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Beta-glucosidase monoclonal antibody and preparing method thereof

A technology of glucosidase and cloning antibody, which is applied in the field of β-glucosidase monoclonal antibody and its preparation, which can solve the problems of difficult detection and easy volatilization of hydrocyanic acid, etc., and achieve high stability, high titer and high specificity sexual effect

Inactive Publication Date: 2016-09-21
TROPICAL CORP STRAIN RESOURCE INST CHINESE ACAD OF TROPICAL AGRI SCI
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AI Technical Summary

Problems solved by technology

Anne VintherMorant et al found that after the cassava tissue is damaged, the cyanogenic glucoside located in the vacuole interacts with the β-glucosidase from the milk duct or cell wall and is degraded into hydrocyanic acid, which is easy to volatilize , it is not easy to detect

Method used

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  • Beta-glucosidase monoclonal antibody and preparing method thereof
  • Beta-glucosidase monoclonal antibody and preparing method thereof
  • Beta-glucosidase monoclonal antibody and preparing method thereof

Examples

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Embodiment 1

[0025] The preparation method of β-glucosidase monoclonal antibody provided by the present invention has the following steps:

[0026] S1: Take antigen β-glucosidase injection to immunize mice:

[0027] Take out 250μl QuickAntibody-mouse 5w (5-week mouse rapid immunization adjuvant) under aseptic conditions, mix it with 250μl normal saline containing 250μg antigen β-glucosidase, and immunize mice by intramuscular injection in the hind leg of the mouse. β-glucosidase 50μg / mouse was injected to immunize mice. In this experiment, each mouse was injected with 100μl, and 5 mice were injected altogether; on the 21st day, each mouse was immunized with another booster shot in the same manner as above.

[0028] In this example, a 5-week mouse rapid immune adjuvant is used to replace the traditional Freund’s adjuvant, so that the β-glucosidase antigen can be slowly released in the mouse body. The adjuvant does not need to be emulsified, does not destroy the antigen structure, and avoids The d...

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Abstract

The invention provides a preparing method for a beta-glucosidase monoclonal antibody. The method comprises the steps of 1, injecting antigen beta-glucosidase into immunized mice; 2, measuring the serum titer of each immunized mouse with the indirect ELISA method; 3, selecting the mice with serum titer higher than 20000, and fusing splenocytes with myeloma cells; 4, conducting culture with HAT culture solution and conducting indirect ELISA test screening to obtain positive hybridoma cells; 5, cloning the positive hybridoma cells with the limiting dilution method till positive monoclonal rate is 100%; 6, preparing the beta-glucosidase monoclonal antibody with the mouse in-vivo ascites induction method; 7, purifying the beta-glucosidase monoclonal antibody. The beta-glucosidase monoclonal antibody prepared with the method has high titer, stability and specificity.

Description

Technical field [0001] The invention relates to the technical field of monoclonal antibodies, in particular to a β-glucosidase monoclonal antibody and a preparation method thereof. Background technique [0002] β-Glucosidase (β-Glucosidase, EC3.2.1.21), also known as β-D-glucosidase glucohydrolase, generally has a molecular weight between 40-250Kd. The relative molecular weight of β-glucosidase from different sources is determined by its molecular weight. The structure and composition differ greatly. It can hydrolyze the non-reducing β-D-glucosidic bond at the end and release β-D-glucose and the corresponding ligand at the same time. In 1837, Liebig and Wohler were first discovered in bitter almonds. Later studies found that β-glucosidase exists in many plants, insects, yeast, Aspergillus, Trichoderma and bacteria in nature. It participates in the carbohydrate metabolism of the organism and plays an important role in maintaining the normal physiological functions of the organi...

Claims

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Application Information

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IPC IPC(8): C07K16/40
CPCC07K16/40
Inventor 余厚美张振文林立铭王琴飞徐媛
Owner TROPICAL CORP STRAIN RESOURCE INST CHINESE ACAD OF TROPICAL AGRI SCI
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