This research addresses the development of flow cytometer-based immunoreactive bead (IRB) assay for the detection of specific IgG and IgM antibodies to rDNA-produced HIV antigens. Uniform polystyrene beads are coated with four assay antigens (p31, gp120, p24 and gp41), with a different sized bead used for each antigen. The coated beads are incubated with diluted serum samples and the bound immunoglobulin identified by using fluorescein isothiocyanate-F (ab')2 anti-human IgG or IgM. The stained beads are then analyzed with FACSCAN (Becton- Dickenson) according to bead size. Beads with different fluorescein conjugates can also be separated and counted at a rate of 20,000 beads in less than 20 seconds. Using the Western blot assay as the standard, the IRB assay exhibited 100% sensitivity and 100% specificity. Furthermore, 61 of the 62 enzyme immunoassay (EIA) false positive, WB- negative specimens were negative by IRB assay. Of 18 EIA-positive WB- equivocal sera, 14 were identified as HIV-negative and 1 as HIV-positive by IRB assay. Thus, the assay is sensitive, specific, rapid, and adaptable to automation. Most importantly, detection of HIV infection may be made much earlier by an assay such as this based on IgM detection than by other currently available assays. The development of an automated blood screening system (ABSS) practically useful in blood centers.
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