Multiple-virus-obtaining process for proliferating porcine reproductive and respiratory syndrome virus by Marc (rhesus monkey kidney cell)-145 cell cultured by microcarriers

A technology for microcarrier culture and respiratory syndrome, which is applied in the field of biotechnology and pharmaceuticals, can solve the problem of high price of imported microcarriers, achieve the effect of reducing production costs and improving utilization

Inactive Publication Date: 2014-02-05
乔自林 +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Microcarrier suspension culture technology will gradually replace traditional technology, but the price of imported microcarriers is higher

Method used

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  • Multiple-virus-obtaining process for proliferating porcine reproductive and respiratory syndrome virus by Marc (rhesus monkey kidney cell)-145 cell cultured by microcarriers
  • Multiple-virus-obtaining process for proliferating porcine reproductive and respiratory syndrome virus by Marc (rhesus monkey kidney cell)-145 cell cultured by microcarriers

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Experimental program
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Embodiment 1

[0022] 1. Materials:

[0023] 1) Cell line: Marc-145 cells imported from CCTCC;

[0024] 2) DMEM culture medium, Invitrigen Corporation;

[0025] 3) Trypsin, Invitrigen Corporation;

[0026] 4) Microcarrier: Cytodex-1, GE Company, containing 4.3×10 6 carrier; GF-240, Lanzhou Bailing Biotechnology Co., Ltd., containing 3.1×10 6 a carrier;

[0027] 5) Newborn bovine serum, Lanzhou Minhai Biological Engineering Co., Ltd.;

[0028] 6) PRRS virus (TJM-F92 strain), provided by China Veterinary Drug Control Institute.

[0029] 2. Instruments and equipment:

[0030] 1) CO 2 Incubator, Thermo Fisher Scientific Corporation, 3111;

[0031] 2) Rotary bottle culture machine, Lanfei Biochemical Equipment Factory, SHXJ-Ⅵ;

[0032] 3) Phase contrast inverted microscope, OLYMPUSCKX41;

[0033] 4) Bioreactor, NBS, 310, the total volume of the culture tank is 5L, and the final culture volume is 3.5L.

[0034] 3. Others:

[0035] Kjeldahl cell culture flask (T-75).

[0036] 4. Method...

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Abstract

The invention discloses a multiple-virus-obtaining process for proliferating a porcine reproductive and respiratory syndrome (PRRS) virus by a Marc (rhesus monkey kidney cell)-145 cell cultured by microcarriers. The multiple-virus-obtaining process comprises the following steps: culturing the Marc-145 cell by using two microcarriers of 2-10 g / L of Cytodex-1 or GF-240; inoculating PRRS virus liquids by using 0.01-0.1 of M.O.I (Multiplicity Of Infection); separating the PRRS virus liquids into a contrast group and an experimental group; sampling and measuring TCID50 (Tissue Culture Infective Dose) at 24 hours, 48 hours and 72 hours after viruses are inoculated in the contrast group; and respectively obtaining a virus supernate every 24-28 hours after viruses are inoculated in the experimental group, and then supplementing a maintenance medium to the virus supernate for continuous culturing to obtain the virus liquids twice or three times. The multiple-virus-obtaining process for proliferating the porcine reproductive and respiratory syndrome virus by the Marc-145 cell cultured by the microcarriers, which is disclosed by the invention, has the beneficial effects that the qualified virus liquid obtained in a multiple-virus-obtaining manner when the Marc-145 cell is used for proliferating the porcine reproductive and respiratory syndrome virus under microcarrier culture conditions is more than twice of that obtained in a normal virus-obtaining manner, the utilization rate of the microcarriers and seed cells is increased and the production cost of vaccines is reduced.

Description

technical field [0001] The invention relates to the field of biotechnology and pharmacy, in particular to a technique for repeatedly collecting poison of porcine reproductive and respiratory syndrome virus by culturing Marc-145 cells with microcarriers. Background technique [0002] Marc-145 cells are rhesus monkey embryonic kidney epithelial cells, which are widely used in the production of porcine reproductive and respiratory syndrome vaccines. At present, domestic manufacturers use the traditional spinner bottle culture process to produce the vaccine. Microcarrier suspension culture technology will gradually replace traditional technology, but the price of imported microcarriers is higher. Considering the production cost, multiple virus harvesting processes and the use of domestic gelatin microcarriers can increase the harvested amount of virus liquid and reduce production costs, and have application prospects in actual production. Contents of the invention [0003] T...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00C12R1/93
Inventor 乔自林令世鑫沈武玲冯若飞李明生冯玉萍马忠仁
Owner 乔自林
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