Selective staining of immature hemopoietic cells with merocyanine 540 in flow cytometry

Belloc, Françis; Lacombe, Francis; Bernard, Philippe; Dachary, Dominique; Boisseau, M. R.

doi:10.1002/cyto.990090104

Cited by 8 publications

(2 citation statements)

References 9 publications

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“…8:189-198. composition, it stands to reason that this probe may be useful in examining membrane structure and differentiation-associated changes in the phospholipid bilayer of primitive HPC. Previous studies have suggested that MC540 can be used to isolate populations of quiescent and metabolically active cells (16). In our hands, when normal and CML BM CD34 1 cells were fractionated into groups ranging in MC540 fluorescence from dim to bright, the MC540 D im fraction from each cell type contained the highest percentage of cells within the G 0 /G 1 phases of cell cycle, whereas the MC540 B right fraction contained the highest percentage of mitotically active cells.…”

Section: A B C Dmentioning

confidence: 64%

“…Both mature erythrocytes (14,15) and lymphocyte s (9) from peripheral blood have been described as not staining or impermeable to the membrane probe. Mononuclear bone marrow (BM) cells, which exhibit a high degree of MC540 uptake, are enriched for colony forming unit±granulocyte macrophage (CFU-GM) and cells in the S/G 2 1 M phases of the cell cycle (16), which suggests that quiescent, primitive HPC would display a significantly lower uptake of MC540. Using the photoexcitation property of MC540, relative sensitivities have been established for murine (13) and human (17) HPC.…”

Section: C 540 Uptake Could Be Used To Distinguish Between Quiescenmentioning

confidence: 99%

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Use of Merocyanine 540 for the Isolation of Quiescent, Primitive Human Bone Marrow Hematopoietic Progenitor Cells

Pyatt¹,

Jenski²,

Allen³

et al. 1999

Journal of Hematotherapy

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Merocyanine 540 (MC540) is a membrane probe that inserts preferentially into loosely packed domains in the phospholipid bilayer of intact cells. Previous experiments have demonstrated that MC540 will bind to human bone marrow (BM) hematopoietic progenitor cells (HPC). Fractions of mononuclear BM cells expressing high MC540 fluorescence have been shown to be enriched for myeloid progenitors and cells residing in the S/G2 + M phases of the cell cycle. We rationalized that MC540 uptake could be used to distinguish between quiescent and metabolically active cells and, therefore, to fractionate normal and leukemic BM cells and normal mobilized peripheral blood (MPB) cells into functionally distinct groups of progenitors. BM and MPB cells were separated into fractions ranging in fluorescence from MC540Bright to MC540Dim. Cell cycle analysis of these fractions revealed that the MC540Dim fraction of normal and CML BM CD34+ cells constituted the most quiescent fraction, and the MC540Bright fractions from these cell types contained the most actively cycling cells. However, no differences in the percentage of cells in G/G1 were observed between MC540Bright and MC540Dim fractions of MPB CD34+ cells. To investigate if these cell cycle status differences translated into distinct functional properties, the hematopoietic potential of BM CD34+MC540Bright and CD34+MC540Dim cell fractions was analyzed in vitro in long-term BM cultures and limiting dilution analysis (LDA) assays. CD34+MC540Dim cells produced more total and committed progenitor cells in long-term cultures than did the CD34+MC540Bright fraction. The CD34+MC540Dim fraction also contained a 2-fold higher number of long-term hematopoietic culture-initiating cells (LTHCIC) than the CD34+MC540Bright fraction, as defined by LDA assays. These data demonstrate that MC540 can be a useful probe for the isolation of primitive HPC from some hematopoietic tissues and may assist in monitoring structural changes in the phospholipid bilayer during proliferation and differentiation of HPC.

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Section: A B C Dmentioning

confidence: 64%