Flow Cytometry and Optical Morphology
The Flow Cytometry/Optical Morphology Shared Resource of the Mayo Clinic Cancer Center combines two complimentary technologies to provide optimal high-speed population based analysis and morphological information for Cancer Center members. The goal of this shared resource is to provide high-end instrumentation, expertise, and training related to these two areas.
- Flow Cytometry
- Flow cytometry utilizes a combination of fluidic, optical, and electronic components to analyze individual cell characteristics and collect the data generated for further analysis. In flow cytometry, cells in liquid suspension that have been labeled with a fluorescent marker, often conjugated to an antibody, are made to flow single file at high rates of speed (up to thousands of cells per second) through focused laser beams. Forward (low angle) scattered light for relative size, side (ninety-degree angle) scattered light for relative granularity, and fluorescence signals are collected by photodetectors. Electrical impulses, proportional to the amount of incident light striking the detector are converted to digital signals and displayed through specialized software or stored for future analysis. Optical emission filters in front of the detectors allow for the detection of specifically selected wavelengths of fluorescence. In this way, multiple dye combinations can be used in a single sample.
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James E. Tarara
Manager
- Optical Morphology
- The term 'optical morphology' is used to describe a number of techniques related to image production and image analysis. Typically, samples are prepared from tissues or cell cultures, mounted on glass slides with coverslips or in various culture dishes. Viewing is done on light microscopes capable of a variety of image production techniques including brightfield, darkfield, phase contrast, differential interference contrast, and fluorescence. Advanced image analysis programs are included in the facility. Confocal laser scanning microscopes are provided for high-resolution fluorescence imaging and optical sectioning for three dimensional image reconstruction. Other specialized instruments are used for time dependent image capture, total internal reflection fluorescence, and microinjection.
Facilities, Equipment & Staffing
Primary location: Guggenheim Building, Rochester, Minn.
Telephone: 507-284-4241
Director: Richard G. Vile, Ph.D.
Manager: James E. Tarara
Flow Cytometry
- FACScan (single laser) flow cytometer (BD Biosciences)
- Two FACSCalibur (dual laser) flow cytometers with sample loaders
- FACSCanto (dual laser) flow cytometer with sample loader
- LSR-2 (five laser) flow cytometer with high throughput sampler (HTS) for sampling from multi-well plates
- FACSVantage SE (dual laser) cell sorter
- FACSAria (five laser) cell sorter
- Data analysis software
In addition to bulk sorting both cell sorters are equipped with the capability to sort single cells into multi-well plates.
Optical Morphology
- Three Zeiss LSM510 confocal microscopes, one equipped with a Meta detector for spectral fingerprinting
- LSM 5 Live confocal microscope for faster acquisition
- Axioplan 2 upright microscope, with high-resolution digital color camera and acquisition software
- Axiovert 200M inverted microscope equipped with an Apotome module for optical sectioning
- Axiovert 200M inverted microscope equipped for total internal reflection fluorescence (TIRF) microscopy
- Axiovert S100 inverted microscope equipped for microinjection
- Macroscope
- KS400 image analysis software
- Axiovision image analysis software
In addition to instrumentation, the Flow Cytometry/Optical Morphology Resource provides training on all instrumentation for those interested. Further support includes technical assistance with data analysis and interpretation, protocol development and implementation, and the development of customized programs for image analysis.
