Abstract A novel substrate solution, dye-coupling composition and method for determining gamma-glutamyl transpeptidase in biological fluids have been developed. The substrate solution is an aqueous buffered solution containing an L-gamma-glutamyl-p-nitroanilide substrate, glycylglycine, sodium nitrite, and optionally, magnesium chloride hexahydrate. The dye-coupling composition for use in conjunction with the substrate solution comprises a mineral acid reagent solution of 8-hydroxy quinoline sulfate containing a water soluble, nonionic emulsifier, adjusted to a pH of between 0.8 and 1.3. In the determination of gamma-glutamyl transpeptidase, the biological test fluid is incubated with the substrate solution for a predetermined period of time; when incubation is completed, the dye-coupling composition is added to initiate the dye-forming reaction; an alkaline solution which will impart a pH of between 10.5 and 12 is then added to promote color development. The final alkaline solution optionally may contain ethylenediaminetetraacetic acid, tetrasodium salt. The optical density of the total reaction solution is a measure of gamma-glutamyltranspeptidase concentration. The method of the invention is accurate and approximately 4.5 times more sensitive than the standard test method currently being used.
A DIAGNOSTIC DYE-COUPLING COMPOSITION FOR THE DETERMINATION OF GAMMA-GLUTAMYL TRANSPEPTIDASE CONCENTRATIONS IN BIOLOGICAL FLUIDS COMPRISING AN AQUEOUS MINERAL ACID SOLUTION CONTAINING: A. FROM ABOUT 11.8 TO ABOUT 29.5 MICROMOLES OF 8-HYDROXY QUINOLINE SULFATE PER MILLILITER OF BIOLOGICAL FLUID; AND B. FROM ABOUT 1% TO ABOUT 5% BY VOLUME, BASED ON THE TOTAL VOLUME OF THE MINERAL ACID SOLUTION, OF A WATER SOLUBLE NONIONIC EMULSIFIER; SAID MINERAL ACID SOLUTION BEING ADJUSTED TO A PH OF FROM ABOUT 0.8 TO ABOUT 1.3.
Claim 2
Dependent Claim
80 words
A DIAGNOSTIC DYE-COUPLING COMPOSITION FOR THE DETERMINATION OF GAMMA-GLUTAMYL TRANSPEPTIDASE CONCENTRATIONS IN BIOLOGICAL FLUIDS COMPRISING AN AQUEOUS MINERAL ACID SOLUTION CONTAINING: A. FROM ABOUT 11.8 TO ABOUT 29.5 MICROMOLES OF 8-HYDROXY QUINOLINE SULFATE PER MILLILITER OF BIOLOGICAL FLUID; AND B. FROM ABOUT 1% TO ABOUT 5% BY VOLUME, BASED ON THE TOTAL VOLUME OF THE MINERAL ACID SOLUTION, OF A WATER SOLUBLE NONIONIC EMULSIFIER; SAID MINERAL ACID SOLUTION BEING ADJUSTED TO A PH OF FROM ABOUT 0.8 TO ABOUT 1.3.
Claim 3
Dependent Claim
33 words
A diagnostic dye-coupling composition according to claim 1 wherein the mineral acid solution contains about 14.8 micromoles of 8-hydroxy quinoline sulfate per milliliter of biological fluid and has a pH of about 1.1.
Claim 4
Dependent Claim
38 words
A diagnostic dye-coupling composition according to claim 2 wherein the mineral acid solution contains about 1% by volume, based on the total volume of the mineral acid solution, of ethoxylated tridecyl alcohol as the water soluble nonionic emulsifier.
Claim 5
Dependent Claim
17 words
A diagnostic dye-coupling composition according to claim 1 wherein the mineral acid in an aqueous hydrochloric acid.