diagnosis of-aids

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Diagnosis of AIDS

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Page 1: Diagnosis of-aids

Diagnosis of

AIDS

Page 2: Diagnosis of-aids

AIDS A Global Concern

TODAY THERE IS NO REGION OF THE

WORLD UNTOUCHED BY PANDEMIC OF

AIDS

Page 3: Diagnosis of-aids

Majority of persons infected with HIV present with AIDS soon or later.

Page 4: Diagnosis of-aids

Why AIDS is more important TODAY

Apart from Men, there is a increasing burden of

Infections in Women.

Implication on Mother to Child Transmission

Female Adolescents are infected 3 - 6

times higher than male counterparts.

AIDS has turned out to be Social

Problem.

Page 5: Diagnosis of-aids

Implication of AIDS

Being diagnosed as having HIV/AIDS has life time repercussions.

Every body understands AIDS as a life threatening condition.

A casual way of dealing as being HIV + will have Moral, Legal, and Social consequences.

Page 6: Diagnosis of-aids

Prompt and Precise Diagnosis of AIDS make great impact on Humanity

Page 7: Diagnosis of-aids

Importance of precise Diagnosis of AIDS

All patients should be informed about

consequences of Testing for HIV infection,

Legality of respective National Laws should be

followed.

A pretest and posttest counseling is Growing

importance, even in Developing countries.

Today’s debate on testing for HIV status as

matter of Health protocol ?

Page 8: Diagnosis of-aids

Counseling Improves the confidence

Page 9: Diagnosis of-aids

Purpose of Testing for HIV Infection

A matter of great importance as in Blood donors to 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 Child transmission.

Patient's presenting with opportunistic infections.

Page 10: Diagnosis of-aids

Diagnosis of HIV/ AIDS

Basic Tests.

1.We can detect Antigen or Antibody

2.We can detect Antigen and Antibody.

3.Majority of Laboratories depend on Commercial Kits.

4.Devloped Nations going for Molecular and more precise Methods.

Page 11: Diagnosis of-aids

Screening Tests for HIV/AIDS Detection

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 even

for Diagnosis.

Page 12: Diagnosis of-aids

Confirmatory Tests in AIDS

It is important to confirm all screening tests

with Confirmatory tests, or we brand some

one without infection as infected,

Confirmatory tests differentiates false

reactive tests. and identifies truly infected

or not.

Page 13: Diagnosis of-aids

Differences of Screening and Confirmatory Tests.

Screening test

High degree of

Sensitivity

Few false negatives

Confirmatory test

High degree of

specificity

Few / No false positive

results.

Page 14: Diagnosis of-aids

Why patients should we tested with Screening and Confirmatory tests.

Before you declare a person infected with

HIV / AIDS you should perform both

methods.

Faulty testing methods can lead to

catastrophic consequences, and legal

litigations.

Page 15: Diagnosis of-aids

Choosing Screening Testing methods

Our aim to obtain 100 % specific results ?

But impossible.

As few may be infected and do not have sufficient antibodies to detect in the testing methods.

Depend on time tested methods.

Explore the reputation of the suppliers.

Page 16: Diagnosis of-aids

ELISA METHOD

Universally accepted test ,most popular

even in the developing nations.

Useful in large scale screening.

A common method used in Blood banks in

mass screening of Human blood.

Page 17: Diagnosis of-aids

ELISA method – A ELISA plate

Page 18: Diagnosis of-aids

Sequence of Events in ELISA test

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Different Methods of ELISA testing.

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Why ELISA is very trust worthy.

Easy to perform

Free from Radiation.

Relatively specific

Sensitive.

With scientific advances we have advanced to new generation of testing methods

Today we depend on 3rd and 4th generation ELISA methodologies.

Page 21: Diagnosis of-aids

Significance of ELISA

Most reliable screening test for HIV infection.

50% of the infected will show positive reactivity in < 22 days.

95% show reactivity < 6 weeks

Sensitivity > 99.9 %

But needs confirmation with Western Blot.

Page 22: Diagnosis of-aids

Advances in ELISA Methods.

1st and 2nd generation ELISA methods have

become obsolete we have to switch to

Antigen sandwich ELISA protocols ( called

as 3rd generation ELISA ).

Many Nations have switched to 3rd

generation methods.

Page 23: Diagnosis of-aids

What is 3rd Generation ELISA

Helps in detection of early infections,

Can detect all classes of antibodies

including Ig M

All isotypes of antibodies can be captured.

But proving to be expensive to developing

Nations.

Page 24: Diagnosis of-aids

Need for 4th Generation ELISA

Methods

The quest for better Diagnosis of AIDS

continues

Page 25: Diagnosis of-aids

What is 4th generation ELISA.

A new generation Method in ELISA

Can detect both Antigen and Antibody in

the same run ( in the same ELISA Plate )

Helpful to make early diagnosis at least in

few cases.

Page 26: Diagnosis of-aids

Determination of P 24 Antigen by ELISA

Helps in the early detection of HIV infection.

