Ultrasensitive detection of unstained proteins in acrylamide gels by native UV fluorescence

Rögener J, Lutter P, Reinhardt R, Blüggel M, Meyer HE, Anselmetti D. Ultrasensitive detection of unstained proteins in acrylamide gels by native UV fluorescence. Analytical chemistry. 2003;75(1):157-159.Visualization of proteins inside acrylamide and other gels usually relies on different staining methods. To omit the protein-staining procedure, we visualized unstained proteins inside acrylamide gels by laser excitation with ultraviolet (UV) light (280 nm, 35 mJ/cm²) and directly detected native UV fluorescence. In one-dimensional gels, a detection limit as low as 1 ng for bovine serum albumin and 5 ng for other proteins with a linear dynamic range (2.7 orders of magnitude) comparable to state of the art fluorescent dyes could be achieved. In addition, the application of this method to 20 µg of a whole cell lysate separated in a two-dimensional gel showed more than 600 spots. Since protein labeling always represents a serious obstacle in protein identification technologies, the working efficiency with our procedure can be considered as a significant improvement for protein visualization and reproducibility in proteomics