The cidal effect of lysolecithin on mycoplasmas and acholeplasmas was studied. Thirty-one mycoplasma strains and species were tested. The growth of all of them was inhibited by 250 μg/ml of lysolecithin and the growth of most by 125 μg/ml. T-mycoplasmas were as sensitive to lysolecithin as the other mycoplasmas tested. On the other hand, acholeplasmas were about four-fold less sensitive; several strains of the three acholeplasma species grew even in the presence of 500 μg/ml of lysolecithin. Strain “Squire” of Bovine Group 6 was more susceptible to lysolecithin than the other non-sterol requiring strains tested and in this respect resembled the mycoplasmas. Mycoplasmas were found also to be more susceptible to lecithin than acholeplasmas. The influence of the serum and cholesterol content of the medium on the results of the tests is considered and the importance of maintaining standard conditions is stressed. The possibility of using the susceptibility of mycoplasmas and acholeplasmas to lysolecithin of distinguish between them is discussed. The different amounts of cholesterol in the cell membrane of the organisms of these families might account for the differences observed.