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HIV TESTING PROTOCOLS D.Vijay kumar Dist.Sales Manager AbbottHyderabad.

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Presentation on theme: "HIV TESTING PROTOCOLS D.Vijay kumar Dist.Sales Manager AbbottHyderabad."— Presentation transcript:

1 HIV TESTING PROTOCOLS D.Vijay kumar Dist.Sales Manager AbbottHyderabad.

2 Informed consent prior to testing is essential to deal with issues like-  Confidentiality  Discrimination  Victimization  Psychological harm * HIV testing – voluntary * HIV testing – voluntary Mandatory testing – counterproductive Mandatory testing – counterproductive

3 Purpose of HIV testing

4  Transfusion safety  Epidemiological - Sentinel surveillance  Diagnostic purpose  Voluntary testing  Research

5 PPTCT PPTCT Pregnant HIV infected women can-  make informed decisions about dealing with pregnancy  receive appropriate and timely interventions to decrease MTCT  ensure safe delivery  secure early access to HIV care and treatment  educated in prevention of HIV transmission  receive follow-up health care for self and child Pregnant HIV non infected women can be educated and counselled to remain uninfected

6 Serological Profile of HIV infection

7 Tests for detecting HIV INFECTION  Detection of specific antibodies screening tests: 100% sensitive screening tests: 100% sensitive supplemental/ confirmatory tests- supplemental/ confirmatory tests- sens. > 99.8% sens. > 99.8% spec. > 98.5% spec. > 98.5%  Detection of specific antigens

8 Sensitivity – Accuracy with which a test can confirm the presence of an infection. Test with high sensitivity – few false negatives TP TP Sensitivity = x 100 TP + FN TP + FN SELECTION OF TESTS : BASED ON SENSITIVITY,SPECIFICITY,EFFICIENCY,PPV & NPV

9 SPECIFICITY- Accuracy with which the test can confirm the absence of an infection Accuracy with which the test can confirm the absence of an infection test with high specificity – few false positives test with high specificity – few false positives used for diagnosing infection in an individual used for diagnosing infection in an individual TN TN Specificity = X 100 TN + FP TN + FP

10 Efficiency - ability of a test to correctly identify - all positives as positives all positives as positives all negatives as negatives all negatives as negatives TP + TN TP + TN Efficiency = X 100 TP + FN + TN + FP TP + FN + TN + FP

11 Predictive values PPV – identifies ACTUALLY infected individuals PPV – identifies ACTUALLY infected individuals TP TP X 100 X 100 TP + FP TP + FP NPV – identifies ACTUAL non infected NPV – identifies ACTUAL non infected TN TN X 100 X 100 TN + FN TN + FN

12 ELISA  Most common screening test  Indirect solid phase enzyme linked immunosorbent assay/ EIA  Used in blood banks/ tertiary labs  Fourth generation ELISA decreases window period DISADVANTAGES DISADVANTAGES  Many false positives  Few false negatives  Time consuming  Needs infrastructure & tech. expertise

13 PRINCIPLE OF INDIRECT ELISA Enzyme conjugated Anti-HIV antibody in specimen Antigen On solid phase Substrate

14 ELISA cont.  False positives: auto immune diseases auto immune diseases multiple pregnancies multiple pregnancies hematologic malignancies hematologic malignancies primary biliary cirrhosis primary biliary cirrhosis alcoholic hepatitis alcoholic hepatitis CRF CRF  False negatives window period window period immunosuppressive therapy immunosuppressive therapy malignant disorders malignant disorders late stage disease late stage disease technical errors technical errors

15 Rapid tests  Dot blot assay- immuno concentration method Retroquic -line assay Retroquic -line assay Tridot - dot assay Tridot - dot assay  Immunocomb assay – dipstick/comb ELISA based ELISA based HIV comb HIV comb coomb AIDS coomb AIDS  Immunochromatography – lateral flow assays Determine,Unigold,Hemastrip Determine,Unigold,Hemastrip  Particle aggluttination – Capillus,Serodia  ELISA based - EIA

16 Comb AIDS  Dot immunoassay for HIV 1&2 using whole blood,serum or plasma. Comb with 8 teeth- Megenta red spot  Synthetic& recombinant peptides used  Two spots-- Control spot & test spot. NON-REACTIVE REACTIVE For HIV-1 &2 INVALID TEST

17 TRIDOT  HIV 1 – gp 41, gp 120.  HIV 2 – gp 36.  Highly specific. C 12

18 RETROQUIC HIV

19 HIV EIA Comb

20  Rapid Visual EIA Test  Detection of Antibodies to HIV-1 (including subgroups O & C) and HIV-2  Sensitivity (100%)  Specificity (99.9%) NON-REACTIVE HIV-1 HIV-2 HIV-1&2 REACTIVE INVALID TEST INTERPRETATION OF RESULTS

21 SUPPLEMENTAL TESTS  1.Rapid tests.  2.Western blot.  3.Immunoblot.  4.Line immunoassay.  WB/IB/LIA: highly specific but Expensive Labour intensive Needs expertise Equivocal/indeterminate results.

