High-contrast imaging of chlorophyll-containing protein complexes separated by native polyacrylamide gel electrophoresis

Abstract

Electrophoregrams with bands of green chlorophyll-containing protein complexes separated by native polyacrylamide gel electrophoresis (PAGE) are usually photographed in white light or scanned by a narrow beam of monochromatic light using densitometers. Present CCD cameras allow taking densitograms of a whole gel surface with many samples. However, commercial CCD imaging systems use mostly white or broad-band light scanning for densitometry measurements and are not constructed for the measurements by monochromatic light. In this work we present an application of a home-made two-dimensional CCD imaging system for monochromatic densitometry of chlorophyll-containing proteins separated by PAGE. A very mild Deriphate-PAGE according to Peter and Thornber (1991) was used for the separation. We scanned the gel at 675 and 650 nm, i.e. at red absorption maximum of chlorophyll a and b, respectively. Scanning of green bands in the gel by monochromatic light, corresponding with absorption maxima of chlorophylls, increased markedly a contrast of the bands. This allows a better visualization of faint-green bands in the gel. Moreover, a quotient of the gel densitograms scanned at 675 and 650 nm allows to distinguish the bands containing proteins with different chlorophyll a/b ratio, i.e. mainly photosystem cores and light-harvesting complexes. An application of this method to high-temperature changes in the content of chlorophyll-containing proteins is shown