Figure 2

Antibodies from immunized animals recognize mouse BP180/ CXVII and activate complement ex vivo . Serum was obtained from immunized animals at different time points. Immune rabbit (a) and sheep (b) antibodies bound to the basal membrane of the mouse skin. In contrast, antibodies obtained from a rabbit before the first immunization (c) did not recognize the antigen in situ. When incubated with mouse split skin, IgG from the immune sera of both the rabbit (d) and sheep (e) bound to the epidermal side of the 1 NaCl-split skin (magnifications 200x). (f) Pathogenic antibodies in contrast to control antibodies recognized recombinant forms of BP180/ CXVII by immunoblotting; in the left hand panel: 1, GST and 2, GST-mCXVII-EC1 incubated with immune rabbit serum; 3, GST-mCXVII-EC1 incubated with control rabbit antibodies as a negative control; in the right hand panel: 4, GST-mCXVII-EC3 and 5, GST-mCXVII-EC7 incubated with immune rabbit serum. BP180/ CXVII-specific antibodies from the rabbit (g) and sheep (h) activated complement when incubated on cryosections of murine skin (magnifications 200x).