Immunohistochemical Analysis of Rac and p-Src

Immunohistochemistry was used to measure the levels of Rac and p-Src in the cells. After PBS washing, rehydrated culture dishes were incubated with primary antibodies against Rac (ab155938) and p-Src (ab40660) purchased from Abcam (Tokyo, Japan) diluted at 1:100 in Ab Diluent (Dako ChemMate; Dako, Japan) overnight at room temperature. For staining with Alexa Fluor 488 anti-rabbit IgG (Invitrogen, Carlsbad, CA, USA), secondary antibodies were diluted at 1:200 in Ab Diluent and added for 60 min at room temperature in the dark. Digital images were taken on a BIOREVO microscope equipped with a confocal microscopy system (BZ-9000, Keyence, Japan) as described previously^{28 (link)}.