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目的建立和优化人骨肉瘤蛋白质组双向电泳图像分析方法。方法对第一向双向电泳的关键因素与环节(如样品处理、电泳参数和凝胶浓度等)进行一系列优化。进一步采用双向凝胶电泳分离骨肉瘤组和正常组总蛋白质,银染显色,Image Master图像分析系统分析电泳图谱。结果重复性实验结果显示同组样品在3次不同实验中所的蛋白质斑点数目相对标准差。(变异系数平均值%):骨肉瘤组和正常组分别为23.00±10.11、20.33±9.90。(变异系数范围%):骨肉瘤组和正常组分别为3.80-6.89、2.70-6.89。同一蛋白质斑点在3次实验中等电点、分子量的相对标准差分别为8.93%±1.17%,10.16%±2.02%和10.87%±3.86%。获得了优于传统实验方法的分辨率和可重复性的双向凝胶电泳图谱。结论优化的双向凝胶电泳图象分析技术适合于骨肉瘤蛋白质组研究。
Objective To establish and optimize two-dimensional electrophoresis image analysis method of human osteosarcoma proteome. Methods The key factors and links (such as sample processing, electrophoresis parameters and gel concentration) of the first two-dimensional electrophoresis were optimized. The total protein of osteosarcoma group and normal group was further separated by two-dimensional gel electrophoresis, and silver staining was performed. Image analysis system was used to analyze the electropherogram. Results The results of repetitive experiments showed that the number of protein spots in the same group of samples in three different experiments was relatively standard deviation. (% Mean value of variation coefficient): The osteosarcoma group and the normal group were respectively 23.00 ± 10.11 and 20.33 ± 9.90. (% Of variation coefficient): Osteosarcoma group and normal group were 3.80-6.89,2.70-6.89 respectively. The same protein spot in the three experiments isoelectric point, the relative standard deviation of the molecular weight were 8.93% ± 1.17%, 10.16% ± 2.02% and 10.87% ± 3.86%. Two-dimensional gel electrophoresis patterns were obtained with better resolution and repeatability than the traditional experimental methods. Conclusion The optimized two-dimensional gel electrophoresis image analysis technique is suitable for osteosarcoma proteome research.