Isolation Of Porcine Alveolar Epithelial TypeⅡ Cell And Research Of HP-PRRSV Infection Experiments

Alveolar epithelial typeⅡcells(alveolar epithelial typeⅡcell)is one of the main cells of mammalian alveolar epithelium,is the main cell synthesis and secretion of alveolar surface active substance.Alveolar epithelial typeⅡcells(AECⅡ)isthe ancestor of almost lung cells.Porcine reproductive and respiratory syndrome virus is a kind of virus have capsule membrane,single strand positive RNA viruses,classification in arteritis virus,arteritis virus genera,which is the pathogens of Porcine reproductive and respiratory disorder syndrome(PRRS).The main cells infected by PRRSV are macrophages and lymphocytes of the immune system.In the lungs,PRRSV infection of target cells PAM is recognized as the preferred the research object,and PRRSV infection PAECⅡ research remains weak.This experiment by lavage piglet lungs and then use the method of immune adhesion purification PAECⅡ,determined PAECⅡ purity by alkaline phosphatase staining and electron microscope.Finally HP-PRRSV infection PAECⅡas the research object,detection the different time points N protein changes in the cells after PRRSV infection,it can explore HP-PRRSV infection PAECⅡ.The results are as follows:1.Separated PAECⅡby lung lavage and elastase digest method,then the immune adhesion method is adopted to purificate the PAECⅡ,by alkaline phosphatase staining and transmission electron microscopy(sem)for determination of purity and characteristics.Results show that the elastase digestion and IgG adhesive can produce high purity PAECⅡ,observed PAEC Ⅱ under electron microscope find surface microvilli and lamellar corpuscle,These specific structure prove separation cells are PAECⅡ.2.After HP-PRRSV infect PAECⅡ 0 h,12 h,24 h,the cells detected by indirect immunofluorescence test and western blot experiment.The indirect immunofluorescence experiment at 12 h,24h detected the green fluorescence in the cell,the western blot experiment in 12 h,24h detected the N protein expression of PRRSV.By indirect immunofluorescence test and western blot experiments proved that HP-PRRSV can infect PAECⅡ.