Anti acetylated α tubulin 6 11b 1

A previously published protocol was followed (44 (link),56 (link)). Primary antibodies were: Custom-made anti-Wtip (1:100 dilution; Covance, Inc., Denver, PA, USA), anti-γ-tubulin (GTU-88; 1:800 dilution; Sigma-Aldrich), anti-acetylated α-tubulin (6-11B-1; 1:800 dilution; Sigma-Aldrich), and anti-centrin (20H5; 1:1,000 dilution; EMD Millipore, Billerica, MA, USA). The following secondary antibodies were obtained from Thermo Fisher Scientific, Inc.: Goat anti-rabbit Alexa Fluor^®546 (IgG [H+L]), goat anti-mouse Alexa Fluor^®488 (IgG2b), goat anti-mouse Alexa Fluor^®488 [IgG1 (γ1)], goat anti-mouse Alexa Fluor^®488 [IgG2a (γ2a)] and goat anti-mouse Alexa Fluor^®546 [IgG1 (γ1)]. Whole-mount immunohistochemistry samples were dehydrated with a graded series of methanol, embedded in JB4 resin (Polysciences, Inc., Warrington, PA, USA), and were cut into 5–7 µm sections using an RN2255 microtome (Leica Technology, Exton, PA, USA). The sections were stained with DAPI (Kirkegaard & Perry Laboratories, Inc., Gaithersburg, MD, USA), mounted in Fluorescent Mounting Media (Kirkegaard & Perry Laboratories, Inc.), and were imaged with a FV-1000 confocal laser-scanning microscope (Olympus America, Inc., Center Valley, PA, USA).