2.4  IFA
When IPEC-J2 cells grown in 96-well microplates reached ∼80% confluence, they were mock infected or infected with PDCoV at an MOI of 0.1 TCID50 per cell. After a 48-h cultivation at 37 °C with 5% CO2, the cells were fixed with prechilled 100% ethanol for 15 min at room temperature. After washing with PBS, the cells were probed with the mAb 1A3 (1:5000 dilution) at 37 °C for 1 h. Following another washing step, 1000-fold-diluted Alexa Fluor 488-conjugated goat antimouse IgG (Thermo Fisher Scientific) was added to the cells and incubated at 37 °C for 1 h. After washing thrice with PBS, cells were observed with an Eclipse Ci-S microscope (Nikon Corp., Tokyo, Japan).