Encephalitozoonosis is a common infectious disease caused by Encephalitozoon cuniculi in rabbits. The purpose of this study was to examine the levels of antibody in urines of experimentally E. cuniculi-infected rabbits. Humoral immune response and spore excretion in urines were also examined. Rabbits were orally infected with E. cuniculi, and sera and urines were collected. Levels of serum IgG, IgM, IgA and urinary IgG were examined. The presences of spores in urines were examined by quantitative PCR. Specific IgG in sera started to be detected at day 20 after infection and kept high level during the course of experiment. Serum IgM started to be detected 1 week earlier than that of serum IgG and decreased to almost undetectable level by day 100 after infection. However, the level of IgM increased gradually again thereafter. Serum IgA was transiently detected between 20 and 70 days after infection and kept undetectable level thereafter. Specific IgG in urines started to be detected from 60 days after infection. The amount of spore DNA was correlated with the amount of IgG in urine. In conclusion, the detections of serum and urinary antibodies were useful for the prediction of stage of latent infection in encephalitozoonosis.