Brilliant blue G-250

Brilliant Blue G-250 is a dye commonly used for the visualization of proteins separated by SDS-PAGE, offering a simple staining procedure and high quantitation. In the Bradford protein assay, protein concentrations are determined by the absorbance at 595 nm due to the binding of Brilliant Blue G-250 to proteins. Brilliant Blue G-250 is a safe highly selective P2×7R antagonist with promising consequent inactivation of NLRP3 inflammasome^[1][2][3]. In Vitro: Staining Proteins in Polyacrylamide Gel Electrophoresis with Brilliant Blue G-250^[1]
1. Solution Preparation
Coomassie staining solution:
(1) Dissolve 100 g aluminum sulfate (14-18 hydrate) in 2000 mL Milli-Q purified water. Add 200 mL ethanol (96%) and mix well.
(2) Add 0.4 g Brilliant Blue G-250 and stir until dissolved.
(3) Slowly add 47 mL phosphoric acid (85%) while stirring.
(4) Finally, adjust the volume to 2000 mL with Milli-Q purified water.
Note: Do not filter the solution. It should appear colloidal with suspended particles.
Destaining solution:
Prepare 2000 mL Milli-Q purified water, adding 200 mL ethanol (96%) and 47 mL phosphoric acid (85%).
2. Staining Procedure
(1) After protein electrophoresis, carefully remove the gel from the glass plates. Wash the gel with Milli-Q purified water 3 times for 10 minutes each to remove SDS. Shake the Coomassie staining solution before use to disperse the colloidal particles. Immerse the gel in the staining solution and agitate on a shaker for 2-12 hours.
(2) Protein spots will begin to appear after 10 minutes, and 80% of staining will be completed within 2 hours. For optimal results, overnight staining is recommended.
(3) After staining, remove the Coomassie solution and rinse the gel twice with Milli-Q purified water. Place the gel in the destaining solution and agitate for 10-60 minutes to remove excess dye.
3. Rinsing and Storage
Rinse the gel twice with Milli-Q purified water to restore its original thickness. The gel can be stored in a refrigerator with an acidic solution to prevent mold contamination.
4. Notes
(1) Ensure the gel is thoroughly washed before staining, as residual SDS will interfere with dye-protein binding.
(2) The staining solution can be reused as long as particles remain.
(3) Store the staining solution in a dark bottle to extend its shelf life.