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Canine parvovirus single-domain antibody, and preparation method and application thereof

A technology of canine parvovirus and single-domain antibody, which is applied in the direction of antibodies, antiviral agents, antiviral immunoglobulin, etc., can solve the problems of difficult to stimulate T cell immune response, complex molecular structure of whole antibody, low VHH immunogenicity, etc. problem, to achieve the effect of shortening the treatment cycle, high affinity and small molecular weight

Inactive Publication Date: 2014-03-26
SHANXI AGRI UNIV
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Problems solved by technology

At present, antibodies are limited to cell surface targets in the treatment of diseases. Full antibodies are difficult to express in cells due to their complex molecular structure, which limits their application in cells. VHH has low immunogenicity and is difficult to stimulate T cell immune response. , can be used as a therapeutic antibody for long-term administration

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  • Canine parvovirus single-domain antibody, and preparation method and application thereof
  • Canine parvovirus single-domain antibody, and preparation method and application thereof
  • Canine parvovirus single-domain antibody, and preparation method and application thereof

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Embodiment Construction

[0022] The present invention will be further described in detail below by way of examples. It should be understood that the following examples are only used to explain the present invention more clearly, but shall not limit the protection scope of the present invention. Within the scope of protection defined by the claims of the present invention, changes, modifications or equivalent changes without substantial changes shall fall within the scope of protection of the present invention.

[0023] 1. Canine parvovirus inactivated vaccine immunized alpaca

[0024] Put the CPV virus liquid into the conical flask, add analytical pure formaldehyde to dilute to a final concentration of 0.25wt%, shake well while adding, add a stopper and wrap it with sterilized gauze, and act at 37°C for 24-48 hours; remove the stopper and wrap it with gauze The mouth of the Erlenmeyer flask was shaken at intervals until the smell of formaldehyde dissipated.

[0025] Equal amount of virus liquid is e...

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Abstract

The invention discloses a canine parvovirus single-domain antibody, and a preparation method and an application thereof. The canine parvovirus single-domain antibody has a nucleotide sequence as shown in SEQIDNO.1. The preparation method comprises the steps: immunizing an alpaca by a canine parvovirus inactivated vaccine, and taking peripheral blood to obtain alpaca anti-canine parvovirus serum; extracting RNA in the peripheral blood, and carrying out reverse transcription to generate cDNA; utilizing specific primers for amplification, and purifying a VHH fragment; connecting the VHH fragment with a phagemid vector, transforming TG1, and constructing a canine parvovirus single-domain antibody library; and enriching the antibody library by utilizing a canine parvovirus structural protein VP2, introducing into an expression bacterial strain E.coil HB2151, and utilizing IPTG to induce the canine parvovirus single-domain antibody VHH having the biological activity to be expressed. The prepared canine parvovirus single-domain antibody has the detection activity of 15 [mu]g / ml and the detection sensitivity of 250 ng / ml, and can be used as drugs applied for treating dogs with canine parvovirus disease.

Description

technical field [0001] The invention belongs to the technical field of clinical veterinary medicine and antibody engineering, and relates to a virus antibody, in particular to a canine parvovirus single-domain antibody mainly relying on the heavy chain antibody of alpaca to realize its biological expression, and a preparation method of the antibody and apply. Background technique [0002] Canine parvovirus disease is caused by canine parvovirus (Canine parvovirus, CPV). It is characterized by high morbidity, high mortality, and strong infectivity. It can occur all year round. Hemorrhagic enteritis type and myocarditis type canine parvovirus disease The mortality rates are 10% to 50% and 60% to 100%, respectively, which can cause serious economic losses. Vaccines to prevent canine parvovirus mainly include inactivated vaccines, attenuated vaccines and multiple vaccines, and optimized immunization procedures can achieve better immune effects. However, testing and investigati...

Claims

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Application Information

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IPC IPC(8): C07K16/08C12N15/63A61K39/42A61P31/20
Inventor 段智变孙耀贵程志学员世宇詹俊杰杨妍丽李小茜
Owner SHANXI AGRI UNIV
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