Equipment and Services
Conventional microscopy, utilizing standard techniques in brightfield, darkfield, fluorescence, phase contrast and differential interference contrast, is available on a variety of instruments in the main Core Facility room 287. Multiple upright and inverted microscopes capable of low to high magnification documentation are available with individual image capture workstations networked to the Imaging server. Sensitive monochrome cameras and one-shot digital color cameras are available on each system. Real-time stitching software allows high resolution imaging of larger samples.
Live-Cell Time-Lapse Microscopy
Using the Nikon TE300 inverted microscope in room 287 with its ImagePro workstation, 5D time-lapse studies of cells grown in culture can be accomplished (X, Y, time, wavelength and location). Imaging combinations using phase contrast and fluorescence illumination are possible, taken at short or long intervals, from several minutes to several days. The environmental chamber has temperature and CO2 control and is monitored continuously. Cells are normally presented in multi-well plates or 35mm glass-bottom dishes, but other vessels can be accommodated as needed. Common uses for this system include scratch (wound) assays and cellular proliferation studies.
For higher resolution options, including 3D timelapse, The Leica SP5 II confocal microscope (see below) is also capable of multi-day timelapse, with CO2 and temperature control.
A Leica TCS SP5 II scanning laser confocal system with resonant scanner and environmental chamber is also available for individual and assisted imaging. This instrument allows investigators to carry out high-resolution, single cell observations, spectral separations, thick specimen analysis and co-localization studies in both fixed and dynamically interacting cell populations. It supports studies designed for both single timepoint examinations as well as multi-dimensional time-lapse observations. This equipment will allow long-term, environmentally controlled, minimmaly photo-toxic, reduced noise, high resolution microscopy, and the ability to carry out FRET, FRAP, confocal reflectance, 3D and 4D tracking, and temporal, spectral and spatial analysis.
This system has 9 laser lines and 3 spectral PMT detectors, as well as 2 HyD detectors for superior sensitivity and single photon capability. Sample preparation is generally done by the interested lab and analysis of the captured images can be done with the Leica analysis package or with 3rd party options available in the Core.
2-photon microscopy continues where traditional confocal leaves off. With laser excitations in the longer wavelengths, 2P allows deeper imaging and reduced phototoxicity. This system was designed to be used for intravital imaging, with tissues such as inguinal lymph nodes, but it can also used with cell cultures in acceptable vessels, tissue slides, explanted specimens such as lymph nodes and brain slices, kept viable in heated media. This system has the ability to resolve non-labeled structural details with 2nd harmonics (SHG), as well as capture emission wavelengths simultaneously with three other detectors. Thick, fixed specimens, such as spheroids and skin reconstructs can benefit from the increased penetration and imaging capabilities of this system, as well as fixed cultures in deep collagen matrices.
Low Magnification and Photomacrography
Specimens that are too large for a standard microscope require flexible equipment configurations for illumination and imaging conditions that vary depending on the specimen. We have the ability to image anything from whole mount slides to entire Petri dishes, as well as gross specimen photography of excised biological tissues using our stereomicroscopes or digital SLR-based macro equipment. Transmitted or reflected light, as well as fluorescence can be accommodated with our systems.
Small Animal Whole Body Luminescence and Fluorescence Imaging
The Perkin-Elmer IVIS 200 whole body imager is designed for detection of weak fluorescent or bioluminescent signals deep in living mice for non-invasive longitudinal studies of growing (or regressing) tumors in mouse models. The system is housed in the Institute’s barrier mouse facility and is only available to those researchers with approved protocols and animals housed in the colony. The integrated photon counting software can normalize the data for direct comparison to future studies or to similar instruments anywhere in the world.
Specialty Image Capture
The facility maintains equipment to provide “traditional” photographic support, such as photography of small animals (mice), gels and blots, experimental set-ups, copy stand specimens, catalogue items, or even human groups and portraits when needed. Special equipment for unusual situations such as GFP fluorescence in large specimens like adult mice is available as well. The Nikon D200 digital camera has a variety of interchangeable lenses, depending on the situation, and special portable studio lighting is also available as needed.
Image Analysis and Presentation Figures
Capturing images is only the first step in scientific documentation. The images also need to be analyzed to extract meaningful data and prepared for proper presentation in either hard copy or electronic publication. The facility maintains multiple workstations and a variety of software packages for
this purpose with internal support and training from the staff, or custom support available from the suppliers for special cases. Custom macros for automating analytical functions can be developed with our staff for your specific image sets. Help with creating publication figures and cover art is also available as needed.
Imaging Support Services
All successful experiments begin with an understanding of the capabilities of the existing equipment and software. The facility staff constantly work with researchers to help design protocols that can be effectively accomplished with the resources at hand. Choice of fluorophores, freezing and fixation techniques, thickness of specimens, and other issues can all negatively affect results if the instrumentation is not considered in advance. The facility staff also takes great care to make users aware of the dangers of unacceptable image manipulations that can alter the results of their experiments.
All facility equipment is available to be used unassisted or with assistance. Users must first be trained for each specific piece of equipment before they can be certified by the Facility for unassisted usage. Some equipment requires significant training to attain proficiency, so the staff can always work with a user directly to obtain the desired results. Training also includes image processing and analysis, as well as digital imaging ethics. Training is mandatory prior to unassisted use of all Facility equipment.
In-Lab Microscopy Assistance and Maintenance
In addition to providing core instrumentation and services, the facility staff is also available to help support microscopy instrumentation elsewhere in the Institute. Common maintenance issues such as cleaning and bulb replacement in individual labs are easily supported and response time is more immediate from the Imaging Facility than from outside vendors. Consolidation of outdated and discarded microscopes by the Facility also creates a repository of used equipment, which can easily be redistributed as needed throughout the Institute