Simple and reliable protein quantification The BCA protein assay is based on a biuret reaction, which is the reduction of Cu2+ to Cu+ by proteins in an alkaline solution with concentration-dependent detection of the monovalent copper ions. Bicinchoninic acid is a chromogenic reagent that chelates the reduced copper, producing a purple complex with strong absorbance at 562 nm (Smith 1985, Wiechelman 1988). This assay can be used to quantify protein concentration with a wide variety of samples and can be performed in minutes.
The Novagen® BCA Protein Assay Kit can be used to determine protein concentration in the range of 20-2000 µg/ml in either a standard assay or microassay configuration. Kit components are sufficient to complete 500 standard-size reactions (50 µl protein sample plus 1 ml reagent) or 2500 micro-scale reactions (25 µl protein sample plus 200 µl reagent). A BSA standard (2 mg/ml) is provided for convenient and reliable preparation of standard curves.
This assay is robust and can be performed in the presence of many compounds. Some reagents, including chelating agents, strong acids or bases, and reducing agents, interfere with the reduction and chelating reactions on which this assay depends (Brown 1989). The BCA assay is compatible with the following Novagen protein extraction and lysis reagents: BugBuster® Protein Extraction Reagent, PopCulture® Reagent, CytoBuster™ Protein Extraction Reagent, Reportasol™ Extraction Buffer, and Insect PopCulture Reagent. Options for the removal or dilution of interfering substances are described in the kit literature. References:
Smith, P.K., et al. 1985. Anal. Biochem. 150, 76. Wiechelman, K., et al. 1988. Anal. Biochem. 175, 231. Brown, R., et al. 1989. Anal. Biochem. 180, 136. Components:
| • | 500 ml | BCA Solution
| | • | 15 ml | 4% Cupric Sulfate
| | • | 3 x 1 ml | BSA Standard, 2 mg/ml |
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