Figure 10.

The typical images of filopodia and lamellipodia. Cultured cells were fixed in 3.5% (w/v) paraformaldehyde, permeabilized in 0.2% (v/v) Triton X-100 and blocked in 2% (w/v) bovine serum albumin (BSA). The cells were incubated with anti-cortactin antibody at 4 °C overnight, followed by Alexa Fluor 594-conjugated IgG (Thermo Fisher Scientific, Yokohama, Japan) as the secondary antibody and Acti-stain 488 phalloidin (Cytoskeleton) for actin-fiber staining. After incubation, SlowFade gold antifade reagent with 4′,6-diamidino-2-phenylindole (Invitrogen/Life Technologies) was added to the cells. The specimens were observed by fluorescence microscopy (Olympus IX73; Olympus, Tokyo, Japan). We determined lamellipodia formation by evaluating fluorescent actin fibers and cortactin co-localization at the cell periphery. (A) Arrow heads indicate filopodia which are slender cytoplasmic projection that extended beyond the leading edge in migrating cells. (B) It is known that actin and cortactin co-localizes at lamellipodia. These images show the immunofluorescence staining of actin or cortactin, in HSC4 cells. Arrow heads indicate lamellipodia.