Light Microscopes and Image Analysis Workstations

Light Microscope & image analysis workstations

Location: LSN 429 (see map)
Contact: Doug Cromey, 626-2824
Administration: Cellular & Molecular Medicine
Fees for use: Currently there are none.

Definiens image analysis workstation

Location: UACC 0930 (see map)
Contact:  Dr. Raghunand, 626-6041
Administration: University of Arizona Cancer Center
Fees for use:  No cost if you provide the operator, $30/hr if lab staff do the analysis

Reminder: SWEHSC Investigators (including their staff and students) should contact the Cellular Imaging Facility Core directly for advice regarding experimental design, training, and technical support for this instrument.

 

Light microscopic & image capture capabilities:

  • The Olympus IMT-2 inverted microscope is equipped with a digital camera that can capture 1280x1024, 10 bit (1024 intensity or grayscale values) images of brightfield, phase contrast, DIC and fluorescence (FITC cube) imaging modes. The microscope has motorized controllers for shutters and x-y-z stage control. The automated stage has holders for microscope slides, 35mm culture dishes and 96 well plates (note: DIC imaging uses polarization and cannot be performed through plastic). A BiopTechs Delta T3 system for keeping 35mm culture dishes at 37°C enables the system to be able to capture time-lapse images of cultured cells over periods of several hours. A copy of Microsoft Excel™ is available on the computer to assist in transferring the numerical data.
  • The Leica DMI6000B inverted microscope (July 2013) is equiped with a 16bit sCMOS Hamamatsu Flash 4.0 greyscale camera (4Mpix) capable of 30fps and a single chip Leica DFC450 color CCD camera (5Mpix).  The microscope is fully motorized and can be controlled almost completely from software.  The stage can hold microscope slides, small culture dishes (35mm, 70mm) and multiwell plates.  The microscope can capture images in brightfield (transmitted light), DIC (selected objectives), POL, fluorescence (cubes for dyes similar to DAPI, FITC/GFP, Rhodamine/TRITC, CY5).  The phase contrast light path does not currently go to a camera, but we could change that if necessary.

The Leica computer is a workstation class Windows 7/64 computer with quad-core Intel i7 CPUs, 64GB of RAM, solid state drive scratch disk space and a 2TB hard disk drive.

Image analysis & measurement capabilities:

The workstations use either SimplePCI software 6.x (C•Imaging Systems, by Hamamatsu, Inc.) or the newer HCImage 4.x (Hamamatsu Corporation, Sewickley, PA). The Leica workstation also has LAS-AF 3.x, which is primarily useful for image acquisition only.

Digital images captured from any appropriately calibrated source (e.g. confocal, TEM, SEM, and other microscopes or scanned images) can be analyzed by the software. The software has the capability to measure a large number of parameters related to size, shape, intensity and position. In many instances the software can configured to be acquire measurements in a semi-automated mode. Basic statistical measurements are included in the software. Numerical data can be exported as Microsoft Excel™ spreadsheets.

The software has been used to measure/capture:
• The distance traveled by cultured cells over time (time lapse)
• The length of dendrites (confocal images)
• Calculate pH based on using color ratios from SNARF dyes (Zeiss confocal images)
• Point counting in lung tissue (light microscope images)
• Image montages of whole cross-sections that were later used for 3D reconstruction or poster presentations (light microscope images)
• Create time-lapse movies for presentations (light microscope images)
• Count fluorescently stained nuclei to measure migration in a matrix (fluorescence microscope images)
• Measure sizes of intestinal villi in histologic samples (light microscope images)
• Measure the area of lipid droplets found in Oil Red O stained histologic samples (light microscope images)
• 3D rotations of multi-level/Z-stack images (light microscope images)
• Automatically count the number of cells that crossed the membrane in transwell plates (24 well plates).
• Analysis of changes in melanized skin moles to assess the success of cancer treatment (macro photography).

 

Definiens Tissue Studio - image analysis capabilities

The Arizona Cancer Center's Cancer Imaging Shared service offers access to Definiens Tissue Studio 3.0 for biomarker and morphological profiling in histological images. It is an easy to use, robust, and powerful tool to automatically detect regions of interest based on user-defined criteria. It can distinguish cells and sub-cellular objects within these target regions, and determine morphology and expression profiles per individual cell or cell compartment. It is compatible with brightfield or immunofluorescence images, whole tissue slides or tissue microarrays.

For more information about the software, or to receive training, please contact Dr. Raghunand

More information about Definiens Tissue Studio 3.0 (software vendor site).