Optimization of conditions of two-dimensional gel electrophoresis for detection of BHK-21 cellular protein sample

Conditions of two-dimensional gel electrophoresis for detection of BHK-21 cellular protein sample were optimized from four aspects including cell processing,sample loading quantity,concentration of polyacrylamide gel in SDS-PAGE and staining method,to provide comparative proteomics technical foundation for foot-and-mouth virus infected cell study.Results showed that compared with the method of freezing and thawing,ultrasonic method had a better effect on cells breaking.And 100 μg sample loading quantity was appropriate for the 7 cm IPG strip of pH3-10 with coomassie brilliant blue stain,while 10% polyacrylamide gel could separate more protein points than other concentrations of polyacrylamide gels.Silver stain was superior to coomassie brilliant blue staining.After optimization of the conditions,the acquired maps,which could be used for proteomics study of BHK-21 cells infected with FMDV,possessed many advantages,such as more and clearer protein spots,good reproducibility,reduced delay phenomenon,and complied with the requirements of the software analysis.