Microscopy and Imaging Shared Resource

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Cytogenetics Services

Equipment (List by Imaging Method)

Equipment (List by Microscope)

Multiple fluorescence, confocal and automated microscopes allow high resolution microscopy required for visualizing molecules within intracellular compartments and tissues. They also enable digital documentation of histochemical, immunofluorescence, and in-situ hybridization experiments. Live automated imaging of cells or tissues is possible on confocal and widefield microscopes equipped with 37°C/CO2 chambers.

A microinjection system on an inverted microscope allows direct cellular injections of peptides, proteins, and plasmids. The behavior of microinjected cells can then be recorded using brightfield or fluorescence microscopies.

The MISR supports image acquisition and analysis with Metamorph Image Analysis and Volocity Visualization software for 2D and 3D image stacks with acquisition and stand alone analysis workstations. Writable DVD/CD-ROM and external eSATA hard drives are available for image storage and transfer. Automated collection of digital images such as image montaging are possible with a motorized stage. Image analysis macros facilitate automated or semi-automated quantification of cell or tissue features.

Details of services and available equipment is listed below. A price list is available in Research Building room W407 or on the Intranet site.

Conventional Light Microscopy

    • Nikon SMZ-1500 EPI-Fluorescence Digital Stereoscope System
      For stereoscopic visualization of tissues expressing GFP or RFP. Filter for Quantum Dots available. Otimized for monitoring of invasion/metastasis of tumor cells in tissues. Motorized Z-stage acquisition allows for 3D image collection.

    • Olympus IX71 Inverted Microscope System with Eppendorf Femtojet Microinjection
      For routine brightfield/Nomaski imaging and injection of samples. The system is equipped with two precision three axis micromanipulators and so is suitable for injecting adherent cells and cell in suspension. Samples can be image with 10X and 20X brightfield/Nomarski objectives.

    • Molecular Devices ImageXpressMICRO
      This system is used for automated screening of samples using fluorophores with blue, green, red or far red wavelength emissions using Metamorph software for image analysis. Screening programs for quantification of the following morphological features are available: Translocation, Count Nuclei, Granularity, Transfluor, Live/Dead Application, Cell Health, Mitotic Index, Multiwavelength Cell Scoring, Monopole Detection, and Cell Cycle Application Modules. In addition, custom analysis can be set up using the Metamorph software. Samples can be imaged using 4X, 10X, 20X and 40X lenses. This system does not have environmental control, or brightfield imaging properties.

      Available by collaboration with Dr. Louis Weiner: contact Dr. Sandra Jablonski (saj24@georgetown.edu).

Total Internal Reflection Microscopy

  • Olympus IX81 Total Internal Reflection Microscope (TIRF)
    For imaging plasma membrane in contact with the coverglass chamber slide. Five laser line imaging capacity, with DualView for simultaneous imaging in two colors (GFP, RFP, CFP, YFP, far red). TIRF and interference reflection microscopy (IRM) can be carried out sequentially. IRM permits visualization of cell contacts in living cells. The TIRF microscope (TIRFM) utilizes an optical phenomenon called Total Internal Reflection (TIR), which occurs when light travels from a higher to a lower refractive index medium at an angle greater than the critical angle. Beyond this angle of total reflection, the electromagnetic field of the incoming/ reflected light extends into the Z direction and is termed the evanescent field. One property of an evanescent field is that the magnetic strength of the field decreases exponentionally with distance, and so its effects extend only a few hundred nanometers into the second medium. The portion of the specimen within the evanescent field, usually the cell membrane, can be excited to emit fluorescence which consequently can be seen or recorded. TIRFM allows enhanced imaging of cell-surface proteins by substantially reducing background fluorescence from inside the cell.

Confocal Microscopy

  • Nikon Eclipse TE-300 Spinning Disk Timelapse Microscope System
    For live cell imaging of mutant GFP and RFP chimeras as well as fluorophore conjugated antibodies for recognizing extracellular epitopes. DIC optics and 3D reconstruction and 3D analysis using Volocity Visualization and Quantification are available.

    Multi-well chambered, time lapse imaging using bright field or fluorescence microscopy is superb for following cell migration or cell division within a tissue culture environment.

  • Zeiss 510LSM/ META/ NLO live imaging, multiphoton microscope
    With configuration #23 / META NLO + VIS, Argon, Dual HeNe, META and 2 PMTs, stage top environmental chamber for CO2 and O2 control, TiSapphire laser and FLIM hardware, this microscope is capable of FRET (sensitized emission and fluorescence lifetime emission (FLIM)), multispectral imaging, FRAP, time lapse, and 3D imaging for studies of subcellular localization and dynamic protein interactions.

    Time Correlated Single Photon Counting (TCSPC) for measurement of fluorescence lifetime intensity (FLIM) of fluorophores, e.g. donor molecules of FRET pairs in an experiment measuring protein-protein interactions.

Electron Microscopy

Limited technical assistance or training (by appointment, pj42@georgetown.edu) includes using the TEM microscope as well as sample preparation for conventional negatively stained specimens, thin-sectioned embedded samples and immuno-EM.
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Image Analysis & Deconvolution

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Other Equipment and Technical Support

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