to prewarm ProCell M and

CleanCell M before use
after opening at 2‑8 °C 8 weeks
▪  03005712190, ProbeWash M, 12 x 70 mL cleaning solution for run
on Elecsys 2010 and cobas e 411 up to 5 hours finalization and rinsing during reagent change
at 20‑25 °C
▪  12102137001, AssayTip/AssayCup Combimagazine M,
on MODULAR ANALYTICS E170, use only once 48 magazines x 84 reaction vessels or pipette tips, waste bags
cobas e 601 and cobas e 602 ▪  03023150001, WasteLiner, waste bags
Store calibrators upright in order to prevent the calibrator solution from ▪  03027651001, SysClean Adapter M
adhering to the snap‑cap. Accessories for all analyzers:
Specimen collection and preparation ▪  11298500316, Elecsys SysClean, 5 x 100 mL system cleaning
Only the specimens listed below were tested and found acceptable. solution
Serum collected using standard sampling tubes or tubes containing Assay
separating gel. For optimum performance of the assay follow the directions given in this
K3-EDTA plasma. document for the analyzer concerned. Refer to the appropriate operator’s
Criterion: Recovery within 90‑110 % of serum value or slope manual for analyzer‑specific assay instructions.
0.9‑1.1 + intercept within < ± 2x analytical sensitivity (LDL) + coefficient of Resuspension of the microparticles takes place automatically prior to use.
correlation > 0.95. Read in the test‑specific parameters via the reagent barcode. If in
If plasma treated with lithium heparin, sodium citrate or sodium exceptional cases the barcode cannot be read, enter the 15‑digit sequence
fluoride/potassium oxalate is used, the values obtained are 25 % lower than of numbers.
those obtained from serum. Bring the cooled reagents to approximately 20 °C and place on the reagent
Do not use lithium heparin plasma tubes containing separating gel. disk (20 °C) of the analyzer. Avoid foam formation. The system
automatically regulates the temperature of the reagents and the
Stable for 6 days at 2-8 °C, 3 months at -20 °C. The samples may be frozen opening/closing of the bottles.
6 times.
Place the calibrators in the sample zone.
The sample types listed were tested with a selection of sample collection
tubes that were commercially available at the time of testing, i.e. not all All the information necessary for calibrating the assay is automatically read
available tubes of all manufacturers were tested. Sample collection systems into the analyzer.
from various manufacturers may contain differing materials which could After calibration has been performed, store the calibrators at 2‑8 °C or
affect the test results in some cases. When processing samples in primary discard (MODULAR ANALYTICS E170, cobas e  601 and cobas e  602
tubes (sample collection systems), follow the instructions of the tube analyzers).
manufacturer.
Calibration
Centrifuge samples containing precipitates and frozen samples before Traceability: This method has been standardized against the 1st WHO
performing the assay. Reference Standard 1977.
Do not use heat‑inactivated samples. Every Elecsys Anti-HBs reagent set has a barcoded label containing
Do not use samples and controls stabilized with azide. specific information for calibration of the particular reagent lot. The
Ensure the samples, calibrators and controls are at 20‑25 °C prior to predefined master curve is adapted to the analyzer using A‑HBS Cal1 and
measurement. A‑HBS Cal2.
Due to possible evaporation effects, samples and calibrators on the Calibration frequency: Calibration must be performed once per reagent lot
analyzers should be analyzed/measured within 2 hours. using A‑HBS Cal1, A‑HBS Cal2 and fresh reagent (i.e. not more than
24 hours since the reagent kit was registered on the analyzer).
Materials provided
Renewed calibration is recommended as follows:
See “Reagents – working solutions” section for reagents.
▪ after 1 month (28 days) when using the same reagent lot
▪ 2 x 6 bottle labels
▪ after 7 days (when using the same reagent kit on the analyzer)
Materials required (but not provided)
▪ as required: e.g. quality control findings with PreciControl Anti‑HBs
▪  11876317122, PreciControl Anti-HBs, for 8 x 1.3 mL each of outside the defined limits
PreciControl Anti-HBs 1 and 2
▪ more frequently when this is required by pertinent regulations
▪  11732277122, Diluent Universal, 2 x 16 mL sample diluent or
 03183971122, Diluent Universal, 2 x 36 mL sample diluent Range (in IU/L) for the calibrators: 4‑15 for calibrator 1 (A‑HBS Cal1) and
350‑600 for calibrator 2 (A‑HBS Cal2).
