General information
The Imaging Cores - Electron location is Room 410 - Life Sciences North (LSN) building on the University of Arizona campus.
Please send an email to toninop@arizona.edu to request general information. For a TEM consultation on your research project you need to make a request in iLab and a planning session will be coordinated. Once the in person appointment or Zoom meeting is scheduled please be sure to provide background literature with the type of data you intend to collect.
To request services in the Imaging Cores - Electron facility you need to be registered in iLab system. This will allow you to make requests, schedule equipment, get quotes, billing and project management, accordingly.
Please click on this link https://ua.ilab.agilent.com/landing/3645 and follow the instructions provided here.
Samples submission
Samples must be fixed and transferred to the buffer (0.1 M PBS or PIPES, pH 7.2 - 7.4) prior submission to the core facility.
Fixation will depend on the specific biological material you are working with (e.g. cells, tissue, microorganisms, etc.). To perform the primary fixation please follow the guidelines for preparation of suspensions (e.g. cells, bacteria, virus) or tissues, accordingly.
Fixed samples are received in the core from Monday to Wednesday (9 am to 12 pm).
Samples Processing and Imaging
The workflow of samples processing for TEM involves several steps: primary fixation (performed in your lab), post-fixation, dehydration, infiltration and embedding in resin (performed in the core). Then ultramicrotomy provides thick sections (0.3 - 1 µm) stained with Toluidine blue suitable for light microscopy, and ultrathin (70 - 90 nm) sections stained with uranyl acetate or KMnO4 and lead citrate, ready for TEM imaging in the FEI Tecnai G2 Spirit BT transmission electron microscope.
In general, to complete the experiments workflow it depends on the type, number of samples and the procedure requested (TEM, IEM, negative staining, SEM). Usually from specimen preparation to imaging, TEM takes about 2.5 weeks, IEM 3.5 weeks and negative staining 1 week.
Training on an individual basis is provided on the transmission electron microscope or the ultramicrotomes upon request, and this usually requires 3 1-hour-sessions on each equipment.
USB devices are not allowed to save images. Data will be saved on the Tecnai TEM support computer in a specific folder under the PI name/user name/acquisition date and after finishing the session this will be transferred to Box (internal users) or Dropbox (external users).
For SEM processing, the sample is fixed (performed in your lab), and followed by post-fixation, dehydration and chemically dry using HMDS (performed in the core). Further imaging is undertaken prior schedule in the W.M. Keck Center for Nano-Scale Imaging, which house a SEM instrument.
Image Analysis
There are several free open source image analysis software available for your data, such as Fiji/ImageJ (commonly used), Cell Profiler, Ilastick, Orbit, Icy and Cytomine.
Yes, quantitative analysis and/or interpretation of results are performed upon request through iLab. For quantitation is required a previous discussion of the parameters to be analyzed on your samples.