Question

The Bradford Assay is one of the most popular protein assays, but it is not the only one. Briefly describe any of the alternative assays that are available. If you were setting up your own lab, which would you prefer to use? Briefly state why.

Answer 1

Protein assays types:

1. Bradford assay (most popular one)

2. Lowry assay

Lowry protein assay includes the measuring of the protein concentration (0.01–1.0 mg/mL )by colourimetric methoids using Folin’ reagent which consist of copper with phospotungistic acid and phosphomolybdic acid.

In this assay sulfur containing amino acid, cysteine also has its reacting impact on the reacting agent.

This biochemical assay used to know the ultimate protein conc. in the given sample using absorbance studies at a particular wavelength. This test is very sensitive compared to the biuret test more precautions must be taken.

Bradford protein assay is considered as a major more advantageous than other lowry protein assays or biuret protein assays. It is spectroscopic method in which the amino acid concentration is main important measure of protein that need to be assessed. A dye called Coomessie brilliant blue G-250 is used in acidic conditions, the proteins reacts with the dye and changes the color from red to blue. The crucial interactions involved here are the breaking of the protein tertiary structure and finally result in the exposure of the hydrophobic groups. The vanderwall’s forces of attraction operates between the non-polar regions within the peptide by providing positive charge (zwitter ionic amino functional groups) to the negative end of the dye . Finally the amino acid with the dye formed a strong complex which produces a colour via absorbance phenomenon according Beer’s-Lambert’s law we can study the exact conch of protein.

Lowry assay is less sensitive to the proteins and more susceptible to other chemical within the solution, but Bradford assay is more sensitive and also possess low susceptibility to any other chemicals within the assay for examples detergents (Sodium dodecyl sulfate)