▪  11776576322, CalSet Vials, 2 x 56 empty snap-cap bottles
Quality control
▪ General laboratory equipment
For quality control, use PreciControl Anti‑HBs.
▪ Elecsys 2010, MODULAR ANALYTICS E170 or cobas e analyzer Controls for the various concentration ranges should be run individually at
Accessories for Elecsys 2010 and cobas e 411 analyzers: least once every 24 hours when the test is in use, once per reagent kit, and
▪  11662988122, ProCell, 6 x 380 mL system buffer following each calibration.
▪  11662970122, CleanCell, 6 x 380 mL measuring cell cleaning The control intervals and limits should be adapted to each laboratory’s
solution individual requirements. Values obtained should fall within the defined
limits. Each laboratory should establish corrective measures to be taken if
▪  11930346122, Elecsys SysWash, 1 x 500 mL washwater additive values fall outside the defined limits.
▪  11933159001, Adapter for SysClean If necessary, repeat the measurement of the samples concerned.
▪  11706802001, Elecsys 2010 AssayCup, 60 x 60 reaction vessels Follow the applicable government regulations and local guidelines for
quality control.
▪  11706799001, Elecsys 2010 AssayTip, 30 x 120 pipette tips
Accessories for MODULAR ANALYTICS E170, cobas e 601 and Calculation
cobas e 602 analyzers: The analyzer automatically calculates the analyte concentration of each
sample in IU/L.
▪  04880340190, ProCell M, 2 x 2 L system buffer
▪  04880293190, CleanCell M, 2 x 2 L measuring cell cleaning
solution
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Anti-HBs
Antibody to hepatitis B surface antigen (anti-HBs)

Limitations - interference After manual dilution, multiply the result by the dilution factor.
The assay is unaffected by icterus (bilirubin < 513 µmol/L or < 30 mg/dL), After dilution by the analyzers, the MODULAR ANALYTICS E170,
hemolysis (Hb < 0.93 mmol/L or < 1.5 g/dL), lipemia (Intralipid Elecsys 2010 and cobas e software automatically takes the dilution into
< 1500 mg/dL) and biotin (< 123 nmol/L or < 30 ng/mL). account when calculating the sample concentration.
Criterion: Recovery within ± 10 % of initial value. Manual dilution can also be made with negative human serum.
Samples should not be taken from patients receiving therapy with high Note: Antibodies to HBsAg are heterogenous. In some isolated cases, this
biotin doses (i.e. > 5 mg/day) until at least 8 hours following the last biotin may lead to non‑linear dilution behavior.
administration.
Expected values
No interference was observed from rheumatoid factors up to a
concentration of 2100 IU/mL. Interpretation of the results
A high-dose hook effectc) may be observed for samples with anti‑HBs Samples with concentrations < 10 IU/L are considered non‑reactive in the
concentrations > 150000 IU/L. In rare cases a high‑dose hook effect Elecsys Anti‑HBs assay.
< 150000 IU/L cannot be excluded. In case of an unexpected low result, i.e. Samples with concentrations ≥ 10 IU/L are considered reactive in the
after re‑vaccination, the sample should be diluted 1:100 (refer to chapter Elecsys Anti‑HBs assay.
“Dilution”) and tested again. Note: Due to the diversity of the antibodies, the measured anti‑HBs value
In vitro tests were performed on 17 commonly used pharmaceuticals. No can vary depending on the testing procedure used. Results obtained from a
interference with the assay was found. single sample using tests from different manufacturers can therefore differ
In rare cases, interference due to extremely high titers of antibodies to by up to a factor of 4 (or even a factor of 10 in rare cases).8 If there is a
streptavidin and ruthenium can occur. The test contains additives which change in the assay procedure used during the monitoring of vaccination
minimize these effects. protection, then the anti‑HBs values obtained upon changing over to the
new procedure must be confirmed by parallel measurements with both
MODULAR ANALYTICS E170, cobas e  601 and cobas e  602 analyzers: methods.
