The revised algorithm recommends initial testing with an HIV-1/2 antigen/antibody combination immunoassay which, if reactive, is followed by supplemental testing with an HIV-1/2 antibody differentiation assay. Specimens negative or indeterminate by the HIV-1/2 antibody differentiation assay undergo an HIV-1 nucleic acid test (NAT). The HIV Laboratory Diagnostic Testing Algorithm offers several advantages over the conventional algorithm of HIV antibody screening followed by Western blot confirmation of repeatedly reactive results, including earlier detection of HIV infections and the ability to accurately classify HIV-1 and HIV-2 infections.
Step 1: HIV-1/2 Antigen/Antibody Immunoassay
If the result of this test is “nonreactive,” the interpretation is HIV-1 antigen and HIV-1/HIV-2 antibodies were not detected; there is no laboratory evidence of HIV infection (Table 1). No further testing of the specimen is indicated. If the result is “reactive,” this indicates possible presence of HIV-1 antibodies, HIV-2 antibodies and/or HIV-1 p24 antigen. This is considered a presumptive positive result and additional supplemental testing is required.
It is strongly recommended that laboratories use a 4th generation IA as the initial test; however, some laboratories may elect to use 3rd generation IAs. Although less sensitive than 4th generation IAs, 3rd generation IAs will detect some HIV-1 infections before the Western blot becomes positive. If a 3rd generation IA is used in step 1, the remainder of the algorithm should be followed because the HIV-1/HIV-2 antibody differentiation assay and HIV-1 NAT assay, in combination, provide fewer false negative results than the Western blot for specimens collected early after infection. If laboratories using 3rd generation IAs continue to perform Western blot testing for confirmation, specimens with a negative or indeterminate Western blot result should reflex to an HIV-1 NAT as outlined in step 3 of the algorithm. Reports should include a limitation statement noting the decreased sensitivity of 4 Association of Public Health Laboratories.
Step 2: HIV-1/HIV-2 Antibody Differentiation Immunoassay
If the initial HIV-1/2 immunoassay (step 1) is reactive and the result of HIV-1/HIV-2 antibody differentiation immunoassay is “reactive” for HIV-1 or HIV-2 antibodies, the interpretation is “positive for HIV-1 antibodies; laboratory evidence of HIV-1 infection is present” or “positive for HIV-2 antibodies; laboratory evidence of HIV-2 infection is present” respectively. No further testing of the specimen is required and clinical follow-up is recommended. If the result is “reactive” for both HIV-1 and HIV-2 antibodies (i.e. HIV positive, undifferentiated), the interpretation is “positive for HIV antibodies; unable to differentiate between HIV-1 and HIV-2 antibodies.” Reactive results are consistent with laboratory evidence of HIV infection and clinical follow-up is recommended. The clinician may consider additional testing for HIV-1 RNA and HIV-2 RNA or DNA to verify or rule-out the presence of HIV-1, HIV-2, or HIV-1/HIV-2 dual infection. If the result of the HIV-1/HIV-2 antibody differentiation test is “nonreactive” or “indeterminate,” testing of the specimen should reflex to an HIV-1 NAT (step 3).
Step 3: HIV-1 NAT
If the initial HIV-1/2 immunoassay (step 1) is reactive and HIV-1 RNA is detected, the final interpretation is “positive for HIV-1; laboratory evidence of HIV-1 infection consistent with an acute or early HIV-1 infection.” Clinical follow-up is recommended. If HIV-1 RNA is not detected, the final interpretation is “HIV-1 antibodies were not confirmed and HIV-1 RNA is not detected; no laboratory evidence of HIV infection.” The initial HIV-1/2 immunoassay result is a possible false positive. If there is reason to suspect recent HIV-2 infection, or if other clinical symptoms suggest immunosuppression, follow-up testing should be considered. Currently, there are no FDA approved HIV-2 RNA or DNA detection assays and those that are performed are laboratory developed tests. In cases where the HIV-1/HIV-2 antibody differentiation assay (step 2) is “nonreactive” or “indeterminate” and the HIV-1 NAT assay (step 3) is either invalid or could not be performed, the algorithm is incomplete and cannot be fully interpreted. In this situation, the interpretation should be reported as “inconclusive.” Whether the report is written or verbal will be situation dependent. An additional specimen should be requested and the entire algorithm should be repeated.