Western Blot Analysis
Whole protein extracts were diluted to concentrations of 1 mg/mL and separated by SDS-PAGE under reducing conditions (2.5% β-mercaptoethanol). Membranes were incubated with rabbit anti-Ara h 1, Ara h 2 or Ara h 8 polyclonal antibody (1:1,000, 4°C overnight; Indoor Biotechnologies, VA, USA). Bound antibodies were visualized using horseradish peroxidase (HRP)-conjugated donkey anti-rabbit IgG monoclonal antibody (1:1,000, 1h room temperature; BioLegend, CA, USA) and Clarity/Clarity Max enhanced chemiluminescence (ECL) substrates (Bio-Rad Laboratories, Canada). Products were visualized with the ChemiDoc XRS+ Imaging System (Bio-Rad Laboratories).