Human Immunodeficiency Virus Western Blot |
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Specimens that test repeatedly reactive for HIV-1 antibody by ELISA must be retested with a more specific confirmatory test. The Western blot is the most widely used serologic test for distinguishing between true and false positive EIA results.
The Western blot assay detects antibodies that react with specific proteins (antigens) of the HIV-1 virus. The gene products used to interpret Western blot results are shown in the table below.
Gene |
Antigen |
Description |
Immunogenicity |
Detection |
Envelope |
gp160 |
gp precursor |
High |
Earliest |
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gp120 |
outer gp |
High |
Earliest |
| |
gp41 |
transmembrane |
Moderate - High |
2-4 wk later |
Gag |
p55 |
gag precursor |
Moderate |
2-4 wk later |
| |
p24 |
gag protein |
Moderate |
Earliest |
| |
p17 |
gag protein |
Weak |
Earliest |
Polymerase |
p66 |
rev transcriptase |
Moderate |
2-4 wk later |
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p51 |
rev transcriptase |
Moderate |
2-4 wk later |
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p31 |
endonuclease |
Moderate |
3-6 wk later |
A consistent sequence of antibody responses occurs after infection. The earliest antibodies to appear are directed against gp160, gp120, p24, and p17, followed shortly by antibodies to gp41, p55, p66, and p51. Anti-p31 appears later. Antibodies to p24 and p55 decline after the onset of symptoms, while antibodies to envelope glycoproteins persist. Anti-p31 also diminishes, but not to the same extent as anti-p24. In some cases, reactions with gp120 and gp160 may be due to antibodies binding to multimers of gp41.
The criteria established by the Centers for Disease Control and the Association of State and Territorial Public Health Laboratory Directors for interpretation of Western blots are as follows:
Interpretation |
Bands Present |
Positive |
Presence of any 2 bands; p24, gp41, gp120/160 |
Negative |
absence of bands |
Indeterminate |
any other combination of bands |
The majority of indeterminate patterns consist of p17, p24, or p55 alone, or combinations of these 3 bands. Indeterminate patterns may be either false positives or early seroconverters. The complete medical history must be considered in interpreting indeterminate Western blots. Individuals with a positive Western blot lacking the p31 band should be counseled that, although they may be infected, uncertainty exists about this conclusion. These individuals should be further evaluated by HIV RNA PCR testing and follow-up HIV serologic testing. Most seroconversions will be detected in repeat Western blots within 3 months. Persons with stable indeterminate patterns lasting 6 months or more, in the absence of known risk factors and clinical symptoms, may be considered negative for HIV-1 antibodies.
Specimen requirement is one SST tube of blood.
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