Troponin I monoclonal antibody magnetic particles and preparation method thereof, and detection kit

A monoclonal antibody and detection kit technology, applied in the field of in vitro diagnostic medical testing, can solve the problems of unfavorable promotion of troponin I detection, high cost, poor repeatability, etc.

Active Publication Date: 2017-02-01
SHANGHAI KEHUA BIO ENG
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  • Abstract
  • Description
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AI Technical Summary

Problems solved by technology

[0006] Although there are already registered and approved troponin I chemiluminescence immunoassay assay kits in the Chinese market, most of them are imported reagents with high cost, and the preparation methods and related materials of the kits are kept as confidential information and have not been made public. , which is not conducive to the promotion of troponin I detection in community primary hospitals
Domestic troponin I kits have poor reproducibility, difficult to control batch-to-batch differences, low sensitivity, high cross-reaction rate, and poor clinical application prospects

Method used

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  • Troponin I monoclonal antibody magnetic particles and preparation method thereof, and detection kit
  • Troponin I monoclonal antibody magnetic particles and preparation method thereof, and detection kit
  • Troponin I monoclonal antibody magnetic particles and preparation method thereof, and detection kit

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preparation example Construction

[0063] Correspondingly, the present invention also provides a method for preparing the troponin I monoclonal antibody magnetic particles described above, the method comprising the following steps:

[0064] 1) Using biotin derivatives to pre-crosslink magnetic particles;

[0065] 2) coating the biotin derivative pre-crosslinked magnetic particles obtained in step 1) with streptavidin; and

[0066] 3) Blocking the magnetic particles obtained in step 2), and then reacting with troponin I monoclonal antibody-biotin to prepare troponin I monoclonal antibody magnetic particles.

[0067] In specific embodiments, the method may include:

[0068] a) using biotin derivatives to pre-crosslink the magnetic particles, and then coating them with streptavidin;

[0069] b) reacting the magnetic particles treated in step a) with troponin I monoclonal antibody-biotin after being blocked;

[0070] c) Preserving the magnetic particles treated in step b) in a magnetic particle preservation solu...

Embodiment 1

[0142] Example 1. Preparation of Troponin I Monoclonal Antibody Magnetic Particles

[0143] 1) Dissolve and dilute BSA with 10mM PBS to 2mg / mL, add 2mg / mL Biotin aqueous solution to the 2mg / mL BSA solution according to the molar concentration ratio of 1:20, and mix well;

[0144] 2) React the mixture at room temperature for 30 minutes, place it at 2-8°C, and dialyze with 10mM PBS for 16-24 hours;

[0145] 3) Take out the dialysate and dilute BSA-Biotin to 1mg / mL with 10mM PBS;

[0146] 4) Take 10 mg of 1.0 μm magnetic particles whose active functional groups are carboxyl groups, and wash them twice with 50 mM MES buffer at pH 6.0;

[0147] 5) Remove the supernatant after magnetic suction, add 0.5mL of 50mM MES buffer solution with pH 6.0 and mix well, then add 0.5mL of 25mg / mL carbodiimide (EDC) solution, mix well, and react at room temperature for 30 minutes;

[0148] 6) Wash twice with a solution containing 50 mM pH 7.8 Tris-HCl, 150 mM NaCl, and 1% BSA;

[0149] 7) Add 4...

Embodiment 2

[0161] Example 2. Sensitivity Detection of Troponin I Monoclonal Antibody Magnetic Particles

[0162] Using the troponin I monoclonal antibody magnetic particles obtained in Example 1, the sensitivity detection was carried out as described in the materials and methods, and the results are shown in the following table:

[0163]

[0164]

[0165] Note:

[0166] (1) Magnetic particles obtained by directly cross-linking troponin I monoclonal antibody;

[0167] (2) Traditional streptavidin-biotin troponin I monoclonal antibody magnetic particles;

[0168] (3) The multistage amplified streptavidin-biotin troponin I monoclonal antibody magnetic particle of the present invention.

[0169] As can be seen from the above table, using the multistage amplified streptavidin-biotin troponin I monoclonal antibody magnetic particles of the present invention, its sensitivity (S / N value) is higher than that of direct cross-linking and traditional streptavidin - The biotin method improve...

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Abstract

The invention discloses troponin I monoclonal antibody magnetic particles and a preparation method thereof, and a troponin detection kit comprising the troponin I monoclonal antibody magnetic particles. According to a multi-stage amplification magnetic particle crosslinking method, biotin derivative pre-crosslinked magnetic particles are adopted and then are coated with streptavidin, and the particles are closed and react with troponin I monoclonal antibody-biotin, so that the troponin I monoclonal antibody magnetic particles are prepared. The troponin I detection kit adopts a streptavidin biotin multi-stage amplification system and has the advantages of high detection sensitivity, high specificity, small medicine interference, high result stability and the like, so that the troponin I detection kit can be suitable for cardiovascular disease clinical auxiliary diagnosis on a large scale.

Description

technical field [0001] The invention relates to the field of in vitro diagnostic medical testing. Specifically, the present invention relates to a troponin I monoclonal antibody magnetic particle, a preparation method thereof and a troponin I detection kit comprising the same. Background technique [0002] Troponin (Tn) is a special regulatory protein of myocardial tissue, which is released into the blood when severe myocardial injury occurs, and is a serum marker reflecting myocardial injury. Tn is composed of three subunits: TnI, TnT, and TnC. TnC is Ca 2+ TnI is an inhibitory subunit that can inhibit the binding of myosin to actin and prevent muscle contraction. The cTnI (troponin I) content in blood of normal people is generally lower than 0.3ug / L. When muscle cells are damaged, free cTnI is released into the blood first, and as the injury aggravates, cTnI in blood continues to rise. [0003] Acute myocardial infarction (AMI) is irreversible acute myocardial necrosis...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K17/14C07K17/02B01J13/02H01F1/11G01N33/68G01N33/543
CPCB01J13/02C07K17/02C07K17/14G01N33/54326G01N33/6893G01N2333/4712G01N2800/324H01F1/11
Inventor 陈超罗紫臣孙小禁彭珊李基赵亚茹
Owner SHANGHAI KEHUA BIO ENG
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