Applications of BHK-21 cell full-suspension culture technology in production of newcastle disease vaccine

A BHK-21, 1.BHK-21 technology, applied in the field of veterinary biopharmaceuticals, can solve problems such as immune failure, affecting vaccine quality, and foreign pathogen contamination

Active Publication Date: 2015-01-07
CHINA INST OF VETERINARY DRUG CONTROL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But in my country, in the actual production process, due to the restriction of the supply of SPF (specific pathogen-free) chicken embryos and the cost problem, many domestic vaccines are not all made of SPF chicken embryos.
Especially in the preparation of attenuated vaccines for poultry, if the chicken embryos used come from ordinary eggs, it may cause contamination of foreign pathogens, and thus cause the spread and spread of certain diseases
At the same time, the maternal antibody in the chicken embryo can also interfere with the reproduction of the vaccinated virus, directly affect the quality of the vaccine, and often cause immune failure

Method used

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  • Applications of BHK-21 cell full-suspension culture technology in production of newcastle disease vaccine
  • Applications of BHK-21 cell full-suspension culture technology in production of newcastle disease vaccine
  • Applications of BHK-21 cell full-suspension culture technology in production of newcastle disease vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] ——Aclimation of BHK-21 cells in full suspension culture

[0074] 1 material

[0075] 1.1 Cell strain: BHK-21 cells, purchased from China Veterinary Microbiology Culture Collection Management Center.

[0076] 1.2 Medium: DMEM, VP SFM AGT, both are products of Gibco Company.

[0077] 1.3 Bovine serum: product of Hyclone Company.

[0078] 1.4 Instruments and equipment: Centrifuge, Sigma 3-18K type; Suspension cell culture flask (100ml, 500ml), WHEATON product; BioFlo 115 reactor, American NBS Company.

[0079] 2 methods

[0080] 2.1 Use DMEM medium containing 1% to 10% serum at 35 to 38°C in 5% CO 2 BHK-21 cells were cultured in a single-layer adherent manner, and scattered cells in the culture supernatant with weak or non-adherent ability were collected;

[0081] 2.2 5% CO at 35-38°C 2 Suspension culture the collected cells under the above conditions, the suspension culture BHK-21 cells were centrifuged at 100-1000r / min for 5-10min, the centrifuged supernatant was c...

Embodiment 2

[0085] ——Method for Suspension Culture BHK21 Cell Production Newcastle Disease Virus Mukteswar Strain

[0086] 1. Materials

[0087] 1.1 Cell strain: BHK-21 / IVDC strain suspension cells, domesticated and selected by China Veterinary Drug Control Institute.

[0088] 1.2 NDV Mukteswar strains and strains were purchased from China Center for Veterinary Microorganism Culture Collection.

[0089] 1.3 Medium: VP SFM AGT, product of Gibco Company.

[0090] 1.4 Bovine serum: product of Hyclone Company.

[0091] 1.5 Anti-Newcastle disease virus-specific serum was purchased from China Veterinary Microbiological Culture Collection and Management Center.

[0092] 1.6 Instruments and equipment: Centrifuge, Sigma 3-18K type; Suspension cell culture flask (100ml, 500ml), WHEATON product; BioFlo 115 reactor, American NBS Company.

[0093] 2 methods

[0094] 2.1 Cell preparation

[0095] 2.1.1 Inoculate a single layer of BHK-21 cells with the Newcastle disease vaccine production virus stra...

Embodiment 3

[0120] ——Comparison of the suspension culture BHK-21 cell production method for Newcastle disease virus and chicken embryo culture / roller bottle culture method

[0121] 1 material

[0122] 1.1 Newcastle disease virus strain NDV Mukteswar strain was purchased from China Center for Veterinary Microorganism Culture Collection. .

[0123] 1.2 Spinner bottle The specification is 3 liter spinner bottle.

[0124] 1.3 Bioreactor American NBS BioFlo 115 type reactor (total volume of tank 14L).

[0125] 1.4 Cell culture medium Same as Example 1.

[0126] 1.5 Cell maintenance solution Same as Example 1.

[0127] 1.6 chicken embryos 9-11 days old SPF chicken embryos

[0128] 2 methods

[0129] 2.1 Cultivation of the virus in chicken embryos Dilute the virus species (NDVMukteswar strain, purchased from the China Veterinary Microbiology Culture Collection Management Center) with sterile saline (eg 100 times), inoculate the allantoic cavity at 9 to 11 days old SPF chicken embryos, 0.1...

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Abstract

The invention relates to applications of a BHK-21 cell full-suspension culture technology in production of a newcastle disease vaccine. A process of producing the newcastle disease vaccine by BHK-21 cell full-suspension culturing includes steps as follows: (1) a step of viral strain seed selection, namely a step of inoculating a monolayer BHK-21 cell with a newcastle disease vaccine virus seed cultured by a chick embryo, adding a virus maintenance medium, culturing to obtain a newcastle disease vaccine adapted to the BHK-21 cell, and performing system identification; (2) a step of domestication and seed selection of a suspension cell strain, namely a step of domesticating a full-suspension BHK21 cell used for culturing of the newcastle disease vaccine virus and establishing a basic seed; (3) a step of subjecting the suspension cell to enlarged cultivation; (4) a step of virus inoculation and harvest, namely a step of inoculating the newcastle disease vaccine virus adapted to the BHK-21 cell and harvesting a virus solution; and (5) a step of measuring the viral titer of the multiplicated newcastle disease vaccine virus and preparing the vaccine. According to the applications, culturing and production with chick embryos of the newcastle disease vaccine are changed into to large-scale culture and production with mammalian cells of the newcastle disease vaccine, the process of producing the newcastle disease vaccine is simplified, the production cost is reduced, and the yield and quality of the vaccine are largely improved.

Description

technical field [0001] The invention relates to the application of BHK-21 cell full suspension culture technology in the production of Newcastle disease vaccine, belonging to the field of veterinary biopharmaceuticals. Background technique [0002] At present, the production of domestic poultry vaccines mainly relies on the traditional "chicken embryo method" (Chinese Veterinary Pharmacopoeia Committee. Veterinary Pharmacopoeia of the People's Republic of China Part Three 2010 Edition. China Agricultural Press 2011) process. The "chicken embryo method" technique for making vaccines is a long-standing one, first developed in the 1950s, although the technique has evolved to meet various challenges, including yield, automation, capacity, quality assurance and production speed. But in my country, in the actual production process, due to the restriction of the supply of SPF (specific pathogen-free) chicken embryos and the cost problem, many domestic vaccines are not all made by S...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/17A61P31/14C12N7/00C12R1/93
Inventor 吴华伟陈晓春高艳春郎洪武邓永李俊平李慧姣夏业才
Owner CHINA INST OF VETERINARY DRUG CONTROL
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