1.3.3. Staining gels with Coomassie Brilliant Blue
For preparing the staining solution, which at the same time is used for fixing the proteins within the gel matrix:
1. Dissolve 0.25 % (w/v) Coomassie Brilliant Blue R-250 in ethanol, ddH2O and glacial acetic acid [5:5:1 (v/v)].
2. Let the solution stir overnight in the dark.
3. Filter the solution
4. Store in a dark bottle
The staining solution can be reused several times, but do not leave it for prolonged time in the staining dish, as this will cause the evaporation of the ethanol and thus reduce the fixation properties of the solution.
Staining of the gel can be done in a glass dish covered with plastic foil or in a household plastic dish with cover:
1. Immerse the gel in the staining solution.
2. Agitate slowly for 16 h (overnight) at room temperature on a slowly rocking platform or orbital shaker.
3. Remove the staining solution and save it for future use.
3. Destain the gel in the same solution [5:5:1 (v/v) of ethanol, ddH2O and glacial acetic acid] without the dye.
4. Change destaining solution two or three times and continue destaining until blue bands and a clear background are obtained.
Inserting a paper tissue into the destaining dish helps to absorb unbound Coomassie Brilliant Blue.
5. Destained gels can be dried in a vacuum gel-drying system; alternatively they can be stored in wet conditions within a sealed plastic container; add a few drops of glycerol to keep the gel soft.