Image Acquisition in 2-D Electrophoresis

Abstract

Prevalent methods for visualizing proteins resolved by two-dimensional (2-D) gel electrophoresis include autoradiography, silver staining, and Coomassie brilliant blue staining (1,2. The organic dye Coomassie brilliant blue is capable of detecting as little as 100 ng of protein, but this is considerably less sensitive than silver staining or autoradiography (1,2. Silver staining allows detection of low-nanogram amounts of protein, but detection sensitivity of autoradiography can be considerably better, since it depends principally on the specific activity of the radiolabeling method. Other staining techniques are valuable for specific applications in 2-D gel electrophoresis, such as Edman-based protein sequencing, mass spectrometry, and immunoblotting. Many of these applications require stains that do not covalently modify proteins, are relatively sensitive, and are easily reversible (3). Additionally, immunoblotting protocols are often based upon the enzymatic formation of a colored formazan product or the chemiluminescent production of light (4). Thus, image acquisition requires detection of diverse chromogenic, radioactive, and luminescent signals.