期刊
JOVE-JOURNAL OF VISUALIZED EXPERIMENTS
卷 -, 期 137, 页码 -出版社
JOURNAL OF VISUALIZED EXPERIMENTS
DOI: 10.3791/58061
关键词
Immunology and Infection; Issue 137; Myeloid progenitors; common myeloid progenitor (CMP); granulocyte-monocyte progenitor (GMP); monocyte-DC progenitor (MDP); granulocyte progenitor (GP); monocyte progenitor (MP or cMoP); bone marrow; magnetic-activated cell sorting (MACS); fluorescence-activated cell sorting (FACS)
资金
- Board of Governors Regenerative Medicine Institute at Cedars-Sinai Medical Center
- Careers in Immunology fellowship from the American Association of Immunologists
- American Society of Hematology
Myeloid progenitors that yield neutrophils, monocytes and dendritic cells (DCs) can be identified in and isolated from the bone marrow of mice for hematological and immunological analyses. For example, studies of the cellular and molecular properties of myeloid progenitor populations can reveal mechanisms underlying leukemic transformation, or demonstrate how the immune system responds to pathogen exposure. Previously described flow cytometry strategies for myeloid progenitor identification have enabled significant advances in many fields, but the fractions they identify are very heterogeneous. The most commonly used gating strategies define bone marrow fractions that are enriched for the desired populations, but also contain large numbers of contaminating progenitors. Our recent studies have resolved much of this heterogeneity, and the protocol we present here permits the isolation of 6 subpopulations of oligopotent and lineage-committed myeloid progenitors from 2 previously described bone marrow fractions. The protocol describes 3 stages: 1) isolation of bone marrow cells, 2) enrichment for hematopoietic progenitors by magnetic-activated cell sorting (lineage depletion by MACS), and 3) identification of myeloid progenitor subsets by flow cytometry (including fluorescence-activated cell sorting, FACS, if desired). This approach permits progenitor quantification and isolation for a variety of in vitro and in vivo applications, and has already yielded novel insight into pathways and mechanisms of neutrophil, monocyte, and DC differentiation.
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