Biotinylated goat anti human igg

The sensitivity of the antibody 82E1 and the biosimilar Bapineuzumab antibody was further analysed by urea-bicine/bis-tris/Tris/sulphate sodium dodecyl sulphate (SDS)-PAGE followed by western immunoblotting. Stock solutions of synthetic Aβ_1–40 were prepared in sample buffer (0.36 M bis-tris, 0.16 M bicine, 15% wt/vol sucrose, 1% wt/vol SDS and 0.0075% wt/vol bromophenol blue) and stored at −80°C. Different amounts of Aβ_1–40 peptides (25–100 pg and 25–100 ng respectively) were loaded and separated by urea-bicine/bis-tris/Tris/sulphate SDS-PAGE and analysed by immunoblotting as previously described [40 (link)]. The blotting membranes were blocked in 2% enhanced chemiluminescence (ECL) advance blocking agent (GE Healthcare Life Sciences, Little Chalfont, UK) in phosphate-buffered saline (PBS) with 0.075% Tween 20 (PBS-T) for 1h at room temperature and probed with the primary antibody 82E1 or the biosimilar Bapineuzumab antibody (0.5 μg/μl in PBS-T) at 4°C overnight. After three washing steps with PBS-T, biotinylated goat anti-mouse IgG (1:3000 in PBS-T; Life Technologies, Carlsbad, CA, USA) for 82E1 and biotinylated goat anti-human IgG (DAKO, Glostrup, Denmark, 1:3000) for Bapineuzumab, followed by streptavidin-HRP, were employed as secondary reagents. Chemiluminescent signals obtained with ECL Prime Western Blotting Detection Reagent (GE Healthcare Life Sciences) were recorded.