Prior to selecting a service within the Electron Microscopy Shared Resource Laboratory, an initial consultation with Karen Bentley, who has 27 years of teaching and diagnostic experience in Ultrastructural Pathology and Immunoelectron Microscopy, is required. This consultation defines the best and most cost effective experimental strategy and methods to achieve a researcher’s goals.
To set up a consultation, please contact Karen Bentley via email or by phone at (585) 275-1954.
In general, for TEM imaging, fixed tissue or cells are generally processed as follows: post-fixed in osmium tetroxide, processed through a graded series of alcohols, infiltrated into liquid epoxy resin (Epon/Araldite or Spurr), embedded into molds, and polymerized at 70°C.
The epoxy blocks are cut by a technician using an ultramicrotome to cut semi-thin (1.0-2.0 um) sections with a glass knife, which are stained with Toluidine Blue. Examination of sections by light microscopy are necessary to determine the appropriate area to be trimmed and thin (70nm) sectioned for electron microscopy. A diamond knife with attached water filled boat is substituted for the glass one and thin sections are cut and placed onto grids for subsequent staining with heavy metals such as uranyl acetate and lead citrate.
Researchers/graduate students are present when the grids are examined under the TEM with the presence of Karen Bentley who helps guide and interpret the ultrastructural morphology or immunolabeling.
In 2007, Ms. Bentley acquired funds from the Department of Defense/Air Force to purchase a new Hitachi 7650 Analytical TEM with an Erlangshen 11 megapixel digital camera and Gatan software for imaging and morphometric analysis. Additional features of this microscope include the ability to perform “darkfield” STEM and Electron Dispersive X-ray Spectroscopy (EDS) for elemental analysis. Both of these features will provide “high tech” support to established and future funded research projects involving nanoparticles and nanotechnology.
Scanning electron microscopy (SEM) tissue and cell processing and imaging service is also available. The EM Core technical staff utilizes the River Campus/Optics Department’s Zeiss Supra 40VP Field Emission SEM for these types of requests.