Figure 3

Anti-Neu humoral response following rV- neu T vaccination. Panel A: p185 Neu expression in NIH3T3 cells encoding normal rat Neu (LTR-Neu) and in Neu-overexpressing BALB-neu T salivary gland cancer (SALTO) cells by western blotting. A specific polyclonal anti-Neu (C18) antibody was used. NIH3T3 cells were used as negative control. Panel B: Igs from rV-neu T vaccinated mice recognize p185 Neu expressed on the surface of SALTO tumor cells. Immunofluorescence was performed using purified Igs (1μg/ml) of BALB-neu T mice vaccinated with rV-neu T or V-wt. The specific monoclonal antibody anti-Neu Ab4 was used as positive control. Olympus BX51 microscope and I.A.S. version 007 000 Deconvolution 2D software were used to deconvolve z series images of stained native cells. Original magnification x400. Panel C: Serum antibody response of mice upon vaccination with rV-neu T. Numbering identifies immune sera of individual mice. Mouse pre-immune or immune sera were collected prior to vaccination or one week after the second immunization and employed in immunoprecipitation of Neu from LTR-Neu or SALTO tumor cells. p185 Neu specificity was visualized by immunoblotting analysis using receptor-specific polyclonal antibody of immunoprecipitates and compared to direct immunoblotting of LTR-Neu lysate as positive control, or NIH3T3 as negative control.