We think it would be cheaper to make our own from the solid dye, phosphoric acid, and an alcohol. I think that using acid-washed glassware makes sense, to minimize interference from detergents. Just in casually looking at some recipes, I see that some use ethanol and some use methanol. Any other tips or suggestions on the best recipe to use?
I do not wish to discourage you from making your own Bradford reagent to save money, but I would like to offer an alternative way to save money while still using one of the excellent and convenient commercial Bradford reagents: reduce the volume of the assay so that you can use less of the reagent. There are two ways to do this. (1) If you are using a cuvette-based spectrophotometer, use lower volume cuvettes. (2) Use 96-well plates and an absorbance plate reader, using an assay volume of 200 µl. A plate mixer is very helpful. (Actually, even smaller volumes can be achieved with plate readers if desired by using half area plates, but mixing becomes more difficult.)
Dear Chris,
Removing detergents by using glassware that underwent washing by acid is a very good idea. I suggest to use dilute phosphoric acid for the washing since this acid is the one to be used in the preparation of the reagent. Generally 95% ethanol is used in the preparation of this reagent. Di-ionized water and Whatman #1 paper should be used.
For other hints, I suggest that you look at the procedure depicted in the following link:
http://www.jenway.com/adminimages/P09_003A_Bradford_Protein_Assay.pdf
Hoping this will be helpful,
Rafik
I do not wish to discourage you from making your own Bradford reagent to save money, but I would like to offer an alternative way to save money while still using one of the excellent and convenient commercial Bradford reagents: reduce the volume of the assay so that you can use less of the reagent. There are two ways to do this. (1) If you are using a cuvette-based spectrophotometer, use lower volume cuvettes. (2) Use 96-well plates and an absorbance plate reader, using an assay volume of 200 µl. A plate mixer is very helpful. (Actually, even smaller volumes can be achieved with plate readers if desired by using half area plates, but mixing becomes more difficult.)
I like Adam's idea. But you can always give a try for your own Bradford reagent. Its recipe is easily available online.
Dissolve 50mg of Coomassie Blue G250 in 50ml of methanol.
Add 100ml of 85% H3PO4 to the solution from step 1.
Add the solution from step 2 into 500ml of H2O and mix.
Filter to remove and precipitates.
Add an additional 350ml of H2O.
Store at 4oC.
- Badges
- Science topic
- Similar topics
- Chemistry
- Biochemistry