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Bovine type-A clostridium perfringens subunit vaccine and preparation method and application thereof

A technology for Clostridium perfringens and subunit vaccines is applied in chemical instruments and methods, antibacterial drugs, bacterial antigen components, etc., to achieve the effects of simple process, cheap consumables, and high yield

Active Publication Date: 2018-01-19
NOVO BIOTECH CORP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The technical problem to be solved by the present invention is to provide a large-scale industrial production of bovine Clostridium perfringens type A subunit vaccine and its preparation method; The third is to overcome the defects that may exist when the α-toxin is inactivated in the prior art

Method used

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  • Bovine type-A clostridium perfringens subunit vaccine and preparation method and application thereof
  • Bovine type-A clostridium perfringens subunit vaccine and preparation method and application thereof
  • Bovine type-A clostridium perfringens subunit vaccine and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] The preparation of embodiment 1 alpha-toxin protein truncated body

[0023] 1. Selection of truncation position of 1α-toxin protein truncated body

[0024] Alpha toxin is the main pathogenic factor of Clostridium perfringens type A, which can stimulate the body to produce neutralizing antibodies. It itself is a metalloenzyme dependent on the activity of zinc ion phospholipase C (PLC), which can hydrolyze the phosphatidylcholine in the cell membrane, destroy the integrity of the cell membrane, and eventually lead to cell rupture and death. The structure of α-toxin protein (PDB: 1GYG) was analyzed, and its structure can be divided into two domains: the N-terminal structure includes nine tightly connected α-helices, and the C-terminal structure consists of 8 antiparallel β-sheets , N-terminal and C-terminal are connected by a short and flexible linker. The N-terminal domain is completely present in the phospholipase active region, while the C-terminal region containing a...

Embodiment 2

[0118] Example 2 α-toxin protein truncated mouse toxicity test

[0119] Purchase 10 18-20g BALB / C mice and randomly divide them into 2 groups with 5 mice in each group. One group was used as the control group, and 200 μl of PBS was injected into the leg muscles; the other group was injected with 200 μl (the total amount of protein was 200 μg) of α-toxin protein truncated body. Observed continuously for 7 days after injection, and observed mental appetite and measured body weight every day.

[0120] Within 7 days of injection follow-up, the mental appetites of the mice in the two groups were all good, no significant difference was seen, and the weight gain (see image 3 ) and there is no significant difference. This shows that the truncated α-toxin protein prepared by us is safe, and the tolerated amount of the mouse is very high (up to 200 μg at the lowest), which far meets the requirements for vaccine preparation.

Embodiment 3

[0121] Example 3 Preparation of bovine type A Clostridium perfringens subunit vaccine

[0122] Add an appropriate amount of truncated α-toxin protein to ISA 201VG adjuvant (the weight ratio of antigen phase and adjuvant is 1:1, emulsify and store at 4°C after passing the quality inspection; the specific vaccine information is shown in the following table:

[0123]

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Abstract

The invention provides a bovine type-A clostridium perfringens subunit vaccine and a preparation method and an application thereof. The subunit vaccine comprises a bovine type-A clostridium perfringens alpha-toxin protein truncation body and a pharmacologically acceptable adjuvant. The bovine type-A clostridium perfringens alpha-toxin protein truncation body is an antigen determinant amino acid sequence from 255th to 372nd amino acids of alpha-toxin protein. The subunit vaccine has the following advantages that 1) safety problems due to uncompleted inactivation are not existed; 2) the qualityis controllable, and the difference between the batches is not existed; 3) the production equipment and space requirement is low, the expression level is high, and the cost is low; and 4) toxin dispersion risk is not existed, and the safety of operators can be guaranteed.

Description

technical field [0001] The invention relates to the preparation and application of a bovine type A Clostridium perfringens subunit vaccine and belongs to the technical field of veterinary biological products. Background technique [0002] Clostridium perfringens type A is an anaerobic, spore-forming and thermotolerant opportunistic pathogen in nature and in the gut of animals. The bovine enterotoxemia and bovine sudden death syndrome caused by the disease all have the characteristics of acute onset, short course of disease and high mortality. Therefore, drugs often do not have a good therapeutic effect. [0003] Clostridium perfringens type A mainly produces α-toxin, which is the main immunogen and can stimulate the body to produce neutralizing antibodies. Therefore, α-toxin is the main target antigen for subunit vaccine design. However, α-toxin has phospholipase C activity, which can hydrolyze the phosphatidylcholine of the cell membrane, destroy the integrity of the cel...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/08A61P31/04C07K14/33C12N15/70
Inventor 钱泓吴有强张强王娟白志军查银河
Owner NOVO BIOTECH CORP
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