<ul><li>HIV testing </li></ul><ul><li>Methods and Procedures </li></ul>
<ul><li>diagnosis of infection </li></ul><ul><ul><li>acute, recent, established or late stage disease </li></ul></ul><ul><...
<ul><li>Typical Primary HIV-1 Infection </li></ul>symptoms HIV-1 p24 antigen 0 1 2 3 4 5 6 / 2 4 6 8 10 weeks years HIV an...
<ul><li>Types of HIV Assays </li></ul>DIAGNOSIS Virus Detection & Quantification Antibody Antigen Detection RNA modified A...
<ul><li>HIV Test Spectrum </li></ul>DNA PCR RNA PCR p24 Ag 3rd gen ELISA 1st gen ELISA Detuned ELISA 1wk  2wk  3wk  2mo  6...
<ul><li>HIV p24 antigen– serology </li></ul><ul><ul><li>In isolation or Ab/Ag Combo test  </li></ul></ul><ul><ul><li>Diagn...
<ul><li>Principle of Immunoassays </li></ul>SOLID PHASE ANTIGEN SAMPLE ANTIBODY ANTI-HUMAN IMMUNOGLOBULIN WITH DETECTOR
<ul><li>EIAs  including  </li></ul><ul><ul><li>rapid, simple assays </li></ul></ul><ul><ul><li>particle agglutination </li...
<ul><li>Dot/Blot Assays </li></ul>
<ul><li>Particle Agglutination Assays </li></ul>
<ul><li>technically more difficult </li></ul><ul><li>visual interpretation </li></ul><ul><li>lack standardisation </li></u...
<ul><li>Difficulties in interpretation </li></ul><ul><li>Limitations - ‘window period’ </li></ul><ul><li>antibodies appear...
<ul><li>Detection of Ag & Ab in a single test </li></ul><ul><li>utility in primary infection – pre-seroconversion ‘window ...
<ul><li>Chance of false reactivity </li></ul><ul><li>Might need confirmation tests </li></ul><ul><li>Replacement by other ...
<ul><li>Detect HIV-1 & HIV-2 </li></ul><ul><li>Cannot differentiate between HIV types </li></ul><ul><li>Procedural control...
<ul><li>Detects HIV-1 and HIV-2 </li></ul><ul><li>Will differentiate between types 1 and 2 </li></ul><ul><li>Procedural co...
<ul><li>Strategy I :  Test all samples with one EIA </li></ul><ul><li>Strategy II:  Strategy I with all reactives retested...
<ul><li>Use strategy for confirmation </li></ul><ul><ul><li>Use combination of affordable and simple assays </li></ul></ul...
<ul><li>www.ubio.in </li></ul>
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AIDS/HIV Testing Methodology (www.ubio.in)

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HIV testing methodologies: an Introduction

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Transcript of "AIDS/HIV Testing Methodology (www.ubio.in)"

