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CAS:148504-34-1
Appearance:Solid
Storage:Powder : -20℃, 1 year;In solvent(mother liquid): -20℃, 1 month; -80℃, 6 months
Purity:≥95%
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Calcein, AM is a cellular staining reagent that fluorescently labels living cells. It penetrates the cell membrane and enters the cell, where it is sheared by intracellular esterases to form calcein, which is retained inside the cell and emits strong green fluorescence. Calcein, AM has low cytotoxicity compared to other similar reagents (e.g. BCECF, AM and CFDA).Excitation/Emission: 494/515 nm.
Calcein, AM only stains living cells. PI, a nuclear dye that cannot cross the membrane of a living cell, crosses the disordered region of a dead cell membrane and reaches the nucleus, where it is embedded in the cell’s DNA double helix to produce red fluorescence (Ex: 535 nm, Em: 617 nm), so PI stains only dead cells. Since both Calcein and PI-DNA can be excited at 490 nm, both living and dead cells can be observed with fluorescence microscopy. With 545 nm excitation, only dead cells were observed.
Protocol (for reference only)
(1) 1 mM Calcein, AM solution was prepared with DMSO, and 1 ~ 50μM Calcein, AM solution was diluted with PBS.
(2) Calcein, AM solution of 1/10 cell medium volume was added to cell medium.
(3) Cultured cells at 37℃ for 15-30 minutes.
(4) Wash the cells twice with PBS or an appropriate buffer.
(5) The cells were observed by fluorescence microscopy with a filter with an excitation wavelength of 490 nm and an emission wavelength of 515 nm.
Attention:
If Calcein, AM is difficult to enter the cell, a surfactant such as Pluronic F127 can be used.
The medium can also be replaced with 1/10 concentration of Calcein, AM solution.
