Flow cytometry (FCM) is a powerful tool for diagnosing and monitoring hematological malignancies, including multiple myeloma (MM), which is a plasma cell neoplasm characterized by clonal proliferation and high CD38 expression. This has led to the development of therapeutic anti-CD38 monoclonal antibodies (mAbs), such as daratumumab (DARA) and isatuximab (ISA), which have demonstrated significant clinical efficacy and achieved deep responses in both newly diagnosed and relapsed/refractory MM. However, a critical issue remains: therapeutic anti-CD38 mAbs interfere with anti-CD38 detection antibodies used in FCM, complicating accurate evaluation of CD38 expression during treatment and potentially affecting measurable residual disease (MRD) assessment. To accurately detect CD38 expression during DARA or ISA-containing therapies, it is essential to select anti-CD38 Abs that recognize epitopes distinct from those targeted by DARA or ISA. Despite recent advances in treatment, MM remains incurable, largely due to the acquisition of secondary genetic abnormalities such as gain or amplification of chromosome 1q (1q+), which is associated with poor prognosis and resistance to therapy. Recent studies have demonstrated that some MM cells with 1q+ exhibit reduced CD38 expression compared with 1q-negative MM cells, independent of the immature phenotype. FCM is expected to play a pivotal role not only in MRD detection but also in evaluating therapeutic target expression, thereby facilitating more individualized and optimized treatment approaches for MM.

Hematopoietic stem cell transplantation is crucial for treating refractory, hematological diseases. Nearly all autologous transplantations and increasingly, allogeneic transplantations as well, involve a peripheral blood stem cell transplantation (PBSCT). Hematopoietic stem cells are rarely found in the peripheral blood but are mobilized by administering granulocyte colony stimulating factor. The CD34+ cell count serves as an index of the quantity of hematopoietic stem cells in general, the precise quantification of which is crucial for a successful PBSCT. Methods of enumerating CD34+ cells were established in the 1990’s when PBSCT had just been pioneered; however, the result varies by the method used. The 1996 ISHAGE guidelines aimed to standardize the methods, and internal standard beads were introduced in 1998 to enable absolute enumeration. In Japan, however, standardization was promoted only after the first, unrelated PBSCT was performed in 2011. In 2013, The Japan Society of Transfusion Medicine and Cell Therapy established a committee for standardization by performing external quality-assessment studies (2016, 2017) and a comparative study of the CD34+ count in collection and transplantation sites in each case of unrelated PBSCT (2020). Although the standardization of enumeration has improved, the committee continues to hold annual seminars to promote education on the topic. The committee are also seeking collaboration with related organizations to cultivate expertise in the field and have requested the Japanese government to qualify CD34+ cell enumeration for insurance coverage as means of improving the safety of peripheral blood stem cell collection.

Neutrophils play a critical role in host defense by releasing web-like structures composed of DNA and proteins known as neutrophil extracellular traps (NETs) to trap and eliminate pathogens. However, excessive or dysregulated NET production can lead to vascular endothelial injury, thrombosis, and autoimmune responses, contributing to tissue damage. Establishing a reliable and convenient biomarker to quantitatively assess NET dynamics is thus clinically important. We focused on NADPH oxidase 2 (NOX2), a key enzyme involved in NET formation, and developed a novel method to quantitatively evaluate NET dynamics by measuring NOX2 expression using flow cytometry. A strong positive correlation was observed between the NET formation ratio and mean fluorescence intensity (MFI) of NOX2 expression (y = 2.308x + 0.412, r = 0.898), indicating that NOX2 expression quantitatively reflects NET formation. In a sepsis mouse model, the proportion of NOX2-positive neutrophils showed a similar trend to that of the NET formation ratio. These findings suggest that our flow cytometry-based method offers a simple and practical approach for monitoring NET dynamics and has potential clinical utility for diagnosis, disease assessment, and therapeutic monitoring of NET-associated disorders.

[Background] FMS-like tyrosine kinase 3 (FLT3) mutations, which are positive in about 25% of acute myeloid leukemia (AML) patients, are known to be a poor prognostic factor associated with a high relapse rate. In this study, we investigated the clinical characteristics of AML patients with FLT3 mutation-positive AML. [Subjects and Methods] The subjects were 24 patients diagnosed with AML between 2022 and 2024. Blood test data, morphological characteristics of bone marrow smears, cell surface markers, and chromosome and genetic test results were examined. [Results] Seven of the 24 patients were positive for FLT3 mutation, and all of them had FLT3-ITD mutation. Three of the seven FLT3 mutation positive patients showed a marked increase in leukocyte counts, while two patients showed a decrease in leukocyte counts. In cell surface markers, CD34 was deleted in all cases and HLA-DR was down-regulated in three cases. In addition, CD123 and CD25 were both positive in two cases. Chromosomal and genetic examination revealed normal karyotype and negative leukemia chimerism screening except for a case with KMT2A mutation. Three cases of NPM1 mutation were also observed. [Conclusion] In the FLT3 mutation-positive group, there were cases with markedly increased leukocyte counts and negative or decreased expression of both CD34 and HLA-DR as cell surface markers, and positive CD123 and CD25. Furthermore, WT-1 mRNA tended to be higher in the mutation positive group than in the mutation-negative group. These findings suggest the possibility of FLT3 mutation positivity.