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Georgolopoulos G, Iwata M, Psatha N, et al. Unbiased phenotypic identification of functionally distinct hematopoietic progenitors. J Biol Res (Thessalon). 2019;26:4doi: 10.1186/s40709-019-0097-7.
Georgolopoulos, G., Iwata, M., Psatha, N., Yiangou, M., & Vierstra, J. (2019). Unbiased phenotypic identification of functionally distinct hematopoietic progenitors. Journal of biological research (Thessalonike, Greece), 264. https://doi.org/10.1186/s40709-019-0097-7
Georgolopoulos, Grigorios, et al. "Unbiased phenotypic identification of functionally distinct hematopoietic progenitors." Journal of biological research (Thessalonike, Greece) vol. 26 (2019): 4. doi: https://doi.org/10.1186/s40709-019-0097-7
Georgolopoulos G, Iwata M, Psatha N, Yiangou M, Vierstra J. Unbiased phenotypic identification of functionally distinct hematopoietic progenitors. J Biol Res (Thessalon). 2019 Jul 18;26:4. doi: 10.1186/s40709-019-0097-7. eCollection 2019 Dec. PMID: 31360678; PMCID: PMC6639971.
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J Biol Res (Thessalon). 2019 Jul 18;26:4. doi: 10.1186/s40709-019-0097-7. eCollection 2019 Dec.

Unbiased phenotypic identification of functionally distinct hematopoietic progenitors.

Journal of biological research (Thessalonike, Greece)

Grigorios Georgolopoulos, Mineo Iwata, Nikoletta Psatha, Minas Yiangou, Jeff Vierstra

Affiliations

  1. 1Altius Institute for Biomedical Sciences, Seattle, WA 98121 USA.
  2. 2Department of Genetics, Development and Molecular Biology, School of Biology, Aristotle University of Thessaloniki, 541 24 Thessaloniki, Greece.

PMID: 31360678 PMCID: PMC6639971 DOI: 10.1186/s40709-019-0097-7

Abstract

BACKGROUND: Hematopoiesis is a model-system for studying cellular development and differentiation. Phenotypic and functional characterization of hematopoietic progenitors has significantly aided our understanding of the mechanisms that govern fate choice, lineage specification and maturity. Methods for progenitor isolation have historically relied on complex flow-cytometric strategies based on nested, arbitrary gates within defined panels of immunophenotypic markers. The resulted populations are then functionally assessed, although functional homogeneity or absolute linkage between function and phenotype is not always achieved, thus distorting our view on progenitor biology.

METHOD: In this study, we present a protocol for unbiased phenotypic identification and functional characterization which combines index sorting and clonogenic assessment of individual progenitor cells. Single-cells are plated into custom media allowing multiple hematopoietic fates to emerge and are allowed to give rise to unilineage colonies or mixed. After colony identification, lineage potential is assigned to each progenitor and finally the indexed phenotype of the initial cell is recalled and a phenotype is assigned to each functional output.

CONCLUSIONS: Our approach overcomes the limitations of the current protocols expanding beyond the established cell-surface marker panels and abolishing the need for nested gating. Using this method we were able to resolve the relationships of myeloid progenitors according to the revised model of hematopoiesis, as well as identify a novel marker for erythroid progenitors. Finally, this protocol can be applied to the characterization of any progenitor cell with measurable function.

Keywords: Functional characterization; Hematopoietic progenitors; Immunophenotypic identification; Index sorting; Single-cell

Conflict of interest statement

Competing interestsThe authors declare that they have no competing interests.

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