Epididymis-specific Protein Identification

In order to identify the specific changed protein in epididymis under the pathological status, the rat model of Experimental Left Varicocele (ELV) which simulates the varicocele of human was used, and the variety of epididymal specific protein was investigated by proteomics technology including Two-dimensional electrophoresis and Mass Spectrometry. The varied protein in different segment/area of natural epididymis was also identified and characterized synchronously.The object of the study is to search the markers possibly for use in the diagnosis and therapy in male infertility. On the other hand, the study on mechanism of epididymal specific proteins in sperm maturation may provide basis for establishing new method on male fertility regulation.First of all, the efficacy of the ELV in rat model was evaluated and validated. The comparative analysis between both rat strains of SD and Wistar after operation indicated that the former was better than the latter by observation of the altered diameter of internal spermatic vein.The SDS-PAGE analysis indicated that the content of a 22kDa protein was markedly increased in the supernatant of homogenized epididymis from ELV rat than that of from sham-operated rat. The result also shows that the content of 22kDa protein was varied in caput and cauda epididymidis area of rat.The classical route for identification of an unknown protein was taken in this study, in which the 22kDa protein was purified from PAGE gel, and a rabbit anti-22kDa protein antibody was raised. The specifity of the antibody was identified, and it supplied an effective tool for further study.The supernatant of homogenized epididymis was analyzed by two-dimensional gel electrophoresis and protein spots of interest were cut and digested with trypsin for peptide mass spectrometric analysis using matrix-assisted laser desorption ionization time-of flight technique (MALDI-TOF). The Peptide fingerprints were then analyzed using a peptide mass computer database (www.expasy.ch/tools) and Mascot software.