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Method for producing porcine reproductive and respiratory syndrome viruses

A technology for respiratory syndrome and pig reproduction, applied in biochemical equipment and methods, viruses/phages, microorganisms, etc., can solve the problems of long time required for virus liquid harvest, high labor intensity of workers, hidden dangers of vaccine safety, etc., to save The effect of freezing and thawing time, reducing labor intensity and reducing allergens

Active Publication Date: 2013-03-20
WENS FOODSTUFF GRP CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The current culture of porcine blue ear disease virus (PRRSV) leaves a large amount of bovine serum and other allergens in the crude antigen, which brings certain safety hazards to the use of vaccines
In addition, it takes a long time to harvest the virus liquid and the procedures are relatively cumbersome, resulting in high labor intensity for workers

Method used

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  • Method for producing porcine reproductive and respiratory syndrome viruses
  • Method for producing porcine reproductive and respiratory syndrome viruses
  • Method for producing porcine reproductive and respiratory syndrome viruses

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Test group: DMEM (high sugar type, containing 4500mg / L D-glucose, L-glutamine, and 110mg / L sodium pyruvate, without sodium bicarbonate) was added to 1700mL spinner flask and 15000mL spinner flask +10 % Newborn calf serum + 1% double antibodies (penicillin and streptomycin) are used as culture medium to culture Marc-145 cells; after the cells in the spinner flask grow into a monolayer, discard the cell growth medium in the flask and inoculate PRRSV at a multiplicity of infection of 0.01, Adsorbed at 37°C for 1h, added DMEM medium as a maintenance solution, and placed it on a rotating flask machine with a constant temperature chamber at 37°C for 75h rotation. When Marc-145 cells show 75%~80% CPE, collect the cell supernatant to harvest the virus. Measure half of the tissue culture infection dose (TCID 50 ).

[0024] At the same time, a control group was set up, the maintenance solution was DMEM plus 2% calf serum, and other operations and conditions were the same as the tes...

Embodiment 2

[0039] Test group: DMEM (high sugar type, containing 4500mg / L D-glucose, L-glutamine, and 110mg / L sodium pyruvate, without sodium bicarbonate) was added to 1700mL spinner flask and 15000mL spinner flask +10 % Newborn calf serum + 1% double antibodies (penicillin and streptomycin) are used as culture medium to culture Marc-145 cells; after the cells in the spinner flask grow into a monolayer, discard the cell growth medium in the flask and inoculate PRRSV at a multiplicity of infection of 0.01, Adsorbed at 37°C for 1 hour, added DMEM culture medium as a maintenance solution, and placed it on a rotating flask machine in a constant temperature chamber at 37°C for 78 hours. When Marc-145 cells show 75%~80% CPE, collect the cell supernatant to obtain PRRSV.

[0040] Control group: same as the test group, but after collecting the supernatant, put it in the freezer and freeze-thaw repeatedly 3 times to obtain PRRSV.

[0041] When the two bottles of cells of the same batch are harvested, ...

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Abstract

The invention discloses a method for producing and harvesting porcine reproductive and respiratory syndrome viruses (PRRSV), which comprises the following steps: culturing Marc-145 cells in a spinner bottle; after the cells grow into a monolayer, removing a growth-promoting media; inoculating the PRRSV according to 0.01 infection multiplicity (MOI: multiplicity of infection); absorbing for 1 hour at a temperature of 37 DEG C; adding DMEM serving as a maintenance media to carry out expanding propagation of viruses; arranging the spinner bottle on a rotary machine of a 37 DEG C thermostatic chamber to carry out rotation culture for 72 to 84 hours; and when 75 to 80 percent of cytopathic effect occurs, directly collecting the supernatant to obtain the PRRSV. The invention reduces allergen in the rough antigens and improves the vaccine quality. And the time that the Marc-145 cells have CPE is consilient with that before the process is improved, but the virus harvesting time is shortened.

Description

Technical field [0001] The invention belongs to the field of biotechnology, and specifically relates to a virus culture method. Background technique [0002] Reproductive and Respiratory Syndrome (PRRS) has spread to almost all countries and regions in the world where the pig industry is developed since it was discovered in the United States in 1987. The clinical symptoms are mainly characterized by reproductive disorders in sows and respiratory diseases in piglets. Highly pathogenic porcine blue-ear disease is an acute and highly pathogenic disease caused by a variant strain of the porcine reproductive and respiratory syndrome (commonly known as blue-ear disease) virus, which has caused huge economic losses to the world pig industry . Effective vaccine immunization is an important measure to control the disease. In order to prepare a large amount of virus antigens, sufficient virus must be obtained. The virus is usually propagated by propagating on cells. Porcine Reproductive...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00C12N7/02
Inventor 邓雨修宋延华杨明柳李春梅文传洪于琳忠
Owner WENS FOODSTUFF GRP CO LTD