Automated Detection of Dysplasia: Data Mining from Our Hematology Analyzers

Abstract

Myelodysplastic syndromes (MDSs) are clonal hematopoietic diseases of the elderly, characterized by chronic cytopenia, ineffective and dysplastic hematopoiesis, recurrent genetic abnormalities and increased risk of progression to acute myeloid leukemia. Diagnosis on a complete blood count (CBC) can be challenging due to numerous other non-neoplastic causes of cytopenias. New generations of hematology analyzers provide cell population data (CPD) that can be exploited to reliably detect MDSs from a routine CBC. In this review, we first describe the different technologies used to obtain CPD. We then give an overview of the currently available data regarding the performance of CPD for each lineage in the diagnostic workup of MDSs. Adequate exploitation of CPD can yield very strong diagnostic performances allowing for faster diagnosis and reduction of time-consuming slide reviews in the hematology laboratory.

Keywords: cell population data; complete blood count; hematology analyzer; leukocyte differential; myelodysplastic syndrome.

Figure 1

Illustration of the Coulter technology. (a) The Coulter UniCel DxH 800 analyzer measures cell volume (V) through the V impedance and the internal complexity or opacity (OP) of the cell with the C radio frequency. The light scattering is measured at different angles: upper-median-angle light scatter (UMALS), lower-median-angle light scatter (LMALS), low-angle light scatter (LALS) and axial-loss light scatter (AL2). The fifth light scatter channel, median-angle light scatter (MALS), is the sum of the UMALS and LMALS regions. For each parameter, mean (MN) and standard deviation (SD) are collected as cell population data (CPD). Created with BioRender.com. (b) Leukocyte differential: scatterplot of volume (V) versus rotated light scatter (RLS) identifies five subpopulations: neutrophils (NEs) in pink, eosinophils in orange, basophils in white, lymphocytes in blue and monocytes in green. CPD from volume are illustrated: MN-V-NE (dotted arrow) represents the mean channel value of NE population volume, SD-V-NE (vertical arrow) represents the standard deviation of volume for this population. (c) Leukocyte differential: scatterplot of volume (V) versus opacity (OP). Neutrophils, eosinophils and basophils have a similar opacity but basophils have a decreased volume. (d) Nucleated red blood cell (NRBC) plot: scatterplot of rotated low-angle light scatter (RLALS) versus axial-loss light scatter (AL2) showing in blue the leukocytes, in green macroplatelets and platelet clumps (and debris) and a few sparse red events corresponding to NRBCs. (e) Nucleated red blood cell (NRBC) plot: scatterplot of reflected upper-median-angle light scatter (RUMALS) versus AL2 showing in blue the leukocytes, in green the platelets (macroplatelets, platelet clumps) and a few sparse red events corresponding to NRBCs. (f) Scatterplot of volume (V) versus linear light scattering (LLS) showing in green platelets (and debris), in purple reticulocytes and in red mature RBCs. (g) Scatterplot of volume (V) versus opacity (OP) showing in green platelets (and debris), in purple reticulocytes and in red mature RBCs.

Figure 2

Illustration of the Sysmex technology. (a) The Sysmex-XN applies fluorescence-flow cytometry on cells to collect information on internal complexity (side scatter, SSC), nucleic acid content (fluorescence, SFL) and cell size (forward scatter, FSC. Created with BioRender.com. (b) Position of different cell populations on the white blood cell differential scatterplot (WDF). X-axis shows side scatter (SSC) of laser light, Y-axis represents side fluorescence (SFL). The different types of leukocytes are represented: light blue for neutrophils, pink for lymphocytes, green for monocytes, red for eosinophils and dark blue for platelets and debris. Neutrophil scattering parameters are illustrated by the dotted arrow for granularity (SSC) on the X-axis (Ne-X) and the horizontal arrow for neutrophil side scatter area distribution width (Ne-WX). (c) White blood cell differential scatterplot (WDF). X-axis shows side scatter (SSC) of laser light, Y-axis represents forward scatter (FSC). (d) White count and nucleated red blood cell scatterplot (WNR) showing in light blue lysed leukocytes, in yellow basophils. If present, nucleated red blood cells (NRBCs) are identified in purple. X-axis shows side scatter (SSC) of laser light, Y-axis represents side fluorescence (SFL). (e) White count and nucleated red blood cell scatterplot (WNR) showing in light blue lysed leukocytes and in yellow basophils. If present, nucleated red blood cells (NRBCs) are identified in purple. X-axis shows side scatter (SSC) of laser light, Y-axis represents forward scatter (FSC). (f) Reticulocytes scatterplot showing high-, medium- and low-fluorescent reticulocytes in red, orange and purple, respectively, and mature RBCs in dark blue. X-axis shows side scatter (SSC) of laser light, Y-axis represents forward scatter (FSC). (g) Fluorescence platelet count scatterplot showing in green immature platelets (ÏPF) and in light blue platelets. At the top of the graph, red blood cells appear in dark blue.

