Background: Analysis of human monoclonal antibodies (mAbs) developed from HIV-1 infected donors have enormously contributed to the identification of neutralization sensitive epitopes on the HIV-1 envelope glycoprotein.The third variable region (V3) is a crucial target on gp120, primarily due to its involvement in co-receptor (CXCR4 or CCR5) binding and presence of epitopes recognized by broadly neutralizing antibodies.Methods: Thirty-three HIV-1 seropositive drug naive patients (18 males and 15 females) within the age range of 20-57 years (median 033 years) were recruited in this study for mAb production.The mAbs were selected from EBV transformed cultures with conformationally constrained Cholera-toxin-B containing V3C (V3C-CTB) fusion protein.We tested the mAbs for their binding with HIV-1 derived proteins and peptides by ELISA and for neutralization against HIV-1 viruses by TZM-bl assays.Results: We isolated three anti-V3 mAbs, 277, 903 and 904 from the cells of different individuals.The ELISA binding revealed a subtype-C and subtype-A specific binding of antibody 277 and 903 while 904 exhibited cross reactivity also with subtype-B V3.Epitope mapping of mAbs with overlapping V3 peptides showed exclusive binding to V3 crown.The antibodies displayed high and low neutralizing activity against 2/5 tier 1 and 1/6 tier 2 viruses respectively.Overall, we observed a resistance of the tier 2 viruses to neutralization by the anti-V3 mAbs, despite exposure of the epitopes recognized by these antibodies on the native viruses, as determined by intact virion binding assay with two representative subtype-C and B viruses (Du156.12and JRFL).Conclusion: Our study suggests that the anti-V3 antibodies derived from subtype-C infected Indian patients display neutralization potential against tier 1 viruses.Defining the epitope specificities of these mAbs and further experimental manipulations will be helpful in identification of epitopes, unique to clade C or shared with nonclade C viruses, for immunogen design.
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