Immunofluorescence assay using anti-PFI1140w and anti-PF13_0353 antisera. Purified P. falciparum trophozoites were incubated with a 1/500 dilution of antisera recognizing coding sequences of either PFI1140w or PF13_0353. Goat anti-mouse IgG conjugated with Alexa 633 was used as a secondary antibody. Nuclear DNA was labeled with Hoescht 33258. (A) DIC image of trophozoites treated with anti-PFI1140w antiserum (B) nuclear DNA stained with Hoescht 33258 and (C) negative Alexa 633 signal. (D) Merged images of (A–C). (E) DIC image of trophozoites treated with anti-PF13_0353 antiserum, (F) nuclear DNA stained with Hoescht 33258 and (G) Alexa 633 signal recognizing anti-PF13_0353 antibody. (H) Merged images of (E–G). PF13_0353 is localized in or in close proximity to the food vacuole.Ostera G, Tokumasu F, Oliveira F, Sa J, Furuya T, Teixeira C, Dvorak J. Plasmodium falciparum: food vacuole localization of nitric oxide-derived species in intraerythrocytic stages of the malaria parasite. Exp Parasitol. 2008 120:29-38.

Evidence of L-arginine-dependent production of NO in P. falciparum-harboring RBCs. Uninfected RBCs and PfRBCs incubated with the fluorescent dye 4,5-diaminofluorescein diacetate (DAR-4M AM, a specific NO probe) and L-arginine showed intense fluorescence in the parasite (red color), increasing with the developmental stage of the parasite, indicating intra-parasite production of NO (A1-5). Preincubation of PfRBCs with 3 mM L-NAME, a competitive inhibitor of mammalian NOS, caused a significant reduced fluorescence (B). Intense intra-parasite fluorescence from a P. falciparum sample freshly isolated from malaria patients and incubated in vitro during 48 hours ((C), one representative sample).Rey J, Buffet PA, Ciceron L, Milon G, Mercereau-Puijalon O, Safeukui I. Reduced erythrocyte deformability associated with hypoargininemia during Plasmodium falciparum malaria. Sci Rep. 2014 Jan 20;4:3767. PMID: 24441939

Detection of nitric oxide synthase in both healthy and P. falciparum-harboring red blood cells. NADPH-diaphorase activity within healthy RBCs or PfRBCs at different stages of parasite development (A). The panel (A1) represents the negative control (uRBCs or PfRBCs incubated with NBT without NADPH). NADPHd staining (blue color) was observed in both uRBC and parasite (A2). Identification by immunofluorescence assay (IF) of NOS within uRBCs or PfRBCs using rabbit polyclonal antibody against uNOS (B1, green) or mouse monoclonal antibody against eNOS (B2, green). To differentiate infected from uninfected RBCs, parasite nucleus was labeled in blue with Hoechst 33342. A monoclonal antibody against parasiteSERA5 was used to localize the parasitophorous vacuole (red).Rey J, Buffet PA, Ciceron L, Milon G, Mercereau-Puijalon O, Safeukui I. Reduced erythrocyte deformability associated with hypoargininemia during Plasmodium falciparum malaria. Sci Rep. 2014 Jan 20;4:3767.