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Cell line used for separation culture and multiplication of Orf virus (ORFV) as well as preparation method and application of cell line

A pustular virus, isolation and culture technology, applied in the field of veterinary vaccine preparation, can solve the problems of complicated manufacturing process, high production cost, and carrying other pathogens, and achieve the effects of simplifying the production process, maintaining stable quality, and reducing production costs

Active Publication Date: 2014-07-30
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In China, although the attenuated vaccine for sheep infectious impetigo was developed in the Veterinary Research Institute of Qinghai Academy of Animal Husbandry and Veterinary Sciences and Gansu Provincial Animal Husbandry and Veterinary Research Institute, the manufacturing process is complicated and backward, and newborn cattle must be used to breed the attenuated sheep infectious impetigo antigen. Testicular primary cells, but the preparation of such cells is complicated, the production cost is too high, and the production efficiency is low, which brings inconvenience to the large-scale production of sheep infectious impetigo attenuated vaccine by vaccine factories, and it is difficult for enterprises to obtain huge profits
At the same time, the variety of newborn bovine testes used for each breeding of sheep oral antigens is different, which makes it difficult to maintain a stable quality of the attenuated vaccine produced. In addition, newborn bovine testicles have the potential risk of carrying other pathogens

Method used

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  • Cell line used for separation culture and multiplication of Orf virus (ORFV) as well as preparation method and application of cell line
  • Cell line used for separation culture and multiplication of Orf virus (ORFV) as well as preparation method and application of cell line
  • Cell line used for separation culture and multiplication of Orf virus (ORFV) as well as preparation method and application of cell line

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] [Example 1] Isolation, cultivation and identification of Sertoli cells of bovine testis

[0020] 1.1 Isolation and culture of bovine testicular Sertoli cells

[0021] Select healthy male calves born to healthy cows (whose physique is healthy and negative for foot-and-mouth disease, brucellosis, paratuberculosis, and bovine viral diarrhea), and collect the scrotum after slaughter (the root of the scrotum is ligated, and the outside of the scrotum is sterilized with 75% alcohol) , in an incubator, and sent back to the laboratory within 2 hours. Under sterile conditions, take out the newborn bovine testes and put them in a glass dish, wash the testes 2 to 3 times with double-antibody D-hanks, and remove the epididymis, fat pad and testicular tunica. Use ophthalmic tweezers and scissors to divide the testicular parenchyma into small pieces of the same size, gently pipette several times in D-hanks solution, let it stand for 5-10 minutes, remove the supernatant, and obtain s...

Embodiment 2

[0026] [Example 2] Proliferation of sheep infectious impetigo virus

[0027] Ovine infectious impetigo virus can be cultured and proliferated on bovine kidney cell line and primary cells of calf and lamb testis. Newborn fetal bovine myocytes, newborn fetal sheep dermal cells, fetal bovine lung cells, fetal sheep gyroscope cells and fetal sheep myocytes can also be used for the isolation and culture of OPIV. Among them, the bovine kidney cell line (MDBK) is the most commonly used passage cell line, which is often used for the initial isolation and subculture of viruses. The primary cells of neonatal bovine testis are most commonly used in the isolation and culture of OIPV, passage attenuation and production of attenuated vaccine for OIPV. Therefore, the inventor of the present invention selects, three kinds of cells of MDBK cell line, primary cells of newborn bovine testis and Sertoli cells of newborn bovine testis are inoculated with the same strain, the same generation of sh...

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Abstract

The invention discloses a cell line used for separation culture and multiplication of an Orf virus (ORFV) as well as a preparation method and application of the cell line. The cell line disclosed by the invention is a newly born bovine testis supported cell line obtained by separating and purifying newly born bovine testis primary culture cells; research results show that the cell line is more sensitive to inoculation of contagious ovine ecthyma virus, virus titer multiplied by utilizing the cell line is about 1 virus titer higher than that of newly born bovine testis primary culture cells and 1.9 virus titer higher than that of a madin-darby bovine kidney (MDBK) cell line. ORFV is separated by utilizing the cell line, uniformity and stability of virus can be guaranteed, and a risk that the separated viruses carry other pathogens as the primary culture cells are utilized for multiple times can be prevented; besides, the passage cell line is utilized for preparing a vaccine, a production process of the vaccine can be simplified, a production cycle is shortened, production cost is reduced, and stable quality of the prepared vaccine is also guaranteed, so that the cell line has a certain practical significance on large-scale production of the vaccine.

Description

technical field [0001] The present invention relates to a subculture cell line for the propagation of sheep infectious impetigo virus and vaccine preparation and its preparation method and application, in particular to a neonatal bovine testicular Sertoli cell line and a method for using the cell line to propagate and cultivate sheep infectious impetigo virus . The invention belongs to the technical field of animal vaccine preparation. Background technique [0002] Contagious ecthyma (CE), also known as Contagious pustular dermatitis (Contagious pustular dermatitis), commonly known as "Orf", is caused by the parapoxvirus member Orf Virus, ORFV) is an acute, contact and epithelial zoonotic infectious disease of goats, sheep and humans, which causes erythema to form on the skin and mucous membranes of the affected sheep's lips, hooves, breasts, vulva, etc. , papules, abscesses, ulcers, and verrucous thick scabs. The World Organization for Animal Health (OIE) lists the disea...

Claims

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Application Information

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IPC IPC(8): C12N7/00C12N5/071A61K39/275A61P31/20C12R1/91
Inventor 刘湘涛王光祥尚佑军张志东王艳华黎摄儿田宏吴锦艳陈妍逯忠新
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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