P. knowlesi Schizont Protein Characterization

Parasite lysates were prepared from P. knowlesi schizont as described previously [55 (link)]. Recombinant proteins were separated by 13% SDS-PAGE under reducing or non-reducing conditions and stained with 0.25% Coomassie brilliant blue R-250 (Sigma-Aldrich) and used for western-blotting analysis [25 (link)]. The membrane-transferred proteins were reacted with primary rabbit polyclonal serum (1:50) or an anti-His monoclonal antibody (1:2,000, Hilden, Hamburg, Germany) and then reacted with secondary IRDye-labeled goat anti-rabbit or goat anti-mouse antibodies (1:10,000) (LI-COR Bioscience, Lincoln, NE). An Odyssey infrared imaging system (LI-COR Bioscience) and Odyssey software (LI-COR Bioscience) were used to visualize the bands.