1. The presence of a natural anti-human hemolysin, only active against Group I and Group II cells, was demonstrated in over 50 guinea pigs, obtained from different sections of the surrounding country. It was shown that, in a majority of the sera tested, this hemolysin would be a contributing factor to an excess of amboceptor in the reaction, in the quantities of complement used in an antihuman complement fixation reaction. The hemolysin is specific, in as far as it can be absorbed with either Group I or Group Il cells, but cannot be absorbed with Group III or Group IV cells.
2. To determine the effect of the isohemolysins in human serum, twenty sera were tested against cell suspensions of the four human groups. Tests were also run with twenty-six human sera against all four group cell suspensions, with complement that had the natural anti-human hemolysin absorbed from it. In both experiments various degrees of hemolysis were obtained with all but two sera against cell suspensions from Groups I, II, and III, but no hemolysis was obtained with any serum against a Group IV cell suspension.
3. The importance of the human isohemolysins and the natural anti-human hemolysm in complement was demonstrated by performing complement fixation tests upon twenty-six human sera, cell suspensions of all four human groups for each serum tested being used. A marked variation in the reaction occurred with a large percentage of the sera tested. This variation in a single serum might be as great as a negative reading with Group I cells to a double plus reading with Group IV cells.
4. That the isohemolysins, alone, in human serum are of importance in the reaction was shown by running complement fixation tests upon seventeen human sera with the four group cell suspensions and complement from which the natural anti-human hemolysin had been absorbed. Results showed a much weaker reaction, in a large number of the sera, with Group I, Group II, and Group III cells than with Group IV cells.
5. Experiments to determine a relation between the human blood groups and natural anti-sheep hemolysin present in human serum gave negative results. It was shown, however, with twenty-six human sera tested against twenty-three different sheep cell suspensions that there is a wide variation in the activity of the natural anti-sheep amboceptor present in any human serum when it is tested against different sheep cell suspensions. A human serum capable of completely hemolyzing the cells of one sheep may fail to give any hemolysis whatever when tested against another sheep cell suspension.
6. Tests were made for natural anti-sheep hemolysin in guinea pig serum. The serum from sixteen different guinea pigs was tested against a total of twenty-one sheep cell suspensions. A natural anti-sheep hemolysin was present in all the guinea pigs tested against one of the sheep cell suspensions. The same guinea pigs tested against the other sheep cell suspensions failed to show any hemolysis.