CellTrace™ Violet easily diffuses into cells, where it is cleaved by intracellular esterases to yield a highly fluorescent compound. This compound covalently binds to intracellular amines, resulting in stable, well-retained fluorescent staining that can be fixed with aldehyde fixatives. Excess unconjugated reagent diffuses to the extracellular medium, where it can be quenched with complete medium and washed away.
Figure 1. Bright, well-retained dye that partitions evenly between daughter cells facilitates proliferation analysis. (Left) Illustration of proliferation analysis by dye dilution. (Right) Flow cytometric analysis reveals a bright, homogeneous fluorescent signal from the initial population of cells. Subsequent cell divisions result in larger numbers of cells, each with half the fluorescence intensity of its parent cell.
Superior Fluorescent Staining
Long-Term Signal Stability
Simple, Robust Staining Protocol
Easy Multiplexing With Other Fluorophores
CellTrace™ Violet can be used in multicolor flow cytometry experiments to precisely track the proliferation of cells in culture while collecting other data. To demonstrate this application, the superior resolution of the new Attune™ Acoustic Focusing Cytometer was used in a multicolor experiment with CellTrace™ Violet to track ten generations of proliferating lymphocytes (Figure 2). For more information on the Attune™ cytometer, read the article on The Attune™ Acoustic Focusing Cytometer.
Learn more about the CellTrace™ Violet Cell Proliferation Kit at CellTrace™ Reagents for Cell Proliferation.
Figure 2. Multiparameter proliferation analysis of human lymphocytes. Peripheral blood mononuclear cells were isolated from whole blood, stained with 10 µM CellTrace™ Violet, and stimulated with mouse anti–human CD3 and interleukin-2 for 7 days in culture. Cells were stained with SYTOX® AADvanced™ Dead Cell Stain and mouse anti–human CD4 Alexa Fluor® 488 immediately prior to analysis on the Attune™ Acoustic Focusing Cytometer. A gating strategy was employed to limit analysis to live cells (A), single cells (B), and CD4+ cells (C). When the resulting population of cells is displayed on a histogram of CellTrace™ Violet fluorescence intensity (D), each peak represents one generation of proliferating cells. This fluorescence histogram can be further analyzed using proliferation modeling software (ModFit LT™, Verity Software House) to provide statistics and uniquely identify each generation of cells with a different peak color (E).
- Learn More about CellTrace™ Reagents for Cell Proliferation