The purpose of the present investigation is to determine the effect in vitro of sulfanilamide, sulfapyridine and sulfathiazole on C. diphtheriae. Observations are also made as to how long it takes for effective action, and on the effect of sulfanilamide, sulfa-pyridine and sulfathiazole on various serological types of Mitis organisms. Nitti, Phillips and Bovet reported that sulfanilamide retards the development of diphtheria bacilli in culture. In the same year, Rosa treated diphtheria carriers by applying the drug locally but failed in this attempt. Methods and Materials. One to 1,000 dilutions of sulfanilamide and sulfapyridine were prepared from a 1% sulfanilamide solution and Soludagenan (1 g/3 cc) respectively, and standardized by means of the Klett-Summerson photo-electric colorimeter. One to 1,000 dilution of sulfathiazole was prepared by heating aqueous solution containing a definite weight of the chemical. Final dilutions of 1:10,000 or 10 mg % were prepared by adding 0.2 cc of stock solution to 1.8 cc of liver digest broth, pH 7.6, containing 5% horse serum. All strains of C. diphtheriae were local strains isolated from the throats or noses of patients or carriers and were virulent to Chinese hamsters. One-tenth cc of 1:1,000 dilution of a 24-hour broth culture was used as the inoculum of the first 2 experiments, and a loopful of undiluted 24-hour cultures used in the third one. Survival of organisms was determined by growth on blood agar plates. Experiment 1. The results of the experiment are summarized in Table I. It represents one of the 3 identical experiments. Under similar conditions, sulfathiazole seems to have exerted a very definite bactericidal action on representative strains of all the 3 cultural types of C. diphtheriae, while the other 2 were less effective on one Mitis strain than on strains of the other cultural types.