Calcein is a fluorescent dye widely used in biological and medical research as a tracer and indicator. It is particularly valuable as a calcium-sensitive probe, where its fluorescence intensity increases upon binding to calcium ions, allowing for the monitoring of intracellular calcium dynamics. In cell viability assays, Calcein, being membrane-impermeant, is excluded by live cells but enters dead or damaged cells with compromised membranes, enabling differentiation based on fluorescence. Additionally, it is employed in tracing studies to assess fluid flow, leakage, or membrane permeability in biological systems, including blood circulation and vascular studies, as well as industrial and environmental applications. Its low toxicity and strong fluorescence properties make it ideal for imaging and diagnostic purposes in microscopy and flow cytometry.