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Method and kit for investigating humotype semi-cystinol by enzyme biochemical reaction

A technology of homocysteine ​​and chemical reaction, applied in the field of determination of homocysteine ​​content

Inactive Publication Date: 2005-11-09
ZHEJIANG YAKE SCI & TECH +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0028] TCEP is more stable than DTT, but it is rarely used in the determination of HCY

Method used

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  • Method and kit for investigating humotype semi-cystinol by enzyme biochemical reaction
  • Method and kit for investigating humotype semi-cystinol by enzyme biochemical reaction
  • Method and kit for investigating humotype semi-cystinol by enzyme biochemical reaction

Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0036] 1. HCY determination

[0037] 1) Reduction and enzymatic conversion:

[0038] Add 0.025ml 0-1000μM L-isotype to 0.25ml reaction solution (50mM Tris-HCl pH 7.5, containing 20μM pyridoxal phosphate (PLP for short), 0.05% Triton X-100, 1mM TCEP and 0.1U / ml rMETase) Cystine, placed in a 37°C water bath for 30 minutes.

[0039] 2) Determination of hydrogen sulfide:

[0040] Add 0.025ml color developing solution (1N HCl containing 20mM DMPD 2HCl and 30mM FeCl 3 ), and stand at room temperature for 10 minutes after mixing. Using distilled water as a reference, read the absorbance at 670nm.

[0041] 3) Results:

[0042] L-homocysteine ​​linear range 3-1000μM

[0043] 2. Homocysteine ​​(HCY) Biochemical Kit

[0044] Product Name: Homocysteine ​​(HCY) Biochemical Kit

[0045] Model: Biochemical Reagent

[0046] Product code: DC-HCY

[0047] Specifications: Reagent I 25ml, Reagent II 2.5ml, HCY standard 1 bottle, HCY normal serum control 1 bottle, HCY elevated serum cont...

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Abstract

The invention is an enzyme biologic and chemical reaction measure method to mensurate homotype cysteine in a biologic sample. Cysteine can lose its ammonia and be changed into alpha-4-ketone acid, ammonia and sulfureted hydrogen by L-ovi-ammonia acid and Y-dispeling enzyme. Sulfureted hydrogen and fluorescence cpd.DMPD2HCL can create blue product-sub-armour blue whose degree of absorbing light is 670nm, in the situation of Fe3+ and acid. Thus, people can mensurate the chroma of homotype cysteine in a biologic sample by knowing this degree. This method can measure the chroma of homotype cysteine(3-1000 uMs). The invention also involves a reagent box used for implementing the method mentioned above. The box uses liquid double reagents, and needs less quantity of samples(25 microlitre or serum or plasm);In addition, the respond time is short, and the operation is easy. Thus, this method is suitable for a great deal of examinations. The reagent box costs less than other types.

Description

technical field [0001] The invention relates to a technical solution for measuring the content of homocysteine ​​in a biological sample and a reagent manufactured by the solution. In particular, it relates to a method for measuring homocysteine ​​by enzymatic biochemical reaction. Background technique [0002] Tan (2000) reported a method for the determination of homocysteine ​​by enzymatic biochemical reaction. Homocysteine ​​(HCY) is deaminated by homocysteine ​​α, γ-lyase (rHCYase) into α-ketobutyrate and ammonia (NH 4 ) and hydrogen sulfide (H 2 S). Hydrogen sulfide and fluorescein N, N-dibutylphenylene diamine (N, N-Dibutylphenylene diamine, DBPDA) form a pigment phenothiazine chloride compound [3,7-bis(dibutyl amino)phenothiazine-5- ium chloride], the latter can be quantitatively measured at 660nm and its absorbance is proportional to the content of HCY in the sample. [0003] <chemistry num="001"> <chem file="200510053210_cml001.xml" / > < / chemis...

Claims

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Application Information

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IPC IPC(8): G01N21/3577
Inventor 蔡枫
Owner ZHEJIANG YAKE SCI & TECH
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