A crystalline substance capable of suppressing the growth of a variety of mycobacteria in vitro has been isolated from extracts of tissue in acidified dilute ethanol. This inhibitory material was found to be equally active against virulent, attenuated, and avirulent variants of human and bovine tubercle bacilli, but had little or no effect on saprophytic mycobacteria and on several non-acid-fast microorganisms under the conditions of the test. Its inhibitory activity on the growth of tubercle bacilli was essentially independent of the size of the inoculum within the limits studied. The crystalline material appeared to exert a bactericidal action on the susceptible organisms. Tubercle bacilli maintained in the presence of the agent for 4 days failed to grow when transferred to inhibitor-free media. The findings were not appreciably altered by minor variations in the composition of the medium or by shift in its reaction. When certain preparations of whole serum were used in the medium in place of albumin, no antimycobacterial activity was observed; however, this activity was restored by adding bovine albumin (fraction V) to the media containing whole serum. By chemical purification and analysis, the inhibitory material was identified as spermine, an organic base widely distributed in animal tissues.