Image‐Based Flow Cytometry and Angle‐Resolved Light Scattering to Define the Sickling Process

Abstract

Red blood cells (RBCs) from sickle cell patients exposed to a low oxygen tension reveal highly heterogeneous cell morphologies due to the polymerization of sickle hemoglobin (HbS). We show that angle‐resolved light scattering approach with the use of image‐based flow cytometry provides reliable quantitative data to define the change in morphology of large populations of RBCs from sickle cell patients when the cells are exposed for different times to low oxygen. We characterize the RBC morphological profile by means of a set of morphological and physical parameters, which includes cell shape, size, and orientation. These parameters define the cell as discocyte, sickle, elongated, as well as irregularly or abnormal RBC shaped cells, including echinocytes, holly‐leaf, and granular structures. In contrast to microscopy, quick assessment of large numbers of cells provides statistically relevant information of the dynamic process of RBC sickling in time. The use of this approach facilitates the understanding of the processes that define the propensity of sickle blood samples to change their shape, and the ensuing vaso‐occlusive events in the circulation of the patients. Moreover, it assists in the evaluation of treatments that include the use of anti‐sickling agents, gene therapy‐based hemoglobin modifications, as well as other approaches to improve the quality of life of sickle cell patients. © 2019 International Society for Advancement of Cytometry

In Flow Cytometry, Image Is Everything

“…Gu et al present their demonstration of image‐guided cell sorting together with a cell classification system and real‐time isolation of single cells based on nuclear localization of glucocorticoid receptor, particle binding to the cell membrane, and DNA damage induced γ‐H2AX foci using the instrument. Goktas et al report a combination of IFC and angle‐resolved light scattering measurements to study the morphological features of red blood cells in sickle cell patients. Hui et al describe their development of a method, which they refer to as immuno‐flowFISH, for conducting FISH on immunophenotyped whole cells in suspension to detect chromosomal abnormalities in chronic lymphocytic leukemia.…”

Artificial intelligence in imaging flow cytometry

“…In an IFC system, the specimen flows in a capillary and images of the specimen are rapidly captured by a detector ( Figure 1 ). Illumination of the sample is provided by light emitting diodes (LED) or lasers (not shown in the figure) in different configurations, which include transillumination ( Basiji et al, 2007 ), excitation from an angle ( Goktas et al, 2019 ), structured ( Mashayekh et al, 2022 ) or light-sheet illumination ( Gualda et al, 2017 ). The samples flow in a simple straight tube, in more sophisticated capillaries, or in lab-on-chip devices ( Paiè et al, 2018 ).…”