Analysis of MPO deficiency in an MPN patient cohort. (A) Peripheral blood ADVIA dot plots from a patient with MF and normal MPO activity (left), and a patient with MF and acquired MPO deficiency (right). LUC (light blue), neutrophil granulocytes (purple), monocytes (green), lymphocytes (dark blue), eosinophil granulocytes (orange), and platelets/lysed red blood cells (black) are shown. The numbers indicate the percentage of LUC in peripheral blood leukocytes. (B) Percentage of LUC in peripheral blood leukocytes from patients with MF (n = 81), ET (n = 94), and PV (n = 142). Patients with a CALR mutation and MPO deficiency (dark blue dots); patient 1 analyzed in more detail in Figure 2 (light blue dot); and patient 6 with a JAK2 mutation and MPO deficiency (red dot) are shown. (C) Percentage of LUC in peripheral blood leukocytes from patients with stratified according to the presence of a CALR (n = 29) or a JAK2 (n = 43) mutation. (D, left) Intracellular detection of MPO protein by flow cytometry in neutrophil granulocytes of a patient with MF and normal MPO activity (top), and a patient with MF and acquired MPO deficiency (bottom). The numbers indicate the percentage of cells in the gate. (D, right) Detection of MPO protein by confocal microscopy in neutrophil granulocytes of a patient with MF and normal MPO activity (top), and a patient with MF and acquired MPO deficiency (bottom). Scale bar, 5 μm. (E) Relative expression of MPO mRNA measured by qPCR and normalized to that of porphobilinogen deaminase in neutrophil granulocytes of patients with MF and normal MPO activity (n = 32), and patients with MPO deficiency (n = 6). (F) Correlation of the presence of MPO deficiency with the mutant allele burden of CALR. The median in each dot plot is represented as a green or black line. *P < .05; **P < .01; ***P < .001 (Mann–Whitney U test). cyMPO, cytoplasmic MPO.