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Enzyme-labeled immunoassay kit and application of thereof to serum detection

A technology of immunological reagents and enzyme labels, which is applied in measuring devices, peptide preparation methods, and analysis through chemical reactions of materials, etc., can solve problems such as unstable chemiluminescence signals, and achieve a wide range of substrate effects and a wide range of pH applications Wide, high-throughput effects

Inactive Publication Date: 2014-07-02
靖江市人民医院 +1
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Problems solved by technology

[0004] The purpose of the present invention is to provide an enzyme-labeled immunoassay kit, using soybean peroxidase (SBP) as the labeling enzyme, MORPH and SPTZ as enhancers, to solve the problem of luminol oxidation reaction caused by HRP catalysis in the prior art Problems with unstable chemiluminescent signals

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  • Enzyme-labeled immunoassay kit and application of thereof to serum detection

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Embodiment 1

[0032] The implementation process of the enzyme-labeled immunoassay kit of the present invention is as follows: figure 1 shown.

[0033] The PBS involved in this example is 10 mM pH7.4, and the PBST is 0.1% Tween prepared in 10 mM pH7.4 PBS. The SBP used was purchased from Bio-Research Products, USA.

[0034] The first step, preparation of enzyme-labeled antibody

[0035] Using the improved sodium periodate method: 1. Dissolve 1 mg of SBP in 100 μL PBS, add newly prepared 12.8 mg / mL NaIO 4 50 μL of the solution, mix well, and place at room temperature for 15 minutes; 2. After taking it out, add 50 μL of 9 μL / mL ethylene glycol PBS solution, and place at room temperature for 30 minutes; 3. Add human chorionic gonadotropin antibody (β-HCG) containing 2 mg of purified HCG, and mix well , and put it into a dialysis bag, and dialyzed in 50mM pH9.6 carbonate buffer solution at 4°C for 18h to bind SBP and Ab2 antibody; 4. Add 5mg / ml NaBH 4 20 μL of the solution, mix well, and red...

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Abstract

The invention discloses an enzyme-labeled immunoassay kit which comprises an antibody pre-coating reaction plate, an enzyme-labeled antibody and luminous substrate liquid, wherein the enzyme-labeled antibody takes SBP (soybean protease) as marker enzyme; the luminous substrate liquid comprises H2O2, luminol and an enhancer; the enhancer consists of MORPH (4-marlene pyridine) and SPTZ (3-(10'-phenothiazinyl) propyl-1-sulfonate). According to the enzyme-labeled immunoassay kit, the SBP is introduced to serve as the marker enzyme; on one hand, the thermal stability of the enzyme is higher than that of HRP (hypothalamic regulatory peptide), and the enzyme has the advantages of wide substrate working range, high heat resistance, high acid-alkaline stability, wide pH application range and the like; on the other hand, when the enhancers MORPH and SPTZ are used for enhancing chemical illumination, a chemical luminous signal is enhanced by nearly 100 times, and the detection sensitivity is greatly improved.

Description

technical field [0001] The invention relates to the field of immune reagent kits, in particular to an enzyme-labeled immune reagent kit and its application in serum detection. Background technique [0002] ELISA method is a new technology in immunodiagnosis, and has been successfully applied to the immunodiagnosis of infectious diseases, parasitic diseases and non-infectious diseases caused by various pathogenic microorganisms. It has also been applied to the quantitative determination of macromolecular antigens and small molecular antigens, and has the characteristics of sensitivity, specificity, simplicity, rapidity, stability and easy automation. It is not only suitable for the inspection of clinical specimens, but also suitable for serological epidemiological investigations because hundreds or even thousands of specimens can be inspected in one day. Not only can it be used to detect antibodies, but it can also be used to detect circulating antigens in body fluids, so it...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/543G01N21/76C07K1/13
CPCG01N21/763G01N33/54393G01N33/581
Inventor 薛艳春苏娟车彦军王建江毛世琴朱伟田亦平蒋镌翟丽芬陶岚刘松琴沈丽吴慧萍
Owner 靖江市人民医院
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