What is BCA reagent A?

What is BCA reagent A?

BCA Protein Assay Reagent A is a component of the Pierce BCA Protein Assay Kit, a two-component, high-precision, detergent-compatible assay that is used for total protein concentration determination compared to a protein standard.

Does SDS interfere with BCA assay?

The 2-mercaptoethanol will interfere with the BCA assay, and the SDS will interfere with the Bradford assay.

What is the principle of BCA?

The principle of the bicinchoninic acid (BCA) protein assay1 relies on the formation of a Cu2+-protein complex under alkaline conditions, followed by reduction of the Cu2+ to Cu1+. The amount of reduction is proportional to the amount of protein present.

How do you calculate final protein concentration?

Protein concentration can be estimated by measuring the UV absorbance at 280 nm; proteins show a strong peak here due to absorbance from Tryptophan and Tyrosine residues (commonly referred to as A 280). This can readily be converted into the protein concentration using the Beer-Lambert law (see equation below).

How can I start BCA?

Assay

1. Prepare samples containing 0.2 to 50 micrograms protein in microliters.
2. Add 1 ml SWR to each 20 microliters sample and mix. Incubate 30 min. at 60 degrees C.
3. Cool the samples and read at 562 nm. Color will be stable for at least one hour.

Is BCA better than Bradford assay?

Historically, the BCA method is more sensitive than the Bradford method, because the first method is based on protein-copper chelation and secondary detection of the reduced copper. Whereas the Bradford method is based on protein-dye binding and colour shift from 465 to 595 nm.

How does the Pierce BCA protein assay kit work?

Pierce BCA Protein Assay Kit is a two-component, high-precision, detergent-compatible protein assay for determination of protein concentration. Pierce BCA reagents provide accurate determination of protein concentration with most sample types encountered in protein research.

Is the BCA protein assay sensitive to cuprous cation?

The BCA Protein Assay combines the well-known reduction of Cu2+ to Cu1+ by protein in an alkaline medium with the highly sensitive and selective colorimetric detection of the cuprous cation (Cu1+) by bicinchoninic acid.

How is the BCA assay different from dye binding?

Compared to most dye-binding methods, the BCA assay is affected much less by protein compositional differences, providing greater concentration accuracy.

What causes the color of the BCA protein?

The reaction that leads to BCA color formation is strongly influenced by four amino acid residues (cysteine or cystine, tyrosine, and tryptophan) in the amino acid sequence of the protein. However, unlike the Coomassie dye-binding methods, the universal peptide backbone also contributes to color formation,…