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Method for detecting dihyaropteridine reductase activity, detection kit and application thereof

A technology of dihydropteridine and reductase, which is applied to the measurement of color/spectral characteristics, etc., can solve the problems of unclear data segment selection, unstable detection, and large sample consumption, so as to achieve reduced sample consumption, stable reaction, heavy weight, etc. performance improvement effect

Pending Publication Date: 2017-10-20
GUANGZHOU KINGMED DIAGNOSTICS CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] However, our improved UV spectrophotometry can solve the problems of large sample consumption, long measurement time, unstable detection, and unclear selection of data segments in the current method, and can greatly improve the stability of sample detection results, quantitative accuracy and Sample detection throughput, more suitable for clinical practical detection applications

Method used

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  • Method for detecting dihyaropteridine reductase activity, detection kit and application thereof
  • Method for detecting dihyaropteridine reductase activity, detection kit and application thereof
  • Method for detecting dihyaropteridine reductase activity, detection kit and application thereof

Examples

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Embodiment 1

[0062] The embodiment of the present invention provides a method for detecting the activity of erythrocyte dihydropteridine reductase (DHPR), comprising the following steps:

[0063] (1) Pretreatment of samples:

[0064] Punch two blood spots with a diameter of 3mm from the dried blood paper and place them in EP tubes, add 0.2ml of 0.15mol / L KCl solution, shake for 60s, then centrifuge at 14000r / min for 3min, and quickly transfer all the solutions to In another clean EP tube, the sample solution obtained in step (1) is prepared for testing; through the above pretreatment, the red blood cell dihydropteridine reductase in the dried blood paper is extracted and transferred to ensure the extraction efficiency Consistent to ensure the quantitative accuracy of the method.

[0065] (2) Detection of erythrocyte dihydropteridine reductase activity:

[0066] Take 2595 μl of 50mmol / L Tris-HCl buffer solution in EP tube and place it in a beaker filled with 37°C water, put it into a 37°C...

Embodiment 2

[0073] In order to further illustrate the beneficial effects of the present invention, repeat the steps of the embodiment of the present invention 1, the only difference between this implementation and the steps of the embodiment of the present invention 1 is that the step (2) of the embodiment of the present invention 1 needs to carry out "immediately and rapidly upside down timing mixing Homogenize for 15s", but the operation of "immediately and quickly upside down and timed mixing for 15S" was not performed in the step (2) of the present embodiment 2, the measurement results obtained in the present embodiment 2 can be found in figure 2 , the slope plot fluctuates widely and has almost no slope.

Embodiment 3

[0075] In order to further illustrate the beneficial effects of the present invention, the steps of Example 1 are repeated, and in the step (2) of Example 1, "the used immediately and quickly upside down and timed mixing for 15s" is replaced by "the used immediately and quickly upside down and timed mixing for 10s" , "immediately and quickly upside down and timed mixing for 30s" to obtain the measurement results see image 3 , where curves a, b, and c are the mixing times of 10, 15, and 30 s respectively. It can be seen that the slope of the slope graph is too slow if the mixing time is too long, and the mixing time is too short, which is not conducive to operation and uneven mixing .

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Abstract

The invention provides a method for detecting dihyaropteridine reductase activity, a detection kit and application thereof. According to the method for detecting dihyaropteridine reductase activity, provided by the invention, the pretreatment step is simple, the sample usage amount is reduced, a 5mm blood stain is changed into two 3mm blood stains, and the actual clinical application is promoted; due to advanced split charging and preheating of the buffer solution, the detection time of each sample is shortened to 10min, and the detection throughput is greatly improved; due to the improved sample pretreatment in the method provided by the invention, the method is stable, the enzymatic activity curve is smooth, the RSD (Relative Standard Deviation) of within-run precision of quantitative results is less than or equal to 3.37%, the RSD of between-run precision is less than or equal to 7.82%, the reproducibility and accuracy of quantification are greatly improved, the detection time is short, the detection throughput is improved, and the detection requirements on clinical actual samples are met.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and in particular relates to a method for detecting the activity of erythrocyte dihydropteridine reductase, a detection kit and an application thereof. Background technique [0002] Hyperphenylalaninemia (hyperphenylalaninemia, HPA) refers to the blood phenylalanine concentration> 120mmol / L (> 2mg / dL), is the most common congenital, autosomal recessive amino acid metabolism disease. Phenylalanine (phenylalamine, PA) is an essential amino acid for the human body, which participates in the formation of various protein components, but cannot be synthesized in the human body. Under normal circumstances, about 50% of the ingested PA is used to synthesize proteins of various components, and the rest is converted into tyrosine under the action of phenylalanine hydroxylase (PAH), and then processed by other enzymes. The role is transformed into dopa, dopamine, epinephrine, norepinephr...

Claims

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Application Information

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IPC IPC(8): G01N21/31
CPCG01N21/31
Inventor 赵蓓蓓陈智李维佘旭辉刘伟霞程雅婷李卓阳陈静宜董衡沙小花汪慧
Owner GUANGZHOU KINGMED DIAGNOSTICS CENT
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