Antihbs Arc
Antihbs Arc
Antihbs Arc
system
Anti-HBs
7C18
48-8436/R7
B7C180
Read Highlighted Changes Revised November, 2009
Anti-HBs
Customer Service: Contact your local representative or find country specific contact information on www.abbottdiagnostics.com
Package insert instructions must be carefully followed. Reliability of assay results cannot be guaranteed if there are any deviations from the instructions in this package insert.
Manufacturer
See REAGENTS section for a full explanation of symbols used in reagent component naming.
NAME
ARCHITECT Anti-HBs
INTENDED USE
The ARCHITECT Anti-HBs assay is a Chemiluminescent Microparticle Immunoassay (CMIA) for the quantitative determination of antibody to Hepatitis B surface antigen (anti-HBs) in human serum and plasma.
Assay Diluent ARCHITECT i Multi-Assay Manual Diluent (7D82-50) 1 Bottle (100 mL) ARCHITECT i Multi-Assay Manual Diluent containing phosphate buffered saline solution. Preservative: Antimicrobial Agent. Other Reagents ARCHITECT i Pre-Trigger Solution Pre-Trigger Solution containing 1.32% (w/v) hydrogen peroxide. ARCHITECT i Trigger Solution Trigger Solution containing 0.35 N sodium hydroxide. ARCHITECT i Wash Buffer Wash Buffer containing phosphate buffered saline solution. Preservatives: antimicrobial agents.
REAGENTS
Reagent Kit, 100/500 Tests NOTE: Some kit sizes are not available in all countries or for use on all ARCHITECT i Systems. Please contact your local distributor. ARCHITECT Anti-HBs Reagent Kit (7C18) 1 or 4 Bottle(s) (4.56 mL per 100 test bottle/ 16.80 mL per 500 test bottle) Hepatitis B Surface (E. coli, Recombinant) Antigen (Subtypes ad and ay) Coated Microparticles in TRIS buffer with protein stabilizers. Minimum concentration: 0.125% solids. Preservatives: sodium azide and antimicrobial agents. 1 or 4 Bottle(s) (5.9 mL per 100 test bottle/26.3 mL per 500 test bottle) Conjugate: Hepatitis B Surface (E. coli, Recombinant) Antigen (Subtypes ad and ay) Acridinium-Labeled Conjugate in MES buffer with protein stabilizers (Bovine and Human Plasma). Minimum concentration: 0.10 g/mL. Preservatives: sodium azide and antimicrobial agents.
Handling Precautions Do not use reagent kits beyond the expiration date. Do not pool reagents within a reagent kit or between reagent kits. Prior to loading the ARCHITECT Anti-HBs Reagent Kit on the system for the first time, the microparticle bottle requires mixing to resuspend microparticles that may have settled during shipment. For microparticle mixing instructions, refer to the PROCEDURE, Assay Procedure section of this package insert. Septums MUST be used to prevent reagent evaporation and contamination, and to ensure reagent integrity. Reliability of assay results cannot be guaranteed if septums are not used according to the instructions in this package insert. To avoid contamination, wear clean gloves when placing a septum on an uncapped reagent bottle. Once a septum has been placed on an open reagent bottle, do not invert the bottle as this will result in reagent leakage and may compromise assay results. Over time, residual liquids may dry on the septum surface. These are typically dried salts and have no effect on assay efficacy. For a detailed discussion of handling precautions during system operation, refer to the ARCHITECT System Operations Manual, Section 7.
