A sensitive enzyme-linked immunosorbence assay for the c-fos and v-pos oncoproteins

Abstract

The c-fos nuclear oncoprotein is rapidly induced when the growth of normal cells is initiated by mitogens, and it is also synthesised in several cell systems in response to stimuli that do not cause cell proliferation. When expressed inappropriately, c-fos, and its retroviral counterpart v-pos, can transform susceptible cells in vivo and in vitro. We have developed a simple and sensitive ELISA for the c-fos and v-pos proteins. Fos proteins are captured from cell lysates by an antibody specific for an amino-terminal peptide substantially conserved between v-pos and c-pos; the captured proteins are recognised by a second antibody against a different peptide sequence also conserved in the two proteins. The second antibody has been conjugated to alkaline phosphatase to provide an enzyme label; bound alkaline phosphatase is measured with a sensitive cycling enzyme system that generates a coloured end-product. We show that the fos ELISA is immunologically specific and use it to monitor increased c-fos expression in serum-stimulated HeLa cells and human fibroblasts, and in mitogen-stimulated murine thymocytes.