Myelin sheath oligodendroglia cell glycoprotein antibody kit and preparation method thereof

An oligodendrocyte and kit technology, applied in the fields of peptide chemistry and immunology, can solve problems such as elevation and affect the treatment process of patients, and achieve the effects of good sensitivity, high specificity and good stability

Pending Publication Date: 2019-11-22
安徽恩禾生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The amount of MOG-Ab (anti-MOG antibody) in the blood of patients with MOG-Ab disease will increase, and the existing detection methods cannot accurately detect the content of MOG-Ab in the blood of patients, which will affect the treatment process of patients

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] The composition of a myelin oligodendrocyte glycoprotein antibody kit:

[0024] 1) MOG-coated plate: a microwell plate coated with MOG antigen. 48-well / 96-well plate

[0025] 2) One set of MOG calibrator A-F: derived from human serum, concentration 0, 10, 20, 50, 100, 200 ng / ml. 1×0.5ml

[0026] 3) MOG-binding antigen: biotin-labeled human myelin oligodendrocyte glycoprotein (MOG)-binding antigen. 1×2.5ml / 1×5.0ml

[0027] 4) MOG enzyme conjugate: streptomycin-labeled horseradish peroxidase. 1×5.0ml / 2×5.0ml

[0028] 5) MOG quality control substance: derived from human serum, the quality control range is 150ng / ml±22.5ng / ml.

[0029] 1×0.5ml

[0030] 6) Color developer A: the main component is urea peroxide. 1×5.0ml

[0031] 7) Chromogenic agent B: the main ingredient is 3,3,5,5,-tetramethylbenzidine hydrochloride (TMB·2HCl).

[0032] 1×5.0ml

[0033] 8) Concentrated washing solution (25×): Concentrated phosphate buffer solution, diluted 1:24 before use.

[003...

Embodiment 2

[0039] A preparation method of myelin oligodendrocyte glycoprotein antibody kit:

[0040] 1) Preparation of MOG-coated antigen and preparation of MOG-bound antigen:

[0041] The S segment and the P segment were intercepted from the human MOG epitope peptide, respectively, and the S segment and the P segment were transformed into Escherichia coli for cultivation. The bacteria were collected and centrifuged to obtain the recombinant precipitated protein, and the recombinant protein was purified separately. The target proteins were obtained and named as S protein and P protein respectively, the S protein is the MOG-coated antigen, and the P protein is the MOG-binding antigen.

[0042] 2) Preparation of MOG-binding antigen-HRP markers:

[0043] The coupling of MOG-labeled antigen and HRP specifically adopts the NaIO4 oxidation method, including the following steps:

[0044] 2.1) Dialyze the MOG-labeled antigen obtained in step S1 in PBS solution overnight at 4°C;

[0045] 2.2) ...

Embodiment 3

[0091] A method for operating a myelin oligodendrocyte glycoprotein antibody kit, comprising the following steps:

[0092] 1) Preparation of cleaning solution: 25× concentrated cleaning solution stored at 2-8°C may form crystals at the bottom of the bottle. Use a 500ml container to fully dissolve the entire bottle of concentrated cleaning solution in 480ml of distilled or deionized water.

[0093] 2) After the kit is equilibrated to room temperature (20°C-30°C), take out the required microwell strips and fix them on the plate rack, and compile the sequence;

[0094] 3) Adding samples: first add 50ul of the sample to be tested / MOG calibrator / MOG quality control substance, then add 50ul of MOG-bound antigen to the corresponding well, pat and mix well, and set a blank well;

[0095] 4) Incubation: seal the strips with parafilm (to prevent contamination), and incubate at 37°C for 30 minutes;

[0096] 5) Plate washing: take out the reaction plate, shake off all the liquid in the p...

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Abstract

The invention relates to the technical field of polypeptide chemistry and immunology and particularly discloses a myelin sheath oligodendroglia cell glycoprotein antibody kit. The kit is used for detecting the human MOG antibody by adopting a double-antigen sandwich method. The kit comprises a coating plate, a calibration product, a binding antigen, an enzyme conjugate, a quality control product,a color developing agent A, a color developing agent B, stop buffer and concentrated washing solution, wherein the coating plate is coated with an MOG coating antigen, the MOG coating antigen is prepared by coupling an S fragment in the human MOG epitope peptide with a carrier protein, the binding antigen is prepared by coupling a P fragment in the human MOG epitope peptide with a carrier protein,the MOG binding antigen is labeled with biotin, the NCBI (National Center of Biotechnology Information) login number of the human MOG epitope peptide is CAA52617.1. The kit is advantaged in that defects in the prior art are overcome, the content of MOG-Ab in the blood of a patient is accurately determined by adopting the double-antigen sandwich method, a doctor is helped to perform treatment, andthe kit is simple to operate, sensitive and rapid.

Description

technical field [0001] The invention relates to the technical field of polypeptide chemistry and immunology, in particular to a myelin oligodendrocyte glycoprotein antibody kit. Background technique [0002] Myelin oligodendrocyte glycoprotein (MOG) is a member of the immunoglobulin superfamily, composed of 218 amino acids, and is a component of myelin in the central nervous system. MOG is expressed relatively late in the development of the nervous system, which suggests that it may be an important surface marker of oligodendrocyte maturation, and it plays an important role in maintaining the integrity of myelin sheath and promoting the exchange of information between cells. Current studies have shown that anti-MOG antibodies may be associated with immune demyelinating diseases of the central nervous system, such as acute disseminated encephalomyelitis (ADEM), multiphasic disseminated encephalomyelitis (MDEM), optic nerve Pathogenic antibodies to optic neuritis (ON), long s...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68
CPCG01N33/6854G01N33/6893
Inventor 余旭亮
Owner 安徽恩禾生物技术有限公司
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