Lowry protein assay

The Lowry protein assay is a biochemical assay for determining the total level of protein in a solution. The total protein concentration is exhibited by a color change of the sample solution in proportion to protein concentration, which can then be measured using colorimetric techniques. It is named for the biochemist Oliver H. Lowry who developed the reagent in the 1940s. His 1951 paper describing the technique is the most-highly cited paper ever in the scientific literature, cited over 300,000 times.[1][2][3]

Mechanism

The method combines the reactions of copper ions with the peptide bonds under alkaline conditions (the Biuret test) with the oxidation of aromatic protein residues. The Lowry method is best used with protein concentrations of 0.01–1.0 mg/mL and is based on the reaction of Cu+, produced by the oxidation of peptide bonds, with Folin–Ciocalteu reagent (a mixture of phosphotungstic acid and phosphomolybdic acid in the Folin–Ciocalteu reaction). The reaction mechanism is not well understood, but involves reduction of the Folin–Ciocalteu reagent and oxidation of aromatic residues (mainly tryptophan, also tyrosine). Experiments have shown that cysteine is also reactive to the reagent. Therefore, cysteine residues in protein probably also contribute to the absorbance seen in the Lowry Assay.[4] The concentration of the reduced Folin reagent is measured by absorbance at 750 nm.[5] As a result, the total concentration of protein in the sample can be deduced from the concentration of Trp and Tyr residues that reduce the Folin–Ciocalteu reagent.

The method was first proposed by Lowry in 1951. The Bicinchoninic acid assay and the Hartree–Lowry assay are subsequent modifications of the original Lowry procedure.

See also

References

  1. ↑ "The top 100 papers — Nature".
  2. ↑ "The Most Highly Cited Paper in Publishing History: Protein Determination by Oliver H. Lowry — JBC".
  3. ↑ Garfield, E. (1990). "The Most-Cited Papers of All Time, SCI 1945-1988. Part 1A. The SCI Top 100 — Will the Lowry Method Ever Be Obliterated?" (PDF). Current contents. 7: 3–14. Retrieved 2011-02-12.
  4. ↑ Everette, Jace D.; Bryant, Quinton M.; Green, Ashlee M.; Abbey, Yvonne A.; Wangila, Grant W.; Walker, Richard B. (2010). "Thorough Study of Reactivity of Various Compound Classes toward the Folin−Ciocalteu Reagent". J. Agric. Food Chem. 58 (14): 8139. doi:10.1021/jf1005935. PMID 20583841.
  5. ↑ Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (November 1951). "Protein measurement with the Folin phenol reagent". J. Biol. Chem. 193 (1): 265–75. PMID 14907713.
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