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Erythrosensors Cellular Characterization
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For diabetics, continuous glucose monitoring, and the resulting tighter control of glucose levels, ameliorates serious complications from hypoglycemia and hyperglycemia. Today, diabetics measure their blood glucose levels multiple times a day by finger pricks, or use implantable monitoring devices. Still, glucose and other analytes in the blood fluctuate throughout the day and the current monitoring methods are invasive, immunogenic, and/or present biodegradation problems. Using carrier erythrocytes loaded with a fluorescent sensor, we seek to develop a biodegradable, efficient, and potentially cost effective method for long-term monitoring of blood analytes. We aim to reintroduce sensor-loaded erythrocytes to the bloodstream and conserve the erythrocytes lifetime of 120 days in the circulatory system. Here, the efficiency of two loading procedures is compared. Hypotonic dilution employs hypotonic buffer to create transient pores in the erythrocyte membrane, allowing dye entrance and a hypertonic buffer to restore tonicity. Electroporation relies on controlled electrical pulses that results in reversible pores formation to allow cargo entrance. As part of the cellular characterization of loaded erythrocytes, size and hemoglobin content was evaluated. Cell recovery and fluorescence per cell measurements also render optimal loading conditions. Furthermore, AFM and confocal microscopy protocols were implemented to evaluate morphological changes induced by hypotonic dilution. The development of a suitable protocol to engineer carrier erythrocytes has profound and lasting implications in the erythrocytes’ lifespan and sensing capabilities.
Bustamante Lopez, Sandra Catalina (2017). Erythrosensors Cellular Characterization. Doctoral dissertation, Texas A & M University. Available electronically from