Abstract
Ex vivo-generated red blood cells are a promising resource for future safe blood products, manufactured independently of voluntary blood donations. The physiological process of terminal maturation from spheroid reticulocytes to biconcave erythrocytes has not been accomplished yet. A better biomechanical characterization of cultured red blood cells (cRBCs) will be of utmost interest for manufacturer approval and therapeutic application. Here, we introduce a novel optical tweezer (OT) approach to measure the deformation and elasticity of single cells trapped away from the coverslip. To investigate membrane properties dependent on membrane lipid content, two culture conditions of cRBCs were investigated, cRBCPlasma with plasma and cRBCHPL supplemented with human platelet lysate. Biomechanical characterization of cells under optical forces proves the similar features of native RBCs and cRBCHPL, and different characteristics for cRBCPlasma. To confirm these results, we also applied a second technique, digital holographic microscopy (DHM), for cells laid on the surface. OT and DHM provided related results in terms of cell deformation and membrane fluctuations, allowing a reliable discrimination between cultured and native red blood cells. The two techniques are compared and discussed in terms of application and complementarity.
Highlights
The availability, quality and safety of blood products is a growing issue worldwide and the emerging supply bottleneck is hardly manageable with voluntary blood donations alone [1]
New methylene blue staining indicated a maturation grade of both cRBCPlasma and cRBCHPL between native reticulocytes and native RBC (nRBC) according to their stained ribosomal rests
We found that the mean corpuscular hemoglobin cRBCis not associated with the biggest volume (CV = 107, 1 (MCH)
Summary
The availability, quality and safety of blood products is a growing issue worldwide and the emerging supply bottleneck is hardly manageable with voluntary blood donations alone [1]. After the terminal shape change to biconcavity do the mature erythrocytes achieve the crucial flexibility and stability within the blood stream [5] This is necessary to adapt to various osmotic conditions as, while RBCs are biconcave disks at equilibrium, they shrink under hyperosmotic conditions and swell under hyposmotic conditions. 42.61CMF by the height normalization coefficient, c = hm_RBC_Type/hm_nRBC, the CMF values are rearranged in a sequence following the expected trend. These corrected values are in agreement with the measurement results using OOTs, indicating that nRBCs are the most flexible and cRBCPlasma the least, with cRBCHPL in between
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