CD11b^low/−/CD16⁻ phenotype defines the subset responsible for the immune suppression in BM-MDSCs. (A) Flow cytometric evaluation of CD11b and CD16 markers in BM-MDSC or sorted CD11b^low/−/CD16⁻, CD11b⁺/CD16⁻ and CD11b⁺/CD16⁺ cell populations from fresh BM samples (left), structural analysis by May-Grünwald-Giemsa staining (center), and CFSE dilution proliferation assay (right) in which values reported on histograms represent the percentages of cells in the parental, undivided generation. (B) Flow cytometric evaluation of CD11b and CD16 markers in BM-MDSCs or sorted CD11b^low/−/CD16⁻, CD11b⁺/CD16⁻, and CD11b⁺/CD16⁺ cell populations from BM-MDSCs (left), structural analysis by May-Grünwald-Giemsa staining (center), and CFSE dilution proliferation assay (right) in which values reported on histograms represent the percentages of cells in the parental, undivided generation. (C) Suppression of allogenic CFSE⁺ PBMCs activated with anti-CD3 and anti-CD28 and cocultured in the presence of 1:1 ratio of the different populations sorted from human BM-MDSCs. The suppression was calculated, analyzing the number of proliferating cells from generation 3 to generation 10, assumed to be 100% without BM-MDSCs. Mean ± SE of 3 independent experiments. P ≤ .01, Student t test, all comparisons among BM-MDSCs containing cultures versus cultures without BM-MDSCs.