In case the Elecsys HBsAg II/Anti‑HBs and HBeAg/Anti‑HBe assay
combinations are processed, make sure that these assays are entered in Vaccination strategies in certain risk groups are based on the measured
the “Special Wash” section of the system software and “Step1” (wash anti‑HBs concentration.4 Respective recommendations are given by
execute) is checked. Please refer to the operator's manual. national or regional guidelines.
For diagnostic purposes, the results should always be assessed in Specific performance data
conjunction with the patient’s medical history, clinical examination and other Representative performance data on the analyzers are given below.
findings. Results obtained in individual laboratories may differ.
c) High-dose hook effect: A sample with a true concentration clearly above the measuring range, Precision
but found within the measuring range.
Precision was determined using Elecsys reagents, human sera and
Limits and ranges controls.
Measuring range
2.00-1000 IU/L (defined by the detection limit and the maximum of the Elecsys 2010 and cobas e 411 analyzers
master curve). Values below the detection limit are reported as < 2.00 IU/L. Repeatabilityd) Intermediate precisione)
Values above the measuring range are reported as > 1000 IU/L (or up to
100000 IU/L for 100‑fold diluted samples). Sample Mean SD CV Mean SD CV
Detection limit IU/L IU/L % IU/L IU/L %
≤ 2.0 IU/L HSf), negative 8.7 0.57 6.6 7.73 0.76 9.9
The detection limit represents the lowest measurable analyte level that can HS, weakly positive 15.4 0.52 3.4 12.9 1.42 11.0
be distinguished from zero. It is calculated as the value lying two standard
deviations above that of the lowest standard (master calibrator, standard HS, positive 603 9.30 1.5 605 20.6 3.4
1 + 2 SD, repeatability study, n = 21). A‑HBS Cal1 10.7 0.67 6.3 11.0 1.79 16.2
Sensitivity in diluted samples A‑HBS Cal2 498 14.1 2.8 514 18.4 3.6
Two human sera reactive for anti‑HBs were stepwise diluted with serum
negative for anti‑HBs and assayed in duplicate with the Elecsys Anti‑HBs d) Repeatability = within‑run precision (n = 21)
assay and a commercial comparison test: e) Intermediate precision = between‑run (n = 10)
f) HS = human serum, negative
HS 1 Elecsys Comparison HS 2 Elecsys Comparison
Dilution Anti-HBs test Dilution Anti-HBs test MODULAR ANALYTICS E170, cobas e 601 and cobas e 602 analyzers
IU/L IU/L IU/L IU/L Repeatabilityg) Intermediate precisionh)
1 > 1000 > 150 1 > 1000 > 150 Sample Mean SD CV Mean SD CV
1:100 > 1000 > 150 1:100 > 1000 > 150 IU/L IU/L % IU/L IU/L %
1:500 310 > 150 1:500 261 > 150 HS, negative < 2.00 - - < 2.00 - -
1:1000 161 109 1:1000 136 99.0 HS, borderline 9.42 0.41 4.3 8.52 1.18 13.9
1:2500 69.5 34.3 1:2500 59.0 36.5 HS, positive 916 11.2 1.2 910 37.3 4.1
1:5000 36.5 17.9 1:5000 31.8 17.1 PC A‑HBS1i) < 2.00 - - < 2.00 - -
1:10000 19.8 8.4 1:10000 16.5 9.0 PC A‑HBS2 89.6 0.99 1.1 86 4.87 5.7
1:20000 10.3 4.3 1:20000 8.5 4.3 g) Repeatability = within‑run precision (n = 21)
h) Intermediate precision = within‑laboratory (modified protocol (EP5‑A) of the CLSI (Clinical and
Dilution Laboratory Standards Institute): 6 times daily for 10 days (n = 60))
Samples with anti‑HBs concentrations above the measuring range can be i) PC = PreciControl
diluted with Diluent Universal. The recommended dilution is 1:100 (either Analytical specificity
automatically by the MODULAR ANALYTICS E170, Elecsys 2010 and No cross‑reactions with HAV, HCV, HEV, CMV, EBV, HIV, Rubella,
cobas e analyzers or manually). The concentration of the diluted sample Toxoplasma gondii, and Treponema pallidum were observed.
must be > 10 IU/L.

