Coolidal Coomassie staining procedure

Colloidal Coomassie staining according to Neuhoff

(Electrophoresis 1988, 9, 255-262)

Detection limit:  0.7 ng/mm² gel (for normal Coomassie: 20 -100 ng/mm² gel)

1.    Stock solutions:

Solution A:

10% (w/v) ammonium sulphate, 2% (w/v) phosphoric acid in MilliQ water

Solution B:

5% (w/v) Coomassie Brilliant Blue G-250 in MilliQ water

Washing solution:

25% methanol in MilliQ water

2.    Protocol

The gel is incubated for 5 min in water and then stained overnight in staining solution. The detailed protocol is described below:

• The staining solution is prepared by mixing 100 ml of the stock solution A with 2.5 ml stock solution B. Shake strongly this solution for 20 minutes and then add 25 ml pure methanol. Continue shaking the next 20-30 minutes.
• Place the gel in the freshly prepared colloidal Coomassie stain. Stain the gel overnight with gentle shaking.
• Wash the gel with 25 % methanol in MilliQ water for around 1 hour.
• Wash the gel with MilliQ water
• Repeat the last two steps until the protein bands are at the desired contrast against the background of the gel.