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Preparation method and application of passage cell origin ND, IB and AI triple inactivated vaccine

A technology of passaging cells and inactivated vaccines, which is applied in the field of veterinary biological products, can solve the problems of unstable batches and complex processes, and achieve the effects of small quality differences, stable and uniform quality, and stable processes

Active Publication Date: 2018-07-10
ZHAOQING INST OF BIOTECHNOLOGY CO LTD +3
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The invention provides a preparation method and application of a triple inactivated vaccine derived from chicken Newcastle disease, infectious bronchitis and avian influenza with passaged cells, so as to at least solve the problem that the triple inactivated vaccine in the prior art needs to be cultivated with multiple cells and the process is complex , batch instability problem

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] The method for culturing the Newcastle disease virus in Example 1 of the present invention is as follows:

[0035] Step 1: Press 0.35×10 of EB66 cells 6 cells / mL-0.75×10 6 The cell density of cells / mL is inoculated into a bioreactor (Sartorius B Plus) for culture, sampling is counted every day, and the viability is calculated. When the cells grow to 8.0×10 6 cells / mL-16×10 6 cells / mL, when the viability is above 90%, keep 600mL of cells in the reactor for virus inoculation;

[0036] Step 2: Newcastle disease virus was inoculated into the EB66 cells in the reactor at MOI of 0.01, the stirring speed was adjusted to 80 rpm after the inoculation, and the adsorption culture was carried out for 1 hour. After the virus adsorption culture was completed, 1.5 times the original growth culture was added The base volume of fresh production medium, the temperature is adjusted to 33℃, the pH is set to 7.20±0.1 to continue the culture, the stirring speed is adjusted to 120rpm, and other cul...

Embodiment 2

[0040] The method for culturing infectious bronchitis virus in Example 2 of the present invention is as follows:

[0041] Step 1: Press 0.35×10 of EB66 cells 6 cells / mL-0.75×10 6 The cell density of cells / mL is inoculated into a bioreactor (Sartorius B Plus) for culture, and samples are taken every day for counting, and the viability is calculated. When the cells grow to 8.0×10 6 cells / mL-16×10 6 cells / mL, when the viability is above 90%, keep 600mL of cells in the reactor for virus inoculation;

[0042] Step 2: Infectious bronchitis virus inoculates the EB66 cells in the reactor at 0.01 MOI. After inoculation, the stirring speed is adjusted to 80 rpm, and the adsorption culture is performed for 1 hour. After the virus adsorption culture is completed, add 2 A fresh production medium that is twice the original growth medium volume, the temperature is adjusted to 33°C, the pH is set to 7.20±0.1 to continue the culture, the stirring speed is adjusted to 120rpm, and other culture parame...

Embodiment 3

[0046] The cultivation method of the avian influenza virus (H9 subtype) in Example 3 of the present invention is as follows:

[0047] Step 1: Press 0.35×10 of EB66 cells 6 cells / mL-0.75×10 6 The cell density of cells / mL is inoculated into a bioreactor (Sartorius B Plus) for culture, sampling is counted every day, and the viability is calculated. When the cells grow to 8.0×10 6 cells / mL-16×10 6 cells / mL, when the viability is above 90%, keep 600mL of cells in the reactor for virus inoculation;

[0048] Step 2: Avian influenza virus is inoculated to the EB66 cells in the reactor at MOI of 0.01. After inoculation, the stirring speed is adjusted to 80 rpm, and the adsorption culture is carried out for 1 hour. After the virus adsorption culture is completed, add twice the original growth culture The base volume of fresh production medium, the temperature is adjusted to 33℃, the pH is set to 7.20±0.1 to continue the culture, the stirring speed is adjusted to 120rpm, and other culture para...

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Abstract

The invention provides a preparation method and application of a passage cell origin newcastle disease (ND), infectious bronchitis (IB) and avian influenza (AI) triple inactivated vaccine. The methodincludes the steps of: subjecting an EB66 cell to growth culture in a bioreactor for virus inoculation; employing a two-stage culture method to inoculate newcastle disease virus, infectious bronchitisvirus or avian influenza virus to the EB66 cell; after 24h of virus inoculation, harvesting and storing the virus when the virus EID50 reaches a highest level, thus obtaining a virus liquid; repeating the steps to obtain the newcastle disease virus liquid, infectious bronchitis virus liquid and avian influenza virus liquid respectively, and conducting inactivation; mixing the inactivated newcastle disease virus liquid, infectious bronchitis virus liquid and avian influenza virus liquid to prepare the inactivated vaccine. The method provided by the invention employs the EB66 passage cell linefor culture of the newcastle disease virus, infectious bronchitis virus and avian influenza virus, avoids the risk of embryonic heterogenous protein and exogenous virus pollution caused by use of chicken embryos, improves the virus production quality, and simplifies the virus production process.

Description

Technical field [0001] The invention relates to a preparation method and application of a triple inactivated vaccine of ND (Newcastle Disease of Chicken), IB (Infectious Bronchitis), and AI (Avian Influenza H9 Subtype) and its application, in particular to a full suspension culture process and passage cells The method for producing triple inactivated vaccines against chicken Newcastle disease, infectious bronchitis and avian influenza H9 subtype belongs to the technical field of veterinary biological products. Background technique [0002] Newcastle disease, infectious bronchitis and avian influenza, as the three major infectious diseases affecting the poultry industry, lead to the death of a large number of farmed chickens every year and cause significant losses to the development of my country's poultry industry. At present, the prevention and control of these three diseases mostly uses vaccine immunization to achieve good prevention and control effects. The current vaccines ar...

Claims

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Application Information

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IPC IPC(8): A61K39/17A61K39/215A61K39/145C12N7/00A61P31/14A61P31/16C12R1/09
CPCA61K39/12A61K2039/5252A61K2039/70C12N7/00C12N2760/16134C12N2760/16152C12N2760/18134C12N2760/18152C12N2770/20034C12N2770/20052
Inventor 陈瑞爱罗顺李延鹏温良海蔡仕君张晓楠罗琼
Owner ZHAOQING INST OF BIOTECHNOLOGY CO LTD