Diagnosis of-aids
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  • 1. DiagnosisofAIDS
  • 2. AIDSA Global ConcernTODAY THERE IS NO REGION OF THEWORLD UNTOUCHED BY PANDEMIC OFAIDS
  • 3. Majority of persons infected with HIVpresent with AIDS soon or later.
  • 4. Why AIDS is more importantTODAY Apart from Men, there is a increasing burden ofInfections in Women. Implication on Mother to Child Transmission Female Adolescents are infected 3 - 6times higher than male counterparts.AIDS has turned out to be SocialProblem.
  • 5. Implication of AIDS Being diagnosed as having HIV/AIDS haslife time repercussions. Every body understands AIDS as a lifethreatening condition. A casual way of dealing as being HIV + willhave Moral, Legal, and Socialconsequences.
  • 6. Prompt and Precise Diagnosis ofAIDS make great impact on Humanity
  • 7. Importance of precise Diagnosis ofAIDS All patients should be informed aboutconsequences of Testing for HIV infection, Legality of respective National Laws should befollowed. A pretest and posttest counseling is Growingimportance, even in Developing countries. Today’s debate on testing for HIV status asmatter of Health protocol ?
  • 8. Counseling Improves the confidence
  • 9. Purpose of Testing for HIV Infection A matter of great importance as in Blood donorsto prevent infected blood being transfused. To diagnose the patients infected with HIV virus. For surveillance purpose. Persons with high risk behavior. In Pregnant women to prevent Mother to Childtransmission. Patients presenting with opportunistic infections.
  • 10. Diagnosis ofHIV/ AIDSBasic Tests.1.We can detect Antigen or Antibody2.We can detect Antigen and Antibody.3.Majority of Laboratories depend onCommercial Kits.4.Devloped Nations going for Molecular andmore precise Methods.
  • 11. Screening Tests for HIV/AIDSDetection A screening test posses high Sensitivity We rarely miss the Diagnosis in Infected patients. Done as mass screening procedure. Economical. Developing countries depend on these tests evenfor Diagnosis.
  • 12. Confirmatory Tests in AIDS It is important to confirm all screening testswith Confirmatory tests, or we brand someone without infection as infected, Confirmatory tests differentiates falsereactive tests. and identifies truly infectedor not.
  • 13. Differences of Screening andConfirmatory Tests.Screening testHigh degree ofSensitivityFew false negativesConfirmatory testHigh degree ofspecificityFew / No false positiveresults.
  • 14. Why patients should we tested withScreening and Confirmatory tests. Before you declare a person infected withHIV / AIDS you should perform bothmethods. Faulty testing methods can lead tocatastrophic consequences, and legallitigations.
  • 15. Choosing Screening Testing methods Our aim to obtain 100 % specific results ? But impossible. As few may be infected and do not havesufficient antibodies to detect in the testingmethods. Depend on time tested methods. Explore the reputation of the suppliers.
  • 16. ELISA METHOD Universally accepted test ,most populareven in the developing nations. Useful in large scale screening. A common method used in Blood banks inmass screening of Human blood.
  • 17. ELISA method – A ELISA plate
  • 18. Sequence of Events in ELISA test
  • 19. Different Methods of ELISA testing.
  • 20. Why ELISA is very trust worthy. Easy to perform Free from Radiation. Relatively specific Sensitive. With scientific advances we have advanced tonew generation of testing methods Today we depend on 3rd and 4th generationELISA methodologies.
  • 21. Significance of ELISA Most reliable screening test for HIVinfection. 50% of the infected will show positivereactivity in < 22 days. 95% show reactivity < 6 weeks Sensitivity > 99.9 % But needs confirmation with Western Blot.
  • 22. Advances in ELISA Methods.1st and 2nd generation ELISA methods havebecome obsolete we have to switch to Antigen sandwich ELISA protocols ( calledas 3rd generation ELISA ). Many Nations have switched to 3rdgeneration methods.
  • 23. What is 3rd Generation ELISA Helps in detection of early infections, Can detect all classes of antibodiesincluding Ig M All isotypes of antibodies can be captured. But proving to be expensive to developingNations.
  • 24. Need for 4thGeneration ELISAMethodsThe quest for better Diagnosis ofAIDScontinues
  • 25. What is 4th generation ELISA. A new generation Method in ELISA Can detect both Antigen and Antibody inthe same run ( in the same ELISA Plate ) Helpful to make early diagnosis at least infew cases.
