2.10. Fluorescence analysis of cell apoptosis

Cell apoptosis was analyzed with Annexin V-FITC/PI Kit (Nanjing Keygen Biotech, China). Annexin V has a high affinity for the membrane phosphatidylserine which was translocated from the inner face of plasma membrane to the cell surface at early stage of cell apoptosis, and FITC-labeled Annexin V could be used as a fluorescent probe to detect cell early apoptosis by fluorescence microscopy. Propidium iodide (PI) can pass through the membrane of apoptotic and necrotic cells, and can be used to detect cellular late apoptosis. A combination of Annexin V and PI was used to distinguish cellular early and late apoptosis. Briefly, FITC-labeled Annexin V (50 μl/well) and PI (50 μl/well) was added to the cells infected by LiCl-treated PEDV, or PEDV-infected cells treated with LiCl, and incubated at room temperature for 15 min in the dark prior to fluorescence observation. Fluorescence was observed under a Nikon ECLIPSE TS100 fluorescence microscope (Nikon, Japan).