Storage Instructions The ARCHITECT Anti-HBs Reagent Kit, Calibrators, and Controls must be stored at 2-8C in an upright position and may be used immediately after removal from 2-8C storage. When stored and handled as directed, reagents are stable until the expiration date. The ARCHITECT Anti-HBs Reagent Kit may be stored on board the ARCHITECT i System for a maximum of 30 days. After 30 days, the reagent kit must be discarded. For information on tracking onboard time, refer to the ARCHITECT System Operations Manual, Section 5. Reagents may be stored on or off the ARCHITECT i System. If reagents are removed from the system, store them at 2-8C (with septums and replacement caps) in an upright position. For reagents stored off the system, it is recommended they be stored in their original trays and boxes to ensure they remain upright. If the microparticle bottle does not remain upright (with a septum installed) while in refrigerated storage off the system, the reagent kit must be discarded. After reagents are removed from the system, you must initiate a scan to update the onboard stability timer.
Indications of Reagent Deterioration When a control value is out of the specified range, it may indicate deterioration of the reagents or errors in technique. Associated test results are invalid and will require retesting. Assay recalibration may be necessary. For troubleshooting information, refer to the ARCHITECT System Operations Manual, Section 10.
INSTRUMENT PROCEDURE
The ARCHITECT Anti-HBs assay file must be installed on the ARCHITECT i System from the ARCHITECT i assay CD-ROM prior to performing the assay. For detailed information on assay file installation and viewing and editing assay parameters, refer to the ARCHITECT System Operations Manual, Section 2. For information on printing assay parameters, refer to the ARCHITECT System Operations Manual, Section 5. For a detailed description of system procedures, refer to the ARCHITECT System Operations Manual. The default result unit for the ARCHITECT Anti-HBs assay is mIU/mL. An alternate result unit, IU/L, may be selected for reporting results by editing assay parameter Result concentration units to IU/L. The conversion factor used by the system is 1. Human serum (including serum collected in serum separator tubes) or plasma collected in potassium EDTA, sodium citrate, ACD, CPDA-1, and sodium heparin may be used in the ARCHITECT Anti-HBs assay. Liquid anticoagulants may have a dilutional effect resulting in lower concentrations for individual patient samples. Other anticoagulants have not been validated for use with the ARCHITECT Anti-HBs assay. Follow the tube manufacturers processing instructions for serum or plasma collection tubes. The ARCHITECT i System does not provide the capability to verify specimen type. It is the responsibility of the operator to verify the correct specimen types are used in the ARCHITECT Anti-HBs assay. Performance has not been established using cadaver specimens or body fluids other than human serum or plasma. Use caution when handling patient specimens to prevent cross contamination. Use of disposable pipettes or pipette tips is recommended. This assay was designed and validated for use with human serum or plasma from individual patient and donor specimens. Pooled specimens must not be used since the accuracy of their test results has not been validated.
Do not use heat-inactivated specimens. Do not use grossly hemolyzed specimens. For optimal results, inspect all samples for bubbles. Remove bubbles with an applicator stick prior to analysis. Use a new applicator stick for each sample to prevent cross contamination. For optimal results, serum and plasma specimens should be free of fibrin, red blood cells, or other particulate matter. Such specimens may give inconsistent results and must be transferred to a centrifuge tube and centrifuged at least 10,000 RCF (Relative Centrifugal Force) for 10 minutes. Ensure that complete clot formation in serum specimens has taken place prior to centrifugation. Some specimens, especially those from patients receiving anticoagulant or thrombolytic therapy may exhibit increased clotting time. If the specimen is centrifuged before a complete clot forms, the presence of fibrin may cause erroneous results. Specimens from heparinized patients may be partially coagulated and erroneous results could occur due to the presence of fibrin. To prevent this phenomenon, draw the specimen prior to heparin therapy. Gravity separation is not sufficient for specimen preparation. Specimens must be separated from clots or red blood cells using centrifugation as recommended by the tube manufacturer. Specimens may be stored on or off the clot or red blood cells for up to 14 days at 2-8C. If testing will be