QUANTITATIVE PEPTIDE OR PROTEIN ASSAY
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.
1 - 23 . (canceled)
24 . A kit for protein or peptide quantitation comprising:
(a) a complex having the formula:
wherein
each of R 1 and R 2 is independently chosen from C 1 -C 6 straight and branched alkyls, C 6 -C 20 aryl, alkylaryl, and arylalkyls;
each of R 3 and R 4 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; and
each of R 5 and R 6 is
wherein R 7 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 R 3 , R 4 , and R 7 is not H;
(b) acetonitrile; and
(c) copper sulfate.
25 . The kit of claim 24 , wherein the complex is present at a concentration ranging from about 0.04 M to about 0.1 M.
26 . The kit of claim 24 , wherein each of R1 and R2 is independently chosen from methyl, ethyl, propyl, butyl, and phenyl.
27 . The kit of claim 24 , wherein the complex has the formula:
28 . The kit of claim 24 , wherein the acetonitrile is present at a concentration ranging from about 10% to about 50%.
29 . The kit of claim 28 , wherein the acetonitrile is present at a concentration ranging from about 10% to about 30%.
30 . The kit of claim 24 , wherein the copper sulfate is present at a concentration ranging from about 0.25 mM to about 0.5 mM.
31 . The kit of claim 24 , further comprising tartrate.
32 . The kit of claim 31 , wherein the tartrate is present at a concentration ranging from about 5.7 mM to about 22.7 mM.
33 . The kit of claim 31 , where the tartrate is sodium tartarate.
34 . The kit of claim 24 , further comprising sodium bicarbonate.
35 . A composition for protein or peptide quantitation comprising:
(a) a complex having the following general formula:
wherein
each of R 1 and R 2 is independently chosen from C 1 -C 6 straight and branched alkyls, C 6 -C 20 aryl, alkylaryl, and arylalkyls;
each of R 3 and R 4 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; and
each of R 5 and R 6 is
where R 7 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 R 3 , R 4 , and R 7 is not H;
(b) acetonitrile; and
(c) copper sulfate.
36 . The composition of claim 35 , wherein the complex is present at a concentration ranging from about 0.04 M to about 0.1 M.
37 . The kit of claim 35 , wherein each of R 1 and R 2 is independently chosen from methyl, ethyl, propyl, butyl, and phenyl.
38 . The composition of claim 35 , wherein the complex has the formula:
39 . The composition of claim 35 , wherein the acetonitrile is present at a concentration ranging from about 10% to about 50%.
40 . The composition of claim 39 , wherein the acetonitrile is present at a concentration ranging from about 10% to about 30%.
41 . The composition of claim 35 , wherein the copper sulfate is present at a concentration ranging from about 0.25 mM to about 0.5 mM.
42 . The composition of claim 35 , further comprising tartrate.
43 . The composition of claim 42 , wherein the tartrate is present at a concentration ranging from about 5.7 mM to about 22.7 mM.
44 . The composition of claim 42 , where the tartrate is sodium tartrate.
45 . The composition of claim 34 , further comprising sodium bicarbonate.
46 . The composition of claim 35 , wherein the pH ranges from about 12 to about 13.
47 . A kit for protein or peptide quantitation comprising:
(a) a complex having the formula:
(b) acetonitrile; and
(c) copper sulfate.