cd33 magnetic bead  (Miltenyi Biotec)

Journal: Molecular Cancer

Article Title: Myeloid-derived suppressor cells endow stem-like qualities to multiple myeloma cells by inducing piRNA-823 expression and DNMT3B activation

doi: 10.1186/s12943-019-1011-5

Figure Lengend Snippet: G-MDSCs are functionally and clinically relevant in MM. a The gating strategy for G-MDSCs in multi-parameter flow cytometry. G-MDSCs were immunophenotyped as the HLA-DR −/low /CD33 + /CD11b + /CD15 + /CD14 − population (P4, the purple cell population). b CD8 + T cells were cultured with G-MDSCs at different ratios for 72 h. Then T cell proliferation was detected using the CCK-8 assay. The data represent the means ± SD of triplicate cultures, and the results are representative of 3 different experiments. c Annexin V-FITC/PI staining of CD8 + T cells was performed 72 h after co-culture with G-MDSCs. CD8 + T cells were used as controls. d The levels of cytokines (IFN-γ, TNF-α, and IL-2) in cell culture supernatants were quantified using flow cytometry. e The frequencies of G-MDSCs in PB and BM from healthy donors were compared with MM patients. f Seventy-two MM patients were classified according to ISS. The frequencies of G-MDSCs in BM were detected, and mean values were compared between groups using the Mann–Whitney U rank sum tests. * P < 0.05, ** P < 0.01. g Clinical impact of G-MDSCs on the survival of MM patients. Kaplan-Meier estimates of overall survival were calculated according to the median G-MDSC density

Article Snippet: G-MDSCs from patients or healthy volunteers were purified using CD33, HLA-DR and CD15 magnetic beads (MiltenyiBiotec, Auburn, CA, USA) according to the manufacturer’s protocol, and the purity of the sorted cells was > 95% using flow cytometry.

Techniques: Flow Cytometry, Cell Culture, CCK-8 Assay, Staining, Co-Culture Assay, MANN-WHITNEY