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Reagent kit

A two-site, reagent kit technology, applied in the field of enzyme-labeled immunoassay technology, can solve the problems of high price, difficulty in clinical widespread use, and the sensitivity of general enzyme immunoassay methods that cannot meet the requirements, achieving low cost, low The effect of negative background

Inactive Publication Date: 2005-03-02
SHANGHAI SECOND MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Corresponding imported kits are used in China, which are expensive and difficult to be widely used in clinical practice
[0007] Under normal circumstances, the content of cardiac troponin I in the blood is very low (<0.1 μg / L), and the sensitivity of general enzyme immunoassay methods cannot meet this requirement

Method used

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Embodiment 1

[0019] The antibody, antigen or immune complex is immobilized on the solid phase indirectly through the streptavidin-biotin capture system to establish a plate-type streptavidin two-site sandwich one-step method. Wherein, the plate-type streptavidin solid-phase two-site sandwich one-step method is to use streptavidin to pre-coat the microtiter plate, biotin-label the anti-cTnI monoclonal antibody, and horseradish peroxidase to label another strain of anti-cTnI. cTnI monoclonal antibody.

[0020] Preparation of streptavidin solid phase: take 50 μl streptavidin (1 mg / ml) and add 200 μl distilled water to dilute, then add 50 μl 3mol / l acetic acid, leave it at room temperature for 5 minutes, adjust to pH 6.0 with 2mol / l Tris, then, Dilute to 2 μg / ml with coating buffer (0.05 mol / l, pH 9.5 carbonic acid buffer), add to polystyrene ELISA microwell plate, 120 μl / well overnight at 4°C, wash 3 times with washing solution, add blocking solution ( 0.5% BSA, 0.02mol / l, pH7.2 PBS) at 37°C...

Embodiment 2

[0024] Two-site sandwich one-step ELISA method for quantitative detection of serum cTnI

[0025] As shown in Figure 1, the plate-type streptavidin solid-phase two-site sandwich one-step ELISA created by the present invention

[0026] Embodiment two

[0027] Two-site sandwich one-step ELISA method for quantitative detection of serum cTnI

[0028] As shown in Figure 1, the operation flow chart of the plate-type streptavidin solid-phase two-site sandwich one-step ELISA quantitative detection of serum cTnI created by the present invention, streptavidin pre-coated microwell plates are added to each well of 20 μl antigen standard Or hold the serum sample, then add 100 μl of biotinylated anti-TnI monoclonal antibody (about 2 μg / ml) and horseradish peroxidase-labeled mixture of another anti-TnI monoclonal antibody (pH7.2, 0.02mol / l phosphate buffer solution), react at 37°C for 90 minutes, and wash. Add o-phenylenediamine (OPD) substrate solution, 37°C, 15 minutes, add 2mol / l sulfuric...

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Abstract

A reagent kit for enzyme-linked immunodetection is characterized by that on the basis of ELISA method, the chain avidin-biotin capture system is used to indirectly fix the antibody, antigen, or immune complex to solid phase and the plate-type chain avidin dual-site sandwich one-step ELISA for quantitatively detecting the serum cTnI. Its advantages are simple method, high speed, and low-negative background.

Description

technical field [0001] The plate-type streptavidin solid-phase double-site sandwich one-step kit of the present invention relates to biotechnology, especially an enzyme-labeled immunoassay technology, which applies streptavidin and biotin capture technology to enzyme-linked immunoassay technology, and establishes Plate streptavidin two-site sandwich one-step quantitative enzyme-linked immunosorbent assay (Streptavidin-ELISA) was used to detect serum cTnI levels in patients. Background technique [0002] In my country, with the development of the national economy and the improvement of people's living standards, the morbidity and mortality of cardiovascular diseases are increasing year by year, so it is necessary to strengthen the early diagnosis of AMI. The diagnosis of ischemic heart disease has three aspects: biochemical tests, ECG changes, and persistent chest pain. In the biochemical examination, the indicators that are considered to be effective in diagnosing myocardia...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/02G01N33/53G01N33/535G01N33/543G01N33/577
Inventor 郑佐娅王鸿利
Owner SHANGHAI SECOND MEDICAL UNIV
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