What are the steps of BCA protein assay?

What are the steps of BCA protein assay?

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.

How does the Pierce BCA Protein Assay work?

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 (BCA).

How do you quantify protein in BCA?

The BCA protocol requires a working solution mixed with the sample; when protein is present, the reaction produces a purple color that absorbs light at 562 nm and is quantified with a spectrophotometer. The BCA assay is similar to other protein quantification assays such as Lowry or Bradford assays.

What are protein assays?

Protein assays differ in their chemical basis for detecting protein-specific functional groups. Some assay methods detect peptide bonds, but no assay does this exclusively. Instead, each protein assay detects one or several different particular amino acids with greater sensitivity than others.

What is BCA reagent?

BCA is a weak acid composed of two carboxylated quinoline rings and serves the purpose of the Folin reagent in the Lowry assay, namely, to react with the product (Cu1 + ions) of complexes between copper ions and peptide bonds to produce a purple end product that strongly absorbs at 562 nm (Smith et al., 1985).

What assays determine total protein content?

This unit describes three copper-based assays to quantitate total protein: the biuret method, a variation of the Lowry method (Hartree-Lowry method), and the bicinchoninic acid (BCA) assay. Acid hydrolysis of a protein is coupled with ninhydrin detection to quantitate amino acid content of a sample.

Is BCA good for Career?

Career Prospects BCA graduates have good job prospects both in the government and private sector companies. After successfully passing their BCA course, students can easily find lucrative job opportunities in leading IT companies like Oracle, IBM, Infosys and Wipro.

What are subject in BCA?

Prominent BCA Course Subjects

• Fundamentals of Computers.
• C Programming.
• Operating Systems.
• Multimedia Systems.
• Understanding Organisational Behaviour.
• Data and Database Management Systems.
• Web-Based Application Development.
• Computer Lab and Practical Work.

What is the best protein assay?

Top 5 Protein Quantification Assays

• Bicinchoninic Acid (BCA) This colorimetric, two-step assay was originally developed in 1985 – making it a baby compared with the 64-year-old Lowry assay!
• Bradford.
• Folin-Lowry.
• Kjeldahl.
• Ultraviolet Absorption.

What is the first step for protein assay?

The first step involves formation of a copper-protein complex in alkaline solution. This complex then reduces a phosphomolybdic-phosphotungstate reagent to yield an intense blue color. This assay is much more sensitive than the biuret method but is also more time consuming.

What is the full form of BCA assay?

Bicinchoninic acid (BCA) assay or Smith assay is a copper-based colorimetric assay for total protein quantification.

How does BCA assay work?

BCA Assay. How it works: The BCA assay is another colorimetric assay like the Bradford assay. It makes use of the biuret reaction, in which the protein backbone chelates Cu2+ ions and reduces them to Cu1+ ions. The Cu1+ ions then react with bicinchoninic acid (BCA) to form a purple-colored product that absorbs at 562 nm.

What is a BCA test?

The biuret test (Piotrowski’s test) is a chemical test used for detecting the presence of peptide bonds. In the presence of peptides, a copper(II) ion forms mauve-colored coordination complexes in an alkaline solution. Several variants on the test have been developed, such as the BCA test and the Modified Lowry test.

What is a recombinant immunoblot assay?

The Recombinant ImmunoBlot Assay (RIBA) is a blood test that detects antibodies to the hepatitis C virus (HCV). It was used for many years as a secondary confirmation test if a first-line screening test for HCV (called the ELISA hepatitis C antibody test) came back positive or indeterminant.