Anti aβ 4g8 antibody

Right hemispheres (fixed in 10% formalin) were processed for paraffin embedding. Serial sagittal brain slices of 10µm of thickness were sectioned using a microtome. For Aβ staining, brain sections were deparaffined and treated with 85% formic acid for 5 min at room temperature. Then, sections were incubated overnight with the anti-Aβ 4G8 antibody (specific for human Aβ, Biolegend, 800708, San Diego, CA, USA) diluted 1:1000 in PBS/0.02% v/v Triton X-100 at room temperature. Sections were washed and incubated with goat anti-mouse secondary antibody, Alexa Fluor™ 594 (Thermo Fisher Scientific, Waltham, MA, USA), diluted 1:500 in PBS, for 90 min at room temperature. Stained sections were washed and cover-slipped with ProLong™ Gold Antifade Mountant (P36930, Thermo Fisher Scientific, Waltham, MA, USA). For ThS staining, sections were deparaffined and incubated with 0.1% ThS (Sigma, St. Louis, MO, USA) diluted in 50% ethanol for 10 min. ThS-stained sections were washed two times in 50% ethanol followed by two washes in PBS. Sections were dehydrated in graded ethanol, cleared in xylene, and cover-slipped with DPX mounting medium (Innogenex, San Ramon, CA, USA).

Transgenic 12-month-old APPswe/PSEN1dE9 mice (The Jackson Laboratory) were anesthetized and perfused with 30 ml of ice-cold PBS and then with 4% paraformaldehyde in PBS. Brains were removed and postfixed at 4 °C overnight, followed by 20 and 30% sucrose in PBS at 4 °C overnight. Brains were cut into 30 μm coronal sections with a cryostat (Leitz 1900) at − 20 °C. Immunolabelling was performed using the anti-Aβ 4G8 antibody (1:100, Biolegend, CA). Anti-mouse-IgG conjugated with Alexa Fluor-488 (1:1000, Molecular Probes), were used as secondary antibodies. Finally, sections were washed four times for 10 min with PBS and then incubated with fNDs for 1 h at a concentration 0.1 nM. The sections were then washed four times for 10 min with PBS and mounted with mounting medium DAKO.