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Cell Sorter Core (BCH)

Director: Mathieu, Ronald, Ph.D.

Location: Boston Children’s Hospital, IDDRC Stem Cell Core Facility, Center for Life Science Building, Room 15037, 3 Blackfan Circle, Boston, MA 02115

Summary:

The Stem Cell Core Facility has been part of the IDDRC center for more than 30 years fostering collaboration and innovation among IDDRC investigators and members of the academic research community. It was used to develop chromosome sorting for human chromosome specific recombinant libraries and these techniques were utilized by others to generate large scale libraries for the early genome project. The facility was the first to develop cell sorting of fetal cells in the maternal circulation as a means to detect fetal genetic abnormalities.

Throughout its history the Stem Cell Core has provided careful and accurate cell cycle analysis via DNA content analysis using Hoechst dye uptake. This service is still used today by investigators. The facility was among the first to take advantage of the ability to express GFP in cells transfected with vectors and induced to express GFP in transgenic animals conditionally expressing a specific gene promoter. This ability allowed IDDRC investigators to isolate neurons, muscle cells and other cell types which express GFP from those which do not. IDDRC investigators were among the first to subsequently culture sorted neurons. Based on the extensive experience of the core using Hoechst dye for cell cycle analysis, the core was able to help IDDRC investigators prepare side population cells (tissue derived potential multipotent progenitor cells) based on Hoechst dye efflux. These methods have allowed the isolation of different muscle side population cells and have facilitated experiments that aim to use the cells for treating muscle disease. The interactive and collaborative nature of the core has fostered these developments, and the interests of the core director, advisory committee, manager and IDDRC investigators will continue to make this core as innovative in the future as it has been in the past.

The overall goal of the Stem Cell Core Facility is to provide both IDDRC and non-IDDRC researchers comprehensive analytical flow cytometry and cell sorting services in a timely, dependable and cost-effective manner.

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Instruments

  • Becton Dickinson FACSAria cell sorter ( Flow cytometer sorter )

    The Becton Dickinson FACSAria is a "high speed cell sorter equipped with three lasers: one 488nm blue laser, one 633nm red laser and one 405nm violet laser. This sorter has eleven optical detectors allowing for nine independent fluorescence and two light scatter parameters, four-way sort capabilities, as well as single cell sorting into multiwell plates and slides, a sample and collection tube temperature control option and an aerosol management system.

    Lasers, available detectors and possible fluorochromes:

    Blue 488 nm laser
    530/30: FITC, GFP, Alexa 488
    576/26: PE, RFP
    610/20: PI, PE-Texas Red
    695/40: PerCP, PerCP-Cy5.5
    780/60: PE-Cy7

    Red 633 nm laser
    660/20: APC
    780/60: APC-Cy7

    Violet 407 nm laser
    450/40: DAPI
    530/30: Alexa 430"

  • Becton Dickinson FACSVantage SE cell sorter ( Flow cytometer sorter )

    The Becton Dickinson FACSVantage SE is a "high speed cell sorter equipped with three lasers:
    Two argon-ion lasers, one tuned to 488nm blue and one tuned to 350nm for ultraviolet excitation. The third laser is a helium-neon mixed gas spectrum laser, which provides 635nm red excitation. This instrument has eight optical detectors, allowing for six independent fluorescence and two light scatter parameters to be detected. Two-way sorting is currently available with the potential for up to 96 well plate sorts.

    Lasers, available detectors and possible fluorochromes:

    Blue 488 nm laser
    530/30: FITC, GFP, Alexa 488
    575/25: PE, RFP
    630/20: PI

    Red 635 nm laser
    660/20: APC

    UV 350 nm laser
    400LP: Hoechst Blue
    600LP: Hoechst Red"

Services

  • Cell cycle analysis ( Material analysis service )

    Includes "DNA content analysis using Hoechst dye uptake."

  • Cell sorting service ( Material analysis service )

    Includes "fractionation of cells on the basis of cell surface antigens or intracellular markers, such as GFP or DNA content."

  • Flow cytometry analysis service ( Material analysis service )

    Includes "cellular DNA content and/or quantitative immunofluorescence, intracellular fluorescence."

  • Side population cell sorting ( Material analysis service )

    Includes "separation based on cell surface antigens enriched on side population cells (tissue derived potential multipotent progenitor cells) or Hoechst dye exclusion."

  • Zebrafish cell sorting and analysis ( Material analysis service )

    Includes "separation based on in vivo GFP and RFP expression or cell surface antigens."


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Last updated: 2017-01-16T08:12:24.480-06:00

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