Identification and Isolation of Burst-Forming Unit and Colony-Forming Unit Erythroid Progenitors from Mouse Tissue by Flow Cytometry

• DOI: 10.3791/64373-v
• Publication Date: 2022-11-05T00:00:38Z
• Authors: Aishwarya Swamina...; Yung Hwang; Ashley Winward; Merav Socolovsky
• Keywords: Colony-forming unit; Progenitor

Early erythroid progenitors were originally defined by their colony-forming potential in vitro and classified into burst-forming "units" known as BFU-e CFU-e. Until recently, methods for the direct prospective complete isolation of pure CFU-e from freshly isolated adult mouse bone marrow not available. To address this gap, a single-cell RNA-seq (scRNAseq) dataset was analyzed expression genes coding cell surface markers. This analysis combined with fate assays, allowing development novel flow cytometric approach that identifies allows subsets or spleen. also other progenitor subsets, including enriched basophil/mast megakaryocytic potentials. The method consists labeling fresh spleen cells antibodies directed at Kit CD55. Progenitors express both these markers are then subdivided five principal populations. Population 1 (P1 CFU-e, Kit+ CD55+ CD49fmed/low CD105med/high CD71med/high) contains all may be further P1-low (CD71med CD150high) P1-hi (CD71high CD150low), corresponding to early late respectively; 2 (P2 BFU-e, CD71low progenitors; P3 (P3, CD49fmed/high CD105med/low CD150low CD41low) is 4 (P4, CD150high CD41+) 5 (P5, CD41-) erythroid, cell, (EBMP) erythroid/ megakaryocytic/ basophil-biased multipotential (MPPs). greater precision when analyzing hematopoietic reference transcriptome information each cytometrically population.