US3769173A
Expired / Lapsed
US
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Determination of gamma-glutamyl transpeptidase in biological fluids
Assignee: Warner Lambert Co LLC
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Filing: 1972-08-21
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Granted: 1973-10-30
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Expires: 1992-08-21
Abstract
Abstract A method for determining gamma-glutamyl transpeptidase in biological fluids is described which comprises incubating the test fluid with an aqueous buffered solution of an L-gammaglutamyl-p-nitroanilide substrate, glycylglycine, sodium nitrite, and optionally, magnesium chloride hexahydrate, for a predetermined period of time; adding, after incubation is completed, a mineral acid reagent solution of 8-hydroxy quinoline sulfate containing a water soluble, nonionic emulsifier, the mineral acid solution having a pH of between 0.8 and 1.3; adding an alkaline solution which will impart a pH of between 10.5 and 12, in the total reaction solution, said alkaline solution optionally containing ethylenediaminetetraacetic acid, tetrasodium salt; and reading the optical density of the total reaction solution as a measure of gamma-glutamyltranspeptidase concentration in the test sample of biological fluid. The method of the invention is accurate and approximately 4.5 times more sensitive than the standard test method currently being used.
Inventors
- 👤 J Carroll
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-03 04:57
CPC Classifications
C; C12; C12Q; C12Q1/00; C12Q1/48; C12Q1/34; C12Q1/37; C12Q2337/00; C12Q2337/10; C12Q2337/12
Claims Analysis
Total Claims: 10
Claim 1
Independent Claim
25 words
A method according to claim 1 wherein the substrate solution of (A) is buffered to a pH of about 8.1 with a (2-amino-2-methyl-propane-1,3-diol)-HCl buffer system.
Claim 2
Dependent Claim
33 words
A method according to claim 2 wherein the mineral acid solution of (C) contains about 14.8 micromoles of 8-hydroxy quinoline sulfate per milliliter of biological fluid and has a pH of about 1.1.
Claim 3
Dependent Claim
39 words
A method according to claim 3 wherein the mineral acid solution of (C) contains about 1 percent by volume, based on the total volume of the mineral acid solution of ethoxylated tridecyl alcohol as the water soluble nonionic emulsifier.
Claim 4
Dependent Claim
57 words
A method according to claim 1 wherein the substrate solution of (A) additionally contains at least 100 micromoles of magnesium chloride hexahydrate per milliliter of biological fluid, and wherein the alkaline solution of (D) additionally contains at least 6.5 micromoles of ethylenediaminetetraacetic acid, tetrasodium salt per micromole of magnesium chloride hexahydrate in the substrate solution of (A).
Claim 5
Dependent Claim
25 words
A method according to claim 5 wherein the substrate solution of (A) is buffered to a pH of about 8.1 with a (2-amino-2-methyl-propane-1,3-diol)-HCl buffer system.
Claim 6
Dependent Claim
33 words
A method according to claim 6 wherein the mineral acid solution of (C) contains about 14.8 micromoles of 8-hydroxy quinoline sulfate per milliliter of biological fluid and has a pH of about 1.1.
Claim 7
Dependent Claim
39 words
A method according to claim 7 wherein the mineral acid solution of (C) contains about 1 percent by volume, based on the total volume of the mineral acid solution, of ethoxylated tridecyl alcohol as the water soluble nonionic emulsifier.
Claim 8
Dependent Claim
256 words
A colorimetric method for the determination of gamma-glutamyl transpeptidase concentrations in biological fluids which comprises: A. Incubating the sample of the biological fluid with an aqueous substrate solution, said substrate solution comprising about 45 micromoles of L-gamma-glutamyl-p-nitroanilide per milliliter of biological fluid, about 400 micromoles of glycylglycine per milliliter of biological fluid, about 72.4 micromoles of sodium nitrite per milliliter of biological fluid, and about 100 micromoles of magnesium chloride hexahydrate per milliliter of biological fluid, in an aqueous solution buffered to maintain a pH of about 8.1 with a (2-amino-2-methyl-propane-1,3-diol)-HCl buffer system; B. Continuing the incubation of solution (A) for about ten minutes at about 37*C to release p-nitroaniline as a result of enzymatic activity; C. Adding to solution (B), at the end of the incubation time, an aqueous hydrochloric acid solution containing about 14.8 micromoles of 8-hydroxy quinoline sulfate per milliliter of biological fluid, and about 1 percent by volume, based on the total volume of the hydrochloric acid solution, of ethoxylated tridecyl alcohol, said solution having a pH of about 1.1; D. Allowing from about 15 seconds to about 10 minutes to elapse and adding to solution (C) a sufficient amount of an aqueous sodium hydroxide solution to provide a final pH of about 10.5 in the reaction solution, said sodium hydroxide solution containing about 6.5 micromoles of ethylenediaminetetraacetic acid, tetrasodium salt per micromole of magnesium chloride hexahydrate in the substrate solution A; and E. Reading the optical density of solution (D) as a measure of gamma-glutamyl transpeptidase concentration in the biological fluid.
Claim 9
Dependent Claim
27 words
A method according to claim 1 wherein the substrate solution of (A) is buffered to a pH of about 8.1 with a tris(hydroxy-methyl)amino methane hydrochloride buffer system.
Claim 10
Dependent Claim
27 words
A method according to claim 6 wherein the substrate solution of (A) is buffered to a pH of aBout 8.1 with a tris(hydroxy-methyl)amino methane hydrochloride buffer system.
References Cited (Backward Citations)
US3703441A
Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1976-10-29 | 1977-04-30 | 1977-10-30 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1980-10-29 | 1981-04-30 | 1981-10-30 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1984-10-29 | 1985-04-30 | 1985-10-30 | $7700.00 | Window Closed / Lapsed |