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A kind of 5'-nucleotidase detection kit and its preparation method

A technology for detecting kits and nucleotidases, which is applied in the direction of biochemical equipment and methods, and the determination/inspection of microorganisms, and can solve the problems of low precision and stability of 5'-nucleotidase detection kits

Active Publication Date: 2019-05-14
JILIN GETEIN BIOTECH CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] The purpose of the present invention is to provide a 5'-nucleotidase detection kit and a preparation method thereof in order to solve the problems of low precision and stability of the existing 5'-nucleotidase detection kits

Method used

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  • A kind of 5'-nucleotidase detection kit and its preparation method
  • A kind of 5'-nucleotidase detection kit and its preparation method
  • A kind of 5'-nucleotidase detection kit and its preparation method

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preparation example Construction

[0034] The present invention also provides a method for preparing a 5'-nucleotidase detection kit, including:

[0035] Step 1: Preparation of reagent R1:

[0036] 1. Add deionized water to the reaction vessel first, then add buffer I, stir to dissolve, then add potassium dihydrogen phosphate, EDTA·2K, 4-aminoantipyrine, Proclin300 and AES in sequence, stir to dissolve completely, preferably Adjust the pH value of the solution to 7.0-7.7 with an alkali solution at 20°C, the alkali solution is preferably KOH, and the concentration of KOH is preferably 4M to obtain a mixed solution;

[0037] 2. Add A90 to the mixed solution obtained in step 1. After there are no insoluble particles in the solution, add MgCl2·6H2O and stir and mix, then use a pipette to add TritonX-305 to the above reaction vessel, stir and mix, and finally Add purine nucleoside phosphorylase, xanthine oxidase and peroxidase, dilute the volume with deionized water, stir evenly to obtain reagent R1;

[0038] Step 2: Prepa...

Embodiment 1

[0043] Preparation of reagent R1: (1L)

[0044] 1) Measure 1L of deionized water with a graduated cylinder, add 0.9L of deionized water into the beaker, and save the remaining water;

[0045] 2) Weigh 40mmol of PIPES and 80mmol of KOH into the above beaker, stir with a glass rod until completely dissolved;

[0046] 3) Weigh 15mmol of potassium dihydrogen phosphate, 1mmol of EDTA·2K, 1mmol of 4-aminoantipyrine, 0.02ml of Proclin300, and 0.5g of AES into the large beaker and stir until completely dissolved;

[0047] 4) Adjust the pH of the above solution to 7.6 with 4M KOH at 20°C;

[0048] 5) Measure 0.1ml of A90 into the above beaker and stir and mix;

[0049] 6) After the solution in the above beaker has no insoluble particles, weigh 3mmol of MgCl 2 ·6H 2 Add O to the above beaker, gently stir and mix;

[0050] 7) Use a pipette to suck 0.1ml of TritonX-305 into the above beaker, gently stir and mix;

[0051] 8) Finally, add 0.2KU of purine nucleoside phosphorylase, 0.6KU of xanthine oxida...

Embodiment 2

[0060] preparation:

[0061] Preparation of reagent R1: (1L)

[0062] 1) Measure 1L of deionized water with a graduated cylinder, add 0.9L of deionized water into the beaker, and save the remaining water;

[0063] 2) Weigh 40mmol of KH 2 PO 4 , 70mmol KOH was added to the above beaker, stirred with a glass rod until completely dissolved;

[0064] 3) Weigh 2mmol of EDTA·2K, 2mmol of 4-aminoantipyrine, 0.02ml of Proclin300, and 1.0g of AES into the above-mentioned large beaker, and stir until completely dissolved;

[0065] 4) Adjust the pH of the above solution to 7.6 with 4M KOH at 20°C;

[0066] 5) Measure 0.2ml of A90 and add it to the above beaker, stir and mix;

[0067] 6) After the solution in the above beaker is free of insoluble particles, weigh 6mmol of MgCl 2 ·6H 2 Add O to the above beaker, gently stir and mix;

[0068] 7) Use a pipette to pipette 0.2ml of TritonX-305 into the above beaker, gently stir and mix;

[0069] 8) Finally, add 0.2KU of purine nucleoside phosphorylase, 0.6K...

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Abstract

The present invention provides a 5'-nucleotidase detection kit and a preparation method thereof, belongs to the field of in-vitro diagnostic reagents, and solves the low precision and stability problems of a 5'-nucleotidase detection kit in the prior art. The kit comprises a reagent R1 and a reagent R2. The present invention also provides the preparation method of the 5'-nucleotidase detection kit. The R1 reagent contains a compound stabilizer comprising TritonX-305 and AES, the compound stabilizer can improve the stability of the reagent, and also can improve the dispersion of the reagent, reduce the absorbance difference between two times of blank calibration, and improve the kit precision; A90 contained in the R1 can effectively prevent precipitation of magnesium phosphate, expensive 18-crown ether-6 is replaced, the stability of the reagent is improved, reagent blank absorbance change rate is reduced, and the cost is reduced greatly; by addition of glycine and EDTA. 2K into the reagent R2, detection sensitivity and precision of the kit can be improved, and by addition of the glycine, reagent blank absorbance is reduced.

Description

Technical field [0001] The invention relates to the field of in vitro diagnostic reagents, in particular to a 5'-nucleotidase detection kit and a preparation method thereof. Background technique [0002] The 5'-nucleotidase (5'-Nucleotidase, 5'-NT) assay is mainly used for the diagnosis of hepatobiliary system diseases and the differential diagnosis of bone diseases. The increase of serum 5'-NT activity is mainly seen in hepatobiliary diseases, such as obstructive jaundice, liver cancer, hepatitis, etc. Its activity can be increased by 2 to 6 times, and it is positively correlated with the severity of the disease, and its activity change is consistent with ALP. However, skeletal system diseases, such as tumor metastasis, osteitis deformity, rickets, hyperparathyroidism, etc., usually have elevated ALP activity while 5'-NT is normal. Therefore, the simultaneous determination of ALP and 5'-NT is helpful for the differential diagnosis of hepatobiliary and skeletal diseases. [0003]...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/48C12Q1/44C12Q1/28C12Q1/26
CPCC12Q1/26C12Q1/28C12Q1/44C12Q1/48C12Q2326/96
Inventor 苏恩本徐李鹏刘玲张晓杨
Owner JILIN GETEIN BIOTECH CO LTD
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