GFP fluorescence in parasites expressing GFP-2xFYVE. This construct has been used in various organisms to monitor the intracellular localization of PI3P. Colocalization of GFP-2xFYVE and the food vacuole membrane marker CRT using rabbit anti-CRT. Serial images of a Z-stack acquisition (0.38 mm step) are displayed. Arrowheads indicate protrusions from the food vacuole membrane that are colabeled with CRT and GFP-2xFYVE, while arrows indicate zones of intense GFP-2xFYVE staining in the absence of the food vacuole marker. A corresponding phase-contrast image is shown in the lower left corner. Fluorescence was associated mainly with the membrane of the food vacuole.Tawk L, Chicanne G, Dubremetz JF, Richard V, Payrastre B, Vial HJ, Roy C, Wengelnik K. Phosphatidylinositol 3-phosphate, an essential lipid in Plasmodium, localizes to the food vacuole membrane and the apicoplast. Eukaryot Cell. 2010 9:1519-30.

Intracellular localization of PfPI3K and its trafficking to the host. (A) IFA was performed for localizing PfPI3K, FCP (i) and VARC (ii). FCP (red) is located inside the food vacuole of mature trophozoites. PfPI3K (green) exhibits “vesicular” staining on PVM/PM (black arrows) and the host erythrocyte (white arrows) and in the food vacuole, which can be identified by the presence of black hemozoin. (ii) PfPI3K colocalizes with VARC at the erythrocyte surface. PfPI3K localization. Lower row: Early trophozoites on thin blood smears were used for IFA to assess the localization of PfPI3K (green). The parasite nuclei (blue) are stained with DAPI. The arrows indicate PfPI3K near the host RBC membrane.Vaid A, Ranjan R, Smythe WA, Hoppe HC, Sharma P. PfPI3K, a phosphatidylinositol-3 kinase from Plasmodium falciparum, is exported to the host erythrocyte and is involved in hemoglobin trafficking. Blood. 2010 115:2500-2507.

Colocalization of GFP-2xFYVE with the apicoplast. This construct has been used in various organisms to monitor the intracellular localization of PI3P. GFP-2xFYVE expressing parasites at the indicated developmental stages were incubated with rabbit anti-ACP (acyl carrier protein) as an apicoplast marker. Nuclei were stained with Hoechst stain (Hoe). Fluorescence was associated with the apicoplast.Tawk L, Chicanne G, Dubremetz JF, Richard V, Payrastre B, Vial HJ, Roy C, Wengelnik K. Phosphatidylinositol 3-phosphate, an essential lipid in Plasmodium, localizes to the food vacuole membrane and the apicoplast. Eukaryot Cell. 2010 9:1519-30.

Localization of GFP-2xFYVE by cryoimmunoelectron microscopy. This construct has been used in various organisms to monitor the intracellular localization of PI3PK. (A) Section through a young schizont showing most of the gold label on or near the food vacuole membrane (arrows). Arrowheads point to what should be considered the background. Hz, hemozoin crystals; N, parasite nucleus; RBC, erythrocyte cytosol. (B and C) The membranes of electron-lucent (B) (arrow) and electron-dense (C) (asterisks) vesicles neighboring the food vacuole (Hz) are strongly labeled. Cytostomal vesicles (Cy) are not labeled. (D to E) General (D) and close-up (D’ and E) views of young schizonts showing the strong labeling of the apicoplast, mostly within the lumen. The four membranes of the apicoplast are clearly identified (E, arrow). A, apicoplast; P, parasite.Tawk L, Chicanne G, Dubremetz JF, Richard V, Payrastre B, Vial HJ, Roy C, Wengelnik K. Phosphatidylinositol 3-phosphate, an essential lipid in Plasmodium, localizes to the food vacuole membrane and the apicoplast. Eukaryot Cell. 2010 9:1519-30.