For fluorescence microscopy, C. albicans cells were fixed in 50 mM thimerosal and stained for exposed β-glucan (1.5 µg/ml Fc-Dectin-1 plus anti-human IgG conjugated to Alexafluor 488; green) and exposed mannan (25 µg/ml Concanavalin A conjugated to Alexafluor 647; red). All samples were examined by phase differential interference contrast (DIC) and fluorescence microscopy using a Zeiss Axioplan 2 microscope. Images were recorded digitally using Openlab v 4.04 (Improvision, Coventry, UK) with a Hamamatsu C4742- 95 digital camera (Hamamatsu Photonics, Japan). Fluorescence was quantified using and processed using Openlab (Openlab v 4.04: Improvision, Coventry, UK).
C. albicans cell walls were examined by high-pressure freeze substitution transmission electron microscopy (TEM). C. albicans SC5314 cells were incubated for 6 h at 37 °C with extracts from the small intestine, cecum or large intestine (above). These fungal cells were then processed for TEM as described previously (Ene et al., 2012 (link)), cutting ultrathin (100 nm) sections. Samples were imaged with a Philips CM10 transmission microscope (FEI, United Kingdom) equipped with a Gatan Bioscan 792 camera, and the images recorded using a Digital Micrograph (Gatan, Abingdon Oxon, United Kingdom).
C. albicans cell walls were examined by high-pressure freeze substitution transmission electron microscopy (TEM). C. albicans SC5314 cells were incubated for 6 h at 37 °C with extracts from the small intestine, cecum or large intestine (above). These fungal cells were then processed for TEM as described previously (Ene et al., 2012 (link)), cutting ultrathin (100 nm) sections. Samples were imaged with a Philips CM10 transmission microscope (FEI, United Kingdom) equipped with a Gatan Bioscan 792 camera, and the images recorded using a Digital Micrograph (Gatan, Abingdon Oxon, United Kingdom).