A new method to assign immunodetected spots in the complex two‐dimensional electrophoresis pattern

A new, easy method for the immunodetection of specific antigens in two-dimensional electrophoresis (2-DE) is described. Areas of 2-DE gels containing antigens of interest are electrophoretically transferred to polyvinylidene difluoride membranes, immunostained with specific antibodies using Fast Red or 5-bromo-4-chloro-3-indolyl phosphate/nitro blue tetrazolium as detection systems and counterstained with Coomassie Brilliant Blue. In contrast to conventional methods, it is possible to use this procedure to exactly assign immunoreacting proteins on a single blot to their corresponding and surrounding blue-stained protein spots.