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
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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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