Removal of anti-cell antibodies. (A) Effective removal of anti-cell antibodies in sera of immunized animals. Sera of rabbits were incubated with human 293 cells for 4 h at room temperature with moderate shaking. Three rounds of incubation were sufficient to remove all significant anti-cell activity, measured by staining of human 293 T cells with dilutions of sera and PE-labeled anti-rabbit antibody. Mean fluorescence intensity (MFI) of a representative serum is shown. (B) Anti-MPER antibodies 2F5 and 4E10, but not anti-gp41 Abs induced by immunization were depleted by incubation with cells. Serial dilutions of 2F5, 4E10 and immune sera in PBS were incubated three times with an equal volume of HEK293F cells. Supernatants were read out in a gp41 ELISA with MPER peptide as coating reagent, and anti-human-HRP coupled antibody as secondary antibody.