Analysis on musculature proteome of rat fetus by two-dimensional gel electrophoresis

Objective To establish protocol of two-dimensional Gel Electrophoresis for analysing the proteome of musculature of Wistar rat fetus. Methods Proteins extracted from tibia-fibulae musculature isolated from 21-day rat fetus were loaded on immobile pH gradient gels to process the first dimension isoelectric focusing electrophoresis and then transfer to SDS polyacrlamide gel for the second dimension electrophoresis.Following the electrophoresis Coomassie Brilliant Blue staining or silver staining was performed,and the images were scanned and analyzed with ImageMaster 5.0 software package.One set or protein spots matching from three separate gels were cut for MALDI-TOF mass spectrometry analysis. Results Images with good reproducibility of the proteome of musculature were obtained after tow dimensional electrophoresis.The reproducibility of 2D gel electrophoresis was verified through comparing of peptide mass fingerprints form matching protein spots of three separate gels which suggested a same protein(light chain 1 of skeletal muscle myotonin). Conclusion The utilization of 2D electrophoresis can satisfactorily separate muscular proteins from rat embryos and achieve a high resolution and good reproducibility for protein identification.