Fig. 4.
Germ-line IGHV1-53 antibodies are polyreactive and do not neutralize HIV. (A) The IGHV1-53 MZ antibodies were tested for polyreactivity by binding to chromatin (Left) and cardiolipin (Center) by ELISA or to HEp-2 cells (Right) by indirect immunofluorescence staining. Mouse IgM hybridomas specific for NP (B1-8) or H-2b (3-83) were used as isotype controls. (B) The IGHV1-53 MZ antibodies were tested for CD4 binding site recognition by ELISA against RSC3. VRC01 was used as a positive control. (C) Neutralization ability of IGHV1-53 MZ antibodies was determined using a TZM-bl assay. Mouse IGHV1-53 antibodies were incubated with JR-FL pseudovirus for 30 min before culturing with TZM-bl cells for 48 h. Infection was determined with the Beta-Glo assay and quantitated on a luminometer. Percent infection was calculated in relation to values from wells that received no antibody. Human b12 was used as a positive control for neutralization, and a mouse IgM specific for LPS served as a negative control.