Two-dimensional Electrophoresis Reseach Of Normal Human Seminal Plasma Protein In Liquify Process

Introduction:Established the method of prepare human seminal plasmaprotein for 2-DE and the 2-DE map of board range pH 3-10 and thenarrower range pH 4-7 immobilized gradient strip (IPG, 24 cm) in ourinstitute。Separate the protein of these samples by 2-DE and analyse the2-DE map to discovery the change in the liquid process, and find theregulation of protein transformation in this process. Intend to find thefunction of the protein in seminal plasma at the area of malereproduction.Material:Choose 13 healthy and fertile donors randomly and get one semensample from each of the donor, the semen samples bymasturbation after 4-6 days abstinence, these donors were testified thatthey all have the normal fertility by the AID. All samples were obtainedunder informed consent using forms approved by the National HumanReproduction Ethic Investigation Committee. The microbe examinationof the semen samples of these donors before and after the donation isnegative.Get the semen samples and liquefied 0min, 10min, 30min, 60min and theexactly liquefied time in microscope. Methods:1.Compare the efficiency of the protein solubilization between theTCA/ acetone precipitate and the desalt/chaotrope and choose theefficient protein solubilization method.2.Use the 2-DE to separate protein spots and use the differentialproteomics method to analyse the differential between each liquefy time.3.Compare the 2-DE map of seminal plasma protein from the exactlyliquified time in microscope and the 2-DE map of each different liquifiedtime.Results:1.Compare the efficiency of the protein solubilization between theTCA/ acetone precipitate and the desalt/chaotrope and determine theTCA/acetone precipitateis more efficiency.2.Established the human seminal plasma protein 2-DE map of thenarrower range pH 4-7 immobilized gradient strip (IPG; 24 cm) and boardrange pH 3-10。Get the 5 different protein 2-DE map at different liquefy time of humanseminal plasma.3.Analyse the seminal plasma protein 2-DE maps of different liquefiedtime and use Image master^tm 5.0 obtain 36 differential protein spots;Compare the 2-DE map of seminal plasma protein from the exactlyliquified time in microscope and the 2-DE map of each different liquified time and find no significant deviation.4.23 protein spots wera identify by Mass Spectrometry, find 14 proteinspots degraded and 4 proteins spots amount is increase in seminal plasmaliquefy transformation process.Conclusions:In our research, we established the normal human seminal plasma protein2-DE map for each time in liquefied process.Differential proteomeanaslysis was perform on the base of 2-DE map and ImageMaster^TMsoftware,about 36 differential protein spots was found and find 14 proteinfragments degraded and 4 proteins fragments amount is increased inseminal plasma liquefy transformation process.