Description
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The kit is designed for protein quantification in biological samples.
Background: The BCA method combines the biuret reaction with the colorimetric detection of the monovalent copper ion by bicinchoninic acid (BCA). In this reaction, protein reduces Cu2+ to Cu+ in an alkaline environment .After the reduction of the divalent copper ion, Cu+ reacts with BCA and the purple colored reaction product of this assay is formed by the chelation of two molecules of BCA with one cuprous ion that has an absorbance at 562 nm. The first reaction occurs at lower temperatures and is the result of copper and BCA interaction with aminoacid residues in the protein. The absorbance increases linearly with increasing protein concentration over a broad working range (20-2000 µg/ µl). At elevated temperatures, the peptide bond is responsible for color development. Hence performing the assay at 37°C or 60°C versus room temperature increases the sensitivity and reduces the variation in the response of the assay as a function of protein composition. When possible, the assay should be incubated at 60°C since, after reaction is complete, the absorbance does not increase appreciably, whereas after cooling samples incubated at 37°C to room temperature, the blank continues to increase in absorbance at ~2.3% every 10 min.The BCA assay has many advantages over other protein determination techniques due to there is less susceptibility to detergents, is easy to use and the color complex is stable.Components that interfere with the BCA protein quantification kit either lead to the reduction of Cu2+ (as DTT) or copper chelators (as EGTA). Generally, these are not critical components of buffers and can be easily removed or omitted prior to the assay.
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