Aim： To extend the analysis of the proteome of human spermatozoa and establish a 2-D gel electrophoresis （2-DE） reference map of human spermatozoal proteins in a pH range of 3.5-9.0. Methods： In order to reveal mo...Aim： To extend the analysis of the proteome of human spermatozoa and establish a 2-D gel electrophoresis （2-DE） reference map of human spermatozoal proteins in a pH range of 3.5-9.0. Methods： In order to reveal more protein spots, immobilized pH gradient strips （24 cm） of broad range of pH 3-10 and the narrower range of pH 6-9, as well as different overlapping narrow range pH immobilized pH gradient （IPG） strips, including 3.5-4.5, 4.0-5.0, 4.5-5.5, 5.0-6.0 and 5.5-6.7, were used. After 2-DE, several visually identical spots between the different pH range 2-D gel pairs were cut from the gels and confirmed by mass spectrometry and used as landmarks for computer analysis. Results： The 2-D reference map with pH value from 3.5 to 9.0 was synthesized by using the ImageMaster analysis software. The overlapping spots were excluded, so that every spot was counted only once. A total of 3 872 different protein spots were identified from the reference map, an approximately 3-fold increase compared to the broad range pH 3-10 IPG strip （1 306 spots）. Conclusion： The present 2-D pattern is a high resolution 2-D reference map for human fertile spermatozoal protein spots. A comprehensive knowledge of the protein composition of human spermatozoa is very meaningful in studying dysregulation of male fertility.展开更多
基金This work was supported by two grants from the National Natural Science Foundation of China （No. 30170480 and No. 30470884）. We would like to thank Professor Guang-Yin Lu for enthusiastic reading and amending of the manuscript. We would like to thank Mr Ji-Xian Xiong, College of Life Science at the Hunan Normal University for enthusiastic support of technology and theory of MS. We are also very grateful to collaborators in our laboratories for help and valuable discussions and suggestions during the course of this work.
文摘Aim： To extend the analysis of the proteome of human spermatozoa and establish a 2-D gel electrophoresis （2-DE） reference map of human spermatozoal proteins in a pH range of 3.5-9.0. Methods： In order to reveal more protein spots, immobilized pH gradient strips （24 cm） of broad range of pH 3-10 and the narrower range of pH 6-9, as well as different overlapping narrow range pH immobilized pH gradient （IPG） strips, including 3.5-4.5, 4.0-5.0, 4.5-5.5, 5.0-6.0 and 5.5-6.7, were used. After 2-DE, several visually identical spots between the different pH range 2-D gel pairs were cut from the gels and confirmed by mass spectrometry and used as landmarks for computer analysis. Results： The 2-D reference map with pH value from 3.5 to 9.0 was synthesized by using the ImageMaster analysis software. The overlapping spots were excluded, so that every spot was counted only once. A total of 3 872 different protein spots were identified from the reference map, an approximately 3-fold increase compared to the broad range pH 3-10 IPG strip （1 306 spots）. Conclusion： The present 2-D pattern is a high resolution 2-D reference map for human fertile spermatozoal protein spots. A comprehensive knowledge of the protein composition of human spermatozoa is very meaningful in studying dysregulation of male fertility.
