I can still remember quite a few of years ago when I needed to quantify protein concentration by the traditional Bradford assay. The preparation work was critical: all materials must be handy – protein samples and standards on ice, buffer blank, clean cuvettes, kimwipes, spectrophotometer (already warmed up), wash bottle, pipetmans, pipet tips, waste bottle, timer, Coomassie Blue G-250 solution and distilled water. The key is to perform the assay quickly, as the absorbance readings will increase with time (in every single minute). And this could be a problem for rookies, especially when there were a great number of samples.
This review does not aim at comparing the Bio-Rad DC protein assay with the traditional Bradford assay – in fact, they are different assays. The Bio-Rad DC protein assay is a modified Lowry assay. The best part of this kit is revealed in section 1, page 1 of the user manual: ‘The reaction reaches 90% of its maximum color development within 15 minutes thereby saving valuable time, and the color changes NOT MORE THAN 5% IN 1 HOUR OR 10% IN 2 HOUR after the addition of reagents.’
By adapting their microplate assay protocol, I have recovered from the previous nightmare. I must admit that the microplate format and ELISA reader have also played a huge part. But anyway, for the two methods I have used, the Bio-Rad DC reagents would be my choice. Although they cost more than the homemade Bradford dye, they are worth their price. Before you use the reagent, check its compatibility with your buffers. Common lysis buffer reagents like SDS, Triton X-100, Tween 20, NP-40, NaN3, EDTA, Tris, etc. are compatible. An exception is 2-mercaptoethanol.
Add the BSA standards, buffer blank and samples to the microplate. Usually, I perform replicates. Then add the reagents from the kit. Make sure there are no bubbles when mixing. Incubate at room temperature for 15 minutes. To verify their claim, I once read a plate at 15 minutes and then at 2 hours. On average, the change in OD750 readings was 5.4% (SD±2%), well fitted to their statement. In other words, there is no need for haste after you mix all the components.
A draw back is the relative short shelf-life of the reagents. The manufacturer guarantees all reagents will be good for 6 months from the date of purchase. Anyway, I think nobody would throw away the remaining reagents sharply after 6 months, right? But what will happen then? I have some data record that may give you an idea: OD750 of buffer blank in March 06 = 0.064, September 06 = 0.114, September 07 = 0.121.
In general, this kit is sensitive and easy to use. Although the blank readings gradually increase after 6 months, it can still tell you the relative abundance of your samples as you subtract all readings with the blank. At the end of the day, this user-friendly product will win the heart of customers.