Measurement and analysis of ionic leakage profiles in refrigerated human red blood cells using dielectrophoresis and inductively coupled mass spectroscopy

Human red blood cells (RBCs) undergo ionic leakage through passive diffusion during refrigerated storage, affecting their quality and health. We investigated the dynamics of ionic leakage in human RBCs over a 20-day refrigerated storage period using extracellular ion quantification and dielectrophoresis (DEP). Four type O− human blood donors were examined to assess the relationship between extracellular ion concentrations (Na⁺, K⁺, Mg²⁺, Ca²⁺, and Fe²⁺), RBC cytoplasm conductivity, and membrane conductance. A consistent negative correlation between RBC cytoplasm conductivity and membrane conductance, termed the “ionic leakage profile” (ILP), was observed across the 20-day storage period. Specifically, we noted a gradual decline in DEP-measured RBC cytoplasm conductivity alongside an increase in membrane conductance. Further examination of the electrical origins of this ILP using inductively coupled plasma mass spectrometry revealed a relative decrease in extracellular Na⁺ concentration and an increase in K⁺ concentration over the storage period. Correlation of these extracellular ion concentrations with DEP-measured RBC electrical properties demonstrated a direct link between changes in the cytoplasmic and membrane domains and the leakage and transport of K⁺ and Na⁺ ions across the cell membrane. Our analysis suggests that the inverse correlation between RBC cytoplasm and membrane conductance is primarily driven by the passive diffusion of K⁺ from the cytoplasm and the concurrent diffusion of Na⁺ from the extracellular buffer into the membrane, resulting in a conductive reduction in the cytoplasmic domain and a subsequent increase in the membrane. The ILP's consistent negative trend across all donors suggests that it could serve as a metric for quantifying blood bank storage age, predicting the quality and health of refrigerated RBCs.