Simplified silver protein detection and image enhancement methods in polyacrylamide gels

Abstract The use of silver for detection of protein in polyacrylamide gels is becoming widespread. An effort has been made to develop a silver stain which minimizes the time to perform the stain, the amount of silver used, and the complexity of the procedures, and which maximizes the sensitivity. This stain is almost 200‐fold more sensitive than the Coomassie Brilliant Blue R‐250 stain. This silver stain procedure has been shown, with eight purified proteins, to generally be linear over a 40‐fold range in concentration, between 0.005 ng/mm2 to 2.0 ng/mm2. For greater sensitivity, a recycling procedure has been developed. This procedure is capable of amplifying trace proteins which could not be visualized with previous silver stain techniques. Analysis of the kinetics of image development revealed that the silver ion was the limiting factor in image formation. By recycling the gel through silver nitrate solutions, silver ions are replenished in the gel, permitting amplification of further details when the gel is treated with a developing solution. A number of problems inherent in silver stain detection of proteins in polyacrylamide gels are discussed with suggested remedies.