Methods for assaying the alternative complement pathway activity (ACH50) of rainbow trout Salmo gairdneri, ayu Plecoglossus altivelis, carp Cyprinus carpio, tilapia Tilapia nilotica, eel Anguilla japonica, yellowtail Seriola quinqueradiata, porgy Pagrus major and flounder Paralichthys olivaceus sera were investigated.
The ACH50 values of the sera of all fish except eel could be measured by incubating each serum with rabbit red blood cells (target cells) at 20°C in the presence of 10m_M EGTA and 10m_M MgCl₂. The pH values and incubation periods employed were: rainbow trout, pH 7.0, 1.5h; ayu, pH 7.0, 1.5h; carp, pH 7.5, 1.5h; tilapia, pH 7.0, 1.5; yellowtail, pH 7.0, 2.5h; porgy, pH 7.5, 2.0h; flounder, pH 7.5, 2.0h. The hemolytic activity of eel serum was not reduced even in the presence of 10m_M EDTA, suggesting the participation of unknown factor(s) in the hemolysis. Therefore, titration of the ACH50 value of eel serum was not carried out.
ACH50 values (unit/ml, mean±SD, n=3 to 24) of the above 7 species were as follows: rainbow trout, 456±88; ayu, 880±376; carp, 51±14; tilapia, 684±560; yellowtail, 1016±56; porgy, 328±188; flounder, 508±60. These values are 5 to 60 times higher than those reported for mammals, suggesting that the role of the alternative pathway is much more important in fish than in mammals.