  1. 1. <ul><li>HIV testing </li></ul><ul><li>Methods and Procedures </li></ul>
  2. 2. <ul><li>diagnosis of infection </li></ul><ul><ul><li>acute, recent, established or late stage disease </li></ul></ul><ul><li>prognostic markers </li></ul><ul><li>monitoring of ARV therapies </li></ul><ul><ul><li>immunological and virological markers </li></ul></ul><ul><ul><li>toxicities </li></ul></ul><ul><li>diagnosis of opportunistic infections </li></ul><ul><li>drug resistance testing </li></ul><ul><li>Laboratory Tests </li></ul>
  3. 3. <ul><li>Typical Primary HIV-1 Infection </li></ul>symptoms HIV-1 p24 antigen 0 1 2 3 4 5 6 / 2 4 6 8 10 weeks years HIV antibodies Time following infection HIV viral load HIV proviral DNA symptoms ‘ window’ period 1° infection
  4. 4. <ul><li>Types of HIV Assays </li></ul>DIAGNOSIS Virus Detection & Quantification Antibody Antigen Detection RNA modified Ag Viral Culture, phenotyping CD4+ ARV Resistance – genotyping ARV Sensitivity EIA Simple, rapid tests Immunoblots Incident assays MANAGEMENT DNA (RNA)
  5. 5. <ul><li>HIV Test Spectrum </li></ul>DNA PCR RNA PCR p24 Ag 3rd gen ELISA 1st gen ELISA Detuned ELISA 1wk 2wk 3wk 2mo 6mo 1yr 2yr 3yr +8yr early recent / established advanced
  6. 6. <ul><li>HIV p24 antigen– serology </li></ul><ul><ul><li>In isolation or Ab/Ag Combo test </li></ul></ul><ul><ul><li>Diagnosis of primary infection viraemia </li></ul></ul><ul><li>Virus culture/isolation </li></ul><ul><li>Nucleic acid detection (NAT) </li></ul><ul><ul><li>Clinical uses Proviral DNA vs. plasma RNA(viral load) resolution of inconclusive serology/neonatal </li></ul></ul><ul><ul><li>subtyping </li></ul></ul><ul><ul><li>drug resistance monitoring </li></ul></ul><ul><li>Direct Virus Detection </li></ul>
  7. 7. <ul><li>Principle of Immunoassays </li></ul>SOLID PHASE ANTIGEN SAMPLE ANTIBODY ANTI-HUMAN IMMUNOGLOBULIN WITH DETECTOR
  8. 8. <ul><li>EIAs including </li></ul><ul><ul><li>rapid, simple assays </li></ul></ul><ul><ul><li>particle agglutination </li></ul></ul><ul><ul><li>dot/blot </li></ul></ul><ul><ul><li>Western blot </li></ul></ul><ul><ul><li>Antigen and Antibody/Antigen (Ab/Ag) </li></ul></ul><ul><ul><li>Incidence assays </li></ul></ul><ul><li>Direct Virus Detection </li></ul><ul><li>Available Assays </li></ul>
  9. 9. <ul><li>Dot/Blot Assays </li></ul>
  10. 10. <ul><li>Particle Agglutination Assays </li></ul>
  11. 11. <ul><li>technically more difficult </li></ul><ul><li>visual interpretation </li></ul><ul><li>lack standardisation </li></ul><ul><ul><li>Performance </li></ul></ul><ul><ul><li>Interpretation </li></ul></ul><ul><li>‘ Gold Standard’ for confirmation </li></ul><ul><li>Western Blot </li></ul>
  12. 12. <ul><li>Difficulties in interpretation </li></ul><ul><li>Limitations - ‘window period’ </li></ul><ul><li>antibodies appear within 3-4 weeks </li></ul><ul><ul><li>Direct detection for early detection </li></ul></ul><ul><ul><li>HIV p24 antigen </li></ul></ul><ul><ul><li>DNA/RNA (NAT) </li></ul></ul><ul><li>Combo test = earlier detection </li></ul><ul><li>Primary infection + therapy = delayed antibody response </li></ul><ul><li>Incident populations – ‘at risk’ </li></ul><ul><li>Antibody Testing: Limitations </li></ul>
  13. 13. <ul><li>Detection of Ag & Ab in a single test </li></ul><ul><li>utility in primary infection – pre-seroconversion ‘window period’ </li></ul><ul><li>Useful for blood banks </li></ul><ul><li>Automated platforms available </li></ul><ul><li>Ag/Ab Combo Tests </li></ul>Ag & Ab
  14. 14. <ul><li>Chance of false reactivity </li></ul><ul><li>Might need confirmation tests </li></ul><ul><li>Replacement by other assays (especially in the USA) </li></ul><ul><li>More expensive than Antibody assays </li></ul><ul><li>Ag/Ab Assays: Limitations </li></ul>
  15. 15. <ul><li>Detect HIV-1 & HIV-2 </li></ul><ul><li>Cannot differentiate between HIV types </li></ul><ul><li>Procedural control using anti human IgG </li></ul><ul><li>Can test using whole blood, serum or plasma </li></ul><ul><li>Widely available </li></ul><ul><li>No additional reagents required </li></ul><ul><li>Storage at room temperature </li></ul><ul><li>15 minutes to get result </li></ul><ul><li>Simple Assays: HIV Determine </li></ul>
  16. 16. <ul><li>Detects HIV-1 and HIV-2 </li></ul><ul><li>Will differentiate between types 1 and 2 </li></ul><ul><li>Procedural control using anti-human IgG </li></ul><ul><li>Serum/plasma only </li></ul><ul><li>Need additional reagents (included in kit) </li></ul><ul><li>Requires refrigerated storage </li></ul><ul><li>‘ Immunoconcentration’ principle </li></ul><ul><li>15 minutes to result </li></ul><ul><li>BioRad HIV-1/2 Multispot </li></ul>
  17. 17. <ul><li>Strategy I : Test all samples with one EIA </li></ul><ul><li>Strategy II: Strategy I with all reactives retested in a more specific test with different principle and/or antigen. </li></ul><ul><li>Strategy III: Strategy II with reactives tested in a third test differing from the first two tests. </li></ul><ul><li>Transfusion Safety </li></ul><ul><ul><li>Strategy I </li></ul></ul><ul><li>Surveillance </li></ul><ul><ul><li>>10% Strategy I, < 10% Strategy II </li></ul></ul><ul><li>Diagnosis </li></ul><ul><ul><li>>10% Strategy II, < 10% Strategy III </li></ul></ul><ul><li>WHO Test Strategies </li></ul>
  18. 18. <ul><li>Use strategy for confirmation </li></ul><ul><ul><li>Use combination of affordable and simple assays </li></ul></ul><ul><ul><li>Use tests with different principles </li></ul></ul><ul><ul><li>Use different antigen preparations </li></ul></ul><ul><li>Use two or three types of ELISAs or rapid tests </li></ul><ul><li>diagnosis confirmed by second sample </li></ul><ul><li>Use direct detection of virus (PCR) for final confirmation </li></ul><ul><li>Important: Always use a QC sample </li></ul><ul><li>WHO Test Guidelines </li></ul>
  19. 19. <ul><li>www.ubio.in </li></ul>
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