Method for preparing rainbow trout IHN(Infectious Haematopoietic Necrosis) inactivated vaccine

A hematopoietic organ necrosis and inactivated vaccine technology, which is applied in the field of preparation of rainbow trout infectious hematopoietic organ necrosis inactivated vaccine, can solve the problem that a single cell cannot obtain high titer IHNV antigen

Inactive Publication Date: 2014-12-10
LANZHOU WEITESEN BIOTECH
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Problems solved by technology

[0007] The preparation method of the present invention uses RTG-2 cells, CHSE-214 cells and EPC cells to alternately culture rainbow trout infectious hematopoietic necrosis virus (IHNV) at a specific ambient temperature to obtain a large amount of high-titer IHNV antigen and produce inactivated Vaccine; solves the technical problem that high-titer IHNV antigen cannot be obtained with a single cell

Method used

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  • Method for preparing rainbow trout IHN(Infectious Haematopoietic Necrosis) inactivated vaccine
  • Method for preparing rainbow trout IHN(Infectious Haematopoietic Necrosis) inactivated vaccine
  • Method for preparing rainbow trout IHN(Infectious Haematopoietic Necrosis) inactivated vaccine

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Embodiment

[0022] Preparation of rainbow trout infectious hematopoietic necrosis antigen and its animal immunity experiment and cohabitation challenge protection experiment

[0023] 1. Isolation and identification of IHNV

[0024] 1) Isolation of IHNV

[0025] The diseased material was collected from the rainbow trout farm where the epidemic broke out in Yongdeng, Gansu Province. After grinding and soaking in the virus, it was inoculated with RTG-2 cells grown three days later. Blind passage was carried out at 14°C, and the passage to the tenth generation took five days In case of typical lesions, the cytotoxic solution was collected and frozen for future use.

[0026] 2. Identification of IHNV

[0027] 2.1) Identification of IHNV

[0028] Design primers, amplify the partial fragments of IHNV nucleoprotein gene by the method for RT-PCR, the designed amplification primers are:

[0029] Upstream IHNs: 5’ tca agg ggg gag tcc tcg a 3’

[0030] Downstream IHNa...

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Abstract

The invention discloses a method for preparing a rainbow trout IHN (Infectious Haematopoietic Necrosis) inactivated vaccine, which comprises the following steps: carrying out grinding, filtering and poison dipping processing on pancreas and livers of juvenile fish which is attacked, but is still alive, inoculating rainbow trout gonad (RTG) cells, carrying out blind passaging at 14 DEG C, keeping for 5 days when carrying out blind passaging on the tenth generation, and collecting cell poisonous fluid; inoculating chinook salmon embryonic (CHSE) cells, carrying out passaging at 14 DEG C, and raising the culture temperature by 1 DEG C when passaging for 5 generations each time until the culture temperature is raised to 20 DEG C; and adopting epithelioma papulosum cyprini (EPC) cells to continuously carry out passaging under the condition of 20 DEG C, passaging to the twelfth generation to obtain high-titer virus solution and carrying out inactivation to obtain the rainbow trout IHN inactivated vaccine. According to the preparation method, RTG-2, CHSE-214 and EPC cells are utilized to alternately culture rainbow trout IHN viruses at a specific environment temperature so as to obtain a high-titer IHNV (Infectious Hematopoietic Necrosis Virus) antigen and produce the inactivated vaccine; and the technical difficult problem that the high-titer IHNV antigen cannot be obtained through single cells is solved.

Description

technical field [0001] The invention belongs to the technical field of biological products and relates to a preparation method of an inactivated vaccine for infectious hematopoietic organ necrosis of rainbow trout. Background technique [0002] Infectious hematopoietic necrosis (INH) is a viral infectious disease of fish with wide distribution, high mortality and great harm caused by rainbow trout infectious hematopoietic necrosis virus (IHNV) in the rhabdoviridae family. , began to be found in various trout farms in Northeast my country in 1985, and it is currently spreading and intensifying. Moreover, in recent years, my country has introduced a large number of salmon eggs from abroad every year, but due to the imperfect quarantine technology, infectious hematopoietic organ necrosis, which is seriously harmful, has strong transmission power, and is difficult to prevent and control, has been introduced into my country and spread. The developing my country's rainbow trout bre...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/205A61P31/14C12N7/00C12R1/93
Inventor 柳纪省李宝玉杨彬李学瑞兰喜殷相平李志勇张韵方玉萍
Owner LANZHOU WEITESEN BIOTECH
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