Optimization of Immunohistochemical and Fluorescent Antibody Techniques for Localization of Foot-and-Mouth Disease Virus in Animal Tissues

Abstract

Immunohistochemical (IHC) and fluorescent antibody (FA) techniques were optimized for the detection of Foot-and-mouth disease virus (FMDV) structural and nonstructural proteins in frozen and paraformaldehyde-fixed, paraffin-embedded (PFPE) tissues of bovine and porcine origin. Immunohisto-chemical localization of FMDV was compared with 7 detection systems, 8 primary antibodies, and 11 epitope retrieval techniques. All serotypes tested (O, A, Asia, C [cryosection]; O, A, Asia [PFPE]) were localized in association with mature vesicles. Multi-label FA was used in conjunction with IHC and conventional histopathology to characterize vesicle maturation in 4 steers and 2 pigs experimentally infected with FMDV. At the edge of advancing vesicles, a consistent finding was acantholytic degeneration of basal keratinocytes surrounding dermal papillae with suprabasilar clefts and microvesiculation. Progression of microvesiculation led to coalescence with the expanding vesicle. Cells at the leading edge of vesicles were positive for FMDV antigens by IHC and FA. Cell marker profile of these cells by FA was consistent with keratinocytes (i.e., cytokeratin [CK]-positive, S100-negative, MHC-II-negative). In rare instances, CK-negative, MHC-II-positive, and FMDV-positive cells (presumptive dendritic cells or macrophages) were identified within dermis subjacent to vesicles.