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BioLegend’s Cell Labeling Webpage

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Cell labeling probes, such as CFSE, Tag-it dyes, and Zombie dyes, are useful in a variety of applications. These dyes may be used as a method of tracking the migration of a cell in vivo, or to track cell proliferation across multiple generations. Proliferation and cell tracking reagents work through multiple mechanisms to label a cell, which has effects on attributes such as retention of the dye in the cell.

Each of these reagents has its own set of applications, so explore this page to learn more about what our cell tracking reagents can do for you.

Certain cell tracking dyes are also useful for cell proliferation assays because the probes are retained for extended periods of time, and will not transfer to other cells after staining. This means that when stained cells divide, the two daughter cells now each have half the dye content of the parent cell. Thus, by tracking the continual reduction in intensity of fluorescence in each cell population, you can monitor the number of cell divisions the original population has gone through.

Long-Term Tracking and Cell Proliferation

In long-term cell tracking and proliferation assays, the probe must be efficiently retained within the cell. When these dyes passively diffuse into cells, esterase activity cleaves acetoxymethyl esters (AM) or diacetates (DA) from the probe, resulting in a fluorescent molecule. Fluorescence allows identification of healthy cells since there is active esterase. In addition to the esterase mechanism, probes like CFDA-SE and Tag-it Violet™ also contain a succinimidyl ester (SE) reactive group that covalently attaches the probe to lysines on intracellular proteins. Once the fluorophore is attached to the protein, it will not be removed from the cell unless the protein is actively transported out. Depending on the length of the experiment or how fast the cells are dividing, additional dye may be required to ensure detection at the assay end point. However, these probes may be toxic at higher concentrations by interfering with protein function, thus careful titration of the reagent is required. For cell tracking applications where cell health is a concern, consider using an alternate method like labeling the cells with the Zombie dyes, where only the cell’s surface is covalently labeled and there is little effect on cell health.