How does CFSE staining work?
Unlike stains that label the lipid membrane of cells, CellTrace™ CFSE dye easily crosses the plasma membrane and covalently binds inside cells where the stable, well-retained fluorescent dye is designed to provide a consistent signal, even after several days in a cell culture environment.
What does CFSE measure?
CFSE dilution is one of the most commonly used measures of lymphocyte proliferation in immunological studies. The use of CFSE in measuring lymphocyte proliferation began with our studies using the fluorescent dye in lymphocyte migration analyses in mouse models [1].
What is CFSE flow cytometry?
CFSE is a versatile tool for the fluorescent intracellular labeling of live cells. Labeled cells can be assayed using flow cytometry and fluorescent microscopy. The dye is long lasting and well retained within labeled cells. The result is live cells with an intracellular fluorescent label.
What CFSE shows?
As controls, unstimulated CFSE-labeled CD8+ T cells, shows the CFSE intensity of non-divided cells, while the non-labeled cells shows the auto-fluorescence of the cells and the limits of detectable cell divisions.
What does CFSE bind to?
CFSE binds to proteins; stain cells in buffers that have decreased serum concentration (0.1% FBS). Cells which incorporate too much CSFE will have reduced protein function and therefore reduced viability.
Is CFSE toxic to cells?
CFSE can be toxic to cells and non-optimal CFSE labeling conditions can thus hamper proliferation of cells and obscure interpretation of results. While CFSE is commonly used to assess lymphocyte proliferation, CFSE can be toxic and impair cell division.
What Colour is CFSE?
CFSE (Carboxyfluorescein succinimidyl ester) is a membrane-permeable fluorescent dye. As the dye diffuses into the cell, its succinimidyl group covalently binds to intracellular lysine residues and other free primary amines, producing green fluorescence.
Does CFSE stain dead cells?
Thus live cells label with CalcienAM, CFSE, etc. while dead cells do not. It is important to note that unlike cell cycle protocols, this protocol requires that all buffers do not contain detergent or premeabilizing agents (so the PI should be solubilized in PBS/water or the PI will stain all cells.
How long does CFSE last?
CFSE can last several days depending on the level of T cell stimulation and your initial labeling of the T cells. The dye can usually measure 6-7 divisions.