Globozoospermia is a severe form of teratozoospermia characterized by round-headed spermatozoa with an absent acrosome, an aberrant nuclear membrane and midpiece defects. Globozoospermia is diagnosed by the presence o...Globozoospermia is a severe form of teratozoospermia characterized by round-headed spermatozoa with an absent acrosome, an aberrant nuclear membrane and midpiece defects. Globozoospermia is diagnosed by the presence of 100% round-headed spermatozoa on semen analysis, and patients with this condition are absolutely infertile. The objective of this study was to investigate the differences in protein expression between human round- headed and normal spermatozoa. Two-dimensional （2-D） fluorescence difference gel electrophoresis （DIGE） coupled with mass spectrometry （MS） was used in this study. Over 61 protein spots were analysed in each paired normal/round-headed comparison, using DIGE technology along with an internal standard. In total, 35 protein spots identified by tandem mass spectrometry （MS/MS） exhibited significant changes （paired t-test, P 〈 0.05） in the expression level between normal and round-headed spermatozoa. A total of nine proteins were found to be upregulated and 26 proteins were found to be downregulated in round-headed spermatozoa compared with normal spermatozoa. The differentially expressed proteins that we identified may have important roles in a variety of cellular processes and structures, including spermatogenesis, cell skeleton, metabolism and spermatozoa motility.展开更多
Since preparation and solubilization of protein samples are crucial factors in proteome research,the authors established a sequential extraction technique to prepare protein samples from the body wall of the 5th insta...Since preparation and solubilization of protein samples are crucial factors in proteome research,the authors established a sequential extraction technique to prepare protein samples from the body wall of the 5th instar larvae of silkworm.Bombyx mori.Two kinds of protein samples were obtained from the body wall using the method.Between the two types of samples only about 15% proteins were identical;the majority were different,indicating that more species of proteins could be obtained with the sequential extraction method;which will be useful for preparation of protein samples for proteome study.展开更多
Objective To identify the sperm membrane proteins that are associated with antisperm antibody Methods Using antisperm antibody positive serum through unidimensional polyacrylamide gel electrophoresis and 2-dimensi...Objective To identify the sperm membrane proteins that are associated with antisperm antibody Methods Using antisperm antibody positive serum through unidimensional polyacrylamide gel electrophoresis and 2-dimensional gel electrophoresis followed by Western blot analysis to determine the molecular weights (MW) and isoelectric points (pI) of sperm membrane proteins that are associated with antisperm antibody. Results Eight kinds of MW with more than ten sperm membrane proteins can be recognized by antisperm antibody positive serum, of which the MWs and pI were 23 kD, 31 kD, 32 kD, 34 kD, 41 kD, 51 kD, 60 kD, 78 kD and 5.3, 5.5,5.7, 5.0, 5.3, 5.8, 6.0, 5.5～6.2, 4.6,5.1,5.5～5.8 respectively. The identification ratios of the sperm membrane proteins on 78 kD (60.7%), 60 kD (71.4%), 51 kD (14.9%) and 23 kD (14.29%) were higher. Conclusion The sperm membrane proteins with MW of 78 kD, 60 kD, 51 kD and 23 kD were associated with antisperm antibody and immunological infertility. Two- dimensional gel electrophoresis and Western blotting can precisely identify the sperm membrane proteins that are associated with antisperm antibody.展开更多
An improved CBB staining with higher sensitivity than that of the typical CBB staining was reported.The main improvement was using a fixing step of 25% trichloroacetic acid(TCA) before CBB staining.For most proteins s...An improved CBB staining with higher sensitivity than that of the typical CBB staining was reported.The main improvement was using a fixing step of 25% trichloroacetic acid(TCA) before CBB staining.For most proteins studied,the sensitivity of the improved CBB staining was about twice as high as that of the typical method.For basic and low molecular weight proteins such as ribosomal proteins,the sensitivity of this improved staining method was about 3.5-28 times that of the typical method.It was speculated that the improved procedure would be suitable for exact quantitative analysis of proteins fractionated by SDS-PAGE,especially for basic and low molecular weight proteins.On the other hand,this new modified method might be also applied to multidisciplinary studies,such as biological researches and nuclear sciences.展开更多
Regardless of thermodynamic stability, less complex proteome than plasma, convenient and noninvasive sampling of urine, reproducibility and comparability of the data among laboratories are challenges for the urine pro...Regardless of thermodynamic stability, less complex proteome than plasma, convenient and noninvasive sampling of urine, reproducibility and comparability of the data among laboratories are challenges for the urine proteome analysis. Pretreatment of urine with 10 mM EDTA in conjunction with desalting methods to remove interfering molecules significantly improved the presentation of urine proteome on 2-D gels. Statistical analysis of the average ratio of the spot numbers on 2-D gels between EDTA-treated and non-treated samples were 1.35 ± 0.167, 1.26 ± 0.091, and 1.24 ± 0.095 for dialysis, ultrafiltration, and desalting column, respectively, with the p-value of 5.85 × 10–6 for the overall comparison. This result came from the fact that EDTA-treated samples showed better resolutions with less streaks and clearer spots on 2-D gels than the control samples. The results suggest that a simple and non-fractionating EDTA-treatment can be employed to the procedures for the high resolution 2-D gel analysis and other urinary proteomic analysis.展开更多
Proteomics is one of the most active research fields in the post-genomic era. Here we briefly introduce the scientific background of the origination of proteomics and its content, research method. The new developments...Proteomics is one of the most active research fields in the post-genomic era. Here we briefly introduce the scientific background of the origination of proteomics and its content, research method. The new developments of proteomics at the levels of individual plants, tissues, organs and organells, as well as its applications in the area of plant genetic diversity, mutant characterization, and plant physiology, etc are reviewed. At last, the challenge and prospect of proteomics are discussed.展开更多
基金Acknowledgment We thank Beijing Proteome Research Center, （Beijing, China） for its enthusiastic technological support and for the theory of 2-D DIGE. We also thank（Changsha, China） College of Life Sciences at Hunan Normal University for supporting the MS technology. Finally, we are very grateful to our collaborators for their help, as well as their valuable discussions and suggestions during the course of this work. This work was supported by two grants from the National Natural Science Foundation of China （NO. 30170480 and NO. 30470884）.
