CLONING AND BIOLOGICAL CHARACTERIZATION OF HUMAN SINGLE-CHAIN FV FRAGMENTS THAT MEDIATE NEUTRALIZATION OF HIV-1

Objective: To develop recombinant single-chain Fv fragments against HI V-1 gp120. Methods: A panel of human monoclonal antibody Fv fragments were generated against the HIV-1 gp120 by affinity selection from an a ntibody library expressed on the surface of filamentous phage. The lib rary was prepared from peripheral blood lymphocytes of an asymptomatic HIV-1-infected mother with a high neutralization titer. This mother d id not transmit HIV-1 to her offspring (non-transmitter). Heavy and li ght chains were initially amplified separately and combined by splicin g by overlap extension to generate Fv fragments. Results: Several clon es expressing single-chain Fv fragments bind strongly to HIV-1 gp120 a nd several were found to neutralize cell-free HIV-1(IIIB). Gross epito pe mapping suggests that different clones bound to different functiona l regions on the envelope. The clones also exhibited sequence diversit y. Conclusions: This strategy of cloning resulted in the development o f functional human-derived antibody reagents with different anti-HIV-1 biological properties in vitro. These recombinant Fv fragments have p otential utility as immune reagents, as well as in the design of poten tial immunotherapeutics. In addition, these antibody reagents may prov ide information on the relationship between humoral immunity and mater nal-fetal (vertical) HIV-1 transmission.