Transmission Electron Microscopy (TEM)

Electron microscopy represents a reliable method to monitor autophagy stages and it is still indispensable for the characterization of autophagic organelles. To evaluate morphological aspects, pellets were washed and immediately fixed “in situ” with 2.5% glutaraldehyde in 0.1M phosphate buffer for 45 min, post-fixed for 1h in 1% OsO₄, alcohol dehydrated, and embedded in araldite. Thin sectioning was preceded by the analysis of toluidine blue stained semithin sections by light microscopy. Thin sections were stained with uranyl acetate and lead citrate and analyzed with a Philips CM10 transmission electron microscope [84].