What are the common methods for determining protein concentration?

Several methods can be used to determine protein concentration these include: uv absorption, bicinchoninic acid assay (BCA), Bradford assay and fluorescent dye-based methods.

• UV absorption: protein concentration is determined measuring the absorbance at 280 nm. While this method is simple in design, it is highly prone to error due to non-protein components (e.g. nucleic acids) that also absorb at 280 nm.
• Bicinchoninic acid assay (BCA): protein concentration is determined based on the conversion of Cu2+ to Cu1+ by protein under alkaline conditions. Bicinchoninic acid reacts with Cu1+ to produce a colorimetric signal that absorbs light at 540 nm. The amount of color produced is directly proportional to the amount of protein present.
• Bradford assay: protein concentration is determined using Coomassie Brilliant Blue. Under acidic conditions, proteins bind to Coomassie Brilliant Blue resulting in an absorbance shift of the dye from 465 nm (reddish brown) to 595 nm (blue). The increase of absorbance at 595 nm is directly proportional to the amount of protein present in the sample.
• Fluorescent-dye based methods: protein concentration is determined using a fluorescent dye that reacts rapidly with proteins and generates a bright fluorescence signal. The amount of fluorescence produced is directly proportional to the amount of protein present and can be monitored using a fluorescent microplate reader or fluorimeter.