Fluorescent Protein Visualization Immediately After Gel Electrophoresis Using an In-Gel Trichloroethanol Photoreaction with Tryptophan
SDS–polyacrylamide gel electrophoresis (SDS-PAGE) is one of the essential techniques in molecular biology and biochemistry laboratories and requires rapid visualization methods for efficient sample analysis. Proteins on polyacrylamide gels can be visualized within 5 min via the photoreaction of tryptophan with trichloroethanol. This process does not require protein fixation, staining, or destaining. In this method polyacrylamide gels are prepared by adding trichloroethanol before polymerization. After electrophoresis, the gel is immediately activated on a standard UV transilluminator and the fluorescently labeled proteins are imaged. The reaction is based on the photoreaction of trichloroethanol with tryptophan residues within the protein. This generates a visible blue-green fluorescence (∼500 nm) that is accurately imaged. Here we describe the preparation of Tris–glycine and Tris–tricine SDS–polyacrylamide gels with trichloroethanol and the photoreaction and visualization of tryptophan containing proteins.
Key wordsProtein electrophoresis Fluorescent visualization Trichloroethanol Tryptophan photochemistry
We thank Natural Sciences and Engineering Research Council of Canada (NSERC) for funding.
- 18.Gilda JE, Gomes AV (2015) In: Posch A (ed) Proteomic profiling: methods and protocols, vol 1295, pp 381–391Google Scholar