Page 64 - AIDSBK23C

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In the pictured example, the patient sample in lane B is positive while the result for
patient C is negative. The presence of any bands that do not meet the criteria for a positive
result is considered an indeterminate result, as shown in lane A above. Errors in interpretation
occur with sample misidentification, cross-contamination of negative samples by adjacent
positive samples, HLA antibodies in the viral lysate used in the preparation, or misinterpretation
of band patterns. A WB should not be used as an initial HIV screening test because it has a
much higher false positive rate than EIA. Likewise, plasma HIV-1 RNA testing should not be
used for screening because of the false positive rate up to 3% (suggested by an assay yielding a
low plasma viral load).[341]
The WB typically becomes positive 25 to 30 days following initial infection.[327] The
"indeterminate" WB assays may result from repeatedly reactive true positive EIA assays on sera
that are negative to WB in individuals at the early stage of HIV infection when a weak titer of
HIV antibodies may be present. It may occur late in the course of AIDS with loss of core
antibodies with loss of immunologic competence. Up to 10 to 20% of repeatedly reactive EIA
assays can have an indeterminate WB assay.[332] Indeterminate WB assays may also result
from cross-reaction from infection with HIV-2, HTLV, or rare subtype of HIV. Indeterminate
results may occur from nonspecific antibody reactions with autoimmune diseases, vaccination
against influenza, viral infections such as hepatitis, or rabies, multiple pregnancies, recipients of
multiple blood transfusions, and malignancies.[348] Indeterminate WB assays in persons who
are truly HIV negative may occur from contaminating proteins in the viral lysate, from reactivity
due to prior exposure to similar proteins or other viruses, and hyperbilirubinemia.[341,349]
Approximately 3% of persons with indeterminate WB tests subsequently have laboratory
evidence for HIV infection, and most of these persons will have identifiable risk factors for
infection. Indeterminate results can usually be resolved by retesting the patient by EIA assay and
WB. About one-third of persons with an indeterminate WB will not be repeatedly reactive by
EIA assay after retesting in one month. After 6 or more months, most truly HIV-1 infected
persons will be positive. However, an indeterminate WB can persist for years in some persons.
[350,351] Additional testing to resolve indeterminate results can include detection in plasma of
HIV-1 p24 antigen in 75% of early infections, nucleic acid testing (NAT), or HIV-1 RNA, which
can identify virtually all early infections.[348,352]
Indeterminate WB results may also arise from non-specific reactivity of true negative
sera for both EIA and WB assays. Some persons have stable indeterminate patterns and are not
HIV-1 infected. Specific risk factors identified among women for presence of an indeterminate
WB test include parity and also presence of autoantibodies, while in men the risks include a
tetanus booster injection in the past two years or sexual contact with a prostitute.[350]
Correlation of test results with clinical status is necessary.[353,354] Western blot testing can
also be performed for HIV-2 to confirm EIA positive tests.[101] Long term follow-up of persons
donating blood and who have no risk factors for HIV infection reveals no evidence for HIV
infection.[351]
WB testing may aid in identifying infected persons in the early post-seroconversion
period. The WB has very high specificity for identifying persons within 30 days of
seroconversion with the criterion of having 3 or fewer bands, and for persons within 90 days of
seroconversion with the criterion of having 6 or fewer bands.[355]

LINE IMMUNOASSAY.-- The line immunoassay (LIA) methodology is similar to that
of Western blot, but incorporates HIV-1 antigens onto nitrocellulose strips so that each reaction