ESTABLISHMENT OF TWO-DIMENSIONAL GEL ELECTROPHORESIS FOR PROTEOMES OF RAT LUNG TISSUE

Objective To establish the technique of two-dimensional gel electrophoresis(2-DE) for proteomic analysis of rat lung tissue. Methods By combination and optimization of various conditions, a sample preparation method was established The lung cells were sufficiently lysed by using physical and chemical techniques. Protein degradation was minimized by operation at lower temperature, and adding protease inhibitor. Then the insoluble impurities were removed by centrifugation. The first-dimensional electrophoresis was immobilized pH gradients isoelectric focusing and second-dimensional electrophoresis was vertical electrophoresis of SDS-PAGE(T=13%). Results By control and optimizition of many experimental conditions, the tehnique could detet about 720 spots on 18 cm IPG strip(pH3-10 L)for mormal rat lung tissue, and the match rate of protein spots was 72.2% Conclusion The technique of 2-DE which has been established is the basis of work later on.