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Polypeptide for promoting pig body to generate African swine fever virus antigen specific immune response, and application of polypeptide

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

Active Publication Date: 2020-11-17
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 pig body to generate African swine fever virus antigen specific immune response, and application of polypeptide
  • Polypeptide for promoting pig body to generate African swine fever virus antigen specific immune response, and application of polypeptide
  • Polypeptide for promoting pig body to generate African swine fever virus antigen specific immune response, and application of polypeptide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0060] Solid phase synthesis and purity detection of embodiment 1 polypeptide

[0061] The polypeptide of the present invention is as follows:

[0062] SEQ ID NO.1: SMMMVFNQL, AP1;

[0063] SEQ ID NO.2: RGLMEITFML, AP2;

[0064] SEQ ID NO.3: MQWFMTMVI, AP3;

[0065] SEQ ID NO.4: YKLHGMRWF, AP4;

[0066] SEQ ID NO.5: HLLNFTLHCHA, AP5;

[0067] 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 It is the identification chromatogram of AP1, and the peak time is 8.607min; figure 2 It is the mass spectrum of AP1, and the identified molecular weight is 1100.45g / mol, which is consistent with the theoretical value; image 3 It is the identification chromatogram of AP2, and the peak time is 15.534min; Figure 4 It is the mass spectr...

Embodiment 2

[0068] Embodiment 2 porcine lymphocyte proliferation experiment

[0069] 1. Pigs immunized with inactivated ASFV virus.

[0070] 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), and a booster immunization was performed one month later, and the pigs were administered 7 days later. Euthanize and remove the spleen after dissection. Unimmunized healthy pigs were used as negative control group.

[0071] 2. Preparation, culture and proliferation detection of splenocytes.

[0072] 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.

[0073] 2) Then draw the liquid into a 15ml centrifuge tub...

Embodiment 3

[0082] Example 3 Detection of IFN-γ secretion by different subtypes of splenic lymphocytes

[0083] 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.

[0084] For test resul...

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Abstract

The invention provides a polypeptide for promoting a pig body to generate the African swine fever virus antigen specific immune response, and an application of the polypeptide, 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, wherein amino acid sequences of the first polypeptide, the second polypeptide, the third polypeptide, the fourth polypeptide and the fifth polypeptide are shown as SEQ ID NO.1-SEQ ID NO.5 respectively. The polypeptide provided by the invention can significantly promote the proliferation of ASFV sensitized immune cells, and can promote the pig body to generate the ASFV antigen specific immune response by promoting pig CD8<+>T cells to secrete IFN-gamma. After animals are immunized, the immune response of the pig body can be remarkably promoted. The polypeptide or a polypeptide polymer obtained by polypeptide polymerization can be used for preparing subunit vaccines of 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
CPCC07K14/005A61K39/12A61P31/20C12N2710/12022C12N2710/12034A61K2039/552
Inventor 郑海学毛箬青杨帆张忠献王尧河孙德惠周晓丽朱昱茜张克山刘华南刘湘涛
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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