Calcein,AM

CAS#:148504-34-1 Chinese name: 3'-O-acetamide-2',7'-di(carboxyethyl)-4 or 5-carboxyfluorescein, diacetate English name: 3',6'-Di(O-acetyl)-4',5'-bis[N,N-bis(carboxymethyl)aminomethyl]fluorescein, tetraacetoxy -methyl ester Alias: Calcein Acetylmethylester Structural formula: Molecular formula: C46H46N2O23 Molecular weight: 994.86 Nature: 1. Appearance: White or light yellow crystals 2. Purity: ≥95% (HPLC) 3. Product description: Calcein, AM is a cell staining reagent capable of fluorescently labeling living cells. After penetrating the cell membrane and entering the cell, it is cleaved by intracellular esterases to form Calcein, which is then retained within the cell and emits strong green fluorescence. Compared to other similar reagents (such as BCECF, AM, and CFDA), Calcein, AM exhibits very low cytotoxicity. The excitation and emission wavelengths of Calcein are 490 nm and 515 nm, respectively. Calcein, AM only stains live cells. As a nuclear staining dye, PI cannot penetrate the cell membrane of live cells. It penetrates the disordered region of the dead cell membrane and reaches the nucleus, where it binds to the DNA double helix of the cell, producing red fluorescence (excitation: 535 nm, emission: 617 nm). Therefore, PI only stains dead cells. Since both Calcein and PI-DNA can be excited at 490 nm, live and dead cells can be simultaneously observed using a fluorescence microscope. With excitation at 545 nm, only dead cells can be observed. Based on these characteristics, Calcein, AM, and PI are often combined for dual staining of live and dead cells. Due to the varying optimal staining conditions for different cell lines, we recommend determining the appropriate concentrations of Calcein, AM, and PI individually. 4. Usage (1) Prepare a 1 mM Calcein, AM solution using DMSO, and dilute it with PBS to obtain a Calcein, AM solution ranging from 1 to 50 μM. (2) Add 1/10 of the volume of Calcein, AM solution to the cell culture medium. b) (3) Incubate the cells at 37°C for 15 to 30 minutes. (4) Wash the cells twice with PBS or an appropriate buffer solution. (5) Observe cells using a fluorescence microscope equipped with a filter with an excitation wavelength of 490 nm and an emission wavelength of 515 nm.