Detection of Glu-Glu-tagged proteins in mammalian cell culture media by dot immunoblotting

A dot immunoblotting technique has been developed to estimate the relative expression levels of tagged recombinant human proteins in mammalian cell cu lture media. Variations in sample denaturation, blocking agents and membran e composition and treatment were used to optimize the signal-to-noise ratio of the defined procedure. The method is rapid, with sensitivity extending to the low nanomolar range for a number of recombinant proteins. This techn ique should have general utility for antibody-based measurements of other t agged and non-tagged proteins in cell culture media or in biological fluids .