The alternative complement pathway in chickens. Purification of factor B and production of a monospecific antibody against it.

Abstract

Restoring the haemolytic activity of heat-treated chicken serum was used to monitor an initial purification of chicken complement component factor B of the alternative complement pathway. The partially purified factor B was used to prepare a monospecific rabbit antiserum against factor B, and this antiserum was now used to monitor and establish a purification scheme for chicken factor B which could be used for large-scale purifications. The final purification involved precipitation with polyethyleneglycol, ion exchange chromatography on CM 52-cellulose, gel-filtration on Sephadex G 200, elution from a Con A-Sepharose column, and finally ion exchange chromatography on a DE 52-cellulose column. 2.8 mg of factor B was purified from 100 ml of chicken plasma with a yield of 28%. Purity was high as judged from crossed immunoelectrophoresis, SDS-PAGE, and from immunizations of rabbits with the factor B preparation. The purification gave preliminary indications that chicken factor B is a glycoprotein with beta-mobility and with a molecular weight around 95 Kd.