The detection of the cellular immune response against bacteriophage phi-X174 by an adaptation of the haemolytic plaque technique.

In view of the extensive use of bacteriophage as an antigen, ϕX174 was bound to sheep erythrocytes by means of bis-diazotized-benzidine. Phagecoated erythrocytes were used for the detection, by the Jerne technique, of rabbit spleen cells producing antibodies against phage. It was shown that intravenous injection of purified ϕX174 causes the appearance of spleen cells producing specific antibodies. The magnitude of the plaque-forming response was similar to that obtained when the antigen used is the sheep erythrocyte, though in some situations, as in the secondary response, one obtains higher numbers of plaque forming cells. Parallel experiments performed with sera treated with 2-mercaptoethanol or analysed by sucrose gradient ultracentrifugation demonstrated that passive haemolytic activity is present in both 19S and 7S classes of phage-neutralizing antibodies. Treatment of plates with 2-mercaptoethanol resulted in a decrease in the number of plaques showing that most plaque-forming cells are 19S producers.