04855019001V8

Anti-HAV
Total antibodies (IgM and IgG) to the hepatitis A virus
REF 04854977 190 100 tests R2 Anti-HAV Ab~biotin; anti-HAV Ab~Ru(bpy)2+ 3 (black cap), 1 bottle, 8 mL:
Biotinylated monoclonal anti-HAV antibody (mouse) 0.25 µg/mL;
• Indicates analyzers on which the kit can be used
monoclonal anti-HAV antibody (mouse) labeled with ruthenium
MODULAR complex 0.15 µg/mL; HEPES buffer 50 mmol/L, pH 7.2; preservative.
Elecsys 2010 ANALYTICS cobas e 411 cobas e 601 cobas e 602 Cal1 Negative calibrator 1 (white cap), 2 bottles (lyophilized) for 1.0 mL each:
E170 Anti-HAV negative human serum; preservative.
• • • • • Cal2 Positive calibrator 2 (black cap), 2 bottles (lyophilized) for 1.0 mL each:
Anti-HAV (human) approx. 46 IU/L in human serum; preservative.
b) HEPES = [4-(2-hydroxyethyl)-piperazine]-ethanesulfonic acid
English
Precautions and warnings
Intended use For in vitro diagnostic use.
Immunoassay for the in vitro quantitative determination of total antibodies Exercise the normal precautions required for handling all laboratory reagents.
to the hepatitis A virus in human serum and plasma. The anti-HAV assay Disposal of all waste material should be in accordance with local guidelines.
is used as an aid to detect a past or existing hepatitis A infection, and to Safety data sheet available for professional user on request.
observe the immune response after HAV vaccination. All human material should be considered potentially infectious.
The electrochemiluminescence immunoassay “ECLIA” is intended for The calibrators (Cal1 and Cal2) have been prepared exclusively
use on Elecsys and cobas e immunoassay analyzers. from the blood of donors tested individually and shown to be free
from HBsAg and antibodies to HCV and HIV.
Summary The testing methods applied were FDA-approved or cleared in compliance
The hepatitis A virus is an RNA-containing virus that lacks an envelope. It with the European Directive 98/79/EC, Annex II, List A.
belongs to the family of picornaviruses. To date, just one human serotype and The serum containing anti-HAV (Cal2) and the HAV Ag (human; R1)
7 genotypes have been described. The viral capsid consists of 3 proteins were inactivated using β-propiolactone and UV-radiation.
(VP1-VP3) that form an immunodominant structure on the surface of the viral However, as no inactivation or testing method can rule out the potential
particle that is highly conserved between all genotypes. After vaccination or risk of infection with absolute certainty, the material should be treated just
natural infection, the immune response is directed against this structure.1 as carefully as a patient specimen. In the event of exposure the directives
Hepatitis A is the most common form of acute viral hepatitis. It is transmitted of the responsible health authorities should be followed.7,8
by the fecal-oral route. The disease has not been known to take a chronic The reagents may not be used after the stated expiration date.
course, nor does the virus persist in the organism.2 Avoid the formation of foam with all reagents and sample types
The hepatitis A virus is one of the most common causes (10-20 %) (specimens, calibrators, and controls).
of a fulminating course of viral hepatitis.3 Reagent handling
Total anti-HAV is positive at the onset of a hepatitis A infection (IgM). After The reagents in the kit are ready for use (except for Cal1 and Cal2) and
natural infection, anti-HAV IgG antibodies can usually be detected lifelong and are supplied in bottles compatible with the system.
provide protection against the disease if the organism is reinfected.4
Elecsys Anti-HAV Cal1 and Cal2: Dissolve carefully the contents of
Vaccines against hepatitis A and combined vaccines against hepatitis A
one bottle by adding exactly 1.0 mL of distilled or deionized water
and B are available today.5 Upon vaccination against hepatitis A,
and allow to stand closed for 15 minutes to reconstitute. Mix carefully,
anti-HAV IgG antibodies can be detected after 2 weeks. In the case of
avoiding the formation of foam. Transfer the reconstituted calibrator
complete immunization, protection usually lasts for years.6 There is no
into the empty labeled snap-cap bottles supplied.
limit value to define immune protection but anti-HAV values > 10-20 IU/L
are generally considered to be protective against infection. Elecsys 2010 and cobas e 411 analyzers: The reconstituted calibrators
Assays to detect anti-HAV antibodies are used to determine an Cal1 and Cal2 should only be left on the analyzer during calibration at
existing or past hepatitis A infection. They are also used to observe 20-25 °C. After use, close the bottles as soon as possible and store at
the immune response after HAV vaccination. 2-8 °C. Ensure that no calibration solution is trapped in the opened snap-cap.
