FLOW CYTOMETRIC IMMUNOFLUORESCENCE ASSAY FOR DETECTION OF ANTIBODIES TO HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 USING INSOLUBLE PRECURSOR FORMSOF RECOMBINANT POLYPROTEINS AS CARRIERS AND ANTIGENS

A new serological assay, the recombinant flow cytometric immunofluores cence assay (r-FIFA), was developed for the early detection of human i mmunodeficiency virus type 1 (HIV-1) antibodies by using recombinant i nsoluble forms of HIV-1 Gag-p45, Gag-gp41 chimeric protein, gp160, and Pol97 polyprotein as antigens and autologous carriers through flow cy tometry. These recombinant proteins were expressed in insect cells by a baculovirus expression system. Eight anti-HIV-1 seroconversion panel s, a low-titer anti-HIV-1 panel from Boston Biomedica Inc, (BBI), and three HIV-1 seroconversion specimens from the Provincial Health Labora tory of Ontario, Toronto, Ontario, Canada (PHL), were tested and analy zed by r-FIFA. In sensitivity comparisons between r-FIFA and tests lic ensed by the U.S. Food and Drug Administration, which were used to tes t all of the HIV-1 panels from BBI, detection of HIV-1 antibody by r-F IFA was on average greater than 20 days earlier than that by enzyme im munoassay. The sensitivity of r-FIFA has permitted the detection of HI V-1-specific immunoglobulin G (IgG), IgM, and IgA antibodies during se roconversion. A kinetic analysis of HIV-1 antibody production by r-FIF A has shown that either IgG or IgM, or both, can be detected, dependin g on the phase and type of the immune response in the HIV-1-infected i ndividual. Both primary and secondary immune responses were observed d uring this period. The r-FIFA results suggest that implementation of r -FIFA may significantly reduce the ''window'' period from the time of infection to the time of seroconversion, with earlier detection of ant ibodies after initial infection. This would also make it possible for us to understand the immune response and the precise mechanisms of imm unopathogenesis in the early period of HIV-1 infection.