OBJECTIVE: To purify MAb183B2 corresponding antigen of ovarian carcinomas and study its physical and chemical characteristics. METHODS: The strongest positive samples reacted with MAb183B2 were screened from ovarian cancer ascites with indirect enzyme linked immunosorbent assay (ELISA). Then the antigen was purified from ascitic fluid by affinity chromatography with MAb183B2. Nature of the purified antigen was examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and western blot and its reactivity was determined after treatment with NaIO4, trypsin, pronase, neuraminidase, deglycolipid mixture and heating. Two dimensional electrophoresis combined with western blot was then used to isolate and identify the antigen from ovarian cancer cell line SKOV3. RESULTS: The reactivity of the antigen after treated by NaIO4, neuraminidase and deglycolipid mixture was still positive, while it was negative after treated by trypsin, pronase and heating. The antigen proved to be composed of two different subunits of 56,000 and 25,000 by SDS-PAGE, and MAb183B2 could blot the 56,000 band by western blot. The antigen from the SKOV3 cell line displayed microheterogeneity appearing as three spots over a pI range 5.3 approximately 5.8 at 56,000. CONCLUSIONS: The epitope of 183B2-antigen is at its peptide core. The antigen belongs to the Ig super family and might be a new ovarian carcinoma associated antigen.
OBJECTIVE: To purify MAb183B2 corresponding antigen of ovarian carcinomas and study its physical and chemical characteristics. METHODS: The strongest positive samples reacted with MAb183B2 were screened from ovarian cancer ascites with indirect enzyme linked immunosorbent assay (ELISA). Then the antigen was purified from ascitic fluid by affinity chromatography with MAb183B2. Nature of the purified antigen was examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and western blot and its reactivity was determined after treatment with NaIO4, trypsin, pronase, neuraminidase, deglycolipid mixture and heating. Two dimensional electrophoresis combined with western blot was then used to isolate and identify the antigen from ovarian cancer cell line SKOV3. RESULTS: The reactivity of the antigen after treated by NaIO4, neuraminidase and deglycolipid mixture was still positive, while it was negative after treated by trypsin, pronase and heating. The antigen proved to be composed of two different subunits of 56,000 and 25,000 by SDS-PAGE, and MAb183B2 could blot the 56,000 band by western blot. The antigen from the SKOV3 cell line displayed microheterogeneity appearing as three spots over a pI range 5.3 approximately 5.8 at 56,000. CONCLUSIONS: The epitope of 183B2-antigen is at its peptide core. The antigen belongs to the Ig super family and might be a new ovarian carcinoma associated antigen.