Generation and characterization of monoclonal antibodies to p70S6 kinase alpha

Ribosomal kinase S6 (p70S6 kinase) is a key signalling molecule in the init iation of protein synthesis induced by growth factors and hormones. Until n ow, two forms of p70S6 kinase, p70 alpha and p70 beta, have been identified . In this study, the C-terminal region of p70 alpha was expressed in bacter ia as a His-tag fusion protein and purified nearly to homogeneity using aff inity chromatography on Talon Sepharose. These preparations of the recombin ant protein were used to produce monoclonal antibodies against p70 alpha ki nase. Generated antibodies were shown to recognise recombinant and endogeno us p70 alpha kinase in immunoprecipitation and Western blotting. They do no t affect the activity of p70 alpha in in vitro kinase assay and do not cros sreact with p70 beta. Taken together, these antibodies may be of use in stu dies on structure/function analysis of p70S6 kinase alpha.