What are the common total protein quantitation assays?

One common total protein quantitation assay involves the measurement of UV-Vis absorbance at 280 nm. This method estimates protein concentration by measuring the absorbance of aromatic residues like tyrosine and tryptophan at 280 nm using a UV-Vis spectrometer. Another type is the Bicinchoninic Acid Assay (BCA). This two-step colorimetric assay first involves the biuret reaction, where proteins reduce Cu²⁺ to Cu⁺. Then, Cu⁺ reacts with BCA to form a deep purple complex, which absorbs at 562 nm. A third assay is the Bradford Assay. This colorimetric assay uses Coomassie Brilliant Blue dye, which binds to arginine and aromatic residues in proteins. The dye’s absorbance shifts from 470 nm to 595 nm when it binds to proteins. Another type is the Folin-Lowry Assay. This method involves two steps. Proteins first complex with copper, then tyrosine and tryptophan react with the Folin–Ciocalteu reagent to produce a blue-green color that absorbs between 650-750 nm. Another type is the 3-(4-carboxybenzyl)quinoline-2-carboxaldehyde assay (CBQCA) , which is a highly sensitive method with a range of 100 ng to 1500 μg/mL for detecting proteins. This assay uses the fluorogenic reagent CBQCA, which reacts with amines in proteins to produce a detectable signal. Lastly, the Kjeldahl method is used to determine protein content by measuring the nitrogen in a sample. First, the protein sample is broken down using heated sulfuric acid, which turns the nitrogen in the sample into ammonia. Next, the ammonia becomes separated through steam distillation. After that, a chemical reaction with sodium hydroxide is used to measure the amount of ammonia that was released. From this measurement, one can estimate the total amount of protein, based on the assumption that proteins are about 16% nitrogen.