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Preparation method of monoclonal antibody for simultaneous detection of staphylococcal enterotoxins A and B

A technology of monoclonal antibody and staphylococcus, applied in the field of immunology, can solve the problems of long cycle, high cost and low yield

Inactive Publication Date: 2012-01-25
王 小红 +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] In the process of monoclonal antibody preparation, if the traditional single immunization method of immunizing mice with one antigen at a time is changed, a mixed immunization method is used, and then conventional methods are used to screen and prepare common determinants that can target multiple enterotoxin antigens at the same time monoclonal antibody, it is possible to realize the detection of multiple enterotoxins in one reaction
Since mixed immunization is to immunize mice with multiple antigens at one time, a cell fusion experiment is performed to screen monoclonal antibodies against multiple different antigens, which overcomes the traditional monoclonal antibody preparation process, where only one antigen can be prepared for each cell fusion monoclonal antibody, and the shortcomings of high cost, low yield, complicated preparation procedures, and long cycle time have become one of the hotspots in the research of throughput antibody preparation in recent years.

Method used

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Embodiment 1 simultaneously detects the preparation scheme one of SEA and SEB monoclonal antibody

[0021] 1) Animal immunization method one

[0022] One week before the mice were injected with the antigen, 1 mg of BCG was subcutaneously injected on the back of the mice to enhance the body's immune response to the antigen. 15 μg of SEA and 15 μg of SEB were mixed in equal amounts, and the antigen was injected once on the 1st, 20th, 50th, and 80th day respectively, and each immunization dose was about 200-400 μL for each mouse. For the first immunization, the antigen was emulsified with an equal volume of complete Freund's adjuvant, and after each time it was fully emulsified with an equal volume of incomplete Freund's adjuvant to immunize BALB / c mice. For the first and second immunizations, the mice were immunized by multiple subcutaneous injections on the back of the neck, and intraperitoneal injections for subsequent immunizations.

[0023] 2) Simultaneous screening...

Embodiment 2

[0031] Embodiment 2 simultaneously detects the preparation scheme two of SEA and SEB monoclonal antibody

[0032] 1) Animal immunization method 2

[0033] One week before the mice were injected with the antigen, 1 mg of BCG was subcutaneously injected on the back of the mice to enhance the body's immune response to the antigen. 10 μg of SEA and 10 μg of SEB were mixed in equal amounts, and then the antigen was injected once on the 1st, 14th, 28th, and 42nd day respectively, and each immunization dose was about 200-400 μL for each mouse. For the first immunization, the antigen was emulsified with an equal volume of complete Freund's adjuvant, and then immunized subcutaneously at multiple points on the back of the neck of the mouse. Afterwards, each time it was fully emulsified with an equal volume of incomplete Freund's adjuvant, and then intraperitoneally immunized.

[0034] 2) Simultaneous screening for SEA and SEB hybridoma cells

[0035] ① Coating: Dilute SEA and SEB to 1...

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Abstract

The invention relates to a preparation method of a monoclonal antibody for simultaneous detection of staphylococcal enterotoxins A and B (referred to as SEA and SEB). The SEA and SEB commonly caused staphylococcal poisoning are used as immunogens, and the monoclonal antibody for simultaneous detection of SEA and SEB is prepared via a mixed immunization method, a hybridoma technology and screening in an indirect ELISA (enzyme-linked immunosorbent assay, referred to as ELISA) method. The monoclonal antibody can be used for simultaneously detecting the SEA and SEB in food via one-step reaction, and thus the workload of analysis of staphylococcal food poisoning can be reduced.

Description

technical field [0001] The invention belongs to the technical field of immunology and relates to a preparation method of a monoclonal antibody capable of simultaneously detecting type A and type B enterotoxins of Staphylococcus aureus. Background technique [0002] Staphylococcus aureus is an important zoonotic pathogen that widely exists in nature. The enterotoxin produced by it can cause food poisoning by contaminating food. Among the food poisoning caused by Staphylococcus aureus enterotoxin, enterotoxin A (staphylococcal enterotoxin A, referred to as SEA) caused the most food poisoning, in addition, enterotoxin B (staphylococcal enterotoxin B, referred to as SEB) An important toxin for purulent infection, and a biochemical weapon with great lethality. Therefore, establishing a sensitive and rapid method for the detection of enterotoxins is an important research content for food safety testing. [0003] The detection methods of enterotoxin mainly include animal test, im...

Claims

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Application Information

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IPC IPC(8): C12N5/20C07K16/12G01N33/569G01N33/577
Inventor 王小红陈福生梁斌
Owner 王 小红
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