关键词: 鸡气管上皮细胞, 分离, 培养, IBV, 培养特性

Abstract: [Objective] The purpose of this study was to explore and find out appropriate technology of separation and culture for chicken trachea-bronchia epithelium and also to research culture characteristics of IBV in chicken trachea-bronchia epithelium. [Method] By the method of douche, chicken trachea-bronchia epithelium was digested overnight at 4℃ and separated. Then the test of immunohistochemistry of No.8/18 keratins of chicken trachea-bronchia epithelium was done to identify epithelium. The cultured epithelium was infected respectively by IBV-M41 strain(Style Breath ) and IBV-T strain(Style Kidney), and respectively at 12,24,32,48,60,72,84 and 96h after epithelium infected by IBV we observed morphologic changes of epithelium. At last the method of RT-PCR was used to detect IBV mRNA in each group of cells. At the same time the ability of haemagglutination and pathological changes of chicken embryo caused by IBV were tested. [Result] By the method of douche and digestion overnight chicken trachea-bronchia epithelium have been separated and have full morphologic structure and clearly visible cilia and high livingness. Through three-step purification the epithelium has good uniformity and grow faster. Respectively at 72h and 84h after each IBV strain and its corresponding epithelium co-culture, the infected epithelium appeared obvious cell pathologic effect(CPE), and the test of ability of haemagglutination and pathological changes of chicken embryo by IBV were both positive. [Conclusion] We have successfully separated and cultured chicken trachea-bronchia epithelium to fulfill the need of relative deeper research by providing chicken trachea-bronchia epithelium as research model. And both IBV-M41 strain and IBV-T strain may cause chicken trachea-bronchia epithelium to appear CPE, which provide nicer bases for future relative research about IBV laboratory diagnosis, medicine sensitivity test and pathology mechanism caused by IBV.

Key words: Chicken trachea-bronchia, Epithelium, Dissociation culture, IBV, Culture characteristics