delayed more than 14 days, remove serum or plasma from the clot, serum separator, or red blood cells, and store frozen (-20C or colder). Frozen specimens must be mixed THOROUGHLY after thawing by LOW speed vortexing. Centrifuged specimens with a lipid layer on the top must be transferred to a sample cup or secondary tube. Care must be taken to transfer only the clarified specimen without the lipemic material. No qualitative performance differences were observed between experimental controls and the 24 nonreactive or 22 spiked reactive specimens subjected to 4 freeze-thaw cycles. The quantitative performance differences observed were within normal assay variability; however, multiple freeze-thaw cycles should be avoided. When shipped, specimens must be packaged and labeled in compliance with applicable state, federal, and international regulations covering the transport of specimens and infectious substances. Specimens may be shipped ambient, at 2-8C (wet ice), or -20C or colder (dry ice). Do not exceed the storage time limitations listed above. Prior to shipment, it is recommended that specimens be removed from the clot, serum separator, or red blood cells. No qualitative performance differences were observed between experimental controls and the 23 nonreactive or 23 spiked reactive specimens tested with elevated levels of triglycerides ( 3000 mg/dL),* bilirubin ( 20 mg/dL),* and hemoglobin ( 500 mg/dL).* No qualitative performance differences were observed between experimental controls and the 30 nonreactive or 30 spiked reactive specimens tested with red blood cells at 0.4% v/v.* No qualitative performance differences were observed between experimental controls and the 21 nonreactive or 20 spiked reactive specimens tested with elevated levels of protein ( 12 g/dL).* ARCHITECT Anti-HBs Calibrators and Controls should be mixed by gentle inversion prior to use. The quantitative performance differences observed were within normal assay variability.
PROCEDURE
Materials Provided 7C18 ARCHITECT Anti-HBs Reagent Kit Materials Required but not Provided ARCHITECT i System 7C18-01 ARCHITECT Anti-HBs Calibrators 7C18-10 ARCHITECT Anti-HBs Controls 7D82-50 ARCHITECT i ARCHITECT i ARCHITECT i ARCHITECT i ARCHITECT i ARCHITECT i ARCHITECT i ARCHITECT i For information on materials required for maintenance procedures, refer to the ARCHITECT System Operations Manual, Section 9. Pipettes or pipette tips (optional) to deliver the volumes specified on the patient or control order screen. Assay Procedure Before loading the ARCHITECT Anti-HBs Reagent Kit on the system for the first time, the microparticle bottle requires mixing to resuspend microparticles that may have settled during shipment: Invert the microparticle bottle 30 times. Visually inspect the bottle to ensure microparticles are resuspended. If microparticles are still adhered to the bottle, continue to invert the bottle until the microparticles have been completely resuspended. Once the microparticles have been resuspended, remove and discard the cap. Wearing clean gloves, remove a septum from the bag. Carefully snap the septum onto the top of the bottle. If the microparticles do not resuspend, DO NOT USE. Contact your local Abbott representative. Order calibration, if necessary. Order tests. For information on ordering patient specimens, calibrators, and controls, and general operating procedures, refer to the ARCHITECT System Operations Manual, Section 5. Load the ARCHITECT Anti-HBs Reagent Kit on the ARCHITECT i System. Verify that all necessary reagents are present. Ensure that septums are present on all reagent bottles. The minimum sample cup volume is calculated by the system and is printed on the Orderlist report. No more than 10 replicates may be sampled from the same sample cup. To minimize the effects of evaporation verify adequate sample cup volume is present prior to running the test. Priority: 125 L for the first Anti-HBs test plus 75 L for each additional Anti-HBs test from the same sample cup. 3 hours on board: 150 L for the first Anti-HBs test plus 75 L for each additional Anti-HBs test from the same sample cup. > 3 hours on board: additional sample volume is required. Refer to the ARCHITECT System Operations Manual, Section 5, for information on sample evaporation and volumes. If using primary or aliquot tubes, use the sample gauge to ensure sufficient patient specimen is present. ARCHITECT Anti-HBs Calibrators and Controls should be mixed by gentle inversion prior to use. To obtain the recommended volume requirements for the ARCHITECT Anti-HBs Calibrators and Controls, hold the bottles vertically, and dispense 7 drops of each Calibrator (for 2 replicates), or 5 drops of each Control (for 1 replicate) into each respective sample cup.
