Microscopy and Image Analysis Laboratory

The Microscopy and Image Analysis Laboratory has been developed to probe into cellular and subcellular structures using a wide variety of microscopy tools. The imaging capabilities include real-time microscopy, electron microscopy, and epi-fluorescence and polarization microscopy, among others. Automated data and image acquisition, analysis, and storage systems are integrated with various microscopes.

This facility has been successfully applied by CEM researchers to investigate aspects of unique biomedical problems. Examples of such projects are as follows:

• Use of the scanning stage and rendering software to acquire and process images of cellular chemotaxis within a microfabricated micro-flow chamber allowing the creation of an accurate 3D profile under various flow conditions
• Employment of computer-controlled shuttering and image acquisiton of fluorescent membrane integrity probes to quantify the kinetics of activity of an engineered bacteriological pore-forming agent
• Use of a computer-controlled filter wheel allowing measurement of proton concentrations through ratiometric imaging of endosomes in fibroblasts infected with murine leukemia virus to determine the relationship of infection efficiency with endosomal pH
• Viability assessment of fibroblasts exposed to therapeutic agents resulting from the activation of phototoxin-conjugated antibodies using stains specific for live/dead cells
• Morphological evaluation of cryopreservation techniques by analyzing the distribution and co-localization of filamentous actin, tubulin, and chromatin material in mouse oocytes through the use of unique fixation, staining, and imaging techniques

The broad range of microscopy systems available at the CEM includes:

• Transmission electron microscope (Philips 410) kV range [ 40, 60, 80, 100] available for detailed studies of cells and tissues at the subcellular level
• Nikon Eclipse 400 upright, compound instrument with dual-head accessory; brightfield optics only; objectives: 4, 10, 20, 40 60 X, dry; for histology/ cryo section examination
• Nikon Eclipse 800 upright compound microscope; brightfield, phase contrast, differential interference contrast (DIC), fluorescence optics; objectives: 10, 20 40 X (dry); 63 and 100 X (oil); 20x water dipping objective (intra-vital use); Spot 2 monochrome/color digital camera with computer/software; Metamorph (v. 6.0/6.1) computer, software for image processing; and image analysis; for histology, immunochemistry, intra-vital applications
• Zeiss Axiovert 200M inverted, compound microscope; brightfield, phase contrast and fluorescence optics; infinity-corrected objectives: 2.5, 5, 10, 20 40 X (dry); 63X (oil); 1X and 1.6X optivar; motorized stage (z,y,z); environmental chamber/ heating stage capability; external filter wheel (ratio imaging); axiocam (b&w) digital camera; AxioVision 3.1 and 4.0 software availability; many cell and tissue culture applications using software modules (including time lapse, z-stacks, co-localization)
• Zeiss Axiovert 10 inverted compound microscope: brightfield, phase contrast, fluorescence optics; infinity-corrected objectives: 5, 10, 20 40 X (dry), 63, 100X (oil); environmental chamber capability for time lapse; video and digital camera documentation; Zeiss Axiovision computer/software modules; appropriate for cell and tissue culture applications
• Nikon Diaphot 300 inverted, compound microscope; brightfield, phase contrast, DIC and fluorescence capabilities; finite objectives: 4,10, 20, 40 X (dry); 63, 100X (oil); Narishige micromanipulators, microinjection accessories. Microscope dedicated to microinjection studies.
• Nikon Diaphot tmd, inverted, compound microscope; brightfield, phase contrast, DIC, and fluorescence optics; objectives: 4, 10, 20, 40 X (dry); 63 and 100 X oil; DP-10 color digital camera smart card and 35mm camera documentation capabilities
• Nikon TMS inverted, compound unit; brightfield and phase-contrast optics; finite objectives: 10, 20 40 X (dry); Hamamatsu video camera/control box, video monitor and paper printer accessories (for basic cell and tissue culture use)
• Olympus IMT-2 inverted, compound microscope, brightfield, phase contrast optics; finite objectives: 10, 20 40 X, (dry); time-lapse and video documentation accessories; cell and tissue culture; various special applications

Technical Support

The technical staff that operates and maintains this facility is skilled in multiple microscopy techniques including brightfield, DIC, phase contrast, darkfield, various fluorescence techniques, timelapse, intravital microscopy, and 3D rendering and analysis. They are also experienced with many cellular, morphological, and biochemical assays valuable in the design of effective experimental protocols utilizing these systems..