Figure 4.

Immunogenicity of MVF HER-3 peptides and ability of anti-HER-3 vaccine antibodies to recognize recombinant and native HER-3 receptor. Rabbits (n = 2) were immunized with either MVF HER-3 237–269 or MVF HER-3 461–479 every three weeks for a total of three immunizations. Anti-peptide antibody titers were determined by ELISA with plates coated with the appropriate MVF HER-3 peptide immunogen (A). Titers are defined as the reciprocal of the highest dilution of sera that gave an absorbance reading above 0.2 after subtracting the pre-immune sera. Results represent the average titer for two rabbits with error bars representing the SD of the mean. Anti-HER-3 peptide antibodies were purified from high titered rabbit anti-serum on a protein A/G column. The ability of the peptide specific antibodies to recognize recombinant HER-3 was determined by ELISA (B). Results are representative of three independent experiments with duplicate samples (n = 2). Direct binding of the anti-peptide antibodies to the native HER-3 receptor was determined by flow cytometric analysis with BXPC3 and MCF7 cells (C). Cells were treated with 50 ug/mL anti-HER-3 antibodies for 2 h. HER-3 binding was detected with goat anti-rabbit IgG-Alexa fluor 488 secondary antibody. Cells were analyzed on a BD FACS Calibur system. Pre-immune serum was used as a negative control.