Not only does mouse complement (C) have low hemolytic activity, but mouse serum has an inhibiting effect on hemolysis by human C. To purify and identify the putative mouse serum factor inhibiting human C activity, a sequential procedure of fractionated precipitation by PEG, followed by chromatographies with a heparin-Sepharose column, a phenyl-Sepharose column, a Protein G column, and a gel-filtration column was performed. The amino acid sequence analyses of two polypeptides obtained by digestion of the purified serum factor with TPCK-trypsin revealed that it was mouse fibronectin (FN). Highly purified mouse FN, but not human FN, has an inhibiting effect on human C-dependent hemolysis. Moreover, the hemolysis of sensitized rabbit erythrocytes by mouse C was also inhibited by the addition of mouse FN in a dose-dependent fashion, but not by the addition of human FN. These results suggest that FN is the putative internal C inhibitor in the mouse system. (C) 1999 Elsevier Science B.V. All rights reserved.