Only mice with correctly placed catheters were included in the analyses. To test the stability of the antibodies after 6 weeks in vivo (Figure S4A), we collected residual pump contents upon removal from the animals and assessed the antibodies using SDS-PAGE and Coomassie blue staining. Light and heavy chains were intact, with no fragmentation, and retained tau binding activity on western

blot (data not shown). To estimate the concentration of anti-tau antibodies in CSF and serum during the infusion, we administered biotinylated HJ8.5 (HJ8.5B) for 48 hr (∼7.2 μg/day) Selleck Alectinib (Figure S4A). The concentration of free HJ8.5B was 7.3 μg/ml in the CSF and 6.2 μg/ml in the serum, indicating clearance of the antibody from the CNS to the periphery (Figure S4C). see more We also detected HJ8.5B bound to human tau in both CSF and serum, though the concentration was lower than that of free antibody (Figure S4C). To determine the extent of tau pathology in P301S mice after 3 months of treatment, we carried out multiple stains for tau pathology. Brain sections were first assessed by immunostaining with the anti-phospho tau antibody AT8 (Figure 4). AT8 binds phosphorylated residues Ser202 and Thr205 of both mouse and human tau (Figure 4) (Goedert et al., 1995). In mice treated

with PBS and HJ3.4, AT8 strongly stained neuronal cell bodies and the neuropil in multiple brain regions, particularly in the piriform cortex, entorhinal cortex, amygdala, and hippocampus (Figures 4A and 4B). HJ8.5 treatment strongly reduced AT8 staining (Figure 4C), especially in the neuropil. HJ9.3 and HJ9.4 also decreased AT8 staining but the effects were slightly less (Figures 4D and 4E). Quantitative analysis of AT8 staining in piriform cortex (Figure 5A), entorhinal cortex (Figure 5B), and amygdala (Figure 5C) demonstrated a strong but variable reduction in phospho-tau in all

anti-tau antibody-treated mice. HJ8.5 antibody markedly reduced AT8 staining in piriform cortex, entorhinal cortex, and amygdala. HJ9.3 had slightly decreased effects compared to HJ8.5, and HJ9.4 had significant effects in both entorhinal cortex and amygdala but not in the piriform cortex (Figure 5). The hippocampus exhibited much more variable AT8 staining versus other brain regions, predominantly in cell bodies, however and thus was not statistically different in treatment versus control groups (Figure 5D). Because it has been reported that male P301S mice have greater tau pathology than females (Zhang et al., 2012), we also assessed the effect of both gender and treatment (Figure S5). In addition to an effect of treatment, there was significantly more AT8 staining in all brain regions analyzed in male mice (Figure S5C). However, the effects of the antibodies were still highly significant and virtually identical after adjusting for gender (Figure S5D). We also compared the treatment groups versus controls in males and females separately, and the effects of antibody HJ8.5 remained most significant (Figures S5A and S5B).