The microsporidian Encephalitozoon cuniculi is a spore-forming, obligate, intracellular parasitic pathogen with a unique organelle called a polar tube, the extrusion of which is essential for invading a host cell. The polar tube consists of three proteins: polar tube protein 1 (PTP1), PTP2, and PTP3. We established three mouse monoclonal antibodies (MAb1, MAb2, and MAb4) against E. cuniculi PTP1. An enzyme-linked immunosorbent assay (ELISA) indicated that all three MAbs reacted with the outer surface of extruded polar tubes and that they also strongly bound to an intracellular antigen in infected host cells. Two-dimensional (2-D) immunoblot analysis showed that MAb1 and MAb2 recognized PTP1 spots at 52 kDa and some spotty smears at molecular weights of less than 50 kDa, whereas MAb4 recognized only PTP1 spots at 52 kDa. Interestingly, all three MAbs were of the immunoglobulin (Ig) E class, suggesting that, in addition to the highly immunogenic or antigenic nature, the PTP1 antigen may have the potential to induce specific IgE antibody production in mice. These antibodies may be useful in the study of allergenic PTP1 as well in the purification and detection of the PTP1 antigen.