The Donath-Landsteiner reaction was studied using low and high titer antisera and purified antibody, normal and PNH erythrocytes, and human serum complement.

The requirement for complement in both the cold and warm phases of the reaction depended upon the level of antibody used and the sensitivity of the cells to hemolytic antibodies. Complement was not necessary in the cold phase using PNH cells and a potent source of antibody, but complement was required to be present at some stage if hemolysis were to occur.

Optimal conditions for the cold phase were at 1°C. for 30 minutes at pH 7.4. Ca++ ions were required. Hemolysis in the warm phase occurred within one minute, was optimal at 32°C., and required Mg++.

The relation of these observations to previous reports is discussed with respect to discrepant observations on the nature of the Donath-Landsteiner reaction.

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