Measurement of CD34+Cells in Bone Marrow by Flow Cytometry

Abstract

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.