A method for culturing serum-free whole suspension cells of Newcastle disease vii attenuated strain

A technology for cell culture and Newcastle disease, applied in biochemical equipment and methods, viruses, inactivation/attenuation, etc. It can solve the problem that the method is not yet developed, the adaptability of suspension cells is weak, and avian viruses cannot grow and reproduce. It has not been discovered yet and other problems, to achieve the effect of stable virus reproduction, large amount of preparation, and high poisonous price

Active Publication Date: 2022-07-12
WUHAN KEQIAN BIOLOGY CO LTD
View PDF2 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The whole suspension culture of avian virus heterologous cells has been studied at home and abroad, but few of them are actually used for production, and most of them are in the research stage. This is related to the adaptability of viruses to heterologous cells. Different viruses have different effects on heterologous cells. Many avian viruses cannot grow and reproduce on heterologous cells or have not been discovered or the method is not right to be developed
In recent years, there have been many literature and patent reports on the full-suspension cell culture of chicken Newcastle disease virus, mainly on the Lasota strain of Newcastle disease virus. The Lasota strain is a more classic and commonly used virus strain, but its adaptability to suspension cells is weak

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • A method for culturing serum-free whole suspension cells of Newcastle disease vii attenuated strain
  • A method for culturing serum-free whole suspension cells of Newcastle disease vii attenuated strain
  • A method for culturing serum-free whole suspension cells of Newcastle disease vii attenuated strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 Optimal harvest time of virus

[0037] In the method described in this example, the optimal harvesting time of the virus solution is as follows: observe and count the cells under a microscope, and the density is 7.9×10 6 , adjust the cell seeding density to 1.5 × 10 6 The amount of TPCK trypsin added is 1-6ug / ml, the amount of virus inoculum is 0.001-1%, shake cultured at 37°C, and the rotation speed is 100-150r / min, respectively 36h, 41h, 48h, 54h, 60h after inoculation , 72h, the virus liquid was harvested, and the HA titer at each time point was detected. The results are shown in Table 1.

[0038] In this example, the pictures of lesions and normal control cells of the attenuated Newcastle disease gene type VII (strain NF02) in suspension cultured for 48 hours are shown in figure 1 .

[0039] Table 1 HA detection results of virus liquid at different harvest times

[0040] Virus liquid harvest time 36h 41h 48h 54h 60h 72h HA 7 8 9...

Embodiment 2

[0042] Example 2 Optimal seeding density of cells

[0043] This example explores the optimal cell seeding density in the method described, and the steps are as follows: observe and count the cell density to be 8.0×10 6 cells, adjust the cell density to 1.0 × 10 6 , 2.0×10 6 , 3.0×10 6 indivual. TPCK trypsin concentration is 1-6ug / ml, virus inoculation concentration is 0.001-1%, shake culture at 37°C, rotation speed is 100-150r / min, culture for 48-54 hours, harvest, and detect virus HA titer, the results are shown in the table 2.

[0044] Table 2 HA detection of virus liquid obtained by inoculation and culture of different cell densities

[0045] Cell density 1.0x10 6

[0046] The results in Table 2 show that the cell density is 1.0 × 10 6 When inoculated, the virus reproduction ability is the best.

Embodiment 3

[0047] Embodiment 3TPCK pancreatin optimal use concentration

[0048] This example explores the optimal concentration of TPCK trypsin used in the method described in this example. The steps are as follows: adjust the cell density to 1.0×10 according to the needs of the experiment. 6 125ml shake flasks, add TPCK trypsin concentration of 1ug / ml, 2ug / ml, 3ug / ml, 5ug / ml, 6ug / ml, virus inoculation concentration of 0.001-1%, 100 -150r / min cultured for 48-54 hours, harvested, and the virus HA titer was detected. The results are shown in Table 3.

[0049] Table 3 Optimal TPCK trypsin use concentration HA detection

[0050] Pancreatin concentration 1ug / ml 2ug / ml 3ug / ml 5ug / ml 6ug / ml HA 6 9 9 8 8

[0051] The results in Table 3 showed that the best virus reproduction was when the TPCK trypsin concentration was 2-3ug / ml.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention relates to the field of cells and biological products, in particular to a method for culturing serum-free whole suspension cells of a Newcastle disease type VII attenuated strain. In the method of the present invention, the strain of the Newcastle disease virus used is the strain of the Newcastle disease gene type VII virulence weakened by mutation of the F gene site. The method of the invention adopts serum-free full suspension cell culture, which is not affected by insufficient supply of eggs or uncontrollable factors; the operation is simple, the amount of prepared virus is large, and the virus content is high, which overcomes the effect of virus chicken embryo culture due to insufficient supply of eggs. The proliferation of vaccines has led to a stagnation of vaccine production, thereby affecting the availability of vaccines in the market.

Description

technical field [0001] The invention relates to the field of cells and biological products, in particular to a method for culturing serum-free whole suspension cells of a Newcastle disease attenuated strain. Background technique [0002] Newcastle disease is a highly contagious infectious disease caused by a virus. It mainly affects chickens and other poultry and wild birds. It can also be infected in humans, mainly manifested as conjunctivitis. This disease is one of the most serious diseases among chicken diseases, with rapid onset and high mortality, resulting in great economic losses. At present, compulsory immunization is widely carried out in all parts of our country to improve the immunity of chickens and reduce the occurrence of epidemic diseases. [0003] At present, most of the Newcastle disease virus is still propagated by chicken embryos, and the operation process is complicated, including the introduction of breeding eggs, egg stacking, disinfection, hatching b...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00C12N7/04
CPCC12N7/00C12N2760/18121C12N2760/18162C12N2760/18151C12N2760/18134Y02A50/30
Inventor 周明光王鑫程泰烺徐高原张锦军张宏斌金建云陈章表
Owner WUHAN KEQIAN BIOLOGY CO LTD
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap