It is well-established that three types of erythrogenic toxins (ETA, ETB, ETC) are produced by Streptococcus pyogenes (group A streptococci) strains. Culture filtrate concentrates from Streptococcus pyogenes strains T19P (T19, ETA+, ETB+, ETC-), 27337 (T12, B3264, ETA-, ETB+, ETC+), 27252 (T4, ETA-, ETB+, ETC+) and 27195 (T8, ETA-, ETB+, ETC-) were analyzed by preparative isoelectric focusing. These concentrates and the purified erythrogenic toxin type B (ETB) isolated by ion exchange chromatography had mitogenic and pyrogenic activity. Now, it has been found that the mitogenic activity and the pyrogenic activity of this ETB can be separated by preparative isoelectric focusing in Sephadex gels. This means that ETB is not a superantigen as described in literature. The mitogenic and biological activity is caused by traces of ETA (strain T19P), ETC (strains 27252 and 27337) and/or by unknown mitogen(s) (MX, strain 27195) which preferentially stimulate V beta 8+ T cells. The differentiation between ETA (stimulating V beta 12+ but not V beta 8+ or V beta 2+), ETC (stimulating V beta 2+ but not V beta 8+), and MX (stimulating V beta 8+) was done using established leukemic cell lines.