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Method for measuring 5'-nucleotidase activity and diagnostic reagent kit of 5'-nucleotidase

A diagnostic kit and nucleotidase technology, which is applied in the field of 5′-nucleotidase diagnostic kits to measure the activity of 5′-nucleotidase, which can solve the problems that are not suitable for promotion and application, pollute the environment with strong acid, and are difficult to promote. Application and other issues, to achieve the effect of low test cost, accurate test results, and easy promotion and application

Inactive Publication Date: 2006-03-22
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] The present invention will be further described below in conjunction with examples of implementation. Embodiment one (single dose)

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

[0077] Buffer 80mmol / l

[0078] Adenosine 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 negative 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 allow...

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] Adenosine 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] Adenosine 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 minut...

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Abstract

The present invention relates to 5í»-nucleotidase activity detecting method and 5í»-nucleotidase diagnosing reagent kit in medical detecting technology. The 5í»-nucleotidase diagnosing reagent kit includes buffering liquid, adenosine monophosphate, pyruvic acid, phosphoenolpyruvic acid, reduction type coenzyme, pyruvic oxidase, phosphoenolpyruvate carboxylase, malate dehydrogenase and stabilizer. Through mixing the sample and reagent in certain volume ratio to produce enzyme coupling reaction, and detecting in biochemical analyzer the main wavelength absorbency change speed, the activity of 5í»-nucleotidase is calculated. The present invention can obtain the measurement result in biochemical analyzer in high sensitivity, high precision and no contamination of various foreign and internal matters.

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. ...

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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