Open AccessMembrane glycoproteins involved in cell--substratum adhesion.
Author(s) -
Karen A. Knudsen,
Patricia E. Rao,
Caroline H. Damsky,
Clayton A. Buck
Publication year - 1981
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.78.10.6071
Subject(s) - antiserum , glycoprotein , hamster , affinity chromatography , biochemistry , cell adhesion , membrane glycoproteins , adhesion , biology , chemistry , microbiology and biotechnology , cell , antibody , immunology , enzyme , organic chemistry
A combination of immunological and biochemical methods were used to identify surface membrane components involved in cell-substratum adhesion. Broad-spectrum antiserum, prepared against surface membranes from hamster cells, induced reversible rounding and detachment of hamster fibroblasts from a substratum in vitro. This phenomenon was inhibited by Nonidet P-40 extracts of hamster cells. Therefore, an antibody neutralization assay was developed to detect the presence of antigen during the fractionation of Nonidet P-40 extracts of cells. After two differential precipitation steps, anion exchange chromatography, and sequential lectin affinity chromatography, a fraction greatly enriched in ability to block antiserum-induced changes in cell adhesion and appearance was isolated. Analysis of this fraction by NaDodSO4/polyacrylamide gel electrophoresis revealed a highly restricted group of glycoproteins with Mr approximately 140,000. A lectin-purified glycoprotein fraction was used to raise a higher titer antiserum that was able to induce reversible rounding and detachment of cells from a substratum and, when immobilized on an antibody affinity column, was able to bind and release material capable of blocking antiserum-induced cell rounding. These methods have allowed us to focus attention on a restricted group of glycoproteins that are integral constituents of the surface membrane and which play some as yet undetermined role in the process of cell--substratum adhesion.