Screening of Blood for HIV infection along with detection for antibody detection.

Early diagnosis of HIV infection in resource poor nations.

Monitoring Anti-retroviral therapy. But not a replacement for RNA quantization.

Even P 24 Antigens are subject of false positive results.

Page 27: Diagnosis of-aids

Limitation of Screening Tests.

Imperfect Sensitivity,

Sensitivity to detect HIV2 ?

Failing to detect HIV variants.

Other technical errors.

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Confirmatory Serological Tests

All the Screening tests need to be

confirmed before being declared as

HIV infected.

Page 29: Diagnosis of-aids

Confirmatory Serological Tests.

HIV Western Blots.

Line Immunoassay ( LIA )

Immuno fluorescent Assay ( IFA )

Utility is limited due technical and

economical reasons

Page 30: Diagnosis of-aids

WESTEN BLOT TEST

A Gold standard confirmatory test.

When tested combined with ELISA

specificity is > 99.9 %.

Page 31: Diagnosis of-aids

Relation of HIV structure and Western Blot configuration

Page 32: Diagnosis of-aids

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.

Page 33: Diagnosis of-aids

Interpretation of Western Blot.

The antibodies in the serum should react

with at least two of following antigens,

gp 160/120 If does not meet

gp 41 requirements, marked

p 24 as indeterminate.

Page 34: Diagnosis of-aids

Negative Western Blot.

Absence of any reactivity to the

bands is declared as Negative.

False positives are rare.

All doubtful results should be

reported as Negative.

Page 35: Diagnosis of-aids

Profile of Western Blot Test

Page 36: Diagnosis of-aids

Limitation of Western Blot Test

If not designed for HIV 2 inclusion, we miss HIV2

infections,

Can give Indeterminate results in

Pregnancy

After administration of Tetanus

Toxoid.

Autoimmune conditions.

Page 37: Diagnosis of-aids

Rapid tests in the Diagnosis of AIDS

A growing importance

Results can be issued within < 20 minutes.

Limited protocols, and less demanding technical skills.

But needs confirmation with ELISA / Western Blot testing.

Can differentiate HIV 1 and HIV 2.

Page 38: Diagnosis of-aids

Rapid Testing for AIDS with Dot Methods

Page 39: Diagnosis of-aids

Dot / Rapid Methods Useful

In patients in Labor whose Immune status

is not known,

Resource poor establishments.

But needs confirmation with ELISA / WB

Page 40: Diagnosis of-aids

Other supporting tests in AIDS

Several tests will help in supporting

to identify the progression of

Disease.

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Flowcytometry for Estimation of CD 4 lymphocytes

Page 42: Diagnosis of-aids

Use of CD4 counts in Progress of AIDS

CD4 count measurements help for staging

the diseases.

Flow cytometry has become the prominent

tool in estimations of CD4.

Specimens should be promptly processed.

Page 43: Diagnosis of-aids

Uses of Absolute CD4 Lymphocyte counts.

Most widely used predictor of HIV

progression.

Risk of progression to an AIDS defining

illness, opportunistic infections or

malignancy is high, when the counts drop

below 200/mcl.

Page 44: Diagnosis of-aids

Uses of CD4 Lymphocyte percentage.

Estimation of percentages is more useful

than CD4 counts.

When the counts drops < 20% we have to

watch for onset of opportunistic infections

and malignancy.

Page 45: Diagnosis of-aids

Molecular Methods in

Diagnosis of AIDS

Page 46: Diagnosis of-aids

Molecular Methods in Diagnosis of HIV / AIDS.

1. The reverse transcription polymerase

chain reaction.

2. Nucleic acid sequence

amplification. ( NASBA )

3. Branched chain DNA bDNA.

Page 47: Diagnosis of-aids

DNA –PCR ( RT-PCR) popular among Molecular Method

Page 48: Diagnosis of-aids

Polymerase Chain Reaction.-RT- PCR

RT-PCR Need the use of Reverse transcriptase

enzyme that converts RNA to DNA.

The testing is based on artificially replicating

Nucleic acid sequence

of the target ( viral sequence ) so that

Millions to Billions are made available

for detection.

Page 49: Diagnosis of-aids
Page 50: Diagnosis of-aids

HIV Viral Load tests.

The tests measure the amount of actively replicating HIV virus.

Correlates with response to antiretroviral drugs

Most helpful in diagnosis of HIV before Sero conversion.

False positives should be possible with low viremia < 500 copies.

Page 51: Diagnosis of-aids

Alternative Testing Media. From other Specimens.

Saliva Detection of Ig G

Urine Detection of Ig G

Both ELISA and Western Blot

can be performed.

Page 52: Diagnosis of-aids

Reporting of all Positive results is a great concern, Avoid casual reporting

Page 53: Diagnosis of-aids

Post test counseling a Must in all Positive results

Page 54: Diagnosis of-aids

Precautions in HIV Testing.

Casual testing to be curtailed.

To do with a purpose.

HIV positive result has Moral,social,and

Economic consequences.

Post test counseling a must in all HIV+

individuals.