Because of possible evaporation effects, not more than 5 calibration
Test principle procedures per calibrator bottle set should be performed.
Competition principle. Total duration of assay: 18 minutes. MODULAR ANALYTICS E170, cobas e 601 and cobas e 602 analyzers:
• 1st incubation: 50 µL of sample; the sample anti-HAV binds Unless the entire volume is necessary for calibration on the analyzer,
the added HAV antigen. transfer aliquots of the freshly reconstituted calibrator into empty snap-cap
bottles (CalSet Vials). Attach the supplied labels to these additional
• 2nd incubation: After addition of biotinylated antibodies and ruthenium
bottles. Store the aliquots for later use at 2-8 °C.
complexa -labeled antibodies specific for HAV antigen, together with
Perform only one calibration procedure per aliquot.
streptavidin-coated microparticles, the still-free binding sites on the
All information required for correct operation is read in via the
HAV antigens become occupied. The entire complex becomes bound
respective reagent barcodes.
to the solid phase via interaction of biotin and streptavidin.
• The reaction mixture is aspirated into the measuring cell where the Storage and stability
microparticles are magnetically captured onto the surface of the Store at 2-8 °C.
electrode. Unbound substances are then removed with ProCell. Store the Elecsys Anti-HAV reagent kit (M, R1, R2) upright in order to ensure
Application of a voltage to the electrode then induces chemiluminescent complete availability of the microparticles during automatic mixing prior to use.
emission which is measured by a photomultiplier. Stability:
• Results are determined via a calibration curve which is unopened at 2-8 °C up to the stated expiration date
instrument-specifically generated by 2-point calibration and a M, R1, R2 after opening at 2-8 °C 8 weeks
master curve provided via the reagent barcode. on MODULAR ANALYTICS E170,
a) Tris(2,2’-bipyridyl)ruthenium(II)-complex (Ru(bpy)2+
3 )
8 weeks
cobas e 601 and cobas e 602 at 20-25 °C
on Elecsys 2010 and cobas e 411 at 20-25 °C 8 weeks
Reagents - working solutions
lyophilized calibrators up to the stated expiration date
M Streptavidin-coated microparticles (transparent cap), 1 bottle, 6.5 mL:
reconstituted calibrators at 2-8 °C 6 weeks
Streptavidin-coated microparticles 0.72 mg/mL; preservative.
on Elecsys 2010 and cobas e 411 at 20-25 °C up to 5 hours
R1 HAV Ag (gray cap), 1 bottle, 7 mL:
HAV Ag (human) 40 U/mL (Roche units); HEPESb buffer on MODULAR ANALYTICS E170, use once only
50 mmol/L, pH 7.2; preservative. cobas e 601 and cobas e 602

2011-01, V 8 English 1/3 Elecsys and cobas e analyzers

Anti-HAV
Total antibodies (IgM and IgG) to the hepatitis A virus
Store the calibrators upright. Ensure that no calibration solution Resuspension of the microparticles takes place automatically before use.
is trapped in the opened snap-cap. Read in the test-specific parameters via the reagent barcode. If in exceptional
cases the barcode cannot be read, enter the 15-digit sequence of numbers.
Specimen collection and preparation
MODULAR ANALYTICS E170, Elecsys 2010 and cobas e analyzers: Bring
Only the specimens listed below were tested and found acceptable.
the cooled reagents to approx. 20 °C and place on the reagent disk (20 °C) of
Serum collected using standard sampling tubes or tubes
the analyzer. Avoid the formation of foam. The system automatically regulates
containing separating gel.
the temperature of the reagents and the opening/closing of the bottles.
Li-, Na-heparin, K3-EDTA, and sodium citrate plasma.
