Fig. 4

Characterization of different tau aggregates: truncated tau (dGAE), human AD brain-derived tau and ability of DC8E8 antibody to recognise various forms of diseased tau protein. a Kinetics of t-tau fibril formation was monitored over time by Thioflavin T fluorescence until it had reached plateau. t-tau indicates recombinant truncated PHF-core tau (dGAE, 297–391/4R). b FTIR spectroscopy confirmed a change in the structure after in vitro fibrillization of recombinant PHF-core subunit dGAE. The prevalent secondary structure changed from disordered to beta-rich structure. c Dynamic light scattering measurements of t-tau monomer (dGAE 297–391), heparin fibrillized t-tau, human brain-derived sporadic and familial AD tau (sarkosyl-insoluble 2p fraction, sonicated). All three sonicated fibrillary tau species were composed of high-molecular particules with an average radius spanning from 20 to 100 nm. d Non-reducing SDS-PAGE using pan tau antibody DC25 confirmed formation of tau fibrils (297–391/4R). Tau (dGAE 297–391) monomer has been used as a control representing band of one specific size (1 μg, band runs at 10–15 kDa). Heparin-assembled t-tau showed a pattern with more fibrillary species. e Western blot analysis of AD-tau preparations (sporadic and familial form) using microtubule binding domain DC8E8 antibody showed as individual fractions of sarkosyl-insoluble AD-tau preparations (2p, 1 s, 2 s). Both preparations of sarkosyl-insoluble fraction (2p) confirmed the presence of a wide range of assembled tau inclusions with DC8E8 antibody. f Sarkosyl-insoluble high-molecular weight tau isolated by centrifugation partially dissociates on the SDS PAGE and forms a typical A68 triplet. DC8E8 captured all diseased tau forms present in the sarkosyl extract from human AD brain (sporadic, Braak VI). The Western blot was developed with DC25 pan tau antibody. The panel on the right shows the same membrane stained with Ponceau S protein dye with the major bands corresponding to the chains of antibodies used in immunoprecipitation. Six recombinant tau protein isoforms were included for comparison