Quantitative peptide or protein assay
AssigneePierce Biotechnology, Inc.
InventorsPaul Jeffrey Haney, Christopher L. Etienne, Sijian Hou, Erum Raja, Ramesh Ganapathy
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Abstract
Peptide and/or protein quantitation methods, kits, and compositions, particularly useful for mass spectrometry, are provided herein based on a bathocuproine-based composition complex such as bathocuproinedisulfonic acid disodium salt hydrate complex. The methods are one-step rapid absorbance methods using small sample volumes. They produce a robust signal with high signal to background ratio and accurately quantitate even complex peptide mixtures with low variability and high sensitivity.
Claims29
A direct method for determining peptide or protein concentration in a sample, the method for determining peptide or protein concentration comprising
- (a) combining the sample with a quantitation assay reagent composition comprising copper sulfate, acetonitrile present at a concentration ranging from about 10% to about 50%, and a complex to form a mixture, wherein the complex contains the formula:
- wherein
- each of R1 and R2 is independently chosen from C1-C6 straight and branched alkyls, and C6-C20 aryl, alkylaryl, and arylalkyls;
- each of R3 and R4 is independently chosen from the group consisting of hydrogen;
- a sulfonate salt chosen from a salt of sodium, potassium, and lithium;
- a phosphonate salt chosen from a salt of sodium, potassium, and lithium;
- and a carboxylate salt chosen from a salt of sodium, potassium, and lithium;
- and
- each of R5 and R6 is
- wherein R7 is independently chosen from the group consisting of hydrogen;
- sulfonate salts chosen from sodium, potassium, and lithium;
- phosphonate salts chosen from sodium, potassium, and lithium;
- and carboxylate salts chosen from sodium, potassium, and lithium;
- with the proviso that at least one of R3, R4, and R7 is not H;
- (b) incubating the mixture under conditions sufficient to form a colored complex;
- and
- (c) measuring absorbance of the colored complex at a wavelength ranging from 450 nm to 500 nm, wherein the amount of colored complex measured is directly proportional to peptide or protein concentration in the sample.
A direct method for determining peptide or protein concentration in a sample, the method for determining peptide or protein concentration comprising
- (a) combining the sample with a quantitation assay reagent composition comprising copper sulfate, acetonitrile present at a concentration ranging from about 10% to about 50%, and a complex to form a mixture, wherein the complex contains bathocuproinedisulfonic acid disodium salt hydrate:
- (b) incubating the mixture under conditions sufficient to form a colored complex;
- and
- (c) measuring absorbance of the colored complex at a wavelength ranging from 450 nm to 500 nm, wherein the amount of colored complex measured is directly proportional to peptide or protein concentration in the sample.
A direct method for determining peptide or protein concentration in a sample, the method comprising
- (a) combining the sample with a quantitation assay reagent composition comprising
- 50% acetonitrile and
- a complex containing bathocuproinedisulfonic acid disodium salt hydrate:
- (b) incubating the mixture under conditions sufficient to form a colored complex;
- and
- (c) measuring absorbance of the colored complex at a wavelength ranging from 450 nm to 500 nm, wherein the amount of colored complex measured is directly proportional to peptide or protein concentration in the sample.
Dependencies are inferred from the claim text.