Isolation of the Second Component of Complement (C2) from Carp Serum

The second component (C2) of carp complement was isolated from carp serum by a six-step purification procedure: 1) affinity chromatography on Blue-Cellulofine, 2) affinity chromatography on heparin-agarose, 3) gel filtration on Sephadex G-200, 4) TSKgel DEAE-5PW chromatography (HPLC), 5) TSKgel Heparin-5PW chromatography (HPLC), and 6) TSKgel G3O0OSW_XL chromatography (HPLC). Hemolytic activity of carp C2 in column eluates was measured using EAC14 (sheep erythrocytes sensitized with carp antibody, Cl and C4) and heat-inactivated carp serum (deficient in Cl and C2) as reagents for detecting C2. About 0.4 mg of carp C2 was obtained from 100 ml of serum. The final recovery was 9.4%, representing a 799-fold purification. Carp C2 was composed of a single polypeptide chain with molecular weight of 108,000 and was susceptible to heat-inactivation at 45°C. These physicochemical properties of carp C2 closely resemble those reported for mammalian C2.