Small pieces, approximately 2 mm³, of a single, spontaneous adenocarcinoma excised from a retired breeder C₃H/HeJ mouse, were implanted in the left side of the axillary fold of 125 thirteen-week-old male and female C₃H/HeJ mice. The mice were then given weekly injections of a coupled tumor protein antigen which was prepared by extracting the antigen from spontaneous mouse tumors and coupling the antigen with human γ-globulin using bis-diazotized benzidine. Careful weekly examinations for 6 weeks after the initial implantation revealed major differences intumor sizes between the treated and untreated mice.

Tumors (adenocarcinomas) excised from mice treated with coupled tumor protein antigen and from mice that were not treated (controls) were prepared according to standard histological techniques, and stained with hematoxylin and eosin. Examination of the different sections revealed that, although the general histological picture of the canalicular and invasive adenocarcinomas remained unchanged before and after treatment, cells from the nontreated tumors were uniform in shape, size and arrangement while cells from the treated tumors were heteromorphic with some cells showing a tendency to become pyknotic.