Proteins in fluids from the seminiferous tubules and rete testis of rats were studied with a high resolution electrophoretic technique, utilizing a step gradient, polyacrylamide gel system. Both fluids contained a number of specific bands, i.e. proteins not seen in serum or in intratesticular lymph. The rete testis fluid contained more serum proteins than the seminiferous tubule fluid. The albumin—globulin ratio was also greater in the former fluid. Two-dimensional separations showed that the bulk of specific proteins moved as prealbumins. Strong esterase and acid phosphatase activities were located in the region of the specific proteins with the intermediate mobility in acrylamide gel. The origin of these proteins and their significance is discussed in relation to the functional differences between the seminiferous tubules and the rete testis.