The whole-mount sciatic nerves were washed in 0.3% Triton X-100/PBS before blocked in 10% bovine serum albumin (BSA) overnight. They were incubated in mouse anti-GAP43 antibody (1:200, Thermo Fisher Scientific, Waltham, MA, USA) for 48 h at 4 °C. After a stringent wash, anti-mouse IgG (H + L) Alexa Fluor® 594 (1:500, Jackson ImmunoResearch, West Grove, PA, USA) was applied to the nerves and incubated in a dark box for 24 h at 4 °C. The nerves were then washed, mounted on glass slides in Vectashield® (Vector Laboratories, Burlingame, CA, USA), and viewed with an Olympus Fluoview 1000 confocal microscope.
The DRG neurons on coverslips were rinsed briefly with PBS and fixed in 4% paraformaldehyde/PBS for 20 min at room temperature. They were then rinsed and blocked in 2% BSA. Mouse anti-tau antibody (1:500, Sigma-Aldrich, St. Louis, MO, USA) was applied to the coverslips and incubated in a humid chamber for 2 h, followed by anti-mouse IgG (H + L) Alexa Fluor® 594 (1:1000, Jackson ImmunoResearch, West Grove, PA, USA) for 45 min. They were then rinsed and stained for DAPI before being coverslipped for imaging.
The DRG neurons on coverslips were rinsed briefly with PBS and fixed in 4% paraformaldehyde/PBS for 20 min at room temperature. They were then rinsed and blocked in 2% BSA. Mouse anti-tau antibody (1:500, Sigma-Aldrich, St. Louis, MO, USA) was applied to the coverslips and incubated in a humid chamber for 2 h, followed by anti-mouse IgG (H + L) Alexa Fluor® 594 (1:1000, Jackson ImmunoResearch, West Grove, PA, USA) for 45 min. They were then rinsed and stained for DAPI before being coverslipped for imaging.