The expression of IL-7Rα in the common lymphoid progenitor (CLP) population marks initiation and/or commitment to the lymphoid lineage. CLPs hold potential for only B, T and NK cell lymphoid lineages and are defined as Lin−IL-7Rα+c-KitloSca-1lo. Study of lymphocyte development largely relies on access to CLPs or other IL-7Rα+ lymphoid progenitors. Fluorescence-activated cell sorting (FACS) commonly used to isolate lymphoid progenitors is costly, time-consuming and possibly detrimental to cell viability. We describe a fast and efficient method for the isolation of lymphoid progenitors from mouse bone marrow (BM). This method is based on immunomagnetic, column-free cell separation technology (EasySepTM). Using this method, lineage positive cells are first depleted by cross-linking them to magnetic particles using biotinylated antibodies. Next, IL-7Rα+ cells are positively selected from the Lin−/lo population. After enrichment, average purity of Lin−IL-7Rα+ lymphoid progenitors as assessed by flow cytometry is 32 ± 10% (n=18). Lin−IL-7Rα+c-KitloSca-1lo cells are enriched 60-fold from 0.08 ± 0.05% in start BM to 4.6 ± 2.5%. Limiting dilution analysis of enriched cells shows increased frequencies of B cell (1:73, n=12), T cell (1:16, n=5) and NK cell (1:65, n=6) progenitors as compared to non-depleted control BM. This system introduces an easy method for the enrichment of lymphoid progenitors with good viability that will enable study of developmental immunology.
- Copyright © 2011 by The American Association of Immunologists, Inc.