Load samples. For information on loading samples, refer to the ARCHITECT System Operations Manual, Section 5. Press RUN. The ARCHITECT i System performs the following functions: Moves the sample to the aspiration point. Loads a reaction vessel (RV) into the process path. Aspirates and transfers sample into the RV. Advances the RV one position and transfers microparticles into the RV. Mixes, incubates, and washes the reaction mixture. Adds conjugate to the RV. Mixes, incubates, and washes the reaction mixture. Adds Pre-Trigger and Trigger Solutions. Measures chemiluminescent emission to determine the quantity of anti-HBs in the sample. Aspirates contents of RV to liquid waste and unloads RV to solid waste. Calculates the result. For optimal performance, it is important to follow the routine maintenance procedures defined in the ARCHITECT System Operations Manual, Section 9. If your laboratory requires more frequent maintenance, follow those procedures.
Specimen Dilution Procedures Specimens with an anti-HBs value exceeding 1,000 mIU/mL are flagged with the code >1000.00 mIU/mL and may be diluted with either the: 1) Automated Dilution Protocol or 2) Manual Dilution Procedure. 1) Automated Dilution Protocol (for concentrations up to 15,000 mIU/mL) The system performs a 1:15 dilution of the specimen and automatically calculates the concentration of the specimen before dilution and reports the result. 2) Manual Dilution Procedure (for concentrations up to 100,000 mIU/mL) The suggested manual dilution for Anti-HBs is 1:100. It is recommended dilutions not exceed 1:100. For a 1:100 dilution, add 10 L of the patient specimen to 990 L of ARCHITECT i Multi-Assay Manual Diluent (7D82). The operator must enter the dilution factor in the Patient or Control order screen. All assays selected for that order will be diluted. The system will use this dilution factor to automatically calculate the concentration of the sample before dilution and report the result. The concentration reported by the ARCHITECT i System MUST be greater than 10.0 mIU/mL. If the reported concentration is less than 10.0 mIU/mL, make a smaller dilution. Specimens with an anti-HBs value reported as greater than 15,000 mIU/mL may be diluted using either the: 1) Automated Dilution Protocol and the Manual Dilution Procedure or 2) Manual Dilution Procedure. 1) Automated Dilution Protocol and the Manual Dilution Procedure (for concentrations up to 1,500,000 mIU/mL) The suggested manual dilution for Anti-HBs is 1:100. It is recommended dilutions not exceed 1:100. For a 1:100 dilution, add 10 L of the patient specimen to 990 L of ARCHITECT i Multi-Assay Manual Diluent (7D82). Order the Automated Dilution Protocol using the manually diluted 1:100 sample. The concentration reported by the ARCHITECT System MUST be greater than 10.0 mIU/mL. Multiply the result (from the Automated Dilution Protocol) by the manual dilution factor (e.g. 100) to obtain the final sample concentration. If the concentration reported by the ARCHITECT i System is less than 10.0 mIU/mL, make a smaller dilution.
2) Manual Dilution Procedure (for concentrations up to 100,000 mIU/mL) The suggested manual dilution for Anti-HBs is 1:100. It is recommended dilutions not exceed 1:100. For a 1:100 dilution, add 10 L of the patient specimen to 990 L of ARCHITECT i Multi-Assay Manual Diluent (7D82). The operator must enter the dilution factor in the Patient or Control order screen. All assays selected for that order will be diluted. The system will use this dilution factor to automatically calculate the concentration of the sample before dilution and report the result. The concentration reported by the ARCHITECT i System MUST be greater than 10.0 mIU/mL. If the reported concentration is less than 10.0 mIU/mL, make a smaller dilution. For detailed information on ordering dilutions, refer to the ARCHITECT System Operations Manual, Section 5. Calibration To perform an ARCHITECT Anti-HBs calibration, test Calibrators 1 and 2 in duplicate. A single sample of all levels of Anti-HBs controls must be tested to evaluate the assay calibration. Ensure that assay control values are within the concentration ranges specified in the Control package insert. Calibrators should be priority loaded. Calibrator Range: 0 - 1000 mIU/mL. Once an ARCHITECT Anti-HBs calibration is accepted and stored, all subsequent samples may be tested without further calibration unless: A reagent kit with a new lot number is used Controls are out of range For detailed information on how to perform an assay calibration, refer to the ARCHITECT System Operations Manual, Section 6.
