Macrophage activation as measured by increased rate of carbon clearance and spreading of peritoneal macrophage was studied in mice infected with BCG, strain Japan. BCG caused marked increase of the numbers of peritoneal cells and spread macrophages. The increases of spread macrophages reached a peak at the 3rd week of BCG infection introduced by the both routes of intravenous(i.v.) injection and foot pad(f.p.) injection. BCG also enhanced the clearance of carbon. In the case of BCG given i.v., the increase of the rate of carbon clearance was biphasic : an early increase reaching maximum at the 1st week and a late increase reaching maximum at the 3rd week of BCG infection. When BCG given into one foot pad, peak increase was reached at the 5th week. The activation of macrophages as measured by increased levels of carbon clearance and increased numbers of spread macrophages in the mice receiving BCG i.v. was approximately two fold greater than that in the mice receiving BCG by f.p. route. When sheep red blood cells (SRBC) as antigen were injected i.v. into the mice primed with BCG i.v., the optimal interval between BCG priming and subsequent antigen injection varied with the dose of antigen for the induction of the highest level of delayed type hypersensitivity (DTH) to SRBC, but not with the degree of macrophage activation.