In Situ Monitoring of Morphology Changes and Oxygenation State of Human Erythrocytes During Surfactant-Induced Hemolysis.

Erythrocytes, the most abundant blood cells, are a prevalent cell model for the analysis of the membrane-damaging effects of different molecules, including drugs. In response to stimuli, erythrocytes can change their morphology, e.g., shape or volume, which in turns influences their main function to transport oxygen. Membrane active molecules can induce hemolysis, i.e., release of hemoglobin into the blood plasma. Free hemoglobin in the blood circulation is toxic causing serious health problems including vasoconstriction, high blood pressure and kidney damage. Therefore, early recognition of the risk of massive hemolysis is highly important. Here, we investigated surfactant induced hemolysis applying UV-vis spectrophotometry. Saponin, sodium dodecyl sulfate and Triton X-100, detergents known to provoke hemolysis at different concentrations and by different mechanisms, were applied to initiate the process. Whole absorption spectra of erythrocyte suspensions in the range 300-750 nm were recorded every 15 s for following the process in real-time. The hemolysis process, with respect to morphological changes in the erythrocytes and their influence on the oxygenation state of hemoglobin, was characterized by the absorbance at 700 nm, the height relative to the background and the wavelength of the Soret peak. The results suggest that these UV-vis spectrophotometry parameters provide reliable information in real-time; not only about the process of hemolysis itself, but also about pre-hemolytic changes in the erythrocytes, even at sub-hemolytic surfactant concentrations.