Smart sem v05

Dry conidia were transferred to an aluminum specimen mount precoated with sticky carbon conductive adhesive tape and examined with an EVO 18 scanning electron microscope (Zeiss, Oberkochen, Germany). The working temperature of the SEM stage module was − 30 to 50 °C. SEM working conditions were as follows: working distance (WD): 11 mm, electron high tension (EHT): 20.00 kV, signal A: secondary electron (SE1). Several images for each sample were digitally produced and registered at variable magnifications with the Zeiss SEM operating software Smart SEM® V05.06. The statistical analysis was done using the built-in software package in the ImageJ program.

Preparation of the kidneys for scanning electron microscopy was done according to Tanaka et al. (Tanaka et al., 1989 (link)) with modifications. Mice were perfusion-fixed with 1× PBS with 4% paraformaldehyde through the distal abdominal aorta for 3 min. Kidneys were cut in half and stored in 2% glutaraldehyde with 0.1 M sodium cacodylate pH 7.4 at 4°C until further processing. Samples were post-fixed in 1% OsO₄ in the same buffer. After incubation in 30% dimethylformamide at 4°C overnight, freeze fracturing was performed with a cold knife under liquid nitrogen followed by immersion in dimethylformamide at room temperature for 1 h. After serial dehydration in increasing acetone concentrations, samples were dried at critical point (Leica EM CPD300), coated with platinum (Edwards Scancoat) and finally examined on a Zeiss LEO 1530 Gemini scanning electron microscope using the SE2 detector and the Smart SEM V05.06 software (Carl Zeiss Microscopy, Oberkochen, Germany). Measurements of ciliary length were performed with ImageJ.

Film microstructure was characterized by means of a ZE122 SEM Supra 40 scanning electron microscope (Carl Zeiss, Jena, Germany). The cross sections and the surface of films were metalized with a platinum layer prior to observation. The image analysis was performed by means of SmartSEM^® V05.06 software (Carl Zeiss, Jena, Germany).