Summary: The action of trypsin on the fifth component of human complement (C′5) results in the appearance of a chemotactic factor for rabbit neutrophils. The generation of this activity is dependent upon the amount of C′5, the duration of incubation and to a lesser degree, the amount of trypsin. On the basis of analysis in density gradient ultracentrifugation and by gel filtration, the chemotactic factor appears to be a cleavage product of C′5 with an estimated molecular weight of 8500. Trypsin appears to be less effective in generating a chemotactic factor from the third component of human complement (C′3). Chemotactic activity can be also demonstrated in the supernatant from sensitized sheep erythrocytes containing the first four reacting complement components and incubated with sufficient amounts of C′5. In this case, the chemotactic factor sediments very slowly (like the marker cytochrome c) in a sucrose density gradient. The relationship between the chemotactically active C′5 fragment and F(a) C′5 which has anaphylatoxin activity is not presently known. These studies document the second fragmentation product of low molecular weight from the complement system possessing chemotactic activity.