AUTOMATION OF QUANTITATIVE 2-D GEL STAINING

本建议是开发一个完整的集成和自动化系统 用于银染色和扫描二维高分辨率凝胶 提高灵敏度和分辨率， 定量 该系统是研究基因突变所必需的 在细胞和组织中响应药物，异生物质， 伤害、压力和疾病。 在50多个变量中， 发表的银染色的变化很大，使得优化 使用人工程序非常昂贵和繁琐。 此外，本发明还提供了一种方法， 现在实现了定量，需要对每个 由于图像显影的动力学， 对于不同的蛋白质来说是不同的。 我们开发了一个机器人原型 银染系统，并建议其完成与 开发了一种新的系统，用于重复扫描每个凝胶， 发展 这需要一种新的凝胶转运系统，以及一种组合的 包括可要求图像的扫描仪的扫描仪显影系统 以很短的间隔。 预计完成的系统将处理多达 每天40粒。匹配连续图像的软件， 计算作为时间的函数的积分时间，并且，通过 参考校准运行，确定每种蛋白质的量 现在，也会写。 建议的商业应用 潜在的商业应用：一个完全自动化的计算机 优化二维凝胶银染色的系统将是关键因素 在高通量2D蛋白质分析系统的开发中， 对蛋白质组学的发展至关重要。

This proposal is to develop a complete integrated and automated system for silver staining and scanning two-dimensional high resolution gels to both increasing sensitivity and resolving power, and of improving quantitation. This system is required for studies on changes in gene expression in cells and tissues in response to drugs, xenobiotics, injury, stress and disease. The number of variables in the over 50 variations of silver staining published are large, making optimization using manual procedures extremely costly and tedious. In addition, quantitation is now realized to require dynamic measurements on each spot during image development since the kinetics of image development differs for different proteins. We have developed a prototype robotic silver staining system and propose its completion together with the development of a novel system for repeatedly scanning each gel during development. This requires a novel gel transport system, and a combined scanner-development system including a scanner which can require images at short intervals. The completed systems is expected to process up to 40 gels per day. The software required to match successive images, calculate integrated absorbances as a function of time, and, by reference to calibration runs, determine the amount of each protein present, will also be written. PROPOSED COMMERCIAL APPLICATION Potential commercial applications: A completely automatic computerized system for optimizing silver staining of 2D gels will be a key factor in the development of a high throughput 2D protein analysis systems essential for the development of proteomics.