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Polypeptide for promoting swine body to generate African swine fever virus antigen specific immune response and application thereof

一种非洲猪瘟病毒、免疫应答的技术，应用在生物医学领域，能够解决尚未见报道等问题，达到没有毒副作用以及生物安全隐患、促进致敏免疫细胞增殖、促进免疫应答的效果

Active Publication Date: 2022-01-07
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But so far, no fully effective ASFV subunit vaccine has been reported.

Method used

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  • Polypeptide for promoting swine body to generate African swine fever virus antigen specific immune response and application thereof
  • Polypeptide for promoting swine body to generate African swine fever virus antigen specific immune response and application thereof
  • Polypeptide for promoting swine body to generate African swine fever virus antigen specific immune response and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] Solid phase synthesis and purity detection of embodiment 1 polypeptide

[0063] The polypeptide of the present invention is as follows:

[0064] SEQ ID NO. 1: SMAAKIFIV, AP1;

[0065] SEQ ID NO. 2: STQAYNDFL, AP2;

[0066] SEQ ID NO.3: FQMNVSACAW, AP3;

[0067] SEQ ID NO.4: PPTQRVDPA, AP4;

[0068] SEQ ID NO.5: NSTRFTTEDP, AP5;

[0069] The above polypeptides were synthesized by Shanghai Sangon Bioengineering Co., Ltd. The detection wavelength is 214nm. The purity of the final peptide purification product was >98%, and the structure was identified by ESI-MS. For the identification results, see Figure 1 to Figure 10 ,in figure 1 is the identification chromatogram of AP1, figure 2 It is the mass spectrum of AP1, and the identified molecular weight is 979.55g / mol, which is consistent with the theoretical value; image 3 is the identification chromatogram of AP2, Figure 4 It is the mass spectrum of AP2, and the identified molecular weight is 1058.5g / mol, which ...

Embodiment 2

[0070] Embodiment 2 porcine lymphocyte proliferation experiment

[0071] 1. Pigs immunized with inactivated ASFV virus.

[0072] Five 90-day-old male Landrace pigs were immunized with ASFV epidemic strain inactivated virus (from the African swine fever regional laboratory of Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences) (10HID50). Euthanasia was performed and the spleen was removed after dissection. Unimmunized healthy pigs were used as negative control group.

[0073] 2. Preparation, culture and proliferation detection of splenocytes.

[0074] 1) Aseptically treat the collected pig spleen with 75% alcohol, wash it three times with PBS, cut the spleen into small pieces, place them in folded sterile gauze (2 layers), and place them in 5 mL of serum-containing 1640 medium Grind the spleen in a plate.

[0075] 2) Then draw the liquid into a 15ml centrifuge tube, centrifuge at 1000rpm for 5min.

[0076] 3) Discard the supernatant and tap the...

Embodiment 3

[0084] Example 3 Detection of IFN-γ secretion by different subtypes of splenic lymphocytes

[0085] The pig immunization procedure and spleen cell isolation and culture are the same as in Example 2. After obtaining the dispersed splenocytes, prepare a single cell suspension with RPMI1640 complete medium at a concentration of 1×10 6 / ml. Inoculate into a 24-well plate, add 0.2 μg of the polypeptide (AP1-AP5) provided by the present invention to each well, and place in CO 2 Cultivate in the incubator for 60h. Collect the cells in each well, label the cells with porcine CD3, CD4, CD8 and IFN-γ specific antibodies respectively, then wash twice with PBS buffer containing 2% serum, and finally use the washing solution to disperse into a cell suspension, and use the flow Cytometer detection, respectively determine B lymphocytes, CD4 + T lymphocytes, CD8 + T lymphocytes, CD4 + CD8 + The levels of IFN-γ secreted by T lymphocytes and monocyte-macrophages.

[0086] For test resul...

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Abstract

The invention provides a polypeptide for promoting a swine body to generate African swine fever virus antigen specific immune response and application thereof, and belongs to the technical field of biological medicines. The polypeptide comprises one or more of a first polypeptide, a second polypeptide, a third polypeptide, a fourth polypeptide and a fifth polypeptide; the amino acid sequences of the first polypeptide, the second polypeptide, the third polypeptide, the fourth polypeptide and the fifth polypeptide are respectively shown as SEQ ID NO. 1 to SEQ ID NO. 5. The polypeptide provided by the invention can obviously promote ASFV sensitized immune cell proliferation, and can promote the swine body to generate ASFV antigen specific immune response by promoting swine mononuclear macrophages and lymphocytes to secrete IFN-gamma. After animals are immunized, the immune response of the swine body can be obviously promoted. The polypeptide or a polypeptide polymer obtained by polypeptide polymerization can be used for preparing a subunit vaccine of African swine fever virus.

Description

[0001] This application is a divisional application with an application date of August 26, 2020, an application number of CN202010873520.9, and an invention title of "A Class of Polypeptides and Applications for Promoting African Swine Fever Virus Antigen-Specific Immune Response in Pigs". technical field [0002] The invention relates to the technical field of biomedicine, in particular to a class of polypeptides for promoting African swine fever virus antigen-specific immune response in pig bodies and applications thereof. Background technique [0003] African swine fever (African Swine fever, ASF) is an acute, hemorrhagic, severe disease caused by African swine fever virus (ASFV) infecting domestic pigs and various wild boars (such as African wild boars, European wild boars, etc.). infectious disease. African swine fever virus mainly infects immune cells such as macrophages and monocytes. After infection, ASFV escapes the host's innate and inherent immune defense system t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/01A61K39/12A61P31/20
CPCC07K14/005A61K39/12A61P31/20C12N2710/12022C12N2710/12034A61K2039/552Y02A50/30
Inventor 郑海学毛箬青杨帆周晓丽孙德惠刘华南张克山刘湘涛
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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