Develepment of high sensitivity and high resolution techniques for the analysis of myeloma proteins

我们的目标是开发高灵敏度和高分辨率的分析人血浆黏液瘤蛋白的技术。微型二维凝胶电泳法和毛细管电泳法是候选的方法，但我们发现二维凝胶电泳法需要许多步骤的人工处理才能获得分离模式，并且还需要额外的图像分析过程来定量每个分离的蛋白质。然后研究了毛细管电泳法的两种分离原理：毛细管等电聚焦和毛细管十二烷基硫酸钠电泳法。针对毛细管电泳法的两种分离原理，建立了高分辨率、高灵敏度分离人血浆蛋白的条件。在所建立的分离条件下，用毛细管等电聚焦在60分钟内将正常人血浆蛋白分离出约60个峰/肩，用毛细管十二烷基硫酸钠电泳法在70分钟内分离出约30个峰/肩。在分离过程中，主要的血浆蛋白和菌丝体蛋白可以通过它们的紫外光吸收进行鉴定和定量。结果表明，在所建立的分离条件下，毛细管等电聚焦和毛细管十二烷基硫酸钠电泳法对人血浆黏液瘤蛋白的高效分离和分析是非常有效的。

We aimed to develop techniques to analyze human plasma mycloma proteins in high sensitivity and high resolution. Microscale two-dimensional gel clectrophoresis and capillary electrophoresis were the candidates, but we found that two-dimensional gel clectrophoresis requires many steps of manual handling to obtain the separation patterns and additional processes of image analysis arc necessary to quantitate each separated proteins. Then we examined two separation principles of capillary electrophoresis, capillary isoelectric focusing and capillary SDS elcetrophoresis, for the analysis of mycloma proteins. Electrophoretic conditions for high-resolution and high-sensitivity separation of human plasma proteins have been developed for the two separation principles of capillary electrophoresis. Employing the established separation conditions, normal human plasma proteins have been separated as about 60 peaks/shoulders within 60 minutes by capillary isoelectric focusing and as about 30 peaks/shoulders within 70 minutes by capillary SDS electrophoresis. Major plasma proteins and mycloma proteins can be identified and quantitated on-column during separation by their UV absorbance. Our results show that capillary isoelectric focusing and capillary SDS electrophoresis are quite effective for high performance separation and analysis of human plasma mycloma proteins, when the separation conditions we have established were employed.