The apoptosis, necrosis and cell viability assays are designed to stain dissociated cells in culture and have not been validated for organ culture. Annexin V staining of early chicken and mammalian embryos in culture has been reported in the scientific literature. For staining of living tissues, the specimen would need to be thin enough to allow exposure of the cells to the 36 kDa Annexin V protein. Also, damage to cell membranes from dissection or sectioning of tissues could result in high background staining.
No. Fixation, freezing, sectioning, or dissociation of tissues can affect the PS on the outer leaflet and compromise membrane integrity. Both Annexin V and Ethidium Homodimer III rely upon the presence of intact membranes in healthy cells to accurately distinguish healthy cells from apoptotic or necrotic cells. To detect apoptosis in fixed cells and tissues we recommend our TUNEL kits. Currently we are not aware of a fluorescent probe that specifically detects necrotic cells in fixed tissues. Necrotic cells in tissue sections are identified based on morphological criteria.
Yes, cells can be fixed with formaldehyde after staining. Because Annexin V staining is dependent on calcium, all buffers used for washing and fixation should contain 1.25 mM CaCl2. Fixation may increase the background signal from Ethidium Homodimer III. Wash the cells several times to remove unbound Ethidium Homodimer III before fixation.
ViaFluor® SE Cell Proliferation Kits use amine-reactive dyes to covalently label cells throughout the cell cytoplasm and intracellular compartments for fixable fluorescent staining. Cell proliferation dyes are commonly used to monitor cell division by flow cytometry. The dyes also can be used to stably label cells to image cell morphology, or to track cell populations in mixed co-culture experiments.
Non-toxic dyes covalently label cell cytoplasm for fixable staining
Track cell proliferation in vivo or in vitro by dye dilution using flow cytometry
Long term imaging of cell morphology or co-cultures by microscopy
Excellent performance & lower cost compared to leading competitors
ViaFluor® 405 & ViaFluor® 488 SE are much less toxic than CFSE
Lyophilized dye in single use vials
Anhydrous DMSO for dissolving dye
Spectral Properties (Ex/Em after hydrolysis)
ViaFluor® CFSE: 495/515 nm
ViaFluor® 488 SE: 493/532 nm
ViaFluor® 405 SE: 408/452 nm
ViaFluor® SE Dyes for Cell Proliferation Tracking
ViaFluor® SE dyes are membrane permeable compounds that are non-fluorescent until they enters viable cells, where the compound is hydrolyzed by cytoplasmic esterase enzymes to releases the fluorescent amine-reactive dye. The dyes then covalently react with amine groups on intracellular proteins, forming fluorescent conjugates that are retained in the cell, while excess unreacted dye is washed away. Immediately after staining, a single, bright fluorescent population will be detected by flow cytometry. With each cell division, daughter cells inherit roughly half of the fluorescent label, allowing the number of cell divisions that occur after labeling to be detected by the appearance of successively dimmer fluorescent peaks on a flow cytometry histogram compared to cells analyzed immediately after staining. Thus, cell proliferation dyes can be used to track multiple cell divisions of cells grown in culture or injected in vivo after labeling with the ViaFluor® SE dye.
The number of assays that can be performed per kit depends on the dye concentration used (see the product protocol for more information). When used at 1 uM to label 106 cells in one mL, each dye vial can be used for 90-100 labelings.
All three ViaFluor® SE dyes can stain gram-positive bacteria, but not gram-negative bacteria. ViaFluor® CFSE stains the cytoplasm in yeast, but ViaFluor® 405 & ViaFluor® 488 stain the yeast cell periphery. See our Cellular Stains Table for more information on how our dyes stain various organisms.
Kits to covalently label dead cells, allowing cells with permeable plasma membranes to be excluded from analysis in flow cytometry. A wide variety of dye options for standard flow, spectral flow, and fluorescence microscopy.