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.
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.
