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Back to Infections Lab Tests
HIV test
Several tests exist to detect the presence of HIV, the virus that
causes AIDS. They are used for different purposes:
- to test whether an individual is infected with HIV
- to screen donated blood or organs
- to measure the amount of free virus in a patient's blood in order to
monitor the progress of the disease under Wilkinson antiretroviral
drug therapy
Some terminology:
the window of a test is the time period after initial infection during
which the test cannot yet detect the presence of HIV. The shorter the
better.
the sensitivity of a test is the percentage of HIV positive cases that
are correctly identified as positive by the test.
the specificity of a test is the percentage of HIV negative cases that
are correctly identified as negative by the test. The higher the
better.
Antibody tests
The body produces antibodies when infected with HIV, and the antibody
tests detect the presence of these rather than the presence of HIV
itself.
In 1996, the CDC described the case of an HIV-infected man who
persistently tested negative on the antibody tests [1]. This is
considered to be extremely rare.
The ELISA test
The ELISA test is the first screening test commonly employed; it has a
high sensitivity but not a very high specificity. The test proceeds by
the general ELISA method: the patients 400-fold diluted serum is
poured over a plate to which known HIV particles were attached. The
HIV-antibodies in the serum (if present) will bind to the HIV
particles. The rest of the serum is washed away. Then a specially
prepared new antibody, which attaches to antibody and to which an
enzyme was bound, is poured over the plate. The new antibody which did
not attach to any antibody is washed away. The enzyme action causes a
color change. This allows to detect whether any of the new antibody
remained, which is only possible if the patient's serum contained
HIV-antibodies.
The low specificity of the test comes from the fact that sometimes
there are cross-reacting antibodies: these are antibodies which attach
to HIV particles "by accident", even though the body has never
encountered HIV before. If an ELISA test is positive, the result is
commonly confirmed with a Western-Blot test.
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The Western-Blot test
The Western-Blot test uses the general Western Blot procedure.
HIV-infected cells are opened and the contained proteins are entered
into a slab of gel to which a voltage is applied. Different proteins
will move with different velocities in this field, depending on their
size and electrical charge. Once the proteins are well separated, they
are transferred to another plate and the procedure continues similar
to ELISA: the patient's diluted serum is poured over the plate,
HIV-antibodies bind to certain proteins on the plate, others are
washed away, and enzyme-linked antibodies detect the presence of bound
antibody. Since the proteins were separated, it is possible to see
exactly to which HIV proteins the patients has antibodies (which the
regular ELISA test cannot accomplish). The test is considered positive
if antibodies to several major HIV proteins are present.
The combination of ELISA and Western Blot is the standard test
employed for diagnosing HIV infection. All blood and organ donations
are also screened with these tests. The window for the newer versions
of these tests is stated in the literature to be on average 22 days.
The patients gets the results usually 2 days to 2 weeks after the
blood withdrawal.
OraQuick
OraQuick is another antibody-based test; it needs only one drop of
blood from the finger and gives results in 20 minutes. Here the blood
is mixed with a solution and then a measuring strip is inserted; the
solution slowly travels upwards on the strip until it reaches a region
where HIV proteins are attached. If the blood contains HIV antibodies,
they will bind here, later causing a color reaction.
Orasure
Orasure is an HIV test which uses mucosal trasudate from the tissues
of cheeks and gums. It is an antibody test which first employs ELISA,
then Western Blot.
There is also a urine test; it employs both the ELISA and the Western
Blot method.
Home Access Express HIV-1 Test
Home Access Express HIV-1 Test is an approved home test: the patient
collects a drop of blood and mails the sample to a laboratory; the
results are obtained over the phone.
There are also several unapproved immediate home-test products on the
U.S. market.
Antigen tests
Antigen tests directly detect the presence of a part of the virus (an
antigen), by applying specific antibody.
p24 Antigen test
The p24 antigen test detects the presence of the p24 protein of HIV
(also known as CA), a major core protein of the virus. It uses a
variant of the ELISA strategy: a well is coated with monoclonal
antibodies specific to the p24 protein. The patient's blood is poured
over the plate, and p24 protein will stick to the antibody; the rest
is washed away. Then enzyme-linked antibody to p24 is applied to the
plate; the enzyme action then causes a color change if p24 was present
in the sample.
This test is now used routinely to screen blood donations, thus
reducing the window to about 16 days. It is not useful for general
diagnostics, as it has very low sensitivity and only works during a
certain time period after infection; once the body produces antibodies
against HIV, the test becomes useless.
A variant of the p24 test first applies heat to denature the p24
protein and thus separate it from its antibody. This test is used to
monitor the viral load and disease progression, as a cheaper
alternative to the nucleic acid based methods described next.
Nucleic acid based tests
In the Amplicor RT-PCR test, the viral RNA is extracted from the
patient's plasma and is treated with reverse transcriptase so that the
RNA of the virus is transcribed into DNA. The polymerase chain
reaction (PCR) is applied, using two primers thought to be unique to
the virus's genome. After the PCR amplification process is completed,
which takes some time, the resulting amplified segments bind to
specific oligonucleotides bound to the vessel wall and are then made
visible with a probe that's bound to an enzyme. The amount of virus in
the sample can be quantified.
Branched DNA plasma test
In the Quantiplex bDNA or branched DNA test plasma is centrifugated to
concentrate the viruses, which are then opened to release the RNA.
Special oligonucleotides are added which bind to viral RNA and to
certain oligonucleotides bound to the wall of the vessel. In this way,
viral RNA is fastened to the wall. Then new oligonucleotides are added
which bind at several locations to this RNA; and other
oligonucelotides which bind at several locations to those
oligonucleotides. This is done to amplify the signal. Finally,
oligonucleotides that bind to the last set of oligonucleotides and
that are bound to an enzyme are added; the enzyme action causes a
color reaction which allows to quantify the viral RNA in the original
sample. Currently, version 3.0 of the Quantiplex is in use; it is
claimed to detect viral loads as low as 50 per millilitre.
Nucleic acid based tests are now routinely used to determine the viral
load, the amount of free virus in the blood. This is an important
variable when monitoring a drug therapy.
In the U.S. donated blood is also screened with nucleic acid based
tests, shortening the window to about 12 days. Since these tests are
relatively expensive, the blood is screened by first pooling some
10-20 samples, testing these together, and if the pool tests positive,
each sample is retested individually.
Other
The CD4 T-cell count is a procedure where the number of CD4 T-cells in
one microlitre of blood are counted in a standard medical lab test
after a blood draw.
This test does not check for the presence of HIV. Instead, it is used
to estimate viral activity and immune system function in HIV+ people.
This test is also used occasionally to estimate immune system function
for people whose CD4 T cells are impaired for reasons other than HIV
infection, which include several blood diseases, several genetic
disorders, and the side effects of many chemotherapy drugs.
Generally speaking, the lower the number of T cells, the lower the
immune system's function will be. Normal T4 counts are about 1000 CD4+
T cells per microliter, although the counts may fluctuate by 10% or
more even in healthy people, depending on recent infection status,
nutrition, exercise and other factors -- even the time of day. Women
tend to have somewhat lower counts than men.
Symptoms of T4 cell immune collapse are almost never seen until the
number drops below 500. In HIV+ people, AIDS is officially diagnosed
when the count drops below 200 cells.
Similar symptoms of immune collapse are generally seen in people with
very low T4 cell counts, no matter whether this immunosuppression is
caused by HIV or by cancer or by any other disease. However, the
long-term treatment differs substantially, because it needs to address
the cause of the immunosuppression.
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