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Cells were washed in PBS, resuspended in Thiazole Orange (BD Retic-Count, BD Bioscience) containing 4 M Hoechst 33342 (Invitrogen) for parasite RNA and DNA detection respectively ( 2 , 5 mM glucose, 32 mM HEPES pH7.4, 5 mM KCl, 1 mM MgSO4, 125 mM NaCl), resuspended in 0.3 M of intracellular calcium dye x-Rhod-1 AM (Invitrogen) and incubated in the dark at 37C for 30 min