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Danielle Precourt, a master's degree candidate in forensic science at Buffalo State, will present her thesis defense seminar, "Assessment of Calcein Dye Method to Observe Cell Volume Regulation," on Thursday, April 7, at 12:30 p.m. in Science Building 272. Light refreshments will be served before the seminar. This seminar is supported by the Faculty-Student Association. The abstract of her talk appears below.

Abstract
Cell volume regulation (CVR) is a crucial mechanism to the maintenance of homeostasis of cells under significant osmotic changes. Cells initiate a regulatory mechanism to restore to their initial volume when they are swollen or shrunken in a non-isotonic medium. Swollen cells in a hypotonic solution show decreases in cell volume by releasing osmolytes and water through a regulatory volume decrease (RVD). On the contrary, shrunken cells in a hypertonic solution exhibit a regulatory volume increase (RVI) by up-taking osmolytes and water. The whole process of CVR is difficult to understand, because we must understand both cell sensory mechanism of a volume change and complex signal pathways that eventually leads to the activation of ion channels or co-transporters. For a better understanding of CVR, it is therefore necessary to develop a method that can effectively trace changes in cell volume using a large number of cells.

Previously our lab had devised a single-cell microwell array based on photo-crosslinked hydrogel and used it to examine volume regulation of more than a hundred single cells at the same time. This highly productive analysis was made possible by tracing the changes in the fluorescence intensities of single cells in the time-lapse fluorescence micrographs, after those single cells were loaded with calcein, a volume-sensitive fluorescence dye. Others reported that the mechanism of calcein to observe for cell volume change was associated with the quenching effect of calcein by nonspecific intracellular proteins present in cytosol. Thus, when the cell swells, the fluorescence of calcein increases, because the decrease of intracellular protein concentration de-quenches the fluorescence. On the other hand, the cell shrinkage quenches the calcein fluorescence. While I tested the calcein method to trace cell volume change, I occasionally observed contradictory results, i.e., observing a decrease in fluorescence signal for the swelling cells. To understand the unusual behavior of calcein fluorescence, my research has been focused to validate the use of calcein dye in measuring cell volume change in non-isotonic solution. Here I will present my findings on the calcein method and show an alternative way of using the calcein to monitor the CVR.