Anti krt10

Human and mice skin tissues were fixed with 4% paraformaldehyde in PBS, embedded in paraffin, sectioned, and stained with H&E for histopathologic examination. Immunohistochemistry was performed on formalin-fixed, paraffin-embedded skin tissues. The slices were probed overnight with SerpinB7 (Sigma-Aldrich, HPA024200), mKi67 (Abcam, ab16667). For Immunofluorescence, skin frozen sections were cryosectioned, blocked, and then stained with anti-Filaggrin (Biolegend, 905804), anti-KRT10 (Abcam, ab76318), anti-Loricrin (Biolegend, 905101). Images were captured using an Olympus BX600 microscope (Olympus Corporation, Tokyo, Japan) and SPOT Flex camera (Olympus Corporation, Tokyo, Japan) and were analyzed with ImagePro Plus (version 6.0, Media Cybernetics) software. The epithelial thickness and infiltrating cells were calculated in independent regions as described previously [47 (link)].

Tissue was fixed with 4% paraformaldehyde then frozen in Optimal Cutting Temperature Compound (OCT) and sectioned at 20 µM. Sections were blocked with IHC/ICC Blocking Buffer (Invitrogen) with 0.4% Triton X-100 (Sigma), incubated with primary and secondary (1:5000) antibodies and finally mounted in Prolong Gold Antifade Mount with DAPI (ThermoFisher). Antibodies and dilutions Anti-Ki67, 1:100 (Abcam ab15580); Anti-KRT14 1:200 (Thermo Fisher MA1-06323); Anti-CD200, 1:50 (ls-b11638); AQ3 1:200 (Abcam ab125219); Anti-KRT10 1:200 (Abcam ab9025); Anti-KRT40, 1:100 (Abcam ab16113); Anti-KRT5, 1:100 (Abcam ab64081). Immunostained samples were visualized by confocal microscopy (Olympus Fluoview FV3000 Confocal Microscope). Line profile intensity was measured using ImageJ (NIH).

The 4% formaldehyde-fixed and dehydrated skin tissue samples were embedded in paraffin wax and cut into 5-µm-thick sections. The skin sections were incubated with a primary antibody against human KRT10 (1:100; Abcam, Cambridge, UK), followed by incubation with the adequate secondary antibody. After staining, sections were counterstained with hematoxylin to provide contrast. For immunofluorescence staining, the skin tissues were quench-frozen and embedded in OCT. The cryosections (10 µm) were fixed in chilled acetone for 15 min and incubated overnight at 4 °C with anti-S100A9 (1:400; Abcam, Cambridge, UK), anti-HBD-2 (1:400; Abcam, Cambridge, UK), anti-Filaggrin (1:400; Abcam, Cambridge, UK), anti-Involucrin (1:400; Abcam, Cambridge, UK), anti-AO-1 (1:400; Abcam, Cambridge, UK), anti-CLDN-1 (1:400; Abcam, Cambridge, UK), anti-KRT6 (1:400; Abcam, Cambridge, UK), anti-KRT10 (1:400; Abcam, Cambridge, UK), and anti-KRT17 (1:400; Abcam, Cambridge, UK) primary antibodies, followed by Alexa Fluor 488- or 568-conjugated secondary antibodies and DAPI. The immunostained images were obtained using a digital camera (DP74; Olympus, Tokyo, Japan) coupled with an optical microscope (BX53; Olympus, Tokyo, Japan).