variability. Sensitivity A total of 389 specimens from 248 HBV vaccine recipients, 41 individuals recovered from HBV infection, and 100 individuals at risk for HBV infection were tested. Of the 389 specimens, 340 (87.40%) were repeatedly reactive and positive by supplemental testing (Table II). TABLE II Reactivity of the ARCHITECT Anti-HBs Assay in Specimens from HBV Vaccine Recipients, Individuals who have Recovered from HBV Infection, and Individuals at Increased Risk for HBV Infection Number of Number of Positive Number of Repeatedly by Supplemental Specimens Reactive Testing (% of Category Tested (% of total) Repeatedly Reactive) HBV Vaccine 248 245 (98.79%) 245 (100.00%) Recipients Recovered HBV 41 39a (95.12%) 39 (100.00%) Infection 100 56 (56.00%) 56 (100.00%) Increased Risk for HBV Infectionb TOTAL 389 340 (87.40%) 340 (100.00%)
a Two specimens were reactive for anti-HBc and anti-HBe but also
HBV Vaccine Recipient Serial Bleed Panels A total of 90 specimens comprising 15 serial bleed panels from HBV vaccine recipients were tested. The vaccine was administered in three injections over a six-month period. All specimens drawn one month following the third and final injection were reactive by the ARCHITECT Anti-HBs assay. Correlation ARCHITECT Anti-HBs was compared to a commercially available assay for correlation using the Passing-Bablok Regression method 11 and Spearman Rank correlation. 12 The comparison was made using results from 187 specimens from HBV vaccine recipients and individuals who have recovered from HBV infection. The correlation coefficient was 0.906, the slope was 1.07 and the intercept was -3.01. Specificity Three clinical sites tested a total of 1,716 serum and plasma specimens from the following categories: volunteer whole blood donors, matched serum and plasma pairs, random hospital patients, medical conditions unrelated to HBV infection and potentially interfering substances. A total of 259 (15.09%) of the 1,716 specimens were repeatedly reactive, and 254 (98.07%) of the 259 specimens were positive by supplemental testing (Table III). TABLE III Reactivity of the ARCHITECT Anti-HBs Assay in Specimens from Whole Blood Donors, Plasma Specimens from Matched Serum/Plasma Pairs, Hospital Patients, Individuals with Medical Conditions Unrelated to HBV Infection and in Specimens Containing Potentially Interfering Substances Number of Positive Number of by Supplemental Number of Repeatedly Testinga Specimens Reactive (% of Repeatedly Category Tested (% of total) Reactive) Volunteer Whole Blood Donors 1006 154 (15.31%) 151 (98.05%) Plasma Specimens from Matched Serum/Plasma Pairs 50 8 (16.00%) 8 (100.00%) Hospital Patients 500 65 (13.00%) 63 (96.92%) Medical Conditions Unrelated to HBV Infection and Potentially Interfering Substancesb 160 32 (20.00%) 32 (100.00%) TOTAL 1716 259 (15.09%) 254 (98.07%)
a Supplemental testing for anti-HBc, HBsAg and anti-HBe was performed
BIBLIOGRAPHY
