Identification of Secreted and Membrane-Bound Forms of 2D6 Antigens in Luminal Plasma and Spermatozoa from the Caput and Cauda Epididymidis
Proteins in epididymal plasma and detergent extracts of spermatozoa collected by micropuncture were analyzed under nonreducing and reducing conditions (Fig. 3). Under nonreducing conditions, 2D6 McAb recognized a high-molecular-mass protein of ~250 kDa in caput plasma. In cauda plasma a strong reaction was obtained at ~200 kDa, and weaker reactions were obtained with two lower-molecular-mass components at 160 and 48 kDa. In detergent extracts of caput spermatozoa a very weak signal was present only at —250 kDa. This protein, and two additional ones at 48 and 24 kDa, was present in extracts of cauda spermatozoa.
Under reducing conditions a significantly different protein pattern was obtained to that described above. In caput plasma 2D6 McAb recognized proteins at —200 kDa together with two additional components at approximately 60 and 70 kDa. In extracts of caput sperm a signal was obtained only over a protein with a molecular mass of —65 kDa. In contrast, 2D6 McAb bound strongly to several proteins in cauda plasma and cauda sperm extracts. In cauda plasma, proteins of 32, 28, 24, and 20 kDa gave the strongest reaction, whereas in the sperm extracts only the 24-kDa component bound 2D6 McAb. Click Here
FIG. 3. Western blot probed with 2D6 McAb of proteins in luminal plasma and detergent extracts of spermatozoa collected from distal caput and cauda epididymidis. Proteins were separated by SDS-PAGE under reducing and nonreducing conditions as indicated.