Pasteurella multocida toxin monoclonal antibody and blocking ELISA kit thereof

A technology of monoclonal antibody and Pasteurella, applied in bacterial peptides, biological testing, anti-bacterial immunoglobulin, etc., can solve problems such as difficult to develop kits

Pending Publication Date: 2022-01-28
JIANGSU NANNONG HI TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In the existing detection of Pasteurella multocida toxin protein and its antibody, the whole protein is easy to degrade, it is difficult to develop a kit, and there is a contradiction between the sensitivity and specificity of the detection method. Therefore, we propose a Pasteurella multocida toxin monoclonal antibody and kit comprising same

Method used

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  • Pasteurella multocida toxin monoclonal antibody and blocking ELISA kit thereof
  • Pasteurella multocida toxin monoclonal antibody and blocking ELISA kit thereof
  • Pasteurella multocida toxin monoclonal antibody and blocking ELISA kit thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Embodiment 1 Preparation and Identification of Hybridoma Cell Line and the Monoclonal Antibody It Secretes

[0025] 1 Materials and methods

[0026] 1.1 Strains, plasmids, cells and experimental animals

[0027] The pET32a-toxC plasmid is preserved in the laboratory for the expression of PMT-C-terminal truncated protein; resuscitated myeloma cells SP2 / 0 preserved in the laboratory; cell fusion uses BALB / c mice and ICR mice provided by feeder cells in Ordered from Nanjing Qinglongshan Animal Center.

[0028] Wherein, the coding nucleotide sequence of the ToxC gene is shown in SEQ ID NO:4.

[0029] 1.2 Main experimental reagents and consumables

[0030] Purchased His tag 1mL protein purification prepacked column from GE; SDS-PAGE kit was purchased from Yazyme; RPMI Medium 1640 basic (1x) was purchased from Gibco; Freund’s complete adjuvant (FCA) was purchased from Sigma , Freund's incomplete adjuvant (FIA), PEG4000, HAT Media Supplement, HT Media Supplement; FBS serum...

Embodiment 2

[0104] Example 2 Establishment of Pasteurella multocida toxin monoclonal antibody blocking ELISA detection method

[0105] 1 Materials and methods

[0106] 1.1 Experimental animals and reagents

[0107] 2 New Zealand white rabbits of 2-3kg; ISA-15A adjuvant; Escherichia coli containing plasmid PET32a-toxC has been constructed in the laboratory; enzyme-labeled antibody 3E3; 1mL protein purification prepacked column produced by GE company; ELISA detection plate purchased From Costar Company; TMB chromogenic solution was purchased from Beyontian Biological Company; BCA detection kit was purchased from Kangwei Century Company; other chemical reagents were of domestic analytical grade.

[0108] The indirect ELISA method established in our laboratory screened clinical PMT standard negative serum; the prepared rabbit polyclonal antibody was used as a standard positive control; and the reference positive serum kept in the laboratory.

[0109] 1.2 Preparation of antigenic protein and...

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Abstract

The invention discloses a pasteurella multocida toxin monoclonal antibody and a blocking ELISA kit thereof, the kit comprises a coated elisa plate and a horse radish peroxidase labeled monoclonal antibody, the preservation number of a hybridoma cell strain is CCTCC NO: C2021111, the pasteurella multocida toxin monoclonal antibody is secreted by the hybridoma cell strain, the coated elisa plate takes pasteurella multocida toxin C-terminal truncated protein as a coating antigen. The kit disclosed by the invention is used for detecting the toxin-producing pasteurella multocida by taking a blocking ELISA (Enzyme-Linked Immunosorbent Assay) method as a principle, has the advantages of high sensitivity and specificity, good repeatability, rapidness and convenience in detection, easiness in standardization and the like, and is very convenient.

Description

technical field [0001] The invention relates to the technical field of pleuropneumonia, in particular to a pasteurella multocida toxin monoclonal antibody and a blocking ELISA kit. Background technique [0002] Porcine progressive atrophic rhinitis (PAR) is a chronic infectious disease of the respiratory system, and about 25%-50% of the world's pigs are infected; due to the introduction of breeding pigs from abroad, the disease was introduced into my country through multiple channels, resulting in cost The disease is widely spread and popular; it is characterized by rhinitis, facial deformation, atrophy and growth retardation of the turbinate bone, especially the curl under the turbinate bone; it damages the normal structure and function of the respiratory tract, reduces the resistance of the pig body, and is extremely susceptible to other pathogens. Cause respiratory syndrome and increase the death rate of pigs. [0003] Toxigenic Pasteurella multocida (T+Pm) is a bipolar st...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/12C12N5/20G01N33/68G01N33/577G01N33/543C07K14/195
CPCC07K16/12G01N33/68G01N33/577G01N33/543C07K14/195
Inventor 董彦鹏李玉峰王天宇肖澄周琳胡静雅车巧林缪芬芳
Owner JIANGSU NANNONG HI TECH
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