Purification of colorectal cancer-associated antigen by immuno-affinity chromatography

AIM: To purify and characterize the colorectal cancer-associated antigen from cultured colorectal cancer cells. METHODS: The colorectal cancer cell lines that highly expressed the associated antigen were selected by flow cytometry with five specific monoclonal antibodies (mAbs) CYL1-5. Western blot was used to determine the binding ability of five mAbs to the associated antigens released from colorectal cells lysed with single or triplex-detergent, respectively. The mAb with highest binding ability was employed as the ligand for the immuno-affinity chromatography. The antigens purified through immuno-affinity chromatography were identified by Western blot. RESULTS: The associated antigens were highly expressed on the colorectal cancer cell lines Hce-8693. The binding ability of mAb CYL-2 to the antigen was higher than that of CYL-1, CYL-3-5. The purified associated antigen binding to mAb CYL-2 was a heterodimer composed of two subunits with relative molecular mass (M(r)) of 60 x 10(3) and 70 x 10(3), respectively. CONCLUSION: The purified associated-antigen binding to mAb CYL-2 was obtained from the colorectal cancer cell line Hce-8693 through immuno-affinity chromatography with mAb CYL-2.