Analysis of Sarkosyl-insoluble tau from the hindbrain by PHF1 immunoblot.

Quantification of immunoblot signal of human and mouse tau doublet at (A) ~50 kDa and (B) ~25 kDa fragment. (C) The ratio of ~25 kDa fragment to ~50 kDa bands. Panel D is a representative blot showing Baseline (lanes 1–4); Vehicle (lanes 5–8), 30 mg/kg dose group (lanes 9–11), and 40 mg/kg dose group (lanes 12–15). The arrow in panel D indicates the 25 kDa tau fragment containing the ptau 396/404 epitope. The same preparations that were evaluated by ELISA in Fig 1 were resolved by 4–20% SDS-PAGE (Criterion TGX Gel, Bio-Rad Laboratories, Inc., Hercules, CA) run with reducing agent and transferred to a PVDF membrane (Trans-Blot Turbo Transfer System, Bio-Rad). The FluorChem R System (ProteinSimple, San Jose, CA) was used to capture images, and quantification of chemiluminescent signal was performed using AlphaView software (ProteinSimple). 30, 30 mg/kg dose group; 40, 40 mg/kg dose group; BL, Baseline; Veh, Vehicle.