TABLE 1.
Overview of parameters showing inter-cellular heterogeneity as well as basic principle and methodological approaches of their detection in single cells and sub-populations.
| Parameter | Indicator | Method | References |
| Shape/size |
Direct: Shape classification Projected area Perimeter/roughness Sphericity/elongation |
Microscopy: Blood smears, images of living cells (snapshots, time series in flow, microfluidics), Imaging flow cytometers |
Gonzalez-Hidalgo et al., 2015; Quint et al., 2018; Herold-Garcia and Fernandes, 2019 |
| Volume | Confocal microscopy + 3D deconvolution | Sadafi et al., 2019 | |
| Scanning probe microscopy (semi quantitative) | Kihm et al., 2018 | ||
| Indirect: Forward and side light scatter Impedance (coulter principle) | Flow cytometry Coulter counters Multiple Blood analyzers |
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| Density | Direct: Separation according to RBC density | Fractionation in Percoll-, Stractan or similar density gradients | Lutz et al., 1992 |
| Lab-on-a-chip approaches | Catarino et al., 2019 | ||
| Indirect: Swelling- or shrinkage- resistance (e.g., the changes in SS and FS within the swelling test) Single cell rheology | Flow cytometry Cell-flow properties analyzer | Kaul et al., 2008; Barshtein et al., 2016; Fermo et al., 2017 | |
| Membrane surface/EMA test | Flow cytometry | ||
| Free Ca2+/channel activity | Fluorescent dyes for Ca2+ Detection of ionic currents across the membranes of single cells | Flow cytometry and fluorescence microscopy Patch-clamp incl. automated planar chips | Kaestner et al., 2006; Makhro et al., 2013; Wang et al., 2013; Fermo et al., 2017; Rotordam et al., 2019 |
| Redox state and metabolism | Fluorescent dyes for reduced thiols (e.g., thiol tracker, monobromobimane), Fluorescent dyes for N2O3 (DAF-DA), Dyes for detection of H2O2, ONOO, HO* (e.g., H2DCF-DA) Single cell metabolomics (not yet used for red blood cells) | Flow cytometry Fluorescence microscopy Mass-spectrometry |
Jemaa et al., 2017; Gilmore et al., 2019 |
| Hb levels and variance | Antibodies with fluorescent tags Chromicity Sodium metabisulfite (Na2S2O5) and similar deoxygenation-based sickling tests Hemoglobin Distribution Width (HDW) |
Flow cytometry, Fluorescence microscopy Microscopy |
Kunicka et al., 2001; Darrow et al., 2016; Jung et al., 2016 |
| Age | Labeling of cells (biotin conjugated with fluorescent tag or staining with PKH dyes) Reticulocyte count RNA-positive or Transferrin receptor-positive | Flow cytometry, microscopy | Mock et al., 1999; Piva et al., 2010 |