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Method for testing activity of 5'-nucleotidase, and diagnosis kit for 5'-nucleotidase

A diagnostic kit and nucleotidase technology, which is applied in the field of 5′-nucleotidase diagnostic kits to measure 5′-nucleotidase activity, and can solve the problems of unsuitable promotion and application, strong acid pollution, and isotope pollution and other issues to achieve the effect of low test cost, accurate test results and high specificity

Inactive Publication Date: 2006-04-19
王尔中
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The isotope substrate method is complicated and has isotope pollution, and requires an isotope analyzer, so it is difficult to practically popularize and apply
Inorganic phosphorus determination method is a method invented in 1925, the accuracy is not good, strong acid pollutes the environment, etc., it is not suitable for popularization and application

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0075] Embodiment one (single dose)

[0076] The 5'-nucleotidase diagnostic kit of the present embodiment comprises:

[0077] Buffer 80mmol / l

[0078] Uridine monophosphate 1mmol / l

[0079] Pyruvate 1mmol / l

[0080] Phosphoenolpyruvate 1mmol / l

[0081] Reduced coenzyme 0.2mmol / l

[0082] Pyruvate oxidase 6000U / l

[0083] Phosphoenolpyruvate Carboxylase 6000U / l

[0084] Malate dehydrogenase 6000U / l

[0085] Stabilizer 50% (total volume)

[0086] Set on the automatic biochemical analyzer: temperature 37°C, reaction time 10 minutes, test main wavelength 340nm, test sub-wavelength above 405nm, the volume ratio of the tested 5'-nucleotidase sample to the reagent is 1 / 25, The reaction direction is positive reaction, the delay time is 1 minute, the detection time is 2 minutes, and the theoretical K value is -4180.

[0087] After the sample and reagents are added and allowed to mix, the following reactions occur:

[0088] Uridine monophosphate + water 5 nucleotidase Uri...

Embodiment 2

[0096] Embodiment two (two doses)

[0097] The 5'-nucleotidase diagnostic reagent of the present embodiment has:

[0098] Reagent I

[0099] Buffer 100mmol / l

[0100] Pyruvate 6mmol / l

[0101] Phosphoenolpyruvate 5mmol / l

[0102] Reduced coenzyme 0.25mmol / l

[0103] Pyruvate oxidase 13000U / l

[0104] Phosphoenolpyruvate Carboxylase 13000U / l

[0105] Malate dehydrogenase 13000U / l

[0106] Stabilizer 50% (total volume)

[0107] Reagent II

[0108] Buffer 100mmol / l

[0109] Uridine monophosphate 3mmol / l

[0110] Stabilizer 30mmol / l

[0111] When measuring 5′-nucleotidase activity, the temperature is controlled at 30°C, the reaction time is 15 minutes, the main wavelength of the test is 340nm, the secondary wavelength of the test is above 405nm, and the volume ratio of the tested 5′-nucleotidase sample to the reagent is 1 / 25, the reaction direction is negative reaction, the delay time is 1 minute, the detection time is 2 minutes, and the theoretical K value is -4180. ...

Embodiment 3

[0121] Embodiment three (three doses)

[0122] The 5′-nucleotidase diagnostic reagent of the present embodiment is three doses, including:

[0123] Reagent I

[0124] Buffer 120mmol / l

[0125]Pyruvate 10mmol / l

[0126] Phosphoenolpyruvate 8mmol / l

[0127] Reduced coenzyme 0.3mmol / l

[0128] Stabilizer 50mmol / l

[0129] Reagent II

[0130] Buffer 120mmol / l

[0131] Pyruvate oxidase 20000U / l

[0132] Phosphoenolpyruvate Carboxylase 20000U / l

[0133] Malate dehydrogenase 20000U / l

[0134] Stabilizer 50% (total volume)

[0135] Reagent III

[0136] Buffer 120mmol / l

[0137] Uridine monophosphate 5mmol / l

[0138] Stabilizer 50mmol / l

[0139] Set on the automatic biochemical analyzer: temperature 25°C, reaction time 20 minutes, test main wavelength 340nm, test sub-wavelength above 405nm, the volume ratio of the tested 5'-nucleotidase sample to the reagent is 1 / 25, The reaction direction is negative reaction, the delay time is 1 minute, the detection time is 2 minutes,...

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Abstract

A reagent kit for diagnosing 5í»-nucleotidase is composed of buffer liquid, uridine monophosphate, pyruvic acid, phosphoenolpyruvic acid, reduced coenzyme, phosphoneolphruvate carboxylase, malate dehydrogenase and stabilizer. A process for measuring the activity of 5í»-nucleotidase includes such steps as proportionally mixing specimen with reagents, enzyme coupling reaction and biochemically analyzing the final resultant to determine the variation in light absorptivity of master wavelength and in turn the activity of 5í»-nucleotidase. Its advantages are high sensitivity and precision, and no pollution.

Description

technical field [0001] The invention relates to a method for measuring 5'-nucleotidase activity, and meanwhile, the invention also relates to a 5'-nucleotidase diagnostic kit for realizing the method, which belongs to the technical field of medical examination and determination. Background technique [0002] Medical research shows that the increase of 5'-nucleotidase (5NT) is mainly seen in obstructive jaundice, and can also be seen in liver cancer and hepatitis. In cholestasis complicated by cholangitis, primary and secondary biliary cirrhosis, and chronic hepatitis, the rate of 5NT elevation is higher than that of alkaline phosphatase; in liver tumors and hepatic granulomas, the sensitivity of 5NT elevation is higher than that of alkali sexual phosphatase. Because there is no physiological increase in 5'-nucleotidase activity, it is more sensitive and specific than alkaline phosphatase for the diagnosis of infantile liver disease and hepatic cholestasis of pregnancy. The...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/32
Inventor 王尔中
Owner 王尔中
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