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Comparing the New EIAs with Old Standbys: Florida Bureau of Laboratories Verification Data

Comparing the New EIAs with Old Standbys: Florida Bureau of Laboratories Verification Data. HIV Diagnostics: New Developments and Challenges Feb. 28, 2005 Berry Bennett, MPH Retrovirology Section Chief FL. Bureau of Laboratories Jacksonville, FL. Did you know?.

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Comparing the New EIAs with Old Standbys: Florida Bureau of Laboratories Verification Data

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  1. Comparing the New EIAs with Old Standbys: Florida Bureau of Laboratories Verification Data HIV Diagnostics: New Developments and ChallengesFeb. 28, 2005 Berry Bennett, MPHRetrovirology Section ChiefFL. Bureau of LaboratoriesJacksonville, FL.

  2. Did you know? New CLIA standard for laboratories who introduce a new test system (MC or HC) on or after April 24, 2003; “Verification of performance specifications. Each laboratory that introduces an unmodified, FDA-cleared or approved test system must do the following before reporting patient test results.” - 42 CFR 493.1253(b)(1)revised 10/1/2004 (2-year phase-in period?)Interpretive Guidelines “A laboratory may use the manufacturer’s performance specifications as a guideline, but is responsible for verifying the manufacturer’s analytical claims before initiating patient testing.” Analytical claims include but not limited to sensitivity, specificity, accuracy, precision, detection range or interfering substances, if applicable.“For some qualitative tests, the laboratory may verify the manufacturers’ specifications by testing known positive and negative samples to assure that the expected results are obtained.”Check with local AHCA for acceptance of in-house samples, performance panels, or manufacturer’s assistance/panels. Laboratories must determine panel size and construct for their population to verify performance.

  3. Florida’s HIV Screening Change, Effective January 2004 Reasons:1) Indications from the manufacturer that a product substitution most likely would take place.2) Encouragement from the FDA to manufacturers to include HIV-1 Group O sensitivity as well as group M subtype performance.From: BioRad HIV-1/2 synthetic peptide EIATo: BioRad HIV-1/HIV-2 Plus O EIASample size: >1,000 routine fresh serum samples (~1.9% seropositivity)(If construction allowed)Expectations & Concerns:1) Based on BioRad’s seroconversion data, we expected increased sensitivity.2) As with all new assays we were concerned about compromising specificity as sensitivity increases.2) Data suggests the new assay could potentially detect more HIV acute infections.3) We were concerned about an increase number of indeterminate and negative Western Blots.

  4. Florida’s HIV-1/HIV-2 Plus O EIA Initial Verification n = 883* ( 23 r/r, 860 nr )BioRad HIV-1/2 synthetic peptide EIA:P.Insert claim = 100% sensitivity, 99.87% specificityFBOL = 99.0% specificity (9 false pos)BioRad HIV-1/HIV-2 Plus O EIA:P. Insert claim = 100% sensitivity, 99.89% specificityFBOL = 100% sensitivity, 99.3% specificity (6 false pos)* evaluation cut short due to laboratory construction.

  5. BioRad’s Performance Results:

  6. BBI Seroconversion Panel Results: G.S G.S. days since HIV-1/2 HIV-1/2 OTC bDNAMember# 1st bleed SP Plus O HIV-1 copies/ml PRB958-01 0 0.1 0.25 0.4 7 X 101 PRB958-02 2 0.1 0.25 0.4 1 X 103PRB958-03 7 0.1 0.2 0.4 1 X 105PRB958-04 9 0.1 0.2 0.4 3 X 105PRB958-05 15 1.0 2.4 0.4 5 X 105PRB958-06 17 2.6 11.0 0.5 >5 X 105BBIBOLBBI BBIC.B. W Blot PRB958-01 no bands 02 no bands 03 no bands 04 no bands 05 no bands 06 no bands BBI

  7. BBI Seroconversion Panel Results: G.S. G.S days since HIV-1/2 HIV-1/2 OTC PCR Member# 1st bleed SP Plus O HIV-1 copies/ml PRB959-01 0 0.1 0.2 0.3 2 X 105PRB959-02 7 0.1 1.0 0.4 >8 X 105PRB959-03 9 0.2 7.0 0.4 >8 X 105PRB959-04 14 1.8 13.1 1.8 8 X 105PRB959-05 19 7.3 13.1 4.2 5 X 105PRB959-06 21 7.6 13.1 4.7 3 X 105 PRB959-07 267.8 13.1 5.3 >8 X 105BBIBOLBBI BBIC.B. W Blot PRB959-01 no bands 02 no bands 03 no bands 04 p24, gp160 05 p24, gp160 06 p24, gp160 07 p24, gp160 BBI

  8. BioRad’s Performance Results:Seroconversions HIV-1/HIV-2 PLUS O: 21 Days Earlier cutoff

  9. Florida’s Lessons Learned: Most likely one will see increased number of Western Blot indeterminates and/or negatives; a) Poor client redraw/follow up makes it difficult to confirm acute infection sensitivity. b) Original sample submission and/or sample handling may not be satisfactory for NAAT. c) A need for alternative more sensitive confirmation assays. d) A need to monitor automated EIA instrument performance, especially washer calibration. e) Examine unconfirmed repeatedly reactive EIAs for trends such as pregnancy, rheumatoid factor, etc. if such information is provided. Recommend monitoring technical staff’s competency assessment in sample processing to avoid cross contamination in very sensitive assays. BioRad’s 2004 Technical Bulletin addressed sample cross contamination.“ Serological markers such as HBsAg and anti-HIV may be present in serum at very high concentrations, with titers exceeding 1 in 106 in some specimens. Data demonstrates that 3rd generation HIV tests are often more sensitive than Western Blot and/or 2nd generation HIV EIAs.”

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