Place the reconstituted calibrators Cal1 and Cal2 in the sample zone of the
Criterion: Recovery within 90-110 % of serum value or slope 0.9-1.1 + intercept
analyzers. Only keep open during calibration. All information necessary for
within < ± 2 x analytical sensitivity (LDL) + coefficient of correlation > 0.95.
calibration is encoded on the barcoded bottle labels and is read in automatically.
Stable for 7 days at 2-8 °C, 3 months at -20 °C. The samples
After calibration has been performed, store Cal1 and Cal2 at 2-8 °C or discard
may be frozen 6 times.
(MODULAR ANALYTICS E170, cobas e 601 and cobas e 602 analyzers).
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 available Calibration
tubes of all manufacturers were tested. Sample collection systems from Traceability: This method has been standardized against the
various manufacturers may contain differing materials which could affect “Second International Standard for Anti-Hepatitis A, Immunoglobulin,
the test results in some cases. When processing samples in primary tubes Human, NIBSC code: 97/646” of the NIBSC (National Institute
(sample collection systems), follow the instructions of the tube manufacturer. for Biological Standards and Control).
Centrifuge samples containing precipitates and frozen samples before
performing the assay. Do not use heat-inactivated samples. Do not Calibration frequency: Calibration must be performed once per reagent lot
use samples and controls stabilized with azide. using Elecsys Anti-HAV Cal1 and Cal2 solutions, and fresh reagent (i.e. not
Ensure the patients’ samples, calibrators, and controls are at ambient more than 24 hours since the reagent kit was registered on the analyzer).
temperature (20-25 °C) before measurement. Renewed calibration is recommended as follows:
Because of possible evaporation effects, samples, calibrators, and controls • after 1 month (28 days) when using the same reagent lot
on the analyzers should be measured within 2 hours.
• after 7 days (when using the same reagent kit on the analyzer)
Materials provided • as required: e.g. quality control findings with Elecsys PreciControl
See “Reagents - working solutions” section for reagents. Anti-HAV reagents outside the specified limits
• 2 x 6 bottle labels • more frequently when this is required by pertinent regulations
• 4 empty labeled snap-cap bottles
Quality control
For quality control, use Elecsys PreciControl Anti-HAV.
Materials required (but not provided)
The controls 1 and 2 should be run as single determinations at least
• REF 04855043190, PreciControl Anti-HAV, for 2 x 4 mL each once every 24 hours when the test is in use, once per reagent kit, and
of PreciControl Anti-HAV 1 and 2 after every calibration. The control intervals and limits should be adapted
• REF 11361252122, Diluent Hepatitis A, 2 x 15 mL sample diluent to each laboratory’s individual requirements. Values obtained should
• REF 11776576322, CalSet Vials, 2 x 56 empty bottles with snap-caps fall within the defined limits. Each laboratory should establish corrective
• General laboratory equipment measures to be taken if values fall outside the limits.
• Elecsys 2010, MODULAR ANALYTICS E170 or cobas e analyzer If necessary, repeat the measurement of the samples concerned.
• Distilled or deionized water Follow the applicable government regulations and local guidelines
for quality control.
Accessories for Elecsys 2010 and cobas e 411 analyzers: Note:
• REF 11662988122, ProCell, 6 x 380 mL system buffer Always consider the value sheet contained in the kit of the assay/PreciControl
• REF 11662970122, CleanCell, 6 x 380 mL measuring cell cleaning solution and make sure whether target values for specific assay/PreciControl
• REF 11930346122, Elecsys SysWash, 1 x 500 mL washwater additive combinations need to be entered manually.
• REF 11933159001, Adapter for SysClean
Calculation
• REF 11706802001, Elecsys 2010 AssayCup, 60 x 60 reaction vessels
The analyzer automatically calculates the analyte concentration
• REF 11706799001, Elecsys 2010 AssayTip, 30 x 120 pipette tips of each sample in IU/L.
Accessories for MODULAR ANALYTICS E170, cobas e 601 Interpretation of the results:
and cobas e 602 analyzers: Samples with concentrations < 20 IU/L are not reactive in
• REF 04880340190, ProCell M, 2 x 2 L system buffer the Elecsys Anti-HAV test.
• REF 04880293190, CleanCell M, 2 x 2 L measuring cell cleaning solution Samples with concentrations ≥ 20 IU/L are reactive in the Elecsys Anti-HAV test.
