Development of a broadly effective HIV-1 vaccine has been limited because of the inherent structural properties the HIV-1 envelope expressed on the native virion. HIV-1 infection involves a series of stepwise changes in env antigenic structures and epitope presentation during which the virus unfolds and exposes neutralizing epitopes in close proximity to the target cell surface structure. We propose, that it is difficult for the immune system to respond in a higher order integration in which a two step antibody response can modify the envelope to expose and then secondarily recognize exposed neutralizing epitopes. This higher integration must be imposed on the immune system through vaccine design in that immunogens must be designed to induce multiple classes of antibodies which together mediate viral neutralization through an integration of these two steps: """"""""exposure"""""""" and """"""""neutralization."""""""" Analysis of the humoral immune response has been hampered by the fact that protein subunits and antibody assays do not accurately reflect the complex oligomeric structure on native virions. To address these concerns, identify antibodies that react with native primary isolate virions, and facilitate analysis of antibody specificity and viral neutralization, we have designed an assay using whole virions from primary isolates of HIV-1. When serum from HIV-1 infected individuals was tested for IgG antibodies reactive with intact primary isolate virions, 36% of HIV-1 infected individuals captured virus (>300pg/ml). Only 7% of these individuals captured significant quantities of virus (>500pg/ml). By comparison, F240, a gp41 human monoclonal antibody (HMAb) captured more than 2150pg/ml of p24. Clearly antibodies reactive with primary isolate virions are not prevalent in the majority of HIV-1 infected individuals which may be related to the general inability of sera to neutralize primary isolates. We propose to generate HMAb which bind to intact primary isolates of clades A-G to identify broadly reactive antibodies. We further propose to generate antibodies to epitopes exposed on virions following binding of reactive antibodies. Isolation and characterization of HMAb reactive with intact virions from primary isolates should lead to the identification of antigenic epitopes and motifs for vaccine purposes.
