Immunofluorescence Staining of Skin Tissues

Immunofluorescence in tissues was performed as described previously^{16 (link),37 (link)–39 (link)}. Briefly, the skin tissues were embedded, fixed, permeabilized, incubated with primary mouse anti-K5 (1:200; Progen Biotechnik, Heidelberg, Germany), rabbit anti-Filaggrin (1:1000; Abcam, Cambridge, MA), rabbit anti-E-cadherin (1:500; Cell Signaling, Danvers, MA), mouse anti-β-catenin (Cell Signaling, Danvers, MA), rabbit anti-F-actin (Cell Signaling, Danvers, MA) and rabbit anti-ZO-1(Cell Signaling, Danvers, MA), and then incubated with Alexa Fluor 488 F (ab’) 2 fragments of goat anti-guinea pig IgG antibodies and Alexa Fluor 568 of goat anti-rabbit IgG antibodies (Invitrogen, Carlsbad, CA). The cells were then fixed in Prolong Gold Antifade with DAPI (Invitrogen, Carlsbad, CA) to visualize the cell nuclei, and observed under a fluorescence microscope (OlympusIX71, Japan) with a peak excitation wavelength of 340 nm.