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Preparation method of TSH (thyroid stimulating hormone) quantitative detection kit based on time-resolved fluoroimmunoassay

A time-resolved fluorescence and thyrotropin technology, which is applied in the preparation of test samples, measurement devices, and analysis materials, can solve the problems of difficult sensitivity and accurate quantification, and achieve the effect of high detection sensitivity and wide detection range

Inactive Publication Date: 2012-07-04
北京协和洛克生物技术有限责任公司
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Problems solved by technology

Although enzymes are an ideal marker, the sensitivity of the enzymatic method for measuring the OD value of the chromogenic product of the enzyme reaction is difficult to directly and accurately quantify high-concentration analytes
Due to the influence of factors such as scattered light, background fluorescence from different sources, and fluorescence quenching, the traditional fluorescence method with conventional fluorescent molecules as markers generally has an analytical sensitivity of 10 -8 ~10 -11 mol / L, limited to the determination of some analytes and some small molecules that do not require high sensitivity

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  • Preparation method of TSH (thyroid stimulating hormone) quantitative detection kit based on time-resolved fluoroimmunoassay
  • Preparation method of TSH (thyroid stimulating hormone) quantitative detection kit based on time-resolved fluoroimmunoassay
  • Preparation method of TSH (thyroid stimulating hormone) quantitative detection kit based on time-resolved fluoroimmunoassay

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Embodiment Construction

[0030] In order to make the content of the present invention more clear, the following is the key preparation operation process of the reagents in the thyroid stimulating hormone quantitative detection kit (time-resolved fluorescence immunoassay).

[0031] 1. Eu-Anti-TSH labeled and isolated antibody process

[0032] (1) Dissolve Eu with double distilled water 3+ Chelate;

[0033] (2) After being dissolved, take 1mg Eu 3+ The chelate solution was added to 1 mg TSH monoclonal antibody solution, and reacted at room temperature for 48 hours;

[0034] (3) Separation of Eu-Anti-TSH conjugates and free Eu by Sephadex G-50 chromatographic column 3+ ;

[0035] (4) Collect the eluate whose fluorescence count is greater than 1 million, which is the Eu-Anti-TSH conjugate.

[0036] 2. Preparation of enhancement solution

[0037] Main raw material names Glacial acetic acid, Triton X-100 (Triton X-100, polyethylene glycol octyl phenyl ether), 0.1M potassium hydrogen phthalate (KHP), β...

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Abstract

The invention provides a TSH (thyroid stimulating hormone) quantitative detection kit based on time-resolved fluoroimmunoassay, applicable to clinical screening of neonatal hypothyroidism. The preparation method of the kit is characterized by comprising the following steps of: 1, separating an antibody conjugate (Eu-Anti-TSH) from free rare earth ions (Eu<3+>) by using a specific chromatograph column; (2) using a bifunctional chelator (DTTA) for marking, wherein one end of the bifunctional chelator is chelated with rare earth ions (Eu) and the other end of the bifunctional chelator is connected with -NH2 of protein; adding beta-NTA to reinforcing liquid and regulating the concentration so that the fluorescence intensity of a finally obtained reagent is improved; and (4) adding a substitute Proclin300 for sodium azide to an analysis buffer system. Based on the characteristics of the steps in the preparation method, the TSH quantitative detection kit based on time-resolved fluoroimmunoassay can be produced and other in vitro diagnosis detection reagents can be developed.

Description

technical field [0001] The present invention relates to the technical field of immunodiagnostic applications. Specifically, the present invention provides a "double antibody sandwich" using time-resolved fluorescence immunoassay, using Eu-labeled antibodies and antibodies on solid-phase carriers to bind to different sites on antigens Technology, quantitative detection of thyroid-stimulating hormone (TSH) concentration in neonatal whole blood samples collected on 903# filter paper, mainly used for screening of neonatal hypothyroidism. technical background [0002] The theory of "time-resolved fluorescent immunoassay" was first proposed by Finland's Soini and Hemmila in 1979. In 1984, Hemmila determined the technical scheme of time-resolved immunoassay. [0003] At present, the most widely used experimental methods for detecting the concentration of thyrotropin in neonatal blood in China are enzymatic method and fluorescence method. Although enzymes are ideal markers, the sen...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N33/532G01N1/34
Inventor 王晓华
Owner 北京协和洛克生物技术有限责任公司
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