1. Wainwright RB, McMahon BJ, Bulkow LR, et al. Duration of Immunogenicity and Efficacy of Hepatitis B Vaccine in a Yupik Eskimo Population-Preliminary Results of an 8-year Study. In: Hollinger FB, Lemon SM, Margolis HS, editors. Viral Hepatitis and Liver Disease. Baltimore: Williams & Wilkins, 1991:762-6. Ambrosch F, Frisch-Niggemeyer W, Kremsner P, et al. Persistence of Vaccine-induced Antibodies to Hepatitis B Surface Antigen and the Need for Booster Vaccination in Adult Subjects. Postgrad Med J 1987;63(S2):129-35. Krugman S, Giles JP, Hammond J. Viral Hepatitis Type B (MS-2 Strain): Studies on Active Immunization. JAMA 1971;217:41-5. Jilg W, Schmidt M, Deinhardt F. Immune Response to Hepatitis B Revaccination. J Med Virol 1988;24:377-84. US Department of Labor, Occupational Safety and Health Administration, 29 CFR Part 1910.1030, Bloodborne pathogens. US Department of Health and Human Services. Biosafety in Microbiological and Biomedical Laboratories. 5th ed. Washington, DC: US Government Printing Office; January 2007. World Health Organization. Laboratory Biosafety Manual. 3rd ed. Geneva: World Health Organization; 2004. Clinical and Laboratory Standards Institute. Protection of Laboratory Workers from Occupationally Acquired Infections: Approved Guideline Third Edition. CLSI Document M29-A3. Wayne, PA: Clinical and Laboratory Standards Institute; 2005. Box GEP, Hunter WG, Hunter JS. Statistics for Experimenters: An Introduction to Design, Data Analysis, and Model Building. New York: John Wiley & Sons, Inc, 1978:510-39, 571-83. SAS Institute Inc. The MIXED Procedure. In: SAS Technical Report P-229, SAS/STAT Software: Changes and Enhancements, Release 6.07. Cary, NC: SAS Institute Inc, 1992:289-366. Passing H, Bablok W. A New Biometrical Procedure for Testing the Equality of Measurements from Two Different Analytical Methods. J Clin Chem. Clin Biochem. 1983; 21:709-20. Netter J, Wasserman W, Kutner M. Applied Linear Regression Models. Boston: IRWIN, 1989:539-540.
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ARCHITECT is a trademark of Abbott Laboratories in various jurisdictions. The following U.S. Patents are relevant to the ARCHITECT System or its components. There are other such patents and patent applications in the United States and worldwide. 5,468,646 5,543,524 5,545,739 5,565,570 5,669,819 5,783,699
to support the presence of anti-HBs in an ARCHITECT Anti-HBs reactive specimen. Detection of anti-HBs by RIA was also performed. A specimen was defined as anti-HBs positive if one or more of the following HBV markers were detected: anti-HBs (detected by the comparator method or RIA), anti-HBc, HBsAg, or anti-HBe. b Category included the following: anti-CMV positive (10), anti-EBV positive (10), anti-HSV (10), anti-HAV (10), anti-HCV (10), anti-HIV-1 (10), rubella antibody positive (10), toxoplasma antibody positive (10), E. coli infections (10), yeast infections (10), syphilis positive (10), antinuclear antibody positive (10), rheumatoid factor (10), multiple myeloma (10), HBsAg positive (10) and alcoholic liver disease (10). Overall Specificity and Sensitivity Overall specificity and sensitivity were estimated from the results of 2,105 specimens tested with ARCHITECT Anti-HBs at five clinical sites. In order to represent unique specimens, results from the HBV vaccine recipient serial bleed panels and the serum specimens from the matched serum/plasma pairs were excluded from these calculations. The overall specificity was estimated to be 99.67% (1,491/1,496) with a 95% confidence interval of 99.22% to 99.89%. The overall sensitivity was estimated to be 97.54% (594/609) with a 95% confidence interval of 95.97% to 98.62%.
Abbott Ireland Diagnostics Division Finisklin Business Park Sligo Ireland +353-71-9171712 November 2009 1999, 2009 Abbott Laboratories
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