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Fig. 1. Cruzipain and chagasin co-localize to reservosomes and the Golgi apparatus of T. cruzi epimastigotes. Epimastigotes were fixed in 4% paraformaldehyde, adhered on poly-L-lysine-coated glass coverslips and permeabilized with PBS containing 1% NP-40. The parasites were incubated overnight at 4°C with anti-chagasin rabbit antiserum or mouse anti-cruzipain antiserum, followed by incubation with Alexa-543-conjugated anti-rabbit and FITC-conjugated anti-mouse antibodies. (A) Differential-interference-contrast image. (B) Anti-cruzipain labelling. (C) Anti-chagasin labelling. (D) Merged image of anti-cruzipain and anti-chagasin labelling. Arrows and arrowheads, respectively, indicate the posterior and anterior ends of the cell. Scale bar, 2 µm. Epimastigote cryosections were incubated with rabbit anti-chagasin antiserum or mouse anti-cruzipain antiserum at 1:100 and 1:500 dilutions. The sections were subsequently incubated with goat anti-rabbit IgG conjugated to 5 nm gold particles and goat anti-mouse IgG conjugated to 15 nm gold particles (at 1:100), respectively. (E) T. cruzi reservosomes. (F) A higher-magnification image of the interior of a reservosome showing cruzipain and chagasin in close proximity. (G) A reservosome (R) at high magnification, next to the Golgi complex (GC). Dark arrows indicate cruzipain staining, arrowheads indicate chagasin staining and white arrows point to chagasin inside cruzipain-free small vesicles. Scale bars, 0.1 µm (E), 0.4 µm (F) and 0.1 µm (G).
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