Supernatant fluids of mitogen-activated human tonsil lymphocytes contain large amounts of a factor toxic to mouse [fibroblast] L cells. This substance, with a MW of 80,000 .+-. 5000 daltons, is called .alpha.-lymphotoxin (.alpha.-LT), to differentiate it from another toxin elaborated by mitogen-activated human blood lymphocytes, called .beta.-lymphotoxin (.beta.-LT), which differs from .alpha.-LT in size (45,000 .+-. 5000 daltons), antigenicity and stability. Further purification of .alpha.-LT by sequential phosphocellulose and DEAE-cellulose chromatography and polyacrylamide gel electrophoresis (PAGE) identifies a series of cytotoxins differing in ion exchange characteristics and electrophoretic mobilities. The 3 PAGE fractions (PAGE Ia, Ib and II), recovered in 2, 4.6, and 21% yield from the starting serum-free culture supernatant, represent purifications of 24-, 24- and 1851-fold, respectively. Each cytotoxic fraction has a RNase activity. Comparison of RNase and mouse L cell cytotoxic activities of the 3 .alpha.-LT fractions shows that both activities for all 3 fractions have a similar temperature stability pattern and that both are similarly inhibited by DNA (single strand forms better than double strands), by glycerol in 5-20% concentration and by protein denaturing reagents. Mouse L cell toxicity and RNase acitvity are probably mediated by the same substance, which appears to occur in multiple or isozymic forms.