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In\u00a0vitro\u2010generated MDSCs are a convenient tool to study the properties of tumour\u2010associated MDSCs. Here, we compared six protocols for in\u00a0vitro generation of functional mouse MDSCs from bone marrow progenitors. The protocols included granulocyte\u2010macrophage colony\u2010stimulating factor (GM\u2010CSF) alone or in combination with interleukin\u20106 (IL\u20106) or granulocyte colony\u2010stimulating factor (G\u2010CSF), with or without a tumour\u2010conditioned medium (TCM) derived from B16\u2010F10 melanoma. Obtained MDSCs were characterized by morphology, phenotype, gene expression of key immunosuppressive factors, and in\u00a0vitro suppression of T cell proliferation. All tested protocols yielded approximately 25% monocytic and 50% polymorphonuclear MDSCs. Protocols using IL\u20106 generated MDSCs with reduced maturation and differentiation status, upregulated <jats:italic>Arg1<\/jats:italic> and <jats:italic>Nos1<\/jats:italic> mRNA expression, increased levels of Arg\u20101 and TGF\u2010\u03b2 proteins and enhanced ROS production compared to the other protocols. All tested protocols yielded MDSCs that efficiently inhibited T cell proliferation in\u00a0vitro, with some advantage for the GM\u2010CSF and G\u2010CSF\u2009+\u2009GM\u2010CSF protocols. Interestingly, a combination of protocols with B16\u2010F10\u2010derived TCM resulted in the generation of MDSCs with reduced immunosuppressive properties. Our results provide valuable insights into the optimal conditions for in\u00a0vitro generation of MDSCs with specific immunosuppressive properties.<\/jats:p>","DOI":"10.1111\/sji.70001","type":"journal-article","created":{"date-parts":[[2025,1,27]],"date-time":"2025-01-27T07:21:56Z","timestamp":1737962516000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["The impact of cytokines and tumour\u2010conditioned medium on the properties of murine in\u00a0vitro generated myeloid\u2010derived suppressor cells"],"prefix":"10.1111","volume":"101","author":[{"ORCID":"https:\/\/orcid.org\/0009-0008-2075-2599","authenticated-orcid":false,"given":"Mona","family":"Awad","sequence":"first","affiliation":[{"name":"Institute of Chemical Biology and Fundamental Medicine SB RAS  Novosibirsk Russia"},{"name":"Faculty of Natural Sciences Novosibirsk State University  Novosibirsk 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