Western blot analysis of serological responses following immunization with vaccinia viral lysates of melanoma cells

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Abstract
Western blot analysis was used to characterize the antibody response of melanoma patients during immunization with vaccinia‐virus‐induced melanoma cell lysates (VMCL). Strong antibody responses were detectable within 4 weeks from commencement of immunization against different fractions (from 3 to 10) in blots of VMCL. The approximate MW of the main fractions identified were 100, 91, 85, 77, 64‐66, 49, 46, 43 and 34‐36 kDa. Comparison of the reactivity of the sera with extracts of vaccinia virus suggested that the reactivity against VMCL was not against viral antigens in the lysates. Studies on autologous extracts from 7 patients immunized with VMCL indicated that the majority of the fractions identified in extracts of VMCL were also identified in autologous extracts of melanoma cells. This indicated vaccine‐induced responses against the patients' tumor cells and not merely against alloantigens in the vaccine. Sera from the immunized patients appeared to identify a number of fractions of similar MW in extracts of colon and glioma cells, suggesting that the responses were not specific for melanoma. Possible exceptions were reactivity against melanoma fractions of 85, 64‐66 and 36 kDa. Our studies provide evidence for the effectiveness of VMCL in inducing antibody responses against autologous melanoma cells and form a basis for subsequent biochemical and genetic analysis of the antigens identified by antibody responses in immunized patients.