Trypan Blue is an anionic active dye and the most commonly used reagent for identifying dead cells in cell and tissue culture. The staining principle is based on the fact that viable cells possess intact membranes that exclude the dye, preventing its entry; whereas cells that have lost viability or membrane integrity exhibit increased permeability, allowing Trypan Blue to penetrate and stain the cells blue to pale blue. This enables a simple and rapid distinction between live and dead cells. Trypan Blue can also be phagocytosed by macrophages, making it applicable for in vivo macrophage staining.
After staining, cell viability can be quantified by direct microscopic counting or image-based counting, using the formula: Cell Viability (%) = (Total Cells − Blue-Stained Cells) ÷ Total Cells × 100%. The procedure is straightforward, requiring only 3–5 minutes of staining at room temperature.
Note: Prolonged staining should be avoided, as extended incubation may lead to dye uptake by viable cells, resulting in underestimated viability. Trypan Blue is potentially toxic and classified as a suspected carcinogen; appropriate personal protective equipment (lab coat and disposable gloves) should be worn during handling.
For research use only. Not for clinical diagnosis.
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