Laboratory Diagnosis of HIV

Lab diagnosis for HIV infection include test’s for immunodeficiency as well as specific test for HIV.

 A] Immunological test’s –

        Following parameter's are help to establishe the immunodeficiency in HIV infection.

1] Total leucocytes & lymphocyte count. Count usually below 2000 mm3

2] Absolute CD4 + T cell count will be usually less than 200 mm3

T4 : T8 ratio is reversed.

 

3] Thrombocytopenia

 4] Ig G & Ig A level.

 

5] Diminished CMI

 6] Abnormal lymphnodes.

B] Specific test for HIV –

 1] Antigen detection –

Following massive infection as by blood transfusion, virus Ag may be detect able in blood

- after about 2 week’s.

- Major core Ag is P24

- Ig m Ab appear about 4-6 week’s.

- P24 – disappear’s from circulation but antibody bound P24 Ags are continues.

- P24 Ag – present 1st few week’s after infection & in terminal phase.

- It is currently used for screening blood donor in USA along with HIV ELISA.

2] Virus isolation –

Technique – by co – cultivation of patient lymphocyte with uninfected lymphocytes inpresence of interleukin – 2.

 Viral replication detected by – demonstration of reverse transcription activity as well as antigen in system.

3] P.C.R. –Most sensitive & specific test in two forms.

* DNA PCR –Peripheral lymphocytes subject are lysed & proviral DNA amplified using primer pains from relatively constant regions of HIV genome.

* RNA PCR –Can be used for diagnosis as well as monitoring the level of viremia.

 4] Antibody detection – Simplest & most employed technique for diagnosis.

 “Seronegative infective stage is known as the window period.”

• Serological test’s are of two types.

  1] Screening test.

  2] Confirmatory test.

 

▶Screening test –

• Posses high sensitivity.

• Have broad spectrum reactivity.

• Simple to perform.

• Automated for handling large number of samples at a time.

• But not specific.

ELISA –

▶Confirmatory test –

• Western blot – HIV proteins separated according to their electrophoretic mobility by polyacrylamide gel electrophoresis on nitrocellular paper.

Test may be considered +ve even if it shows band against at least two gene band.

Ex – P24 gp41 gp120 or gp160

Western blotting

Separates the components according to their molecular weight the proteins in the gel are transferred to

the sheet of nitrocellulose or nylon by the passageof an electric current probed with Ab & then radiolabeled or

enzyme-linked 2nd Ab.

A position is visualized by means of an

ELISA reaction.