Skin Punch as a Potential Diagnostic Tool for Peripheral Neuropathies of Dogs: Set up of an Indirect Immunofluorescence Protocol on Formalin-Fixed Paraffin-Embedded (FFPE) Biopsy

In dogs, the use of semithin sections or nerve fiber teasing from a nerve biopsy in the diagnosis of peripheral neuropathies fails to identify more specific length-dependent and somatosensitive neuropathies. In humans, skin punch biopsy is used for this purpose, through the identification and count of intraepidermal nerve fibers (IENFs) with indirect immunofluorescence (IIF). However, the current need for frozen samples for this technique limits its routine application in clinical practice. In this study, we set up an IIF protocol to identify IENFs in dogs’ 8 mm skin punches using common formalin-fixed paraffin-embedded sections and the antibody anti-PGP9.5. (1) The effectiveness of the co-localization immunoreaction, (2) the thickness of sections, and (3) the magnification for image acquisition were checked. The best IIF results in terms of sharpness of fiber visualization and possibility to count them were obtained with 10 µm sections, at a ×40 magnification, without co-localization for nuclei and epithelial structures.