The document discusses various laboratory tests for diagnosing HIV infection. It describes specific tests like antigen detection using the p24 antigen test, virus isolation, detection of viral nucleic acids using PCR, and antibody detection using ELISA, rapid tests, and Western blot. It also mentions nonspecific tests involving blood counts and immune markers. The timeline of biomarkers in acute HIV infection is outlined, noting that p24 antigen appears first followed by IgM and IgG antibodies.

1. Labtest for the diagnosisof
HIV
M.ARJUN
18UT25
III BSc Biotechnology

2. SPECIFICTESTSFORH.I.VINFECTION
• Antigen detection: p24 antigen
• Virus isolation
• Detection of viral nucleic acid
• Antibody detection

3. NonespecificTests
• Total and differential leucocyte
count
• T lymphocyte subset assays
• Platelet count
• IgG and lgA levels
• Skin Tests for CMI

4. Antigen Detection:
SPECIFICTESTSFORHIV
INFECTION
• 1.After a single massive infection viral antigen p24 and RT
maybe detected in blood about 2 weeks.Note:
• The p24 antigen is the earliest viralmarker to appear in the
blood.
• lgM antibodies appear in about 4-6 weeks to be followed by
IgG antibodies.

5. • If the infecting dose is small example inneedle-
stick injury the process may be considerably
delayed.
• The appearance of p24 antigenemia and
viremia followed by IgM antibodies response
coincides with the acute illness.
• The p24 antigen capture assay (ELISA) which
uses anti-p24 antibodies as the solid phase and
can be used for this.
• The test is positive in about 30% cases.

6. Virus isolation
• The viruses present in circulation and body fluids, within
lymphocytes or free cells.
• Virus titers parallel p24 titers, being high soon after
infection, low and antibodies-bound during the
asymptomatic period, and again high towards the end.
• The infectivity being highest in "the early phase and when
the person becomes terminally ill

7. • The virus is present in many parts of the
body and can be isolated from the
peripheral lymphocytes by the technique of
co-cultivation of the patient's lymphocytes
with uninfected lymphocytes in the
presence of interleukin-2.
• Viral replication can be detected by RT.
• Note: Because of the risk involved,
virusisolation is to be attempted only in
laboratorywith adequate containment
facilities.

8. Detection of Viral
Nucleic Acid
• As the most sensitive and specific test, PCR has become
the gold standard for diagnosis in all stages of HIV.
PCR
DNA PCR
RNA PCR
• A related test, HIV RNA PCR can be used fordiagnosis
as well as for monitoring the level of viremia.

9. Antibody detection
• 2-8 weeks to months for the antibodies to
appear after infection and during part of this
period, the individual may be highly infectious
known as the window period.
• Infection can be detected during the window
period by the p24 assay.
• Antibody testing will have to be done after 2-6
months to ascertain whether the infection has
occurred or not.
• IgM antibodies diss appear in 8-10 weekswhile
lgG antibodies remain throughout.

10. Specifictestsforthelaboratory
diagnosisof HIVinfection

11. Screening test
• ELISA
• Rapid test
• Simple test
Supplement tests
• Western blot test
• Indirect immunofluorescence test
• Radio immuno-precipitation assay

12. • Direct solid-phase antiglobulin ELISA is the method
most commonly used. The antigen is obtained from HIV
grown in a continuous T-lymphocyte cell line or by
recombinant techniques and should represent all
groups and subtypes of HIV-1 and HIV-2. The antigenic
coated on the microtitre wells or other suitable solid
surfaces. The test serum is added. And if the antibody is
present, it bindsto the antigen.
SCREENINGTests
ELISA tests:

13. • After washing away the unbound
serum,antihuman immunoglobulin linked to
suitable enzymes is added. If the test serum
contains an anti-HIV antibody, a photometrically
detectable color is formed which can be read by
special ELISA readers.
• Rapid Tests: It has been introduced for the
purpose such as a cylinder or cassette ELISA,
immunochromatographic, coated particle
agglutination.
• Simple Tests: They take 1-2 hours and do not
require expensive equipment.

14. ELISATest

15. Western Blot test:
• The confirmatory test commonly employed is
the Western Blot test. In this test HIV proteins
separated according to their electrophoretic
mobility(and molecular weight) by
polyacrylamide gel electrophoresis are blotted
onto strips of nitrocellulose paper. These strips
are reacted with test sera and then with
enzyme-conjugated antiuman globulin.
SUPPLEMENTTESTS

16. • A suitable substrate is then added, which produces a
prominent color band where the specific antibody
has reacted with the separated viral proteins. Bands
will be of p24,p31 & gp41, gp120 or gp160.
• A positive reaction with proteins representing the
three genes gag, pol,env is considered positive even
if it shows bandsagainst at least two of the following
gene:p24, gp41, gp120/160.

17. Westernblotreactivityin 1-HIVSeroconvertor
Western Blot test

18. IndirectImmunofluorescenceTest:
• HIV-infected cells are fixed onto glass slides and
then reacted with serum followed by fluorescein-
conjugated antihuman gamma globulin. In a
positive test, apple-green fluoresce appears when
examined under a fluorescent microscope.

19. NON-SPECIFICTESTS
• Total leukocyte and lymphocyte count to
demonstrate leucopenia and a lymphocyte
count usually below 2000/mm3.T cell subset
assays. Absolute CD4+T cell count will be
usually less than 200/mm3. Platelet count
will show thrombocytopenia.Raised IgA &
lgG levels. Diminished CMI as indicated by
the skin test.Lymph node biopsy showing
profound abnormalities.