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Reagent kit and method for measuring concentration of asymmetric dimethylarginine

A dimethylarginine, asymmetric technology, applied in the field of kits for measuring asymmetric dimethylarginine concentration, can solve the problem of poor anti-interference performance, inability to apply clinical large-flow automated analysis, and long test time, etc. problems to improve accuracy and eliminate non-specific cross-reactions

Inactive Publication Date: 2016-10-05
王学忠
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The technical problem to be solved by the present invention is that in order to overcome the high pressure liquid chromatography and the enzyme-linked immunoassay method, special measuring instruments are required, the operation is complicated, the test takes a long time, it cannot be applied to the automatic analysis of clinical large flow, and the price is expensive; and If the immunoturbidimetric assay method is applied, the detection accuracy of asymmetric dimethylarginine is low and the anti-interference performance is poor. A kit and method for determining the concentration of asymmetric dimethylarginine are provided.

Method used

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  • Reagent kit and method for measuring concentration of asymmetric dimethylarginine
  • Reagent kit and method for measuring concentration of asymmetric dimethylarginine
  • Reagent kit and method for measuring concentration of asymmetric dimethylarginine

Examples

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Embodiment 1

[0040] (1) Preparation of asymmetric dimethylarginine-ovalbumin conjugate

[0041] Add 10mg ovalbumin to 1.0mL 25mmol / L MES (2-morpholineethanesulfonic acid) buffer solution with a pH of 6.1, stir rapidly, add asymmetric dimethylarginine, and slowly drop in 60mmol / L EDAC ( 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride) solution 300 μL, rapid stirring reaction for 2 hours, adding 2.5mL 50mmol / L Tris buffer solution with a pH of 8.2 to terminate the reaction for 30 minutes . Dialyze at 4°C against citrate buffer with pH 6.1 for later use.

[0042] (2) Preparation of Reagent 1

[0043] Take 2 mL of Polymicrospheres CB0267D latex particles with a concentration of 10%, carboxyl groups on the surface, and a diameter of 250 nm, and add them to 5 mL of 20 mM MES buffer solution with pH 6.1. While stirring, add 300 mg of Sulfo-NHS and 30 mg of EDAC, and react at room temperature for 20 Minutes, centrifuge to remove the supernatant, then add 35 mL of pH 8.3 50 mM boric ...

Embodiment 2

[0059] (1) Preparation of asymmetric dimethylarginine-hemocyanin conjugates

[0060] Add 10 mg of hemocyanin to 1.0 mL of 20 mmol / L MES (2-morpholineethanesulfonic acid) buffer solution with a pH of 6.1, stir rapidly, add asymmetric dimethylarginine, and slowly drop in 50 mmol / L EDAC ( 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride) solution 300 μL, rapid stirring reaction for 2 hours, adding 2.5mL 50mmol / L Tris buffer solution with a pH of 8.2 to terminate the reaction for 30 minutes . Dialyze with MES buffer at pH 6.1 at 4°C for later use.

[0061] (2) Preparation of Reagent 1

[0062]Take 2 mL of Polymicrospheres CB0104D latex particles with a concentration of 10% and carboxyl groups on the surface, and add them to 5 mL of 25 mM MES buffer solution with pH 6.1. While stirring, add 300 mg of Sulfo-NHS and 30 mg of EDAC, react at room temperature for 30 minutes, and centrifuge Remove the supernatant, then add 35 mL of pH 8.1 100 mM borate buffer solution conta...

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Abstract

The invention discloses a reagent kit and method for measuring the concentration of asymmetric dimethylarginine. The reagent kit comprises a reagent 1 and a reagent 2, wherein the reagent 1 is prepared from latex particles coated with monoclonal antibodies resisting asymmetric dimethylarginine, and the reagent 2 is prepared from latex particles coated with asymmetric dimethylarginine-inert carrier conjugates. When the reagent kit is adopted for detecting asymmetric dimethylarginine, immune turbidity can be easily improved, detection signals can be easily amplified, the reagent kit has the advantages of being free of special instruments, easy and convenient to operate, high in sensitivity and low in cost, rapid and large-flow clinical sample testing can be achieved, it is made possible that asymmetric dimethylarginine becomes a clinical conventional detection item, and the reagent kit has high economic value.

Description

technical field [0001] The invention belongs to the technical field of substance detection and determination, and in particular relates to a kit and a method for determining the concentration of asymmetric dimethylarginine. Background technique [0002] Asymmetric dimethylarginine (ADMA), a methylated arginine, is an endogenous nitric oxide synthase (Nitric oxide synthase, NOS) inhibitor, which can competitively inhibit the oxidation of Nitrogen (Nitric oxide, NO) synthesis. Asymmetric dimethylarginine is produced by the methylation of intracellular proteins under the action of arginine methyltransferase, followed by hydrolysis. Many cells, such as human endothelial cells, can produce ADMA. A normal human body produces about 300 μmol (about 60 mg) per day, and its concentration in the body is much higher than other endogenous arginine that can also inhibit NOS, so it is considered the most important NOS inhibitor. After ADMA enters the blood circulation from cells, part o...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N21/82
Inventor 不公告发明人
Owner 王学忠
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