Western Blot in Maize

Since its introduction in 1979 (Towbin et al., 1979), protein blotting (Western blot) has become a routine tool in molecular biology research.It is used to detect low amounts of proteins in complex samples or to monitor protein expression and purification.Western blotting includes protein complex separation by gel electrophoresis (SDS-PAGE), transfer to a fixed matrix or membrane and analysis by immunodetection using a specific antibody against the protein of interest.This method provides molecular mass information of individual proteins, and it can be used to distinguish between isoforms and post-translational modification states.There are many variations on the Western blot procedure that can be performed to improve detection, specificity, and quantitative power.In plants, Western blot has to be standardized to get a satisfactory signal and reproducible results, due to the complex nature of cell extracts.Here, I describe a protocol for a semi-quantitative Western blot using total maize protein and specific polyclonal antibodies raised against maize proteins of interest.The data are analyzed with ImageJ.