• REF 03023141001, PC/CC-Cups, 12 cups to prewarm ProCell M Concentrations > 20 IU/L indicate an exisiting or past hepatitis A infection
and CleanCell M before use or the presence of anti-HAV antibodies after hepatitis A vaccination.
• REF 03005712190, ProbeWash M, 12 x 70 mL cleaning solution for Important! The anti-HAV values measured may differ depending on the assay
run finalization and rinsing during reagent change method used. The test result obtained of an individual sample can hence
• REF 12102137001, AssayTip/AssayCup Combimagazine M, 48 magazines vary when using assays of different manufacturers. If there is a change in
x 84 reaction vessels or pipette tips, waste bags the assay procedure during the monitoring of vaccination protection, then
• REF 03023150001, WasteLiner, waste bags the anti-HAV values obtained upon changing over to the new method must
• REF 03027651001, SysClean Adapter M be confirmed by parallel measurements by both methods.

Accessories for all analyzers: Limitations - interference

• REF 11298500316, Elecsys SysClean, 5 x 100 mL system cleaning solution The assay is unaffected by icterus (bilirubin < 855 µmol/L or < 50 mg/dL),
hemolysis (Hb < 0.745 mmol/L or < 1.2 g/dL), lipemia (Intralipid
Assay < 1000 mg/dL), and biotin < 205 nmol/L or < 50 ng/mL.
For optimum performance of the assay follow the directions given in Criterion: Recovery within ± 10 % of initial value.
this document for the analyzer concerned. Refer to the appropriate In patients receiving therapy with high biotin doses (i.e. > 5 mg/day), no sample
operator’s manual for analyzer-specific assay instructions. should be taken until at least 8 hours after the last biotin administration.

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04855019001V8

Anti-HAV
Total antibodies (IgM and IgG) to the hepatitis A virus
No interference was observed from rheumatoid factors up to Analytical specificity
a concentration of 1600 IU/mL. No cross-reactions with HBV, HCV, HIV, CMV, EBV*, HSV, Toxoplasma
In vitro tests were performed on 18 commonly used pharmaceuticals. gondii, Rubella*, and Treponema pallidum* were observed.
No interference with the assay was found. Measurements were performed using a total of 140 anti-HAV negative
In rare cases, interference due to extremely high titers of antibodies to serum and plasma samples which were positive for antibodies to the
analyte-specific antibodies, streptavidin or ruthenium can occur. These above-mentioned pathogens or contained autoantibodies (ANA, AMA).
effects are minimized by suitable test design. * 1 sample out of each group was weakly positive.
Important!
For diagnostic purposes, the results should always be assessed in conjunction Clinical sensitivity
with the patient’s medical history, clinical examination and other findings. In 97 samples of patients with an existing (anti-HAV IgM positive) or
Vaccination against hepatitis A should be considered where there is any past HAV infection, HAV antibodies (concentration > 20 IU/L) were
uncertainty, and in particular if the test results borderline the cutoff (20 IU/L). detected using the Elecsys Anti-HAV test.
165 samples from 46 HAV-vaccinated persons were tested using the Elecsys
Limits and ranges Anti-HAV test and a commercially available comparison test.
Measuring range In all samples the anti-HAV antibody concentration found was
3.00-60 IU/L (defined by the lower detection limit and the maximum of the above the respective limit.
master curve). Values below the detection limit are reported as < 3.00 IU/L.
Values above the measuring range are reported as > 60 IU/L or must be diluted. Clinical specificity
Samples containing anti-HAV IgM may, in isolated cases, show To investigate the specificity, a total of 1301 samples of non-selected
non-linear dilution behavior. blood donors, hospitalized patients, pregnant women, and dialysis
patients where no HAV infection was indicated were tested using
Lower limits of measurement
the Elecsys Anti-HAV test and a comparison test. Using the Elecsys
Lower detection limit
Anti-HAV assay, 1286 gave results < 20 IU/L.
