The addition of an ultrafiltration step to the purification procedure previously described for cholera toxin permitted the preparation of highly purified antigenic toxoids essentially free of somatic antigen(s). The purity of such toxoids is established: (i) by the absence of more than about one part limulus amebocyte lysate (LAL)-positive endotoxin per 105 parts toxoid and (ii) by the inability of the toxoids to elicit a significant rise in rabbit vibriocidal antibody. The antigenicity of the

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... oxoids is demonstrated by their ability to produce the same high levels of rabbit serum antitoxin as are produced by comparable toxoids containing small amounts of somatic antigen. The results also indicate that amounts of somatic antigen on the order of <1 ,ug/1OO jig of toxoid do not exert an adjuvant effect on the toxoid, at least with respect to circulating antitoxin. Other data show that, where present, the ability of somatic antigen to elicit vibriocidal antibody is influenced by the immunization schedule employed and that a correlation exists between the LAL-determined endotoxin content of the toxoids and their ability to stimulate vibriocidal antibody. Somatic antigen-free toxoids, purified and tested by the refinements herein described, were prepared for use in National Institutes of Health-sponsored field trials, and data pertaining to their purity and antigenic properties are presented.