Chemical carcinogen-induced C57BL leukemia EL4 and Gross (G) virus-induced C57BL/6 leukemia Ed♂G2 were fixed with 0.1% formalin at 4° C for 7 days and tested for a) fixation effects and b) preservation of antigenic activity in vitro and in vivo. The formalin concentration was the optimal one in the pretested range of 0.1–10%.1) Trypan blue exclusion in in vitro studies of fixation effects did not correspond with the results of in vivo tests; no incidence of tumor was observed in C57BL/6 mice inoculated with 107 fixed EL4 and E♂G2 cells and in newborn C57BL/6 mice inoculated with 5×106 fixed E♂G2 cells, though many fixed cells excluded trypan blue. 2) In in vitro tests, the amount of H-2b antigen on fixed EL4 and E♂G2 cells, measured by absorption tests, was well preserved up to 28 days' fixation; G surface antigens on fixed E♂G2 cells were preserved up to 7 days. When formalin-treated EL4 or E♂G2 cells were used for immunization in allogeneic or syngeneic systems, antiserum from immunized mice reacted positively with the corresponding viable tumor cells, which demonstrated excellent preservation of H-2b, EL4 tumor-specific, and G surface antigens. The H-2b antigen on EL4 cells and the G surface antigens of E♂2 cells, treated once with 0.1 % formalin and stored at 4° C in Earle's balanced salt solution for 91 and 82 days, respectively, were well preserved, which suggests that the formalin-treated tumor cells may provide a source of stabilized soluble antigens.