22 Western Blot for HIV  Delineates the antibody profile of reactive serum  Used to grade intensity of ab response –Qualitatively –Quantitatively  Procedure based on principle of ELISA WB Rotator platform WB strips

23 Western blot for HIV antibody gp 36 Control band p 24 gp 31 gp 120 & 160 gp 41

24 TESTS DONE IN VCCTC, PPTCT & BLOOD BANK VCCTC & PPTCT: Test – I:HIV Comb / Comb Aids – RS Test – II:Tridot / Retroquic Test – III:EIA Comb FOR SURVEILLANCE: Test – I:HIV Comb / Comb Aids – RS Test – II:Tridot / Retroquic BLOOD BANK: Only Test:Microlisa HIV

25 STRATEGIES/ALGORITHMS OF HIV TESTING 1.Screening ELISA/Rapid tests – used in strategy I, II, III. 1.Screening ELISA/Rapid tests – used in strategy I, II, III. 2.Supplemental tests – E/R & Western Blot. 2.Supplemental tests – E/R & Western Blot. Strategy I (for transfusion/transplantation safety) Strategy I (for transfusion/transplantation safety) one test kit required one test kit required AI AI A1 + A1 - positive negative A1 + A1 - positive negative

26 Strategy/Algorithm II A (for surveillance) 2 test kits required A1 Strategy/Algorithm II A (for surveillance) 2 test kits required A1 A1 + A1 - Report negative A1 + A1 - Report negative A2 A2 A1 + A2 + A1+ A2 - Report positive Report negative A1 + A2 + A1+ A2 - Report positive Report negative

27 Strategy/Algorithm II B (Diagnosis of an individual with AIDS indicator disease symptoms) 3 test kits required A1 Strategy/Algorithm II B (Diagnosis of an individual with AIDS indicator disease symptoms) 3 test kits required A1 A1 + A1 - report negative A1 + A1 - report negative A2 A2 A1+ A2 + A1+ A2 - report positive A1+ A2 + A1+ A2 - report positive A3 A3 A1+ A2 – A3+ A1+ A2 - A3- indeterminate report negative A1+ A2 – A3+ A1+ A2 - A3- indeterminate report negative

28 Strategy/Algorithm III To detect HIV infection in asymptomatic individuals (VCTCs, PPTCTs) 3 test kits required A1 Strategy/Algorithm III To detect HIV infection in asymptomatic individuals (VCTCs, PPTCTs) 3 test kits required A1 A1+ A1 - Report negative A1+ A1 - Report negative A2 A2 A1+ A2 + A1+ A2 – A1+ A2 + A1+ A2 – A3 A3 A1+ A2+ A3+ A1+ A2+ A3- A3 Report positive indeterminate A1+ A2+ A3+ A1+ A2+ A3- A3 Report positive indeterminate A1+ A2- A3+ A1+ A2- A3- Indeterminate report negative A1+ A2- A3+ A1+ A2- A3- Indeterminate report negative

29  INDETERMINATE STATUS: Repeat test after days. Results continue to be indeterminate – WB/PCR refer to NRL.  EQUIVOCAL WB: Rpt. WB after 2 weeks 4 weeks 12 weeks one year. Correlate with high risk behaviour & clinical parameters.

30 TESTS TO DETECT ANTIGENS  P24 Antigen: Uncomplexed in serum, plasma, CSF, cell culture. Indicates Active infection especially in newborn. Resolves equivocal WB. Window period. CNS disease. Immune collapse. Monitoring response to ART. Method: EIA Method: EIA Disadvantages: Expensive. Limited sensitivity. Failure to detect HIV 2. Failure to detect Ag when complexed with Antibody Disadvantages: Expensive. Limited sensitivity. Failure to detect HIV 2. Failure to detect Ag when complexed with Antibody

31 Limitations of antigen detection methods  Not reliable  Expensive  Limited sensitivity-69% in patients with AIDS,15% in neonates  Detection not possible in patients with high anti p24 antibody  Cannot be used as a screening test

32  Highly specific test-more than 95%  Highly sensitive-infants over 1 month  Detects proviral DNA.  Detects both latent viral infection and active viral transcriptipn.  Detects viral load.  Detects both HIV1 & HIV2. PCR

33 PCR AS VIRAL ASSAYS IN INFANTS  Counselling for infant feeding & therapeutic intervention.  First done at 6 weeks.  Not Breast-fed, to say not infected: 2 negative test after 1 month (include 1 at 4 months)  Not done as part of PPTCT in India.  If symptoms occur at < 18 months: go for viral assays.

34 Asymptomatic 18 months-ELISA Non-reactive Reactive Non-reactive Reactive Conditionally consider confirm with a retest Conditionally consider confirm with a retest Non-infected Non-infected Re-test Re-test NR NR HIV Unifected HIV Unifected Diagnosing HIV infection in an infant born to HIV positive mother Protocol 1

35 PROTOCOL 2 SYMPTOMATIC Early symptoms AIDS Early symptoms AIDS Earlier than 12 mon. defined symptoms Viral assay at 6 weeks same as for early Whenever symptoms occur symptoms Retest 1 month later If both are reactive consider reactive reactive ELISA at 12 months Non reactive Reactive Non reactive Reactive Follow protocol 1 conditionally consider HIV +ve Follow protocol 1 conditionally consider HIV +ve Retest at 18 months retest at 18 months Retest at 18 months retest at 18 months Non reactive reactive Non reactive reactive HIV UNIFECTED HIV INFECTED

36 HIV TESTING POLICY IN PPTCT  PARENT: Informed consent of the patient Pre n post test counselling Routine testing with three rapid tests first: highly sensitive test-NR-reported with exception of WP Indeterminate: 1 st test reactive, 2 nd /3 rd NR repeat test after days. WB/PCR: For persistent indeterminate cases.  INFANT OF HIV + MOTHER, ASYMPTOMATIC: ELISA at 18 months—NR—Retest—NR—uninfected. Reactive—consider infected, confirm with retest.  INFANT OF HIV + MOTHER, SYMPTOMATIC: ELISA at 12 months—NR—retest NR—uninfected. ELISA at 12 months—NR—retest NR—uninfected. Reactive—retest R at 18 months—infected. Reactive—retest R at 18 months—infected.

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