Background Measurable residual disease (MRD) is a strong independent poor prognostic factor for acute leukemia. Multiparameter flow cytometry (FCM) is a commonly used MRD detection method. However, FCM MRD detection is not well standardized, and the interpretation is subjective. There are normal/reactive minor cell populations in bone marrow (BM) and peripheral blood (PB), which could be confused with MRD. Methods The FCM data of 231 BM and 44 PB pediatric samples performed in a recent 15‐month period were retrospectively reviewed. These samples were from 56 B‐lymphoblastic leukemia (B‐ALL) patients, 11 T‐lymphoblastic leukemia (T‐ALL) patients, 28 acute myeloid leukemia (AML)/myelodysplastic syndrome (MDS) patients, 44 cytopenia/leukocytosis patients, and five patients with mycosis fungoides. Results There were over 10 normal or reactive minor cell populations identified with certain phenotypes mimicking MRD of acute leukemia. These mimickers included CD19+ NK cells, CD22+ basophils, CD22+ dendritic cells (DCs), and plasma cells for B‐ALL MRD; CD4/8 double‐negative T cells, CD4/8 double‐positive T cells, cytoplasmic CD3+ NK cells, CD2− T cells, CD7− T cells, CD5− gamma delta T cells, CD56+ NKT cells for T‐ALL MRD; CD33+ NK cells, CD117+ NK cells, basophils, plasmacytoid DCs, non‐classical monocytes, CD56+ and/or CD61+ monocytes for AML MRD. Conclusions These data confirm the presence of a variety of normal/reactive minor cell populations that could mimic MRD of acute leukemia by FCM. Recognizing these MRD mimickers is important for correct FCM MRD interpretation.

Novel therapies such as monoclonal antibody or chimeric antigen receptor (CAR) T cell therapy (CAR T) have shown promising results in the management of relapsed/refractory B-lymphoblastic leukemia (B-ALL). CD19, a transmembrane glycoprotein almost always expressed in B-ALL, is a commonly used target by these therapies. Blinatumomab, a bispecific monoclonal antibody T cell engager (BiTE), induces antibody-dependent cellular cytotoxicity against the leukemic cells by binding to CD19 on leukemic cells and CD3 on T cells simultaneously. Tisagenlecleucel (Kymriah), a CAR T targeting CD19, uses patients' own modified T cells to attack CD19+ leukemic cells. The superior efficacy of blinatumomab and tisagenlecleucel has been well demonstrated in relapsed/refractory B-ALL compared with high-dose chemotherapy. Multiparametric flow cytometry (FCM) is commonly used to monitor the therapeutic responses to these treatments. The key strategy for FCM to detect residual or relapsed B-ALL is to find an immature B-cell population phenotypically different from normal B-cell precursors (BCPs, also known as hematogones) in the bone marrow (BM). To investigate if these two anti-CD19 therapies could alter the phenotype of normal BCPs, we retrospectively studied the BCPs in the BM specimens from patients treated with blinatumomab and tisagenlecleucel. We also studied the BCPs from the patients after conventional chemotherapy and hematopoietic stem cell transplantation (HSCT) as a comparison.

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