Fig 1: Tau phosphorylation does not change in Braak II samples compared to Braak 0–I controls. Normalized phospho-tau signals from Braak II and Braak 0–I a) entorhinal cortices (EC), b) Hippocampi (Hip) and c) Temporal cortices (TC). Biotinylated antibodies were used as capture, sulfo-tagged Tau12 was used for detection. None of the observed changes were significant (p > 0.05, t-tests)
Fig 2: Tau PTMs in denatured Braak III–IV samples. Normalized PTM signals from a) Entorhinal cortices (EC), c) hippocampi (Hip), and e) temporal cortices (TC) of Braak stages 0–I and III–IV. b, d, f) Corresponding fold changes (log2) versus significance (-log10(p-value)) of the changes. Phosphorylation at the sites above the red line, which corresponds to p-value = 0.05, is significantly higher in Braak III–IV samples
Fig 3: Tau multimers can be disrupted by boiling in SDS-containing buffer. a) Comparison of tau multimer levels in entorhinal cortices, hippocampi, and temporal cortices between Braak 0–I and Braak III–IV after boiling with SDS-containing buffer. Comparison of total tau levels in b) Entorhinal cortices (EC) c) Hippocampi (Hip) and d) Temporal cortices (TC) between Braak 0–I and Braak III–IV
Fig 4: Immunoblotting using hTg-Tau and Tau-KO brain lysates confirms the specificity for tau for a panel of antibodies. a) Scheme of tau (2N4R) showing the epitopes of the Tau12, Tau5, Tau1, HT7, BT2 and Dako antibodies. b) - d’) Immunoblots demonstrate strong reactivity of all antibodies with tau bands between 40 and 60 kD. Minor non-specific bands present in Tau-KO brain lysates were detected in d), e), i), h), p), x), y) and c’). Isoform-specific antibodies in h), i), j) and k) were validated using a tau ladder with all 6 recombinant human tau isoforms. kDa sizes for the marker are given in e’). Tg: hTg-Tau mouse, KO: Tau-KO mouse brain lysate, TL: recombinant tau ladder
Fig 5: An electrochemiluminescence ELISA detects tau PTMs in hTg-Tau mouse brain lysates. Sandwich ELISA assays using biotinylated antibodies for capture and sulfo-tagged Tau-12 antibody for detection demonstrate strong and specific signals for all antibodies in the panel. a Tau-12 cannot be used as capture and detection antibody at the same time, while all other total tau and tau isoform-specific antibodies in the panel are suitable to detect tau in hTg-Tau brain lysate. b All non-phospho-PTM modifications in the panel can be detected in hTg-Tau brain lysate. c All phospho-specific tau antibodies in the panel give signal from hTg-Tau brain lysate
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