Protocol for quantification of circulating immature granulocytes in patients with a chronic myeloid malignancy using flow cytometry.

Immature granulocytes (iGRANs) circulating in the peripheral blood of patients with a chronic myeloid malignancy, such as chronic myelomonocytic leukemia, are immunosuppressive cells that promote disease progression. Here, we present a multiparameter flow cytometry-based protocol to quantify iGRANs among peripheral blood mononucleated cells (PBMCs). We describe steps for collecting and labeling PBMCs. We describe the gating strategy to quantify iGRANs as a fraction of myeloid cells or as an absolute number of circulating cells, which supports patient stratification and monitoring. For complete details on the use and execution of this protocol, please refer to Deschamps et al.1.