First observations on enzymatic activity and protein content of vesicles separated from rat epididymal fluid.

Fluid of rat cauda epididymidis was obtained by flushing the duct with 0.25 mol l-1 sucrose in 0.01 mol l-1 Tris-HCl buffer pH 7.4. The fluid was centrifuged at 600 x g for 15 min and the sperm free supernatant was centrifuged at 47,000 x g for 1 h. The sediments observed with the electron microscope consisted of a heterogeneous population of membrane-bound vesicles similar to those seen in the intact organ. In the sediment containing the vesicles the activity of beta-galactosidase was mostly unavailable for the substrate showing a high degree of latency: the activity became soluble after a treatment with 0.5% saponin. The activity of N-acetyl-galactosaminadase instead, was mainly available for the substrate and soluble in buffer containing 0.6 mol l-1 KCl. It was then inferred that beta-galactosidase is located inside vesicles with no or little affinity for the membrane, while N-acetylglucosaminadase is bound to the external surface of vesicles. Supernatants and precipitates from suspensions of vesicles in buffered 0.5% saponin were analysed for proteins by gel electrophoresis. The electrophoretic patterns of the sediments were very different from those of supernatants and showed a number of bands greater than that of the latter. The vesicles are believed to arise from the epididymal epithelium, but their physiological role is unknown.