A simple defined basal medium is presented for the study of proteolytic activity, induction and repression, and protease purification with Serratia marcescens. Since the medium contains no protein, it does not interfere with or present artifact to protein assays, column chromatography, or electrophoresis. The medium consists of the basal salts and buffer medium of Bromke and Hammel (1979) plus a carbon-energy source such as glycerol, calcium chloride for the cation requirement for protease activity, and an amino acid, preferably leucine. Growth parameters and proteolytic activities are presented for unsupplemented medium and for the medium supplemented with each of 18 amino acids. Unsupplemented medium completes the logarithmic phase in 12.5 h of incubation and has a constitutive level of proteolytic activity. Supplementation with any amino acid, except cysteine and tryptophan, increases significantly the proteolytic activity, but has a varied effect on growth parameters.