Total and oligomeric tau protein levels are reduced with DOX treatment. (A) Representative western blot (WB) probed for total human tau. (B) Quantification of total tau western blots shows a significant decrease in total human tau in DOX treated rTg4510 and APP × rTg4510 mice. Treatment effect F(1,29) = 127.4, P < 0.0001. (C) Total human tau levels as detected by ELISA were significantly reduced in DOX-treated rTg4510 and APP × rTg4510 CSF compared to naïve genotype controls. Treatment effect F(1,26) = 49.20, P < 0.0001. Two-way ANOVA, Sidak post hoc analysis. (D) SDD-AGE blot probed for total tau shows a reduction in high molecular weight (HMW) and low molecular weight (LMW) tau. Black dashed line separates treatment. Red dashed line separates genotype. Quantification is presented in Supplementary Fig. 3. (E) Total tau densitometry values (a.u. = arbitrary units) down the length of the SDD-AGE blot. (F) The per cent tau in the bins down the length of the blot was calculated for each sample and plotted as a group. APP × rTg4510 DOX samples have a higher percentage of high molecular weight tau and lower percentage of low molecular weight tau than the rTg4510 DOX samples. Treatment effect F(12,84) = 11.85, P < 0.0001. Two-way ANOVA, Sidak post hoc analysis. (G) Naïve rTg4510 and APP × rTg4510 lysate incubated with increasing concentrations of proteinase K (PK) was run on western blot. More <14 kDa fragmented tau in the rTg4510 digested sample compared to the APP × rTg4510 sample (arrowheads) was identified at 1 µg/ml proteinase K. (H and I) Lysates were incubated with 1 µg/ml proteinase K and run on western blot for total tau. Representative blot shows the proteinase K digestions patterns (H). The relative intensity of the <14 kDa tau fragments normalized to total tau in the full lane showed a significant decrease in the amount of small proteinase K fragmented tau in the APP × rTg4510 DOX lysate as compared to the rTg4510 DOX lysate (I). Two-tailed student t-test. (J–L) Proteinase K dose escalation was run on lysate from neurofibrillary tangle containing Braak III human brain with no amyloid-β plaques (amyloid-β−) and with frequent amyloid-β plaques (amyloid-β+). An increase at 2.5 µg/ml proteinase K in >28 kDa tau in the amyloid-β+ digested sample as compared to the amyloid-β− sample (arrowheads and red box) was identified (J). All amyloid-β− and amyloid-β+ samples were incubated with 2.5 µg/ml proteinase K and then run on western blot for total tau. Representative blot shows the proteinase K digestions patterns (K). The ratio of the >28 Da tau bands to total tau in the full lane (L). Two-tailed student t-test. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Each data point represents one mouse or human sample. Mean ± SEM.