Figure 3

Illustration of the Abbot technology. (a) Alinity-hq from Abbott uses seven light scatter detectors to determine various cellular features: axial light loss (ALL) related to size, intermediate-angle scatter (IAS) related to cellular complexity, polarized side scatter (PSS) related to nuclear lobularity/segmentation and depolarized side scatter (DSS) allowing for specific identification of eosinophil granulocytes. The three narrow-angle light scatter detectors IAS1, IAS2 and IAS3 provide information on the volume, hemoglobin content and granularity of red blood cells (RBCs) and platelets. Created with BioRender.com. (b) Leukocyte differential on the axial light loss (ALL) versus intermediate-angle scatter (IAS) scatterplot: neutrophils are shown in yellow, lymphocytes in light blue, monocytes in purple, eosinophils in green, basophils in black and nucleated red blood cells in red, if present. Neutrophil scattering parameters are illustrated by the dotted arrow for granularity (IAS) on the X-axis (NE-IAS-M) and the horizontal arrow for neutrophil side scatter standard deviation (NE-IAS-S). (c) Leukocyte differential on the axial light loss (ALL) versus fluorescence (FL1) scatterplot: neutrophils appear in yellow, lymphocytes in light blue, monocytes in purple, eosinophils in green and basophils in black. Gray dots are non-DNA-containing material such as platelet clumps or lyse-resistant RBCs. (d) Leukocyte differential on polarized side scatter (PSS) versus axial light loss (ALL) scatterplot: neutrophils appear in yellow, lymphocytes in light blue, monocytes in purple, eosinophils in green and basophils in black. (e) Leukocyte differential on depolarized side scatter (DSS) versus polarized side scatter (PSS) allows for specific identification of eosinophil granulocytes in green. (f) RBCs (red) and platelets (yellow) on axial light loss (ALL) versus intermediate-angle scatter 1 (IAS1) scatterplot. (g) RBCs (red) and platelets (yellow) on intermediate-angle scatter 3 (IAS3) versus intermediate-angle scatter 1 (IAS1) scatterplot. (h) RBC scatterplot showing the distribution of cell hemoglobin concentration (CHC) versus volume. Microcytic RBCs (MICs) have a volume inferior to 60 fL, while that of macrocytic RBCs (MACs) is superior to 120 fL. Hypochromic RBCs (HYPs) have a CHC inferior to 28 g/dL and hyperchromic RBCs (HPRs) superior to 41 g/dL. (i) RBC and platelet fluorescence (FL1) versus axial light loss (ALL) scatterplot showing in green reticulocytes separated from mature RBCs in red, and reticulated platelets in blue separated from platelets in orange. (j) Platelets (yellow) axial light loss (ALL) versus intermediate-angle scatter 2 (IAS2) scattergram. (k) Platelets (yellow) polarized side scatter (PSS) versus intermediate-angle scatter (IAS) scattergram.

Figure 4

Illustration of typical MDS cases on DxH 800 and Sysmex-XN. (a) Leukocyte differential plot from DxH 800, showing decreased light scattering in a patient with myelodysplastic syndrome with excess blasts (MDS-EB-1). LMALS-NE-M, UMALS-NE-M and MALS-NE-M were decreased (128, 127 and 131, respectively). (b) Leukocyte differential plot from Sysmex-XN showing an increased Ne-WX (427) in this patient with normal neutrophil count (3.1 × 10⁹/L). This patient had normocytic anemia (Hb 9.7 g/dL, MCV 90 fL) and thrombocytopenia (platelet count: 107 × 10⁹/L), the MDS-CBC score was highly suggestive of MDS (0.755). This patient was diagnosed with MDS with multilineage dysplasia. (c) Fluorescence platelet count scatterplot from Sysmex-XN showing increased IPF% (19%) in this patient with macroplatelets on blood smear. (d–f) Blood film from patient illustrated in (b,c) showing hypogranular neutrophils (d,e, magnification ×100) and a macroplatelet (f, magnification ×50).