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Related Experiment Videos

Side population analysis using a violet-excited cell-permeable DNA binding dye.

William G Telford1, Jolene Bradford, William Godfrey

  • 1Experimental Transplantation and Immunology Branch, National Cancer Institute, Building 10 Room 3-3297, 9000 Rockville Pike, Bethesda, Maryland 20892 USA. telfordw@mail.nih.gov

Stem Cells (Dayton, Ohio)
|December 23, 2006
PubMed
Summary

DyeCycle Violet (DCV) reagent offers a viable alternative to Hoechst 33342 for side population (SP) analysis. This method allows for stem cell identification using common violet lasers, overcoming limitations of UV-dependent Hoechst SP analysis.

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Area of Science:

  • Flow cytometry
  • Stem cell biology
  • Biochemistry

Background:

  • Hoechst 33342 side population (SP) analysis is crucial for identifying stem cells.
  • This method requires expensive UV lasers, limiting its accessibility.
  • Violet lasers are common but provide insufficient excitation for Hoechst 33342.

Purpose of the Study:

  • To identify an alternative fluorescent substrate for SP analysis compatible with violet lasers.
  • To evaluate DyeCycle Violet (DCV) reagent as a substitute for Hoechst 33342.
  • To enable SP analysis on more accessible flow cytometers.

Main Methods:

  • Comparative analysis of Hoechst 33342 and DyeCycle Violet (DCV) for SP identification.
  • Flow cytometry using both UV and violet lasers.

Related Experiment Videos

  • Inhibition assays with fumitremorgin C.
  • Simultaneous immunophenotyping with stem cell markers.

    • Main Results:

    • DCV reagent demonstrated similar emission characteristics to Hoechst 33342.
    • DCV SP analysis yielded comparable results with both violet and UV excitation.
    • DCV SP exhibited the same membrane pump specificity as Hoechst SP, inhibited by fumitremorgin C.
    • DCV SP identified the stem cell lineage(-) Sca-1(+) c-kit(+) population in mouse bone marrow.

    Conclusions:

    • DyeCycle Violet reagent is a promising alternative for Hoechst 33342 in SP analysis.
    • DCV enables side population analysis using common violet laser flow cytometers.
    • This substitution expands the accessibility of stem cell identification techniques.