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  • Title: Establishment of a flow cytometry assay for detecting paroxysmal nocturnal hemoglobinuria-type cells specific to patients with bone marrow failure.
    Author: Hosokawa K, Sugimori C, Ishiyama K, Takamatsu H, Noji H, Shichishima T, Obara N, Chiba S, Ninomiya H, Shirasugi Y, Nakamura Y, Ando K, Ueda Y, Yonemura Y, Kawaguchi T, Nishimura JI, Kanakura Y, Nakao S.
    Journal: Ann Hematol; 2018 Dec; 97(12):2289-2297. PubMed ID: 30039297.
    Abstract:
    Minor populations of glycosylphosphatidylinositol-anchored protein-deficient (GPI[-]) cells in the peripheral blood may have a prognostic value in bone marrow failure (BMF). Our objective is to establish the optimal flow cytometry (FCM) assay that can discriminate GPI(-) populations specific to BMF from those of healthy individuals. To identify a cut-off that discriminates GPI(-) rare cells from GPI(+) cells, we determined a position of the borderline that separates the GPI(-) from GPI(+) cells on a scattergram by testing more than 30 healthy individuals, such that no GPI(-) dot fell into the upper left quadrant where fluorescein-labeled aerolysin (FLAER)-CD11b+ granulocytes and CD55-CD59- glycophorin A+ erythrocytes were positioned. This method allowed us to define ≥ 0.003% CD11b+FLAER- granulocytes and ≥ 0.005% glycophorin A+CD55-CD59- erythrocytes to be specific to BMF patients. Longitudinal cross-validation studies showed minimal (< 0.02%) inter-laboratory differences in the GPI(-) cell percentage. An analysis of 1210 patients with BMF revealed a GPI(-) cell population in 56.3% of patients with aplastic anemia and 18.5% of patients with myelodysplastic syndrome. The GPI(-) granulocyte percentages was 0.003-0.01% in 3.7% of patients. This FCM assay effectively identified an increase in the percentage of GPI(-) rare cells that are specific to BMF patients and allowed different laboratories to accurately detect 0.003-0.01% of pathological GPI(-) cells.
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