Characterization of G-MDSCs based on conventional criteria. (A) BM samples from MM patients (n = 10) and HAs (n = 7) were stained with HLADR-BV421, CD45-OC515, CD15-FITC, CD13-PE, CD33-PerCPCy5.5, CD16-PECy7, CD11b-APC, and CD14-APCH7 mAbs. Cells with a CD11b⁺CD14⁻CD15⁺CD33⁺HLADR⁻ phenotype represent ∼25% of total BM nucleated cells both in HAs and MM patients. (B) Unbiased analysis based on uniform manifold approximation and projection (UMAP) according to expression levels of HLADR, CD45, CD15, CD13, CD33, CD16, CD11b, and CD14 revealed various granulocytic subsets (neutrophils, eosinophils, and basophils) in BM samples from HAs and MM patients. (C) UMAP of the neutrophil population led to the identification of 3 neutrophil maturation stages according to differential expression of CD11b, CD13, and CD16: immature (CD11b⁻CD13^−/loCD16⁻), intermediate (CD11b⁺CD13^−/loCD16⁻), and mature (CD11b⁺CD13⁺CD16⁺) neutrophils. Cellular maturation was confirmed on cytospinned cells from the 3 different populations by evidencing the classic changes in nuclear shape. Images are shown with a ×400 magnification. (D) Frequency of each granulocytic subset was similar between HAs and MM patients. Bars represent the mean and lines the standard deviation.