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Anti-HBs
Antibody to hepatitis B surface antigen (anti-HBs)

Measurements were performed on each of the pathogens listed above Symbols

using ≥ 5 serum or plasma samples which were positive for antibodies to Roche Diagnostics uses the following symbols and signs in addition to
the above‑mentioned pathogens. those listed in the ISO 15223‑1 standard.
Clinical sensitivity
Contents of kit
Samples from various patient groups found to be reactive in a comparison
test were measured with the Elecsys Anti‑HBs assay and confirmed as Analyzers/Instruments on which reagents can be used
anti‑HBs‑positive by additional HBV‑tests.
Reagent
Characterization of Number Elecsys Anti-HBs Sensitivity Calibrator
samples tested Anti‑HBs comparison %
Volume after reconstitution or mixing
Sample groups reactive tests
reactive COBAS, COBAS E, ELECSYS, MODULAR and PRECICONTROL are trademarks of Roche. INTRALIPID is a
Convalescents 203 201 203 99.0 trademark of Fresenius Kabi AB.
All other product names and trademarks are the property of their respective owners.
positive for: a‑HBc,
Significant additions or changes are indicated by a change bar in the margin.
a‑HBe, a‑HBs © 2014, Roche Diagnostics
Drug addicts positive 53 52 52 100
for: a‑HBc, a‑HBs
Hospitalized patients 86 78 80 97.5
Roche Diagnostics GmbH, Sandhofer Strasse 116, D-68305 Mannheim
positive for: a‑HBc, www.roche.com
a‑HBs
Vaccinated persons 133 131 131 100
Total 475 462 466 99.0
Clinical specificity
Samples from blood donors which had not been selected and hospitalized
patients were used to determine the specificity.
Group Number Elecsys Anti‑HBs Specificity
tested false positive %
Blood donors a‑HBs 1469 3 99.8
< 10 IU/L (comparison test)
Hospitalized patients 297 0 100
negative for: a‑HBs, a‑HBc
(comparison tests)
References
1 Lander JJ, Holland PV, Alter HJ, et al. Antibody to hepatitis-associated
antigen. Frequency and patterns of response as detected by
radioimmunoprecipitation. J Am Med Assoc 1972;220:1079-1082.
2 Gold JW, Alter HJ, Holland PV, et al. Passive hemagglutination assay
for antibody to subtypes of hepatitis B antigen.
J Immunol 1974;112;1100-1106.
3 Jilg W, Schmidt M, Deinhardt F. Immune Response to Hepatitis B
Revaccination. J Med Virol 1988;24:377-384.
4 European Consensus Group on Hepatitis B immunity: Are booster
immunisations needed for lifelong hepatitis B immunity? Lancet
2000;355:561-565.
5 Hoofnagle JH, Di Bisceglie AM. Serologic Diagnosis of Acute and
Chronic Viral Hepatitis, Seminars in Liver Disease 1991;11/2:73-83.
6 Occupational Safety and Health Standards: bloodborne pathogens.
(29 CFR Part 1910.1030). Fed. Register.
7 Directive 2000/54/EC of the European Parliament and Council of
18 September 2000 on the protection of workers from risks related to
exposure to biological agents at work.
8 Bornhak H, Jilg W, Hüdig H, et al. Quantitation of anti-HBs in solid
phase immunoassays. What influences the results? Aus: Virushepatitis
A bis E. Diagnose, Therapie Prophylaxe. Kilian Verlag, ISBN:
3-9803688-1-5, 1994;212-221.
For further information, please refer to the appropriate operator’s manual for
the analyzer concerned, the respective application sheets, the product
information and the Method Sheets of all necessary components (if
available in your country).
A point (period/stop) is always used in this Method Sheet as the decimal
separator to mark the border between the integral and the fractional parts of
a decimal numeral. Separators for thousands are not used.

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