An antibiotic was obtained from an unclassified Actinomyces, and by bacterial spectrum it showed a marked similarity to purified streptomycin. Two strains of Staphylococcus aureus (A.T.C.C. 6538 and F.D.A. 209) which were highly resistant to streptomycin were found to be as sensitive to the unknown antibiotic as the original strains. The antibiotic was thus different from streptomycin in spite of their spectral similarities. A strain of S. aureus 6538 was sensitive to penicillin F, G, and X in concs, of 0.063, 0.078 and 0.039 [gamma]/ml., respectively, while a resistant strain required 0.625, 0.625, and 1.25 [gamma]/ml. A strain with acquired resistance against one penicillin is also resistant to the same degree to the other 2 penicillins. Antibiotics can be differentiated by this method, but since 3 penicillins cannot be differentiated by a penicillin G-resistant strain of S. aureus, this method is limited.