Extraction, Antisera Preparation And Immunolocalization Of Sperm Membrane Proteins In Eriocheir Sinensis

Eriocheir sinensis is an unique aquatic treasure as well as important economic crab, which extensively exists throughout South and North of freshwater in China. As one of the famous freshwater species for aquaculture, in recent years, its aquaculture in coastal areas is prosperous. It is having great renown for its excellent biological character, the extremely high nutritional value and the unique flavor all the world. The progress of aquaculture further promotes that of fundamental research, or otherwise, fundamental research accelerates aquaculture, too.This paper is about extraction, antisera preparation and immunolocalization of the sperm membrane proteins in E. sinensis. It provides some references for the further researches on the Fertilization Biology and Developmental Biology. Our study is mainly from the following aspects:1. Male reproductive system of E. sinensis was dissected, spermatophore was obtained from seminal vesicle and then matrix of spermatophores as well as sperms was separated from the spermatophores by using glass homogenizer. Next, sperm membrane proteins of the crab were extracted by two detergent solutions and subjected to SDS Polyacrylamide Gel Electrophoresis(SDS-PAGE). Our results indicated that under the function of double-detergent, the proteins we got had significant differences from those of the demembranated sperms. We found that the molecular weight of 12 extracted proteins was less than 100ku, isoelectric point(pI) ranged from 4.4-6.2, and all of them were glycoproteins. Thus demonstrate that sperm membrane proteins of E. sinensis are lower molecular acidic glycoproteins, which is similar with those in mammalian sperms.2. Lactate dehydrogenase(LDH), is one of the membrane proteins, which is also found on sperm membrane, so we speculated that sperm membrane of E. sinensis may contain this protein. By Native-PAGE and specific staining, we researched on the sperm-specific LDH of E. sinensis, the experiments showed that in the presence of two substrates(sodium lactate and α-hyd roxybutyric acid sodium), we analyzed its serum, gill, heart, muscle, hepatopancreas,...