The lytic and bactericidal actions of polymyxin B on whole cells and spheroplasts of Pseudomonas aeruginosa varied markedly with the suspending media, and there was little correlation between them. Relative rates of lysis of these preparations and also of Bacillus megaterium protoplasts suggested that polymyxin causes progressive damage to the cytoplasmic membrane, such that membrane permeability towards various ions increased as follows: K + > Na + > NO 3 − > Cl − , Ca 2+ , H 2 PO 4 − /HPO 4 2− . Impermeant compounds, such as NaCl and sucrose, protected whole cells against lysis but not against death. It is suggested that lysis of whole cells by polymyxin is a secondary effect, resulting from entry of solutes normally excluded by the cytoplasmic membrane and the fragility of the damaged outer membrane. Because the degree of lysis varies with the external solutes, it should be treated with caution as a descriptor of polymyxin activity.