The proposed orthogonal application of HS glycotyping with canonical CD-marker flow cytometry in heterogenous populations and the differential and dynamic binding of an HS scFv within megakaryocytic and erythroid lineages. (A) The long-standing use of CD markers has helped identify and isolate hematopoietic cell subpopulations from within heterogenous pools (eg, MEP, CMP, and GMP from a mixed pool of hematopoietic progenitor cells). The addition of HS scFvs to this process can help further purify distinct populations. With nearly limitless possibilities of HS-modification patterns, the combinatorial use of HS scFvs holds immense potential in isolating rare subtypes of hematopoietic cells, possibly even at the single-cell level. (B) Schematic of HS3A8 binding during various stages of megakaryocyte and erythroid differentiation. HS3A8 scFv recognizes specific HS motifs containing sulfation marks (eg, 2-O-, 3-O-, and 6-O-sulfation) at certain residues. MEPs contain a variable level of HS3A8 binding, which when separated into high and low binding, helps delineate megakaryocyte vs erythroid lineage commitment. Furthermore, HS3A8 shows dynamic and temporal binding across the different stages of erythroid differentiation, whereas megakaryocytic cells show low HS3A8 binding regardless of the stage of maturation. Baso-E, basophilic erythroblast; CMP, common myeloid progenitors; GMP, granulocyte-monocyte progenitors; Mk, megakaryocyte; MkP, megakaryocyte progenitor; Ortho-E, orthochromatic erythroblast; Poly-E, polychromatophilic erythroblast; Pro-E, proerythroblast.