Method for industrially producing porcine reproductive and respiratory syndrome (PRRS) vaccines by utilizing bioreactor
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A bioreactor, porcine blue-ear disease technology, applied in the direction of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., to achieve the effects of high degree of automation control, easy scale expansion, and small batch-to-batch differences
Active Publication Date: 2011-05-04
WUHAN CHOPPER BIOLOGY
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But utilize Cell Lift type bioreactor, adopt the upside-down type operation technology of step-by-step amplification between bioreactors, carry out the high-density culture of cell to prepare vaccine, have not been reported
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Embodiment 1
[0022] Example 1: Bioreactor: 14L and 40L bioreactors from NBS Company in the United States, the parameters of the bioreactor are: pH 7.2, temperature 37°C, dissolved oxygen 50%, stirring speed 30-100rpm;
[0023] Microcarrier: Cytodex 1 (General Electric Healthcare Life Sciences);
[0027] Cell culture: In 14L bioreactors, Cytodex-1 was added at a concentration of 10g / L, after hydration, washed several times with pH 7.4 phosphate buffer saline PBS, sterilized, and inoculated with Marc-145 cells, Carry out culture; take samples regularly every day to observe the growth of the cells, count the cells, and measure the consumption of glucose. When the density of the cells reaches 1×10 6 / ml, start perfusion...
Embodiment 2
[0031] Embodiment 2: Bioreactors: 14L and 40L bioreactors of NBS Company in the United States, the parameters of the bioreactors are: pH 7.2, temperature 37°C, DO50%, stirring speed 30-100rpm;
[0032] Microcarrier: Cytodex 1 (General Electric Healthcare Life Sciences);
[0036] Cell culture: In 14L bioreactors, Cytodex-1 was added at a concentration of 10g / L, after hydration, washed several times with pH 7.4 phosphate buffer saline PBS, sterilized, and inoculated with Marc-145 cells, Carry out culture; take samples regularly every day to observe the growth of the cells, count the cells, and measure the consumption of glucose. When the density of the cells reaches 1×10 6 perml, start perfusion, and th...
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Abstract
The invention relates to a method for industrially producing porcine reproductive and respiratory syndrome (PRRS) vaccines by utilizing a bioreactor. The method comprises the following steps: (1) sterilizing a microcarrier and the bioreactor, then adding cell growth liquid, inoculating cells for vaccine preparation for culturing, inoculating PRRS viruses when dense monolayers are formed on the cells on the microcarrier and continuing culturing to reproduce the viruses; (2) stopping culturing and harvesting virus suspension when cytopathy reaches more than 80%; and (3) adding a freeze-drying protective agent to the harvested virus suspension to prepare freeze-dried attenuated live vaccines or carrying out ultrafilter concentration as well as virus inactivation on the harvested virus suspension, purifying the inactivated viruses by adopting column chromatography, and adding adjuvants to prepare inactivated vaccines. Compared with the traditional spinner bottle production process, the method has the following advantages: the automation control degree is high, so production can be monitored in real time; the labour is saved, thus reducing the cost; the land for production is few, thus being easy to enlarge the scale of production; and the produced viruses have high titer, so the batch-to-batch variation is small, the product quality is stable and the side reaction is low.
Description
technical field [0001] The invention belongs to the technical field of veterinary biological products, and in particular relates to a method for industrially producing porcine PRRS vaccine by using a bioreactor. Background technique [0002] Porcine reproductive and respiratory syndrome (Porcine Reproductive and Respiratory Syndrome, PRRS), also known as porcine blue ear disease, is a highly contagious infectious disease of pigs caused by porcine reproductive and respiratory syndrome virus (PRRSV), which is harmful to the pig industry very big. At present, PRRS is becoming more and more serious in China, directly and indirectly causing the so-called "pig unknown hyperthermia" that was prevalent in most provinces of my country in 2006, that is, highly pathogenic porcine blue ear disease, resulting in the death of 10 million to 30 million pigs. my country has listed highly pathogenic porcine blue-ear disease as a class of infectious diseases, and vaccine immunization is the m...
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