Fig. 3

Vaccine-induced antibody (VIA) in serum from mice immunized with adenovirus encoding full length human 3 epidermal growth factor receptor (HER3-VIA) induces receptor internalization and downregulation of human epidermal growth factor receptor 3 (HER3), but not epidermal growth factor receptor (EGFR) nor HER2. SKBR3, BT474-M1 (a) and MDA-MB-468 (b) were incubated with 1:100 HER3-VIA or green fluorescence protein (GFP)-VIA at 37 °C for 1 or 3 h, respectively. After washing and permeabilization, RedTM-conjugated anti-mouse IgG (H + L) was used to visualize internalization of the VIA-bound proteins with a fluorescence microscope. SKBR3 cells (c) or MDA-MB-468 cells (d) were incubated with 1:100 HER3-VIA or GFP-VIA at 37 °C for 3 h. Cells were fixed, permeabilized and stained with anti-HER2 antibody (trastuzumab, 40 μg/mL) (c) or anti-EGFR antibody (cetuximab, 40 μg/mL) (d), followed by Cy2-conjugated anti-human IgG antibody