α6β4 integrin is involved in collective motion. (A) A maximum intensity projection image of immunofluorescence against α6 and β4 integrin in a growing colony by confocal microscopy. Bar, 50 µm. (B) Magnification of the area outlined by the white boxes in A. Optical sections show α6β4 integrin on basal and lateral surfaces. Bar, 10 µm. (C) Detergent-insoluble α6β4 integrin on basal surface. α6β4 integrin can be linked with cytokeratins via cytoplasmic proteins, and cytoskeleton-linked α6β4 integrin is detergent insoluble and can be detected even if cells are treated with detergent before fixation (Rabinovitz et al., 1999). In keratinocytes, α6β4 integrin was also detectable when treated with 0.5% Triton X-100 before fixation. Bar, 20 µm. (D) Electron micrographs of the growing and terminal colonies of human epidermal keratinocytes. Arrowheads indicate well-defined electron-dense structures of hemidemosomes in the basal surface. Bar, 500 nm. (E) Effect of inhibitory antibodies against α6 (GoH3) and β4 integrin (ASC-8) on keratinocyte locomotion in keratinocyte aggregates. Two-dimensional keratinocyte aggregates were formed after high density keratinocyte inoculation with antibodies. P-values were calculated by Student’s t test.