Generation and protective efficacy of a cold-adapted attenuated genotype 2b porcine epidemic diarrhea virus

Won, Hokeun; Lee, Dong-Uk; Jang, Guehwan; Noh, Yun-Hee; Lee, Seung-Chul; Choi, Hwan-Won; Yoon, In-Joong; Yoo, Han Sang; Lee, Changhee

Intestinal tissues and other major organs were grossly examined upon necropsy. Small intestinal tissue specimens (< 3-mm thick) collected from each piglet were fixed with 10% formalin for 24 h at RT and embedded in paraffin according to standard laboratory procedures. The formalin-fixed paraffin-embedded tissues were cut in 5-8-μm thick sections on a microtome (Leica, Germany), floated in a 40°C water bath containing distilled water, and transferred to glass slides. The deparaffinized intestinal tissue sections were stained with hematoxylin and eosin (Sigma, USA) to observe histopathological change or were subjected to immunohistochemistry (IHC) for the detection of PEDV antigen using an MAb specific for PEDV N protein as described previously [23, 24] . PEDV antigen detection was semiquantitatively measured throughout the IHC-stained jejunal sections based on the following scoring criteria as described previously [32] : 0 = no signal, 1 = 1%-10% of villous enterocytes within the section showing a positive signal, 2 = 11%-50% of villous enterocytes showing a positive signal, and 3 = greater than 50% of villous enterocytes showing a positive signal.