Immunofluorescence Staining Protocol for α-Tubulin

Immunofluorescence was conducted as described in our previous research (Peng et al., 2019 (link)). Specifically, the cells were fixed in 4% paraformaldehyde for 30 min and then treated with 0.3% Triton X-100 for 5 min. For non-specific blocking, the cells were incubated in 0.2% bovine serum albumin (Calbiochem, San Diego, CA, USA) for 15 min and then incubated with anti-α-tubulin antibody (dilution ratio was 1:100; GeneTex, Inc., North America, GTX628802) at 4°C for 12 h. The cells were stained with anti-mouse IgG fluorescent secondary antibody (dilution ratio was 1:200; Abways Biotechnology Co., Ltd., Shanghai, China, AB0132) for 2 h at room temperature. DNA was stained with Hoechst 33342 (Invitrogen). Fluorescence was imaged using OLYMPUS FV1200 confocal microscope (Olympus, Tokyo, Japan).