文摘Globozoospermia is a severe form of teratozoospermia characterized by round-headed spermatozoa with an absent acrosome, an aberrant nuclear membrane and midpiece defects. Globozoospermia is diagnosed by the presence of 100% round-headed spermatozoa on semen analysis, and patients with this condition are absolutely infertile. The objective of this study was to investigate the differences in protein expression between human round- headed and normal spermatozoa. Two-dimensional （2-D） fluorescence difference gel electrophoresis （DIGE） coupled with mass spectrometry （MS） was used in this study. Over 61 protein spots were analysed in each paired normal/round-headed comparison, using DIGE technology along with an internal standard. In total, 35 protein spots identified by tandem mass spectrometry （MS/MS） exhibited significant changes （paired t-test, P 〈 0.05） in the expression level between normal and round-headed spermatozoa. A total of nine proteins were found to be upregulated and 26 proteins were found to be downregulated in round-headed spermatozoa compared with normal spermatozoa. The differentially expressed proteins that we identified may have important roles in a variety of cellular processes and structures, including spermatogenesis, cell skeleton, metabolism and spermatozoa motility.
文摘Since preparation and solubilization of protein samples are crucial factors in proteome research,the authors established a sequential extraction technique to prepare protein samples from the body wall of the 5th instar larvae of silkworm.Bombyx mori.Two kinds of protein samples were obtained from the body wall using the method.Between the two types of samples only about 15% proteins were identical;the majority were different,indicating that more species of proteins could be obtained with the sequential extraction method;which will be useful for preparation of protein samples for proteome study.
文摘Objective To identify the sperm membrane proteins that are associated with antisperm antibody Methods Using antisperm antibody positive serum through unidimensional polyacrylamide gel electrophoresis and 2-dimensional gel electrophoresis followed by Western blot analysis to determine the molecular weights (MW) and isoelectric points (pI) of sperm membrane proteins that are associated with antisperm antibody. Results Eight kinds of MW with more than ten sperm membrane proteins can be recognized by antisperm antibody positive serum, of which the MWs and pI were 23 kD, 31 kD, 32 kD, 34 kD, 41 kD, 51 kD, 60 kD, 78 kD and 5.3, 5.5,5.7, 5.0, 5.3, 5.8, 6.0, 5.5～6.2, 4.6,5.1,5.5～5.8 respectively. The identification ratios of the sperm membrane proteins on 78 kD (60.7%), 60 kD (71.4%), 51 kD (14.9%) and 23 kD (14.29%) were higher. Conclusion The sperm membrane proteins with MW of 78 kD, 60 kD, 51 kD and 23 kD were associated with antisperm antibody and immunological infertility. Two- dimensional gel electrophoresis and Western blotting can precisely identify the sperm membrane proteins that are associated with antisperm antibody.
文摘An improved CBB staining with higher sensitivity than that of the typical CBB staining was reported.The main improvement was using a fixing step of 25% trichloroacetic acid(TCA) before CBB staining.For most proteins studied,the sensitivity of the improved CBB staining was about twice as high as that of the typical method.For basic and low molecular weight proteins such as ribosomal proteins,the sensitivity of this improved staining method was about 3.5-28 times that of the typical method.It was speculated that the improved procedure would be suitable for exact quantitative analysis of proteins fractionated by SDS-PAGE,especially for basic and low molecular weight proteins.On the other hand,this new modified method might be also applied to multidisciplinary studies,such as biological researches and nuclear sciences.
文摘Regardless of thermodynamic stability, less complex proteome than plasma, convenient and noninvasive sampling of urine, reproducibility and comparability of the data among laboratories are challenges for the urine proteome analysis. Pretreatment of urine with 10 mM EDTA in conjunction with desalting methods to remove interfering molecules significantly improved the presentation of urine proteome on 2-D gels. Statistical analysis of the average ratio of the spot numbers on 2-D gels between EDTA-treated and non-treated samples were 1.35 ± 0.167, 1.26 ± 0.091, and 1.24 ± 0.095 for dialysis, ultrafiltration, and desalting column, respectively, with the p-value of 5.85 × 10–6 for the overall comparison. This result came from the fact that EDTA-treated samples showed better resolutions with less streaks and clearer spots on 2-D gels than the control samples. The results suggest that a simple and non-fractionating EDTA-treatment can be employed to the procedures for the high resolution 2-D gel analysis and other urinary proteomic analysis.
文摘Proteomics is one of the most active research fields in the post-genomic era. Here we briefly introduce the scientific background of the origination of proteomics and its content, research method. The new developments of proteomics at the levels of individual plants, tissues, organs and organells, as well as its applications in the area of plant genetic diversity, mutant characterization, and plant physiology, etc are reviewed. At last, the challenge and prospect of proteomics are discussed.
