Updated and expanded in vivo quantitative testing procedures to determine the efficacy of topical antimicrobial agents are presented. The occlusion test measures the ability of an agent to prevent the expansion of the resident microflora which occurs when an impermeable dressing is applied to the forearm. Measurements are made at 24 and 48 hr. The expanded flora test measures the ability of an agent to suppress a dense population of micro-organisms produced by expansion of the resident flora of the forearm by prior application of an impermeable occlusive dressing. Measurements are made at 6, 24 and 48 hr or after 10 min in the case for agents designed for immediate degerming. The persistence test measures the ability of an agent to establish a reservoir in skin and exert an antimicrobial effect up to 3 days after the last application of the test material. The ecological shift test determines any major alteration in cutaneous microbial ecology following several applications of the material under occlusive dressings. The serum inactivation test determines whether the presence of serum proteins interferes with antimicrobial activity.