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HIV Testing CDC power point edited by M. Myers

HIV Testing CDC power point edited by M. Myers. Message. There are numbers of tests They should be used in combination (strategies) Combinations must be consistent. Laboratory Tests. diagnosis of infection acute, recent, established or late stage disease prognostic markers

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HIV Testing CDC power point edited by M. Myers

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  1. HIV TestingCDC power point edited by M. Myers

  2. Message • There are numbers of tests • They should be used in combination (strategies) • Combinations must be consistent

  3. Laboratory Tests • diagnosis of infection • acute, recent, established or late stage disease • prognostic markers • monitoring of ARV therapies • immunological and virological markers • toxicities • diagnosis of opportunistic infections • drug resistance testing

  4. ‘typical’ primary HIV-1 infection symptoms symptoms HIV proviral DNA HIV antibodies ‘window’ period HIV viral load HIV-1 p24 antigen 01 2 3 4 5 6 / 2 4 6 8 10 1° infection weeks years Time following infection

  5. HIV Assays: MethodologiesFOR THE DIAGNOSIS (DETECTION) Virus Detection EIASimple, rapid tests Immunoblots Incident assays Antibody Antigen Detection + DNA(RNA) DIAGNOSIS MANAGEMENT

  6. HIV Testing -Direct Detection of Virus • HIV antigen– serology • - In isolation • - Diagnosis of primary infection viraemia • Virus culture / isolation • Nucleic acid detection - (NAT) • Clinical uses Proviral DNA vs. plasma RNA (viral load) • resolution of inconclusive serology / neonatal • subtyping • drug resistance monitoring

  7. Available Assays • EIAs including • rapid, simple • particle agglutination, • dot/blot • Western blot • Antigen & Ab/Ag • Incidence assays • Direct Virus Detection

  8. Particle Agglutination

  9. Western Blot • Expensive – $ 80 - 100 • technically more difficult • visual interpretation • lack standardisation • - performance • - interpretation • - indeterminate reactions – resolution of ?? • ‘Gold Standard’ for confirmation

  10. Antibody testing limitations • Difficulties in interpretation • Limitations - ‘window period’ • antibodies appear within 3-4 weeks • Direct detection – HIV p24 antigen or DNA/RNA (NAT) – pre-antibody • Combo test = earlier detection • Primary infection + therapy = delayed antibody response

  11. Ag & Ab Ag/Ab Combo tests Ab & Ag • Detection of Ag & Ab in a single test • utility in primary infection – pre-seroconversion ‘window period’ • Incident populations – ‘at risk’ • Blood bank • Automated platforms available

  12. Issues with Combo Assays • Testing strategies • False reactivity rates • Confirmation strategies • Replacement of other assays (especially in the USA) • Cost • Legal issues

  13. What about simple assays?

  14. HIV Determine test • Detect HIV-1 & HIV-2 • Cannot differentiate • Procedural control – anti Hu IgG • Whole blood or serum/plasma • Widely available • No additional reagents required • Room temperature storage • 15 minutes to result

  15. BioRad HIV-1/2 Multispot • Detects HIV-1 and HIV-2 • Will differentiate 1 and 2 • Procedural control – anti-Hu IgG • Serum / plasma only • Additional reagents (included) • Requires refrigerated storage • ‘Immunoconcentration’ principle • 15 minutes to result

  16. WHO Recommended Strategies • Strategy I Test all samples with one EIA • Strategy II Strategy I with all reactives retested in a more specific test with different principle and/or antigen. • Strategy III Strategy II with reactives tested in a third test differing from the first two tests.

  17. Transfusion safety Surveillance Diagnosis Risk factors No risk factors Strategy I >10% I <10% II Strategy II >10% II <10% III WHO Recommended Testing Strategies

  18. Testing Strategies AIM: To develop the logic used in establishing the use of HIV tests (testing strategies)

  19. Objectives of Testing Strategies • To achieve the correct diagnosis in the most efficient manner • To maintain consistency in testing • To know the predictive value of the testing process • To develop baseline data for assessing changes • To deliver useful results

  20. Aims in Developing HIV Testing Strategies • To arrive at the correct sero-diagnosis • To minimise total testing; thus cost • Minimise samples classed as indeterminate or dual reactors • Detect HIV-1 negative but HIV-2 positive • Follow likely seroconverters (HIV-1 or -2)

  21. Screening Assays • Are used to detect antibody-- specific or nonspecific • Are designed to handle large numbers of samples with rapid throughput • Must be high performance • Should include a full range of HIV antigens

  22. SCREENING TEST, highly sensitive NEG REACTIVE SUPPLEMENTAL TEST, highly sensitive & higher specificity POS NEG IND NEG ADDITIONAL TESTS POS IND Serological Testing Strategy

  23. HIV Testing Strategy HIV1/2 SCREEN NEG SCREENING REACTIVE HIV-1 WB POS NEG IND NEG SUPPLEMENTAL ADDITIONAL TESTS POS IND POINT OF REPORTING

  24. Supplemental Assays • Range of assays that further define sero-status • High Performance (higher specificity)

  25. The Use of Screening Assays • Define samples as negative for a given analyte • Enable high throughput

  26. Predictive Values Positive Predictive Values: The likelihood of a sample identified as a reactive by a test being truly POSITIVE for the analyte used as the basis of the test. True Positives PPV = X 100% True Positives + False Reactives

  27. True Negatives X 100% NPV = True Negatives + False Negatives Predictive Values Negative Predictive Values: The likelihood that a sample identified as a non-reactive by a test is truly NEGATIVE for the analyte used as the basis of the test.

  28. WHO Recommended Strategies • Strategy I Test all samples with one EIA • Strategy II Strategy I with all reactives retested in a more specific test with different principle and/or antigen. • Strategy III Strategy II with reactives tested in a third test differing from the first two tests.

  29. Transfusion safety Surveillance Diagnosis Risk factors No risk factors Strategy I >10% I <10% II Strategy II >10% II <10% III WHO Recommended Testing Strategies

  30. WHO Guidelines Other possibilities • strategy for confirmation • combination of affordable & simple assays • different test principles • different antigen preparations • two or three ELISAs or rapid tests • diagnosis confirmed by second sample • detection of virus (PCR) • antigen detection (limited lab.facilities) • Always use a QC sample

  31. Cost of HIV Testing comparative costs • ELISA (Ab only) - $2 per test • EIA (Ab/Ag combo) - $3.50 • rapid test - $10-20 per test • Western blot $80 - 100 • p24 antigen $30 • PCR - qualitative $80 - 100 • PCR - quantitative (viral load) $90 – 150* • DNA sequencing (resistance) $400 – 700

  32. - - Screening test x1 NEG Eliminates laboratory error R - Screening test x2 - NEG RR or R- + POS Supplemental test NEG Other tests Summary of Testing Strategies

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