Why is it important to mix the Coomassie reagent before use?
Fortunately, gentle mixing completely disperses the dye-dye aggregates. Therefore, it is good practice to mix the Coomassie® Reagent before pipetting and to mix each tube or plate immediately before measuring absorbances.
How long does it take to do a BCA assay?
Choose the right BCA protein assay for your sample
| Rapid Gold BCA Protein Assay | Micro BCA Protein Assay | |
|---|---|---|
| Incubation time and temperature | 5 min at RT | 60 min at 60°C |
| Total assay time | 50 min | 130 min |
| Absorbance | 480 nm | 562 nm |
| Compatible reagents | Detergents | Detergents |
What is BCA testing?
A BCA (body composition analysis, for anyone unfamiliar with the term) is the “process of examining the human body to determine the percentage of body fat compared to muscle tissue.” A BCA is capable of revealing how healthy an individual’s body is, and is a popular resource for trainers, coaches, medical professionals …
How do you dilute BSA for Bradford assay?
Mix 50 μl of the BSA standard solution (2 mg/ml) with 950 μl of diluent and mix well to prepare a 0.1 mg/ml BSA standard solution.
What color change occurs when proteins combine with Coomassie dye under acidic conditions?
In the acidic environment of the reagent, protein binds to the Coomassie dye. This results in a spectral shift from the reddish/brown form of the dye (absorbance maximum at 465 nm) to the blue form of the dye (absorbance maximum at 610 nm).
How does Coomassie blue bind to proteins?
We found that the neutral ionic species of CBB binds to proteins by a combination of hydrophobic interactions and heteropolar bonding with basic amino acids.
What is Reagent A and B in BCA assay?
Reagents. Reagent A: 1 gm sodium bicinchoninate (BCA), 2 gm sodium carbonate, 0.16 gm sodium tartrate, 0.4 gm NaOH, and 0.95 gm sodium bicarbonate, brought to 100 ml with distilled water. Adjust the pH to 11.25 with 10 M NaOH. Reagent B: 0.4 gm cupric sulfate (5 x hydrated) in 10 ml distilled water.
How does BCA assay work?
Like the conventional BCA assay, the Pierce Rapid Gold BCA Protein assay involves the reduction of copper by proteins in an alkaline medium (biuret reaction) to produce sensitive and selective colorimetric detection by a new copper chelator.