Infection of THP-1 Cells with Tachyzoites

Infected HFFs were lysed and filtered through a 3-μm pore-sized membrane to remove host cell debris. Prepared tachyzoites were added to THP-1 cells at a density of 10⁶ cells/mL (parasite/cell ratio 2). After incubation for 24 h at 37°C in an atmosphere containing 5% CO₂, infection was confirmed under the fluorescence microscope. Then, supernatants were separated by centrifugation at 1000g for 10 min and filtration through a 0.22-μm pore-sized filter. The collected supernatants were stored at -20°C. Heat-inactivated tachyzoites were prepared after incubation at 56°C for 30 min as previously described [14 (link), 15 (link)]. For immunocytochemistry, THP-1 cells were labeled with CellTracker^™ Deep Red (Invitrogen) prior to the addition of the tachyzoites.