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Measurement of CD34+ cells in bone marrow by flow cytometry
T M Trischmann1, K G Schepers, C I Civin
1Johns Hopkins Oncology Center, Baltimore, MD 21287.
A procedure is described for the measurement of the %CD34+ progenitor cells in bone marrow using directly conjugated antibodies. Staining cells with anti-CD45.FITC in conjunction with anti-CD34.PE allows the CD45- nucleated red blood cells and the CD45++ lymphocytes and monocytes to be separated from the CD45+ progenitor cells. Granulocytes are separated from the CD34+ cells based on differences in side scatter properties. A gated acquisition of CD34+ cells is used to define the boundaries of the CD34+ population in a plot of forward scatter vs side scatter and in a plot of anti-CD45.FITC vs anti-CD34.PE. Use of these regions during analysis reduces background staining and allows for a consistent identification of a CD34+ population. Acquisition of 50,000 cells provides adequate precision of the %CD34+ measurement. Acquisition and analysis procedures are presented for use of both a Becton Dickinson FACScan flow cytometer and a Coulter EPICS Profile II flow cytometer.
A procedure is described for the measurement of the %CD34+ progenitor cells in bone marrow using directly conjugated antibodies. Staining cells with anti-CD45.FITC in conjunction with anti-CD34.PE allows the CD45- nucleated red blood cells and the CD45++ lymphocytes and monocytes to be separated from the CD45+ progenitor cells. Granulocytes are separated from the CD34+ cells based on differences in side scatter properties. A gated acquisition of CD34+ cells is used to define the boundaries of the CD34+ population in a plot of forward scatter vs side scatter and in a plot of anti-CD45.FITC vs anti-CD34.PE. Use of these regions during analysis reduces background staining and allows for a consistent identification of a CD34+ population. Acquisition of 50,000 cells provides adequate precision of the %CD34+ measurement. Acquisition and analysis procedures are presented for use of both a Becton Dickinson FACScan flow cytometer and a Coulter EPICS Profile II flow cytometer.