Optimization of loading conditions for a violet dye for use in cell proliferation studies (65.9)

Abstract

Abstract Fluorescein diacetate (FDA) and its derivatives, such as carboxyfluorescein succinimidyl ester (CFSE) are nonfluorescent molecules that diffuse into cells and are hydrolyzed by intracellular non-specific esterases to become fluorescent by-products. These fluorescent dye molecules accumulate only in live cells with intact cell membranes and active esterases, while dead cells remain non fluorescent. The precise kinetics of membrane transport and intracellular hydrolysis of FDA and its analogs are related to cellular functions. FDA dyes have been shown to have multiple uses which include, in vitro or in vivo cell tracking experiments and cell proliferation. BD Horizon™ Violet Proliferation Dye (VPD450) dye contains both an esterase-cleavable and an amine-reactive succinimidyl ester group. This dye can be used in multicolor flow cytometry applications with GFP or FITC-labeled cells. Optimization of cell and dye concentrations are important considerations in experimental design with all FDA-based proliferation dyes. A comparison study was performed to determine the effects VPD450 dye loading on cell proliferation, apoptosis, and cell signaling.