Lower detection limit: < 3.0 IU/L
The specificity in this study is 98.85 %. The 95 % confidence
The detection limit represents the lowest measurable analyte level that
range is 98.11-99.35 %.
can be distinguished from zero. It is calculated as the value lying two
standard deviations above that of the lowest standard (master calibrator, References
standard 1 + 2 SD, repeatability study, n = 21). 1. Robertson BH, Nainan OV. Genetic and antigenetic variants of hepatitis A
virus. In: Viral Hepatitis and Liver Disease. Eds: Rizzeto M, Purcell RH,
Dilution
Gerin JL, Verme G, Edizioni Minerva Medica, Turin 1997;14-18.
Samples with anti-HAV concentrations above the measuring range can be
2. Koff RS. Hepatitis A. Lancet 1998;341:1643-1649.
diluted with Elecsys Diluent Hepatitis A. The concentration of the diluted
3. Lemon ML, Days SL. Type A hepatitis. In: Gorbach S, Bartlett JG, Blacklown
sample must be > 20 IU/L. Multiply the results by the dilution factor.
NL (eds). Infectious Diseases. Saunders WB, Philadelphia, 1992;705-708.
Specific performance data 4. Stapleton JT. Host Immune Response to Hepatitis A Virus.
Representative performance data on the analyzers are given below. JID 1995;171(Suppl 1):9-14.
Results obtained in individual laboratories may differ. 5. Ambrosch F, Wiedermann G, André FE, Delem A, Gregor H, Hofmann H,
et al. Clinical and Immunological Investigation of a New Combined
Precision Hepatitis A and Hepatitis B Vaccine. J Med Virol 1994;44:452-456.
Precision was determined using Elecsys reagents, human sera, and controls 6. Dobler G, Nitschko H, Frösner GG, Langer BCA. Hepatitis A:
(repeatability n = 21, intermediate precision n = 10); intermediate precision Medizinische Bedeutung, Klinik, Diagnostik, Aussagewert diagnostischer
on MODULAR ANALYTICS E170 analyzer was determined in a modified Verfahren. In: Frösner G (Hrsg.). Moderne Hepatitisdiagnostik,
protocol (EP5-A) of the CLSI (Clinical and Laboratory Standards Institute): Kilian Verlag 2001;19-30, 2nd edition.
6 times daily for 10 days (n = 60). The following results were obtained: 7. Occupational Safety and Health Standards: bloodborne pathogens.
Elecsys 2010 and cobas e 411 analyzers (29 CFR Part 1910.1030). Fed. Register.
Repeatabilityc Intermediate precision 8. Council Directive (2000/54/EC). Official Journal of the European
Sample Mean SD CV Mean SD CV Communities No. L262 from Oct. 17, 2000.
IU/L IU/L % IU/L IU/L % For further information, please refer to the appropriate operator’s manual
HSd, negative < 3.0 0.29 - < 3.0 0.69 - for the analyzer concerned, the respective application sheets, the product
HS, borderline cutoff 17.6 0.22 1.2 18.4 0.74 4.0 information, and the package inserts of all necessary components.
HS, positive 41.9 0.48 1.2 42.3 1.23 2.9
PCe A-HAV1 20.9 0.40 1.9 21.7 0.81 3.7
PC A-HAV2 35.8 0.84 2.3 36.5 1.02 2.8
COBAS, COBAS E, ELECSYS and MODULAR are trademarks of Roche. INTRALIPID is a trademark of
c) Repeatability = within-run precision Fresenius Kabi AB. Other brand or product names are trademarks of their respective holders.
d) HS = human serum Significant additions or changes are indicated by a change bar in the margin. Changes to reagent barcode
e) PC = PreciControl test parameters which have already been read in should be edited manually.
© 2011, Roche Diagnostics
MODULAR ANALYTICS E170, cobas e 601 and cobas e 602 analyzers
Repeatability Intermediate precision
Sample Mean SD CV Mean SD CV Roche Diagnostics GmbH, Sandhofer Strasse 116, D-68305 Mannheim
www.roche.com
IU/L IU/L % IU/L IU/L %
HS, negative < 3.0 0.34 - < 3.0 0.83 -
HS, borderline cutoff 21.8 0.11 0.5 29.2 0.89 3.0
HS, positive 55.2 0.30 0.5 51.3 1.12 2.2
PC A-HAV1 21.1 0.17 0.8 21.0 0.57 2.7
PC A-HAV2 35.2 0.23 0.7 34.8 0.56 1.6

2011-01, V 8 English 3/3 Elecsys and cobas e analyzers