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Detection kit and preparation method thereof and detection method of troponin T

A detection kit, troponin technology, applied in biological tests, measuring devices, material inspection products, etc., can solve problems such as poor detection sensitivity, and achieve the effects of enhancing sensitivity, improving coating efficiency, and enhancing detection sensitivity

Active Publication Date: 2019-04-09
深圳天辰医疗科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The main purpose of the present invention is to provide a detection kit, aiming at solving technical problems such as poor detection sensitivity in existing chemiluminescent methods

Method used

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  • Detection kit and preparation method thereof and detection method of troponin T
  • Detection kit and preparation method thereof and detection method of troponin T
  • Detection kit and preparation method thereof and detection method of troponin T

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preparation example Construction

[0072] On the other hand, the embodiment of the present invention also provides a method for preparing the detection reagent described above, including:

[0073] S011. Provide gold magnetic particles and troponin T antibodies, react troponin T antibodies with gold magnetic particles, and prepare gold magnetic particles coated with troponin T antibodies;

[0074] Or, provide gold magnetic particles, streptavidin, biotin and troponin T antibody, react streptavidin with gold magnetic particles to prepare streptavidin-coated gold magnetic particles; biotin React with troponin T antibody to prepare biotin-labeled troponin T antibody;

[0075] S012. Provide troponin T antibody and labeled enzyme, react troponin T antibody and labeled enzyme to obtain enzyme-labeled antibody; provide cross-linking reagent, react cross-linking reagent and enzyme-labeled antibody in solution to prepare enzyme labeled antibody polymer.

[0076] Specifically, in step S011, when the magnetic separation ...

Embodiment 1

[0103] This embodiment provides a chemiluminescent detection kit, including: magnetic separation reagents, enzyme-labeled reagents, chemiluminescent substrate solutions and calibration products, the preparation of which includes:

[0104] 1. Preparation of enzyme-labeled reagents

[0105] (1) Select troponin T monoclonal antibody as the detection antibody, add 12mg / mL 2-iminothiolane hydrochloride solution according to 1 / 100 of the volume of the antibody, mix well, and place it at room temperature for 30min to activate treatment to obtain activated antibodies;

[0106] Select alkaline phosphatase as the labeled enzyme, add 7mg / mL Sulfo-SMCC solution according to 1 / 10 of the volume of alkaline phosphatase, mix well, place at room temperature for 25min, and perform activation treatment to obtain activated alkaline phosphatase solution;

[0107] The activated alkaline phosphatase and the activated troponin T monoclonal antibody were mixed and reacted at 4° C. for 15 hours to pr...

Embodiment 2

[0116] This embodiment provides a chemiluminescent detection kit, including: magnetic separation reagents, enzyme-labeled reagents, chemiluminescent substrate solutions and calibration products, the preparation of which includes:

[0117] 1. Preparation of enzyme-labeled reagents

[0118] (1) Select troponin T monoclonal antibody as detection antibody, select horseradish base peroxidase (HRP) as labeling enzyme;

[0119] Centrifuge the HRP and add fresh 0.3M NaIO 4 The solution was stirred at room temperature for 50 minutes in the dark to obtain the hydroformylated HRP; ultrafiltration was performed using 0.05M, pH 4.4 sodium acetate buffer solution, and the ultrafiltration conditions were set at 4°C, 9000rmp, 20min, ultrafiltration 3 times, and the purified Add 1M, pH 9.5 carbonate buffer, adjust the pH to 9.0-9.5, then add troponin T monoclonal antibody, shake at room temperature for 7 hours in the dark; add 0.1ml of newly prepared 9mg / mL of NaBH 4 solution, shake for 2h...

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Abstract

The invention belongs to the technical field of in-vitro detection, and particularly relates to a detection kit and a preparation method and application thereof. The detection kit comprises a magneticseparation reagent and an enzyme-labeled reagent; the magnetic separation reagent is prepared from troponin T antibody-coated gold magnetic particles or prepared from streptavidin-coated gold magnetic particles and a biotin-labeled troponin T antibody; and the enzyme-labeled reagent is prepared from an enzyme-labeled antibody polymer, wherein the enzyme-labeled antibody polymer is prepared from at least two enzyme-labeled antibodies and carbon bridges; the carbon bridges connect any two or more, adjacent or non-adjacent enzyme-labeled antibodies, and each carbon bridge has at least two connecting loci for connecting the enzyme-labeled antibodies; and each enzyme-labeled antibody is prepared from a detection antibody and a labeling enzyme coupled to the detection antibody, and the connecting loci are connected with the detection antibodies and / or the labeling enzymes. When chemiluminescence detection is conducted, a reaction signal value of a chemiluminescence method is amplified, thesensitivity of an antigen captured by the detection reagent can be enhanced, and the detection sensitivity is improved.

Description

technical field [0001] The invention belongs to the technical field of in vitro detection, and in particular relates to a detection kit, a preparation method thereof and a detection method of troponin T. Background technique [0002] Troponin (Tn) is a regulatory protein of muscle tissue contraction, located on the thin myofilament of contractile protein, and plays a very important role in the regulation of muscle contraction and relaxation. Cardiac troponin (cTn) exists only in cardiomyocytes and is a spherical complex composed of three subunits, troponin T (cTnT), troponin I (cTnI) and troponin C (cTnC). molecule. cTnT and cTnI are specific antigens of cardiomyocytes, which are degraded from myocardial fibers when cardiomyocytes are injured. In cardiomyocytes, most cTn is bound to thin myofilaments in the form of complexes, and a small amount of cTn exists in the cytoplasm in free form. Normally, cTn cannot be detected in the peripheral blood of healthy individuals. How...

Claims

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Application Information

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IPC IPC(8): G01N33/68
CPCG01N33/6887
Inventor 汪柯孜唐灿
Owner 深圳天辰医疗科技有限公司
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