Chemiluminescent immunodetection kit for neuronspecific enolase and preparation method of chemiluminescent immunodetection kit

An enolase chemistry and detection kit technology, which is applied in chemiluminescence/bioluminescence, biological testing, and analysis by chemically reacting materials, and can solve the problems of poor repeatability, long detection time, and low sensitivity.

Inactive Publication Date: 2019-01-11
DIRUI MEDICAL TECH CO LTD
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Problems solved by technology

[0008] The purpose of the present invention is to provide a neuron-specific enolase chemiluminescent immunoassay kit and its preparation method in order to solve the problems of long detection time, low sensitivity and poor repeatability of the existing methods for detecting NSE

Method used

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  • Chemiluminescent immunodetection kit for neuronspecific enolase and preparation method of chemiluminescent immunodetection kit
  • Chemiluminescent immunodetection kit for neuronspecific enolase and preparation method of chemiluminescent immunodetection kit
  • Chemiluminescent immunodetection kit for neuronspecific enolase and preparation method of chemiluminescent immunodetection kit

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preparation example Construction

[0044] The present invention also provides a preparation method of the above-mentioned chemiluminescent immunoassay kit for neuron-specific enolase, comprising the following steps:

[0045] Step 1: Preparation of streptavidin magnetic particle suspension

[0046] After mixing the streptavidin magnetic particle solution and TBST solution, place it on a magnetic separator until the supernatant is free of turbidity, discard the supernatant, keep the magnetic particles, and prepare a solid phase reagent in the buffer after washing ;

[0047] The concentration of the streptavidin magnetic particle solution is preferably 50-100 mg / ml; the volume ratio of the streptavidin magnetic particle solution to the TBST solution is preferably (0.5-1): (5-15); The mixing time is preferably 10-15 minutes, the buffer is 50mM MES, 0.05% Tween-20, 0.05% Proclin300, pH6.5 or 100mM PBS, 0.1% Tween-20, 0.1% Proclin300, pH7.2; the concentration of the solid-phase reagent is preferably 0.01% to 1%, mo...

Embodiment 1

[0055] Embodiment 1: the preparation of neuron-specific enolase chemiluminescence immunoassay kit:

[0056] (1) Preparation of streptavidin magnetic particle suspension:

[0057] Take 0.5mL (50mg) of streptavidin magnetic particle solution with a concentration of 100mg / mL, add 10mL of TBST solution and mix well for 10min, place on a magnetic separator until the supernatant is free of turbidity, discard the supernatant, and keep the magnetic particles. After repeated washing for 3 times, use 50mM MES, 0.05% Tween, 0.05% Proclin300, pH 6.5 buffer to prepare a solid-phase reagent with a magnetic bead concentration of 0.05%, and store at 2-8°C.

[0058] (2) Acridine ester labeling process:

[0059] Put 250ug of antibody into a centrifuge tube to ensure that the antibody is at the bottom of the centrifuge tube (centrifuge at room temperature for 20s), then add PBS buffer solution, mix well, add 5μl 2mg / mL acridinium ester DMF solution after mixing, and use a centrifuge Centrifug...

Embodiment 2

[0062] Example 2 Preparation of neuron-specific enolase chemiluminescent immunoassay kit:

[0063] (1) Preparation of streptavidin magnetic particle suspension:

[0064] Take 0.72 ml (72 mg) of streptavidin magnetic particle solution with a concentration of 100 mg / ml, add 15 mL of TBST solution and mix well for 15 minutes, then place it on a magnetic separator until the supernatant is free of turbidity, discard the supernatant, Keep the magnetic particles. After repeated washing for 3 times, a solid-phase reagent with a magnetic bead concentration of 0.072% was prepared in 100mM PBS, 0.1% Tween-20, 0.1% Proclin300, pH7.2 buffer, and stored at 2-8°C.

[0065] (2) Acridine ester labeling process:

[0066] Put 500ug of antibody into a centrifuge tube to ensure that the antibody is at the bottom of the centrifuge tube (centrifuge at room temperature for 30s), then add TRIS flushing solution, mix well, add 15μl 2.5mg / mL acridinium ester DMF solution after mixing, and centrifuge ...

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Abstract

The invention provides a chemiluminescent immunodetection kit for a neuronspecific enolase and a preparation method of the chemiluminescent immunodetection kit and belongs to the technical field of in-vitro detection. The kit comprises a streptavidin magnetic particle suspension, a neuronspecific enolase monoclonal antibody marked by a chemiluminescent marker and a neuronspecific enolase monoclonal antibody marked by a coupling marker. The invention provides the preparation method of the chemiluminescent immunodetection kit for the neuronspecific enolase. The chemiluminescent immunodetection kit for the neuronspecific enolase, provided by the invention, is capable of automatically measuring a sample and directly giving a numerical value, so that human operation errors are reduced; in addition, unmanned operation is realized, so that the time required by clinical detection is shortened; meanwhile, the detection precision is relatively high; and a reagent and an instrument form a sealedsystem, so that small system errors are caused.

Description

technical field [0001] The invention belongs to the technical field of in vitro detection, and relates to a neuron-specific enolase (NSE) chemiluminescent immunoassay kit and a preparation method thereof. Background technique [0002] NSE is the key enzyme that catalyzes the transformation of 2-phosphoglycerate into phosphoenolpyruvate in the tricarboxylic acid cycle of neurons and neuroendocrine cells. The full-length nucleotide sequence of the gene is 2423bp, encoding 434 amino acid residues. The biological half-life may be greater than 20h, the molecular weight is 78kD, and the isoelectric point is pH4.7. It is an acidic protease. NSE is a member of the enolase gene family. Enolase is a dimer composed of three subunits α, β, and γ. Its isozymes are divided into five types: αα, ββ, γγ, αγ, and αβ. The α subunit mainly exists in liver, kidney and other tissues, so it is called non-nervous system enolase (NNE); the β subunit mainly exists in skeletal muscle and cardiac musc...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/573G01N21/76G01N33/532G01N33/543G01N33/577G01N33/58
CPCG01N21/76G01N33/532G01N33/54326G01N33/573G01N33/577G01N33/58G01N2800/164G01N2800/28G01N2800/52
Inventor 李磊孟令敏王凯孙成艳高威
Owner DIRUI MEDICAL TECH CO LTD
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