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Polypeptides for promoting swine body to generate African swine fever virus (ASFV) antigen-specific immune response and application of polypeptides

An African swine fever virus and immune response technology, applied in the field of biomedicine, can solve problems that have not yet been reported, achieve no toxic side effects and hidden dangers of biological safety, promote the proliferation of sensitized immune cells, and promote immune response.

Active Publication Date: 2021-01-12
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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  • Polypeptides for promoting swine body to generate African swine fever virus (ASFV) antigen-specific immune response and application of polypeptides
  • Polypeptides for promoting swine body to generate African swine fever virus (ASFV) antigen-specific immune response and application of polypeptides
  • Polypeptides for promoting swine body to generate African swine fever virus (ASFV) antigen-specific immune response and application of polypeptides

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Solid phase synthesis and purity detection of embodiment 1 polypeptide

[0062] The polypeptide of the present invention is as follows:

[0063] SEQ ID NO. 1: SMAAKIFIV, AP1;

[0064] SEQ ID NO. 2: STQAYNDFL, AP2;

[0065] SEQ ID NO.3: FQMNVSACAW, AP3;

[0066] SEQ ID NO.4: PPTQRVDPA, AP4;

[0067] SEQ ID NO.5: NSTRFTTEDP, AP5;

[0068] 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

[0069] Embodiment 2 porcine lymphocyte proliferation experiment

[0070] 1. Pigs immunized with inactivated ASFV virus.

[0071] 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.

[0072] 2. Preparation, culture and proliferation detection of splenocytes.

[0073] 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.

[0074] 2) Then draw the liquid into a 15ml centrifuge tube, centrifuge at 1000rpm for 5min.

[0075] 3) Discard the supernatant and tap the...

Embodiment 3

[0083] Example 3 Detection of IFN-γ secretion by different subtypes of splenic lymphocytes

[0084] 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.

[0085] For test resul...

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Abstract

The invention provides polypeptides for promoting a swine body to generate an African swine fever virus (ASFV) antigen-specific immune response and application of the polypeptides, and belongs to thetechnical field of biological medicines. The polypeptides comprises one or more of a first polypeptide, a second polypeptide, a third polypeptide, a fourth polypeptide and a fifth polypeptide; the amino acid sequence of the first polypeptide, the amino acid sequence of the second polypeptide, the amino acid sequence of the third polypeptide, the amino acid sequence of the fourth polypeptide and the amino acid sequence of the fifth polypeptide are as shown by SEQ ID NO.1-SEQ ID NO.5 respectively. The polypeptides provided by the invention can obviously promote the proliferation of ASFV sensitized immune cells, and can promote the swine body to generate the ASFV antigen-specific immune response by promoting swine mononuclear macrophages and B and T lymphocytes to secrete interferon-gamma (IFN-gamma). After an animal is immunized, the immune response of the swine pig body can be remarkably promoted. The polypeptides or a polypeptide polymer obtained by polymerization of the polypeptides can be used for preparing subunit vaccines of the African swine fever virus.

Description

technical field [0001] 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 [0002] 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 through different mechanisms, such as type I interferon (IFN) response, apoptosis, inflammatory response and Activation of specific target genes. [0003] Vaccines are one of the most effective ways to prevent African swine fever. Traditional attenuated live vaccines and inactivated vaccine...

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忠献张克山王尧河刘湘涛
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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