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Preparation of immuno-stimulation composition for hemolycin in monad

A technology for Aeromonas hydrophila and immunostimulation, applied in the fields of molecular biology and immunology, can solve the problems of Aeromonas hydrophila complex antigen structure, weak immunogenicity, inconvenient preparation of combined vaccines, etc., to achieve Strong immune protection, stable immune effect, and remarkable synergistic effect

Inactive Publication Date: 2005-02-23
INST OF ANIMAL HUSBANDRY & VETERINARY FUJIAN ACADEMY OF AGRI SCI +3
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  • Abstract
  • Description
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AI Technical Summary

Problems solved by technology

However, due to the large number of serotypes of Aeromonas hydrophila, there are differences in immunogenicity between strains, and the complex antigen structure of Aeromonas hydrophila makes it difficult to prepare combined vaccines. The antigenicity of whole vaccines often changes after inactivation. Furthermore, the whole bacterin vaccine has large particles and is less likely to be absorbed. The above factors have restricted the popularization of the whole bacterium inactivated vaccine.
[0005] The existing genetically engineered subunit vaccines retain the advantages of traditional subunit vaccines, such as: good safety, specific effective immune components, easy industrial production, and low production costs, but the immunogenicity is not strong
The use of adjuvants in subunit vaccines can significantly enhance the immunogenicity of subunit vaccines. Conventional adjuvants such as aluminum hydroxide and propolis, which are commonly used in clinical practice, have limited ability to improve immunogenicity, while Freund's adjuvant has a strong Synergistic effect, but its toxicity and side effects on animals are large, so it is banned from clinical use

Method used

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  • Preparation of immuno-stimulation composition for hemolycin in monad
  • Preparation of immuno-stimulation composition for hemolycin in monad
  • Preparation of immuno-stimulation composition for hemolycin in monad

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] 1. Required materials

[0064] (1) Main reagents

[0065] Quil A was purchased from Beijing Zhongjian Weikang Technology Co., Ltd.; Mega-10 was purchased from Huamei Bioengineering Company.

[0066] (2) Strains

[0067] Aeromonas hydrophila L-316 was isolated and identified by the Fish Disease Center of the Institute of Animal Husbandry and Veterinary Medicine, Fujian Academy of Agricultural Sciences, and its β-hemA recombinant bacteria were constructed.

[0068] (3) Experimental animals

[0069] Eel, purchased from Changle Eel Farm in Fujian Province, size: about 200g.

[0070] Two, the preparation technology of the immunostimulatory complex of hemolysin in Aeromonas hydrophila, has the following steps:

[0071] 1. Extract the recombinant bacteria of Aeromonas hydrophila L316,

[0072] 2. In the culture medium, the bacterial strain is expanded and cultivated to obtain a culture solution,

[0073] (1) inoculate the bacterial strain on common agar medium,

[0074]...

Embodiment 2

[0125] The citric acid in the step of the preparation process of the immunostimulatory complex of hemolysin in Aeromonas hydrophila of Example 1 is replaced with the glycine buffer solution of pH2.5 for preparing the immunostimulatory complex of Aeromonas hydrophila thing;

[0126] Concentrate the toxin to 1 mg / ml and add glycine buffer solution with pH 2.5 for 1 hour;

[0127] Add Quil A adjuvant to a final concentration of 0.05%;

[0128] The remaining experimental materials and experimental procedures are the same as in Example 1. Experimental results: the 40ug dose group has the best effect, and the protection rate reaches 100%, followed by the 80ug dose group. It can be seen that the toxin protein is treated with glycine and citric acid. The effect of the immunostimulatory complex for humoral immunity to immune receptors is basically the same.

[0129] Best practice:

[0130] The steps of the preparation process of the immunostimulatory complex of hemolysin in Aeromona...

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Abstract

An immunostimulating complex with stable and durable immune effect is prepared from hygrophilic monodas through liquid culturing, centrifugal separating of supernatant, treating it by saturated ammonium sulfate solution, suspending its deposit by PBS dialyzing to obtain toxic, concentrating, adding organic acid, adding Mega-10, treating at 37 deg.C for 4 hr, adding Quil A, cholesterol and lecithin, treating by organic acid, and dialyzing.

Description

technical field [0001] The invention relates to the fields of molecular biology and immunology, in particular to a preparation process of the hemolysin immunostimulatory complex in Aeromonas hydrophila. Background technique [0002] Aeromonas hydrophila (Aeromonas hydrophila, Ah) belongs to Vibriaceae Aeromonas genus, is a pathogenic bacterium of humans, fish, amphibians, reptiles and mammals, and can cause fish bacterial sepsis, frog Diseases such as red leg disease and human diarrhea cause a large amount of economic losses every year. [0003] The pathogenic factors of Aeromonas hydrophila include exotoxin, extracellular enzyme, membrane adhesion factor, S layer protein, pili, transferrin and outer membrane protein. The exotoxin of the Ah pathogenic strain is an important pathogenic factor, and its research has been quite in-depth. Different researchers give different names to exotoxins, such as HEC toxin reported by Lu Chengping, enterotoxin reported by Rose et al., aer...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/02A61K39/39A61P31/04C12P21/00
Inventor 方勤美龚晖林天龙
Owner INST OF ANIMAL HUSBANDRY & VETERINARY FUJIAN ACADEMY OF AGRI SCI
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