The Establishment of A Novel Effective Method for the Enrichment of Murine Bone Marrow Erythroblastic Islands
Erythroblastic island(EBI)is the distinct microenvironment for mammalian erythroid terminal differentiation.Investi-gating EBI is essential for gaining a deep understanding of the cellular composition and phenotypic changes of hematopoietic mi-croenvironment under both physiologic and pathologic conditions.The distribution of EBI in bone marrow is relatively scattered.Nowadays,existing enrichment methods exhibit unsatisfactory efficiency.Our research aimed to establish a novel and more effi-cient EBI enrichment method.Drawing on previous research,we introduced innovations and optimizations,developing a novel method centered on dual density gradient sedimentation combined with washing steps to remove non-EBI cell clusters.We com-pared the efficiency of our method with other existing methods using observing cell cluster structure and Wright-Giemsa staining and imaging flow cytometry(IFC).Results showed that few single cells were found in the enriched product of Method 4 through direct microscopy and Wright-Giemsa staining,most of the products were typical EBI structures including a central macrophage and surrounding red blood cells.Using IFC technology to analysis F4/80,CD71 and CD11b in enriching products,about 63.7%of the cell clusters enriched by our method were EBIs.This percentage was higher than the highest proportion obtained in previ-ously reported Method 3,about 38.4%.As previously reported,EBI macrophages were CD11b-.In some of the EBIs,there were CD11b+cells other than erythrocytes and macrophages.These results showed that a significantly enhanced efficiency in our meth-od,particularly in the removal of removing single cells and non-EBI clusters without causing significant destruction of EBI struc-tures.Our modified method is suitable for research with high EBI purity requirements.Our research provided a more efficient method for researching hematopoietic microenvironment study.
万方数据
维普
