Sheep fetal skin fibroblast and separation method and application thereof

Abstract

The invention discloses a sheep fetal skin fibroblast and a separation method and application thereof. The preservation number of the fibroblast is CCTCC No: C2019202. In the separation process of the sheep fetal skin fibroblast SFSFS, digestive juice containing 0.05% trypsin and 0.02% EDTA and IV-type collagenase are adopted for treating sheep fetal skin tissue, and then digestive juice containing 0.01-0.05% trypsin-EDTA is used for treating cells; and 3% fetal calf serum and 10-40ng / ml h-EGF are adopted for replacing 10% fetal calf serum to culture the sheep fetal skin fibroblast, the use amount of the fetal calf serum is greatly reduced, the SFSFS culture cost and the aphtha virus production cost are reduced, and resources are saved. The cell strain can obviously increase the separation rate of aphtha viruses and shorten the separation time, the separated aphtha virus titer and virus copy number are higher, and a tool can be provided for large-scale reproduction of the aphtha viruses and virus infection pathogenesis research.

Description

technical field [0001] The invention relates to the technical field of cell engineering, in particular to a sheep fetal skin fibroblast, and also relates to a separation method and application of the cell. Background technique [0002] Contagious Ethyma (CE), commonly known as sheep mouth sore, is an acute and contagious infectious disease caused by sheep mouth ulcer virus (ORFvirus, ORFV) to which goats, sheep, and mainly lambs are susceptible. Zoonotic diseases that can infect humans and other animals. ORFV is mainly prevalent in Inner Mongolia, Tibet, Xinjiang, Yunnan, Gansu, Sichuan, Ningxia and other sheep-intensive areas. In recent years, it has frequently occurred in Ningxia, Inner Mongolia, Xinjiang, Tibet, Shandong, Hubei, Yunnan, Shaanxi, Heilongjiang, Qinghai and other regions . The typical symptoms of cauliflower-like hyperplasia will appear on the lips, oral mucosa, tongue and nostrils of affected sheep, which seriously affects their feeding and production, an...

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Problems solved by technology

[0003] The preparation of vaccines for the prevention of oral ulcers mainly includes the following aspects: first, through continuous separation and cultivation of primary cells of cattle or sheep, the oral oral ulcer virus is isolated from them, and then continuously passaged for attenuation to prepare attenuated vaccines; second, the current There is also a trend of research and development of inactivated vaccines; third, because the surface of the sheep oral ulcer virus is covered with a capsule shaped like a small tube, which is arranged in a criss-cross pattern to cover its epitope, resulting in less neutralizing antibodies produced in the sheep, and the protection rate for sheep In view of the particularity of this immunology, some researchers used ultrasound and other means to destroy the capsule of ORFV and then prepare a mouth sore vaccine. However, the above situations are limited to laboratory research and have not been used in production large scale use

The main reasons are: the separation of primary cells is time-consuming, laborious and expensive, the number of passages of primary cells is small, the toxicity of the cultured aphthus virus is low, the production volume within a batch is small, and the repeatability between batches is difficult to be consistent due to differences in primary cells

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