A novel PAGE buffer for faster runs and sharper protein bands

Abstract

Since the mid‐70s Laemmli's protocol based on a Tris‐glycine‐SDS buffer system has been widely used for analyzing protein mixtures by PAGE. Laemmli's buffer system allows clear and accurate molecular weight estimation of proteins in complex samples from a wide variety of sources, but at the cost of long electrophoresis time. Modifications and improvements of this method reducing total run‐time are limited to mainly high priced precast gels.In this work, we demonstrate an improved Laemmli method offering significant cut in run‐time and improved sharpening of protein bands. The patent pending modifications apply to the running buffer system which can be run at higher voltages without generating high heat and impacting gel performance. We show that typical mini‐gels (10 x 10 cm gel) cast with the Laemmli recipe can be run 50 to 60% faster and still maintain a wide separation range similar to porosity gradient gels. In fact, a 10% Laemmli discontinuous mini‐gel will provide ultra‐fine band resolution over a wide molecular weight range (10 to 240 kDa) in less than 30 minutes using this unique running buffer. The need for gradient gels may be over since a uniform concentration gel can now fractionate proteins covering a wide range of molecular masses in half the time.