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Determination of 5'-nucleotidase activity and its diagnostic reagent kit of 5'-nucleotidase

A diagnostic kit and nucleotidase technology, applied in the field of determination of 5′-nucleotidase activity, 5′-nucleotidase diagnostic kit, can solve the problems of strong acid pollution, isotope pollution, complex methods, etc.

Inactive Publication Date: 2006-05-31
SUZHOU ANJ BIOTECHNOLOGY CO LTD
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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

[0076] Embodiment one (single agent)

[0077] The 5'-nucleotidase diagnostic kit of the present embodiment comprises:

[0078] Buffer 80mmol / l

[0079] Inosine monophosphate 1mmol / l

[0080] Adenosine triphosphate 2mmol / l

[0081] Phosphoenolpyruvate 2mmol / l

[0082] Reduced coenzyme 0.2mmol / l

[0083] Inosine kinase 4000U / l

[0084] Pyruvate kinase 6000U / l

[0085] Lactate dehydrogenase 6000U / l

[0086] Stabilizer 50% (total volume)

[0087] 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.

[0088] After the sample and reagents are added and allowed to mix, the following reactions occur:

[0089] Inosine monophosphate + water 5′-nucleotidase Inosin...

Embodiment 2

[0096] Embodiment two (two doses)

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

[0098] Buffer 100mmol / l

[0099] Adenosine triphosphate 5mmol / l

[0100] Phosphoenolpyruvate 5mmol / l

[0101] Reduced coenzyme 0.25mmol / l

[0102] Inosine kinase 7000U / l

[0103] Pyruvate kinase 8000U / l

[0104] Lactate dehydrogenase 13000U / l

[0105] Stabilizer 50% (total volume)

[0106] Reagent II

[0107] Buffer 100mmol / l

[0108] Inosine monophosphate 6mmol / l

[0109] Stabilizer 20mmol / l

[0110] 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.

[0111] The specific me...

Embodiment 3

[0119] Embodiment three (three doses)

[0120] The 5′-nucleotidase diagnostic reagent of the present embodiment is three doses, including:

[0121] Reagent I

[0122] Buffer 120mmol / l

[0123] Adenosine triphosphate 8mmol / l

[0124] Phosphoenolpyruvate 8mmol / l

[0125] Reduced coenzyme 0.3mmol / l

[0126] Stabilizer 20mmol / l

[0127] Reagent II

[0128] Buffer 120mmol / l

[0129] Inosine kinase 10000U / l

[0130] Pyruvate Kinase 10000U / l

[0131] Lactate dehydrogenase 20000U / l

[0132] Stabilizer 50% (total volume)

[0133] Reagent III

[0134] Buffer 120mmol / l

[0135] Inosine monophosphate 10mmol / l

[0136] Stabilizer 20mmol / l

[0137] 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, a...

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Abstract

The invention is about a method of measuring the activation of 5í»-phosphonuclease, and it also concerns the reagent box of 5í»-phosphonucleasediagnosis. This invention belongs to the field of medical testing and measuring technology. The reagent box is consisted of buffer solution, carnine monophosphate, adenosine triphosphate, phosphoenolpyruvate, reduced coenzyme, carnine kinase, pyruvate kinase, lactate dehydrogenase and stabilizer. Firstly, we cause an enzyme-coupled reaction through mixing the sample and the reagent according to a certain proportion of volume; secondly, put the final reactant under the biochemical analyzer and test the absorbance variational situation (speed) of dominant wavelength; then we can get theactivation of 5í»-phosphonuclease. By using this invention, we can get the necessary measuring result with high sensitiveness and fine precision through biochemical analyzer, and the result would not be contaminated by material of internal and exogenous sources. Thus, this method can be conveniently promoted and applied.

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): C12Q1/48
Inventor 王尔中
Owner SUZHOU ANJ BIOTECHNOLOGY CO LTD
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