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Adenosine deaminase activity determination method and adenosine deaminase diagnosi kit

A technology of adenosine deaminase and diagnostic kit, which is applied in the field of determination of adenosine deaminase activity and adenosine deaminase diagnostic kit, can solve the problems of reduced test accuracy of reagents, urgent, inconvenient testing, etc.

Inactive Publication Date: 2006-05-10
王尔中
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

One of its disadvantages is that this system also offsets part of the adenosine deaminase reagent (adenosine deaminase coupled glutamate dehydrogenase reaction must Reduced nicotinamide coenzyme is oxidized to oxidized nicotinamide coenzyme), which greatly reduces the accuracy of the reagent test
The second disadvantage is that the reagent must be reacted for a period of time before the formal test in order to produce enough reduced nicotinamide coenzyme, which results in slow results and brings great inconvenience to the test

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0090] Embodiment one (single agent)

[0091] The adenosine deaminase diagnostic kit of the present embodiment comprises:

[0092] Buffer 80mmol / l

[0093] Adenosine 2mmol / l

[0094] Glutamic acid 5mmol / l

[0095] Reduced coenzyme 0.2mmol / l

[0096] Adenosine triphosphate 2mmol / l

[0097] Pyruvate 1mmol / l

[0098] Phosphoenolpyruvate 1mmol / l

[0099] Glutamine Synthetase 4000U / l

[0100] Pyruvate oxidase 6000U / l

[0101] Phosphoenolpyruvate Carboxylase 6000U / l

[0102] Malate dehydrogenase 6000U / l

[0103] Stabilizer 50% (total volume)

[0104] Set on the automatic biochemical analyzer: temperature 37°C, reaction time 10 minutes, test main wavelength 340nm, test sub-wavelength above 405nm, volume ratio of tested adenosine deaminase sample to reagent is 1 / 25, reaction direction It is a negative reaction, the delay time is 1 minute, the detection time is 2 minutes, and the theoretical K value is -4180.

[0105] After the sample and reagents are added and allowed to m...

Embodiment 2

[0114] Embodiment two (two doses)

[0115] The adenosine deaminase diagnostic reagent of the present embodiment has:

[0116] Reagent I

[0117] Buffer 100mmol / l

[0118] Glutamic acid 8mmol / l

[0119] Reduced coenzyme 0.25mmol / l

[0120] Adenosine triphosphate 5mmol / l

[0121] Pyruvate 6mmol / l

[0122] Phosphoenolpyruvate 5mmol / l

[0123] Glutamine Synthetase 7000U / l

[0124] Pyruvate oxidase 9000U / l

[0125] Phosphoenolpyruvate Carboxylase 9000U / l

[0126] Malate dehydrogenase 13000U / l

[0127] Stabilizer 50% (total volume)

[0128] Reagent II

[0129] Buffer 100mmol / l

[0130] Adenosine 6mmol / l

[0131] Stabilizer 20mmol / l

[0132] When measuring the activity of adenosine deaminase, the temperature is controlled at 30°C, the reaction time is 15 minutes, the main wavelength of the test is 340nm, and the secondary wavelength of the test is above 405nm. The volume ratio of the tested adenosine deaminase sample to the reagent is 1 / 25. The direction is negative re...

Embodiment 3

[0142] Embodiment three (three doses)

[0143] The adenosine deaminase diagnostic reagent of the present embodiment is three doses, has:

[0144] Reagent I

[0145] Buffer 120mmol / l

[0146] Glutamic acid 10mmol / l

[0147] Reduced coenzyme 0.3mmol / l

[0148] Adenosine triphosphate 8mmol / l

[0149] Pyruvate 10mmol / l

[0150] Phosphoenolpyruvate 8mmol / l

[0151] Stabilizer 20mmol / l

[0152] Reagent II

[0153] Buffer 120mmol / l

[0154] Glutamine Synthetase 10000U / l

[0155] Pyruvate oxidase 12000U / l

[0156] Phosphoenolpyruvate Carboxylase 12000U / l

[0157] Malate dehydrogenase 20000U / l

[0158] Stabilizer 50% (total volume)

[0159] Reagent III

[0160] Buffer 120mmol / l

[0161] Adenosine 10mmol / l

[0162] Stabilizer 20mmol / l

[0163] Set on the automatic biochemical analyzer: temperature 25°C, reaction time 20 minutes, test main wavelength 340nm, test sub-wavelength above 405nm, volume ratio of tested adenosine deaminase sample to reagent is 1 / 25, reaction dir...

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Abstract

The invention relates to a method for determining the activity of adenosine deaminase, and also the reagent kit for adenosine deaminase diagnosis. The reagent kit comprises cushioning solution, adenosine, glutacid, deacidized type coenzyme, adenosine triphosphate, pyroracemic acid, phosphoenolpyruvate phosphatase, glutamine synthetase, pyruvic oxidase, phosphoenolpyruvate pyruvate carboxylase, malic dehydrogenase, and stabilizer. By mixing sample and reagent of a predetermiend volumetric ratio, generating coupling reaction between them, subjecting the final reactant to biochemiscal analyser, the main wavelength absorbancy variance ratio (speed) can be detected, and the activity of the adenosine deaminase can thus be measured. The method of the invention can be used to obtain the needed measurement result purely through biochemical analytic instruments, and advantages of the method include higher sensibility, better accuracy, less susceptibility to contamination of internal or external materials, and easy application.

Description

technical field [0001] The invention relates to a method for measuring the activity of adenosine deaminase, and at the same time, the invention also relates to a diagnostic kit for adenosine deaminase for realizing the method, which belongs to the technical field of medical examination and determination. Background technique [0002] Medical research shows that adenosine deaminase is a nucleic acid metabolizing enzyme that has an important relationship with the body's cellular immune activity. Therefore, the determination of adenosine deaminase activity is used as an important diagnostic marker for the differential diagnosis of benign and malignant hydrothorax, cerebrospinal fluid, severe combined immunodeficiency disease (SCID), acute and chronic hepatitis, liver cirrhosis, liver cancer and other diseases. [0003] The methods for determining the activity of adenosine deaminase mainly include radionuclide method, physical method and biochemical method. According to the app...

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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