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

开发用于分析骨髓瘤蛋白的高灵敏度和高分辨率技术

基本信息

批准号：
09640731
负责人：
MANABE Takashi
金额：
$ 2.05万
依托单位：
Ehime University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

项目摘要

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.

我们的目标是开发一种高灵敏度和高分辨率的分析人血浆骨髓瘤蛋白的技术。微尺度的二维凝胶电泳和毛细管电泳是候选的，但我们发现二维凝胶电泳需要许多步骤的人工处理来获得分离模式，并且需要额外的图像分析过程来定量每个分离的蛋白质。在此基础上，研究了毛细管电泳等电聚焦和毛细管SDS电泳两种分离原理，用于骨髓瘤蛋白的分析。针对毛细管电泳的两种分离原理，建立了高分辨率和高灵敏度分离人血浆蛋白的电泳条件。在建立的分离条件下，正常人血浆蛋白经毛细管等电聚焦在60分钟内分离出约60个峰/肩，经毛细管SDS电泳在70分钟内分离出约30个峰/肩。主要的血浆蛋白和骨髓瘤蛋白可以在分离过程中通过其紫外吸光度在柱上进行鉴定和定量。结果表明，在建立的分离条件下，毛细管等电聚焦和毛细管SDS电泳对人血浆骨髓瘤蛋白的高效分离和分析是非常有效的。