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There are numerous reports on the studies of the flocculation between tetanus toxin and antitoxins1)2), but only a few of them are successful, because this flocculation is disturbed by many other antigen-antibody reactions unlike the case of diphtheria. Recently, Goldie3) concluded that •gby selecting, diluting, blending and heating certain lots of refined tetanus antitoxin, it is possible to prepare a standard tetanus antitoxin giving only a single and specific flocculation zone•h. Moloney4) reported that it is also possible to obtain a standard antitoxin giving single zone by absorbing the peptic digested tetanus antitoxin with both heat-stable and heat-labile non-toxic antigens. According to Mueller and Miller's reports), if a highly potent toxin is used as antigen, the initial flocculation zone will show true toxin-antitoxin reaction. They also suggested that it would be possible to titrate the antitoxin during the immunization process of horse for antitoxin production. In authours' experience,6) it was also possible to titrate tetanus antitoxins and toxoids with the reference standard antitoxin which was absorbed by Moloney's method4). Though the titration of antitoxins by flocculation was rather difficult, even when highly potent antigen was used, it was possible to obtain good results by selecting suitable antigen. The purpose of this paper is to titrate tetanus antitoxin by direct flocculation method, and to study the relationship between animal test and flocculation test for practical purpose.

STUDIES ON THE TITRATION OF TETANUS ANTITOXINS BY FLOCCULATION II. PREPARATION OF THE MEDIUM TO GET THE TOXIN SUITABLE FOR FLOCCULATION