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Canine distemper virus H protein nano antibody and preparation process thereof

A technology of canine distemper virus and nano-antibody, which is applied in the direction of antiviral immunoglobulin, DNA preparation, immunoglobulin, etc., can solve the problems of cumbersome expression system and high antibody production cost, so as to avoid cross-reaction, low cost, The effect of small molecular weight

Pending Publication Date: 2022-03-15
SHANDONG VOCATIONAL ANIMAL SCI & VETERINARY COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Aiming at the deficiencies of the prior art, the present invention provides a canine distemper virus H protein nanobody antibody and its preparation process, which solves the problems of high production cost and cumbersome expression system of the existing antibody against canine distemper H protein

Method used

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  • Canine distemper virus H protein nano antibody and preparation process thereof
  • Canine distemper virus H protein nano antibody and preparation process thereof
  • Canine distemper virus H protein nano antibody and preparation process thereof

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Embodiment 1

[0032] Such as figure 2 As shown, the embodiment of the present invention provides a canine distemper virus H protein nanobody, the nanobody uses H protein as a target, and the amino acid sequence of the H protein nanobody is shown in SEQ ID No.1.

[0033] The nanobody is coded by a DNA molecule, and the nucleotide sequence of the DNA molecule is shown in SEQ ID No.2.

[0034] A preparation process of canine distemper virus H protein nanobody, comprising the following steps:

[0035] S1. Bactrian camels immunized with inactivated vaccine

[0036]Inject 3mL of inactivated canine distemper vaccine into the neck of Bactrian camels. The injection method is subcutaneous injection. The interval between each immunization injection is two weeks. Blood 100mL;

[0037] S2. Isolation of cells

[0038] Add 3mL of blood and 3mL of PBS solution into the centrifuge tube to dilute by blowing and mixing, then take a new centrifuge tube and add 5mL of ficoll separation solution dropwise, a...

Embodiment 2

[0051] Such as figure 2 As shown, the embodiment of the present invention provides a canine distemper virus H protein nanobody, the nanobody uses H protein as a target, and the amino acid sequence of the H protein nanobody is shown in SEQ ID No.1.

[0052] The nanobody is coded by a DNA molecule, and the nucleotide sequence of the DNA molecule is shown in SEQ ID No.2.

[0053] A preparation process of canine distemper virus H protein nanobody, comprising the following steps:

[0054] S1. Bactrian camels immunized with inactivated vaccine

[0055] Inject 2 mL of inactivated canine distemper vaccine into the neck of Bactrian camels, the injection method is subcutaneous injection, and the interval between each immunization injection is two weeks. Blood 100mL;

[0056] S2. Isolation of cells

[0057] Add 2mL of blood and 2mL of PBS solution into the centrifuge tube to dilute by blowing and mixing, then take a new centrifuge tube and add 3mL of ficoll separation solution dropw...

Embodiment 3

[0070] Such as figure 2 As shown, the embodiment of the present invention provides a canine distemper virus H protein nanobody, the nanobody uses H protein as a target, and the amino acid sequence of the H protein nanobody is shown in SEQ ID No.1.

[0071] The nanobody is coded by a DNA molecule, and the nucleotide sequence of the DNA molecule is shown in SEQ ID No.2.

[0072] A preparation process of canine distemper virus H protein nanobody, comprising the following steps:

[0073] S1. Bactrian camels immunized with inactivated vaccine

[0074] Inject 2.5mL of canine distemper inactivated vaccine into the neck of Bactrian camels. The injection method is subcutaneous injection. The interval between each immunization injection is two weeks. After the fifth immunization injection, use a syringe to collect in the neck vein of Bactrian camels Peripheral blood 100mL;

[0075] S2. Isolation of cells

[0076] Add 2.5mL of blood and 2.5mL of PBS solution into the centrifuge tube...

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Abstract

The invention provides a canine distemper virus H protein nano antibody and a preparation process thereof, and relates to the technical field of nano antibodies. The preparation process of the canine distemper virus H protein nano antibody comprises the following steps: S1, immunizing a bactrian camel with an inactivated vaccine; s2, separating cells; s3, amplifying a VHH gene and constructing a display vector; s4, constructing a nano antibody library and measuring the capacity of the library; s5, measuring the titer of the recombinant bacteriophage; s6, elutriating H protein specific recombinant bacteriophage; enriching the specific recombinant bacteriophage of the S7H protein; the method comprises the following steps: immunizing a camel with a canine distemper inactivated vaccine, separating peripheral blood lymphocytes of the camel, amplifying a VHH gene of a bactrian camel by using a nested PCR (Polymerase Chain Reaction) method, connecting the VHH gene into a pCANTAB 5E phage vector, constructing a phage display library, and screening by using a phage display technology to obtain three H protein specific nano antibodies, tests prove that the nano antibodies can specifically react with H protein, and have the characteristics of small molecular weight, good stability and high affinity.

Description

technical field [0001] The invention relates to the technical field of nano-antibodies, in particular to a canine distemper virus H protein nano-antibody and a preparation process thereof. Background technique [0002] Canine distemper is one of the most serious infectious diseases of canines, mustelidae and raccoons caused by canine distemper virus; canine distemper is a pantropic virus, which can infect many tissues and organs, and only through clinical symptoms Observation and pathological changes are difficult to diagnose canine distemper; canine distemper genome encodes nucleocapsid protein (N), phosphoprotein (P), matrix membrane protein (M), large protein (L), fusion protein (F) and hemagglutinin (H) 6 proteins, and the H protein is necessary for the invasion of the host during the process of canine distemper infection, and is also the main antigen for the production of neutralizing antibodies, as well as the most variable structural antigen. [0003] At present, the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/10C12N15/13C12N15/10
CPCC07K16/1027C07K16/005C12N15/10C12N15/1037C07K2317/569C07K2317/22C07K2317/92C07K2317/94C07K2317/33C12Q2531/113
Inventor 朱光王译晨朱绍辉肖发沂齐艳君王小伟张彤晨柏诚昊
Owner SHANDONG VOCATIONAL ANIMAL SCI & VETERINARY COLLEGE
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