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.

Dielectrophoretic detection of electrical property changes of stored human red blood cells.

The ability to transport and store a large human blood inventory for transfusions is an essential requirement for medical institutions. Thus, there is an important need for rapid and low-cost characterization tools for analyzing the properties of human red blood cells (RBCs) while in storage. In this study, we investigate the ability to use dielectrophoresis (DEP) for measuring the storage-induced changes in RBC electrical properties. Fresh human blood was collected, suspended in K2-EDTA anticoagulant, and stored in a blood bank refrigerator for a period of 20 days. Cells were removed from storage at 5-day intervals and subjected to a glutaraldehyde crosslinking reaction to "freeze" cells at their ionic equilibrium at that point in time and prevent ion leakage during DEP analysis. The DEP behavior of RBCs was analyzed in a high permittivity DEP buffer using a three-dimensional DEP chip (3DEP) and also compared to measurements taken with a 2D quadrupole electrode array. The DEP analysis confirms that RBC electrical property changes occur during storage and are only discernable with the use of the cell crosslinking reaction above a glutaraldehyde fixation concentration of 1.0 w/v%. In particular, cytoplasm conductivity was observed to decrease by more than 75% while the RBC membrane conductance was observed to increase by more than 1000% over a period of 20 days. These results show that the presented combination of chemical crosslinking and DEP can be used as rapid characterization tool for monitoring electrical properties changes of human RBCs while subjected to refrigeration in blood bank storage.