Hallo Tomas, below please find a standard recipe:

Dissolve 100 mg Coomassie Brilliant Blue G-250 (100 mg) in 50 ml 95% ethanol (dilute from pro analysis ethanol), add 100 ml 85% (w/v) phosphoric acid. Dilute to 1 liter when the dye has completely dissolved, and filter through Whatman #1 paper. Final concentrations in the reagent: 0.01% (w/v) Coomassie Brilliant Blue G-250, 4.7% (w/v) ethanol, 8.5% (w/v) phosphoric acid. For standard macroassay (1 ml cuvettes), just use 200 microliter of your sample appropriately diluted plus 800 microliter of the reagent. Only precautions: Use milli q water and use very-very clean harware. If the solution is blue from the beginning then some impurities may have occured in the process. I wish to you good luck, although I feel that for critical reagents like Bradford, a commercial source may be more guaranteed.

Hallo Tomas, below please find a standard recipe:

Dissolve 100 mg Coomassie Brilliant Blue G-250 (100 mg) in 50 ml 95% ethanol (dilute from pro analysis ethanol), add 100 ml 85% (w/v) phosphoric acid. Dilute to 1 liter when the dye has completely dissolved, and filter through Whatman #1 paper. Final concentrations in the reagent: 0.01% (w/v) Coomassie Brilliant Blue G-250, 4.7% (w/v) ethanol, 8.5% (w/v) phosphoric acid. For standard macroassay (1 ml cuvettes), just use 200 microliter of your sample appropriately diluted plus 800 microliter of the reagent. Only precautions: Use milli q water and use very-very clean harware. If the solution is blue from the beginning then some impurities may have occured in the process. I wish to you good luck, although I feel that for critical reagents like Bradford, a commercial source may be more guaranteed.

I agree with the above, except I filter the required amount with a 0.45 um just before use.

Does the brownish color of bradford reagent cause any problem in the case that there is no protein to change its color to blue? And also, is it possible to use R-250 instead of G-250? Cuz I think they both will be blue in PH more than 2, is it right?

Is it possible to use another acid instead of phosphoric acid? I'm really unable to find it...

I used the recipe to prepare Bradford reagent. But the colour is very light, almost like transparent. Can anyone guess why it turned out to be like this? I added exact amount and accurately mixed everything. Previously also, I made it in the same way but the colour seemed fine. After final preparation, this time, the colour didn't appear. It'll be of great help if anyone can find the reason to it.

I have used this recepie to make Bradford solution but we have got some greenish colour instead of brown like the commercial preparation kindly suggest me to make it right ?