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
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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
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