The bovine corneal SGP-complex is related to the tissue form of type VI collagen

A polyclonal antiserum was prepared in rabbits against the structural glycoprotein (SGP) complex previously isolated from a bacterial collagenase digest of bovine corneal stroma (R. Alper, Curr. Eye Res. 2:479, 1983). Direct and indirect enzyme-linked immunosorbent assays indicated that the antiserum was specific for the SGP-complex and did not react with Types I, III and IV collagen, fibronectin, laminin or actin. Immunoblot experiments indicated that the antiserum reacted with all of the components of the SGP-complex as well as with the cell matrix laid down by bovine keratocytes in culture. An attempt was made to isolate individual antibodies from the antiserum by selective elution from immunoblots of the components of the SGP-complex separated by SDS-PAGE. It was found that regardless of the protein band from which the antibody was eluted, every antibody isolated reacted with every protein component of the SGP-complex suggesting that the SGP-complex may have been derived from a single precursor protein and that the observed heterogeneity of the SGP-complex may have been the result of proteolytic breakdown of the protein held together by disulfide bonds. When the anti-SGP antiserum was used to immunoprecipitate 14C-proline labeled proteins from the media of bovine keratocytes in culture, the major protein observed had a Mr of about 140,000 daltons, similar to that of GP-140 also known as CL-glycoprotein. These proteins have been shown to represent the tissue form of Type VI collagen. To test the hypothesis that the SGP-complex may be related to the GP-140 (CL-glycoprotein), ELISA and immunoblotting studies were performed comparing the properties of the anti-SGP serum with those of a polyclonal antibody specific for Type VI collagen. The SGP-complex reacted positively by ELISA with the anti-human Type VI collagen antiserum and, conversely, human Type VI collagen gave a positive ELISA reaction with an antiserum against the SGP-complex. The anti-human Type VI collagen antiserum reacted with most of the major components of the SGP-complex on immunoblots of SDS-PAGE gels. These data indicate that the SGP-complex is related to and probably is derived from the tissue form of Type VI collagen.