Varying degrees of naturally occurring immunoglobulin (Ig) M antibodies to Toxoplasma gondii were observed in almost all of adult Japanese populations negative for IgG antibodies as determined by an enzyme-linked immunosorbent assay (ELISA). The antibody activity was reduced by absorption of sera with Toxoplasma soluble antigen or by 2-mercapto-ethanol treatment of the IgM fraction after sucrose density gradient centrifugation. Umbilical cord sera had almost no detectable IgM antibodies, and the ELISA value in infants increased with age until 1 year. Most paired sera collected from adults at a 1-year interval showed constant IgM antibody levels, suggesting retention of natural IgM antibodies throughout their lives without changing ELISA values of ≥ 0·1/year. Polar staining was observed in immunofluorescent studies for 14 serum samples with natural IgM antibody levels of ≥ 0·5, while samples obtained from a pregnant woman with natural IgM antibody levels of ≥ 0·9 reacted to the whole organism as was observed in positive sera of acute patients. Western blotting analysis revealed that natural IgM antibodies were directed to multiple antigen molecules. The band pattern differed slightly from one serum to another, but almost consistent patterns were observed in sera sequentially obtained from an adult over 4 years.