Abstract
We propose a method for characterization of mature red blood cells (RBCs) morphology, based on measurement of light-scattering patterns (LSPs) of individual RBCs with the scanning flow cytometer and on solution of the inverse light-scattering (ILS) problem for each LSP. We considered a RBC shape model, corresponding to the minimal bending energy of the membrane with isotropic elasticity, and constructed an analytical approximation, which allows rapid simulation of the shape, given the diameter and minimal and maximal thicknesses. The ILS problem was solved by the nearest-neighbor interpolation using a preliminary calculated database of 250,000 theoretical LSPs. For each RBC in blood sample we determined three abovementioned shape characteristics and refractive index, which also allows us to calculate volume, surface area, sphericity index, spontaneous curvature, hemoglobin concentration and content.
Highlights
Determination of such red blood cell (RBC) characteristics as volume, shape, and hemoglobin concentration allows one to assess a blood status and to reveal a number of diseases such as anemia, hereditary spherocytosis, hereditary elliptocytosis, sickle cells disease, and myeloproliferative disorders [1]
We developed a method for characterization of mature RBCs, based on measurement of light-scattering patterns (LSPs) from single cells in flow with the scanning flow cytometer (SFC) and subsequent solution of the inverse light-scattering (ILS) problem for each LSP
The median uncertainties of measured diameter and thickness were 0.25 μm and 0.07 μm, respectively. Such spatial resolution opens a way for precise studies of processes involving RBCs, i.e. lysis, activation, etc
Summary
Determination of such red blood cell (RBC) characteristics as volume, shape, and hemoglobin concentration allows one to assess a blood status and to reveal a number of diseases such as anemia, hereditary spherocytosis, hereditary elliptocytosis, sickle cells disease, and myeloproliferative disorders [1]. The main limitation of these methods is the need to chemically spherize RBCs prior to their characterization This requires additional probe preparation, and only the cell volume, but not the shape can be estimated. A number of interference methods do allow measuring volume, shape, and hemoglobin content (refractive index) of individual RBCs [6,7,8]. They are based on simplifying assumptions in the light-scattering theory (have hard-to-control accuracy) and are not yet ready for highthroughput applications, such as routine clinical blood analysis. For applications involving many instances of the shape, i.e. for fitting a LSP to solve the ILS problem, the shape model needs to be both realistic and quickly computable from a small number of characteristics
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