Hello all,
My lecturer wants a bradford protocol using with G-250 CBB dye. We will use this on spectrophotometer. I already found many of them but they are same with this link: http://www.ruf.rice.edu/~bioslabs/methods/protein/bradford.html
If anyone know different version of this procedure, may I have new version?
Thanks in advance. Best regards.
I like the Bradford reagent concentrate from Bio-Rad. It has excellent sensitivity and is extremely stable when stored in the refrigerator.
Dear Evren,
I attach several protocols on Bradford method and its modifications (including the Bio-Rad protocol just mentioned by Dr. Shapiro). Hope you will find sonmething you need.
Thanks for your answers & help. I will check them tomorrow. Thank you very much. Best regards.
Edit: I checked these protocols but except one of them, are not suitable for me. My purpose is not buy pre-made Bradford reagent, I want to make it myself, like "homemade". One of the protocol is different (Bradford method.mht) than I have and found so far. I am very thankful of your help & interest. I am still looking for the issue and still open for new advices & knowladge sources.
Thank you, best regards.
Hi, I also use the Bradford protein assay nowadays for my routine protein content determinations. However I found the following recipe from this link http://www.my-whiteboard.com/home-made-bradford-reagent-for-protein-assay/: Dissolve 100 mg Coomassie Brilliant Blue G-250 (100 mg) in 50 ml 95% 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.
The secret is to have the dye thoroughly dissolved. We do this overnight and then filter the reagent. Be sure no particulates are present in the working solution.
For the protein assay, try this
protein solution 20 uL
extraction buffer (EB) 30 uL
Homemade Bradford rgt 2.5 mL
for Blank: 50 uL EB
Bradford rgt 2.5 mL
Hello, I am very thankful of your reply. I will also discuss these informations with my lecturer.
I am still waiting for if any new & different protocols available. Best regards.
I wonder EB meaning? If you can explain, i will be glad. Thank you very much for your kindly and helpful answer.
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