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Figure 5 | BMC Microbiology

Figure 5

From: The Vacuolar ATPase from Entamoeba histolytica: Molecular cloning of the gene encoding for the B subunit and subcellular localization of the protein

Figure 5

Localization of EhV-ATPase B subunit in trophozoites during RBC phagocytosis. Trophozoites were incubated with diamine bencidine-stained RBCs for 5 min (a-j), washed with PBS and fixed with paraformaldehyde. Then, cells were treated with anti-rEhBvac21 antibodies and FITC-labeled goat anti-mouse antibodies, and observed through a laser confocal microscope as described. (a, e, i). Immunodetection of EhV-ATPaseB observed in green channel. (b, f) Cells observed in red channel. (c, g) Merge of fluorescent signals observed in (a, b) and (e, f), respectively. (d, h, j, l) Cells observed by light microscopy. (k) Cells incubated only with secondary antibodies as negative control and observed in green channel. (m, n) Electrophoresis in 1.0% agarose gels of RT-PCR gene products of Ehvma2 (m) and actin (n) in trophozoites at 0 min (lane 2) and 30 min (lane 3) of erythrophagocytosis. Lane 1, 100 bp size molecular ladder. Arrowheads, EhV-ATPaseB found in plasma membranes. Arrows, EhV-ATPaseB polypeptide found in cytoplasmic vesicles. Dotted arrows, EhV-ATPaseB polypeptide found in phagocytic vacuoles containing RBCs. Asterisks, huge concentrations of EhV-ATPaseB found within trophozoites.

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