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Magnetic micro-particle chemiluminiscence detection kit for determining content of human Golgi body protein

A Golgi and kit technology, applied in chemiluminescence/bioluminescence, biological testing, measurement devices, etc., can solve the problems of low automation, complicated and tedious operations, and stability to be improved in ELISA, and avoid Missing detection phenomenon, wide linear range, the effect of improving signal strength and sensitivity

Active Publication Date: 2021-01-22
BEIJING LEADMAN BIOCHEM
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  • Description
  • Claims
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Problems solved by technology

However, there are certain defects in these methods, such as the limitation of rare earth elements in the up-transfer luminescence method, the low degree of automation and complicated operation of the enzyme-linked immunosorbent assay, and the stability of the magnetic particle chemiluminescence immunoassay reagent of the horseradish peroxidase system. Sex needs to be improved

Method used

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  • Magnetic micro-particle chemiluminiscence detection kit for determining content of human Golgi body protein

Examples

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preparation example Construction

[0058] A method for preparing a magnetic particle chemiluminescence detection kit for measuring the protein content of human Golgi apparatus, comprising the following steps:

[0059] Preparation of reagent R1: 1) Prepare buffer solution according to the content of reagent R1 buffer component, adjust pH; 2) Prepare fluorescein isothiocyanate-labeled anti-Golgi protein monoclonal antibody; 3) Label fluorescein isothiocyanate The anti-Golgi protein monoclonal antibody was diluted with R1 buffer.

[0060] Preparation of reagent R2: 1) Prepare buffer according to the content of reagent R2 buffer components, and adjust pH; 2) Prepare alkaline phosphatase-labeled Golgi protein polyclonal antibody; 3) Alkaline phosphatase-labeled Golgi protein polyclonal antibody Cloned antibodies were diluted with R2 buffer.

[0061] Preparation of magnetic separation reagents: 1) Prepare magnetic particles coated with anti-fluorescein isothiocyanate monoclonal antibody; 2) Prepare buffer solution a...

Embodiment 1

[0079] Example 1 A magnetic particle chemiluminescence detection kit for determining the protein content of human Golgi apparatus includes R1 reagent, R2 reagent, magnetic separation reagent, calibrator solution series and substrate solution.

[0080] In this example, the R1 reagents include: 1) R1 monoclonal antibody: fluorescein isothiocyanate (FITC)-labeled anti-Golgi protein monoclonal antibody, the concentration is 0.5 μg / mL; 2) buffer: Tris, the concentration 12.04g / L; sodium azide, the concentration is 0.987g / L; sodium chloride, the concentration is 5.79g / L, bovine serum albumin, the concentration is 9.86g / L; 8-aniline-1-naphthalenesulfonate ammonium Salt, the concentration is 0.986g / L; the rest is deionized water. The buffer pH of the R1 reagent is preferably 7.0.

[0081] In this example, the R2 reagents include: 1) R2 polyclonal antibody: alkaline phosphatase-labeled polyclonal antibody to Golgi protein at a concentration of 0.5 μg / ml; 2) buffer: commercialized AP C...

Embodiment 2

[0085] Example 2 Preparation and measurement method of magnetic particle chemiluminescence detection kit for measuring human Golgi protein content

[0086] (1) First, add the 20 μl calibrator series of Example 1 (concentrations are 0, 20, 50, 200, 500, 1000 ng / ml) and 50 μL reagent R1 and 50 μL reagent R2 of Example 1 respectively into the reaction tube medium, mix and incubate at 37°C for 10 minutes;

[0087] (2) Combine the above reagent series with 25 μL of the magnetic separation reagent of Example 1, and then continue to incubate at 37° C. for 10 min;

[0088] (3) Wash 3 times with cleaning solution to remove unbound antibodies and impurities;

[0089] (4) Add 150 μL of the luminescence substrate solution in Example 1, and use the Leadman self-developed chemiluminescence detector to measure the relative luminescence intensity (RLU) after ALP catalyzes the luminescence of the substrate. The results are shown in the following table:

[0090] Table 1

[0091] Ca...

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Abstract

The invention relates to a magnetic micro-particle chemiluminiscence detection kit for determining the content of Golgi body protein of a human body. The kit comprises a Golgi body protein R1 reagent,a Golgi body protein R2 reagent, a magnetic separation reagent, a calibrator solution series and a chemiluminiscence substrate solution, wherein the Golgi body protein R1 reagent is an anti-Golgi body protein mouse monoclonal antibody diluent marked by fluorescein isothiocyanate, the Golgi body protein R2 reagent is an alkaline phosphatase marked Golgi body protein polyclonal antibody diluent, the magnetic separation reagent is a magnetic micro-particle diluent coated by an anti-fluorescein isothiocyanate mouse monoclonal antibody, the Golgi body protein calibrator solution is composed of a synthesized Golgi body protein antigen and a buffer solution, and the luminescent substrate solution is a Tris-HCl buffer solution which is catalyzed by alkaline phosphatase and contains dioxacycloethane. According to the invention, the signal intensity and sensitivity of immune response are greatly improved, and a more accurate, accurate, convenient, rapid and simple method is provided for the detection of the Golgi body protein.

Description

technical field [0001] The invention relates to the field of medical diagnostic reagents, in particular to a magnetic particle chemiluminescence detection kit for detecting human Golgi body protein content using magnetic particle chemiluminescence technology and antigen-antibody combination technology. Background technique [0002] Golgi protein (GP73) is a transmembrane Golgi glycoprotein including 400 amino acid residues, located in the 9q21.33 chromosome genome, also known as GOLM1 or GOLPH2. GP73 is composed of inner and outer C-terminals and hydrophobic N-3, and has a complete transmembrane structure. In normal people, GP73 is only expressed in gallbladder epithelial cells, almost not expressed in liver cells, but has a higher expression level in bile duct epithelial cells. When the liver is inflamed, the level of GP73 in liver cells increases, and the expression level of GP73 in liver cancer cells increases significantly. Relevant data show that the regulation mechan...

Claims

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Application Information

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IPC IPC(8): G01N33/574G01N33/576G01N33/58G01N33/577G01N33/543G01N33/533G01N21/76
CPCG01N33/57438G01N33/57496G01N33/576G01N33/582G01N33/577G01N33/54326G01N33/533G01N21/76G01N2333/47Y02A50/30
Inventor 李晓娟袁方马道一王鹏
Owner BEIJING LEADMAN BIOCHEM
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