  • 26. Determination of P 24 Antigen byELISA Helps in the early detection of HIV infection. Screening of Blood for HIV infection along withdetection for antibody detection. Early diagnosis of HIV infection in resource poornations. Monitoring Anti-retroviral therapy. But not areplacement for RNA quantization. Even P 24 Antigens are subject of false positiveresults.
  • 27. Limitation of Screening Tests. Imperfect Sensitivity, Sensitivity to detect HIV2 ? Failing to detect HIV variants. Other technical errors.
  • 28. ConfirmatorySerological TestsAll the Screening tests need to beconfirmed before being declared asHIV infected.
  • 29. Confirmatory Serological Tests. HIV Western Blots. Line Immunoassay ( LIA ) Immuno fluorescent Assay ( IFA )Utility is limited due technical andeconomical reasons
  • 30. WESTEN BLOT TEST A Gold standard confirmatory test. When tested combined with ELISAspecificity is > 99.9 %.
  • 31. Relation of HIV structure and WesternBlot configuration
  • 32. Principle of Western Blot HIV viral antigens are separated as;gp160,gp120,p66,p55,p51,gp41,p31,p24,p17, and p15.Interpretation depends on presence /absence of reactivity to specific antigens.
  • 33. Interpretation of Western Blot. The antibodies in the serum should reactwith at least two of following antigens,gp 160/120 If does not meetgp 41 requirements, markedp 24 as indeterminate.
  • 34. Negative Western Blot. Absence of any reactivity to thebands is declared as Negative.False positives are rare.All doubtful results should bereported as Negative.
  • 35. Profile of Western Blot Test
  • 36. Limitation of Western Blot Test If not designed for HIV 2 inclusion, we miss HIV2infections, Can give Indeterminate results inPregnancyAfter administration of TetanusToxoid.Autoimmune conditions.
  • 37. Rapid tests in the Diagnosis of AIDS A growing importance Results can be issued within < 20 minutes. Limited protocols, and less demandingtechnical skills. But needs confirmation with ELISA /Western Blot testing. Can differentiate HIV 1 and HIV 2.
  • 38. Rapid Testing for AIDS with DotMethods
  • 39. Dot / Rapid MethodsUseful In patients in Labor whose Immune statusis not known, Resource poor establishments. But needs confirmation with ELISA / WB
  • 40. Other supportingtests in AIDSSeveral tests will help in supportingto identify the progression ofDisease.
  • 41. Flowcytometry for Estimation ofCD 4 lymphocytes
  • 42. Use of CD4 counts in Progress ofAIDS CD4 count measurements help for stagingthe diseases. Flow cytometry has become the prominenttool in estimations of CD4. Specimens should be promptly processed.
  • 43. Uses of Absolute CD4 Lymphocytecounts. Most widely used predictor of HIVprogression. Risk of progression to an AIDS definingillness, opportunistic infections ormalignancy is high, when the counts dropbelow 200/mcl.
  • 44. Uses of CD4 Lymphocyte percentage. Estimation of percentages is more usefulthan CD4 counts. When the counts drops < 20% we have towatch for onset of opportunistic infectionsand malignancy.
  • 45. Molecular MethodsinDiagnosis of AIDS
  • 46. Molecular Methods in Diagnosis ofHIV / AIDS.1. The reverse transcription polymerasechain reaction.2. Nucleic acid sequenceamplification. ( NASBA )3. Branched chain DNA bDNA.
  • 47. DNA –PCR ( RT-PCR) popularamong Molecular Method
  • 48. Polymerase Chain Reaction.-RT- PCR RT-PCR Need the use of Reverse transcriptaseenzyme that converts RNA to DNA. The testing is based on artificially replicatingNucleic acid sequenceof the target ( viral sequence ) so thatMillions to Billions are made availablefor detection.
  • 49. HIV Viral Load tests. The tests measure the amount of activelyreplicating HIV virus. Correlates with response to antiretroviraldrugs Most helpful in diagnosis of HIV beforeSero conversion. False positives should be possible with lowviremia < 500 copies.
  • 50. Alternative Testing Media.From other Specimens. Saliva Detection of Ig G Urine Detection of Ig GBoth ELISA and Western Blotcan be performed.
  • 51. Reporting of all Positive results is agreat concern, Avoid casual reporting
  • 52. Post test counseling a Must in allPositive results
  • 53. Precautions in HIV Testing. Casual testing to be curtailed. To do with a purpose. HIV positive result has Moral,social,andEconomic consequences. Post test counseling a must in all HIV+individuals.