Orius camera

Tissue preparation and image acquisition were as previously described (Vladar et al., 2012 (link); Vladar et al., 2015 (link)). Proximal airway direction was tracked throughout the procedure. Tracheas were fixed in 2% glutaraldehyde, 4% paraformaldehyde in 0.1 M NaCacodylate buffer, pH 7.4 at 4°C overnight. Samples were osmicated, stained with uranyl acetate, then dehydrated with a graded ethanol series and infiltrated with EMbed-812 (Electron Microscopy Sciences). 80–100 nm sections were mounted onto copper grids and analyzed with a JEOL JEM-1400 microscope using a Gatan Orius Camera.

Samples were prepared
by incubating at 37 °C for 96 h 100 μL of phosphate buffer
solutions 10 mM and 100 mM NaCL, pH 7.4 containing Aβ_1-42 100 μM or Aβ_1-42 100 μM: Aβ_8-20 100 μM. After incubation, 3 μL of each
sample was deposited onto a copper grid and allowed to adsorb for
5 min before the grid was rinsed with H₂O twice. Samples
were dried overnight and then stained with uranyl acetate. TEM micrographs
were acquired using JEOL JEM 2010F using a 2 K × 2 K Gatan ORIUS
camera. Samples have been observed in low magnification using the
image formed inside the transmitted beam on the focal plane. This
method is useful in low-magnification detection of weak objects, without
any change in the focus configuration of the microscope. By switching
between the out-of-focus image in the focal plane and the in-focus
image in the image plane, it is possible to scan a large area of the
grid with enhanced detection capability and easily come back to the
normal image configuration.

For EM analysis, wing imaginal discs and pupal wings were fixed in a mixture of 4% paraformaldehyde and 2% glutaraldehyde in 0.1 M sodium cacodylate buffer, pH 7.3 overnight at RT. Samples were post-fixed in 1% osmium tetroxide in 0.1 M PBS for 1 hr at RT, stained with uranyl acetate, dehydrated with a graded ethanol series and embedded in EMbed-812 (Electron Microscopy Sciences). Ultrathin sections were cut and analyzed with a JEOL JEM-1400 microscope using a Gatan Orius Camera.