Coomassie brilliant blue method

The Coomassie Brilliant Blue method, also known as the Bradford assay, was developed in 1976 by Marion M. Bradford. This technique is widely used for protein quantification due to its simplicity and high sensitivity. The method relies on the binding of a dye, Coomassie Brilliant Blue G-250, to proteins under acidic conditions. When this binding occurs, the dye undergoes a shift in its maximum absorbance wavelength from 465 nm to 595 nm, and the solution changes color from brownish-black to blue. This color change allows for easy detection and quantification of protein concentration. The principle behind the Bradford method involves the interaction between the dye and specific amino acid residues in proteins, particularly basic amino acids like arginine and aromatic ones such as tryptophan and tyrosine. The absorbance at 595 nm (A595) is directly proportional to the protein concentration, making it a reliable measurement tool. One of the main advantages of the Bradford method is its high sensitivity—approximately four times more sensitive than the Lowry method. It can detect as little as 1 µg of protein, which makes it ideal for low-concentration samples. Additionally, the procedure is quick and straightforward, requiring only one reagent and taking about 5 minutes to complete. The dye-protein complex forms rapidly within 2 minutes, and the color stabilizes within an hour, allowing for flexible timing compared to other methods that require strict time control. Another benefit is the reduced number of interfering substances. Unlike the Lowry method, the Bradford method is not affected by ions such as K⁺, Na⁺, Mg²⁺, or common buffers like Tris, sugars, glycerol, or mercaptoethanol. However, there are still some potential interferents, including detergents like Triton X-100, sodium dodecyl sulfate (SDS), and 0.1N NaOH. These can affect the accuracy of the results. Despite its benefits, the Bradford method has limitations. The response varies depending on the protein’s composition, especially the levels of arginine and aromatic amino acids. Therefore, bovine gamma-globulin is often used as a standard to minimize variability. Also, the standard curve may exhibit slight non-linearity, so Beer's Law cannot be applied directly, and a calibration curve must be used instead. Overall, the Coomassie Brilliant Blue method remains a popular choice for protein quantification due to its speed, sensitivity, and ease of use, despite some limitations in specificity and interference.

Magnetic Lash Box

Magnetic Lash Box,Magnetic Lashes,Diamond Eyelash Boxes,Magnetic Lashes Case

Zhengzhou Cuka Electronic Commerce Co., Ltd. , https://www.cukalashes.com