State-of-the-art two-dimensional gel electrophoresis: a key tool of proteomics research.
Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) is the most popular and versatile method of protein separation among a rapidly growing array of proteomics technologies. Based on two distinct procedures, it combines isoelectric focusing (IEF), which separates proteins according to their isoelectric point (pI), and SDS-PAGE, which separates them further according to their molecular mass. At present, 2D-PAGE is capable of simultaneously detecting and quantifying up to several thousand protein spots in the same gel image. Here we provide comprehensive step-by-step instructions for the application of a standardized 2D-PAGE protocol to a sample of human plasma or cerebrospinal fluid (CSF). The method can be easily adapted to any type of sample. This four-day protocol provides detailed information on how to apply complex biological fluids to an immobilized dry strip gel, cast home-made gradient acrylamide gels, run the gels, and perform standard staining methods. A troubleshooting guide is also included.
Carrette
Odile
O
Biomedical Proteomics Research Group, Department of Structural Biology and Bioinformatics, Faculty of Medicine, Geneva University, 1 rue Michel Servet CH-1211 Geneva 4, Switzerland.
Burkhard
Pierre R
PR
Sanchez
Jean-Charles
JC
Hochstrasser
Denis F
DF
D001798:Blood Proteins / Q000737:chemistry; D002555:Cerebrospinal Fluid / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D006801:Humans; D040901:Proteomics / Q000379:methods*
D001798:Blood Proteins