Cell Tracking / Labeling
|
Dye |
Excitation |
Emission |
Function |
Benefits |
Limitations |
Links |
CFSE
|
494 nm
|
521 nm
|
Labels cytoplasmic proteins
|
• Good for short- & long-term labeling of cells (days – weeks) for flow cytometry or microscopy
• Can be used to track cell division history (see Cell Proliferation Dyes)
|
• Labeling cells with too much dye may inhibit function & significantly increases compensation values (flow cytometry)
• Cannot be used in combination with FITC, Alexa Fluor® 488, or GFP
|
65-0850
|
Cell Proliferation Dye eFluor® 670
|
647 nm
|
670 nm
|
Labels cellular proteins
|
• Good for short- & long-term labeling of cells (days – weeks) for flow cytometry or microscopy
• Can be used to track cell division history (see Cell Proliferation Dyes)
• Compatible with GFP
|
• Labeling cells with too much dye may inhibit function & significantly increases compensation values (flow cytometry)
• Cannot be used in combination with APC or Alexa Fluor® 647
|
65-0840
|
CellVue® Jade
|
478 nm
|
508 nm
|
Labels cell membrane
|
• Good for short term labeling of cells (1-7 days) for flow cytometry or microscopy
|
• Labeling cells with too much dye or for too long will kill cells
• Dye can be transferred to other cells through trogocytosis
• Does not divide evenly between daughter cells for tracking cell division
• Cannot be used in combination with FITC, Alexa Fluor® 488, or GFP
|
88-0876
|
CellVue® Lavender
|
420 nm
|
461 nm
|
Labels cell membrane
|
• Good for short term labeling of cells (1-2 days) for flow cytometry or microscopy
|
• The dimmest of the CellVue dyes
• Labeling cells with too much dye or for too long will kill cells
• Dye can be transferred to other cells through trogocytosis
• Does not divide evenly between daughter cells for tracking cell division
• Cannot be used in combination with eFluor® 450 or Pacific Blue®
|
88-0873
|
CellVue® Maroon
|
647 nm
|
667 nm
|
Labels cell membrane
|
• Good for short term labeling of cells (1-7 days) for flow cytometry or microscopy
|
• Labeling cells with too much dye or for too long will kill cells
• Dye can be transferred to other cells through trogocytosis
• Does not divide evenly between daughter cells for tracking cell division
• May not be compatible with other 633 nm-excitable fluorochromes
|
88-0870
|
CellVue® Burgundy
|
683 nm
|
707 nm
|
Labels cell membrane
|
• Good for short term labeling of cells (1-7 days) for flow cytometry or microscopy
|
• Labeling cells with too much dye or for too long will kill cells
• Dye can be transferred to other cells through trogocytosis
• Does not divide evenly between daughter cells for tracking cell division
• May not be compatible with other 633 nm-excitable fluorochromes
|
88-0872
|
CellVue® Plum
|
652 nm
|
671 nm
|
Labels cell membrane
|
• Good for short term labeling of cells (1-7 days) for flow cytometry or microscopy
|
• Labeling cells with too much dye or for too long will kill cells
• Dye can be transferred to other cells through trogocytosis
• Does not divide evenly between daughter cells for tracking cell division
• May not be compatible with other 633 nm-excitable fluorochromes
|
88-0871
|
CellVue® NIR780
|
745 nm
|
776 nm
|
Labels cell membrane
|
• Good for short term labeling of cells (1-7 days) for flow cytometry (can be excited by the red laser line) or microscopy
|
• Labeling cells with too much dye or for too long will kill cells
• Dye can be transferred to other cells through trogocytosis
• Does not divide evenly between daughter cells for tracking cell division
• Cannot be used in combination with APC-eFluor® 780 or APC-Alexa Fluor® 750
|
88-0875
|
CellVue® NIR815
|
786 nm
|
814 nm
|
Labels cell membrane
|
• Good for short term labeling of cells (1-7 days) for in vivo imaging
|
• Labeling cells with too much dye or for too long will kill cells
• Dye can be transferred to other cells through trogocytosis
|
88-0874
|
Cell Proliferation / Cell Division
|
Dye |
Excitation |
Emission |
Function |
Benefits |
Limitations |
Links |
CFSE
|
494 nm
|
521 nm
|
Labels cytoplasmic proteins
|
• Can identify up to 8 individual cell divisions as dye is divided equally between daughter cells
|
• Labeling cells with too much dye may inhibit function & significantly increases compensation values
• Cannot be used in combination with FITC, Alexa Fluor® 488, or GFP
|
65-0850
|
Cell Proliferation Dye eFluor® 670
|
647 nm
|
660 nm
|
Labels cellular proteins
|
• Can identify up to 6 individual cell divisions as dye is divided equally between daughter cells
• Compatible with GFP
|
• Labeling cells with too much dye may inhibit function & also significantly increases compensation values
• Cannot be used in combination with APC or Alexa Fluor® 647
|
65-0840
|
Cell Viability Dyes
|
Dye |
Excitation |
Emission |
Function |
Benefits |
Limitation |
Links |
Fixable Viability Dye eFluor® 450
|
405 nm
|
450 nm
|
Labels dead cells
|
• Permanently labels dead cells allowing them to be excluded from analysis even when the cell preparation is being analyzed by flow cytometry for intracellular targets
|
• Cannot be used in combination with Pacific Blue® or eFluor® 450
|
65-0863
|
Fixable Viability Dye eFluor® 660
|
633 nm
|
660 nm
|
Labels dead cells
|
• Permanently labels dead cells allowing them to be excluded from analysis even when the cell preparation is being analyzed by flow cytometry for intracellular targets
|
• Cannot be used in combination with APC or Alexa Fluor® 647
|
65-0864
|
Fixable Viability Dye eFluor® 780
|
633 nm
|
780 nm
|
Labels dead cells
|
• Permanently labels dead cells allowing them to be excluded from analysis even when the cell preparation is being analyzed by flow cytometry for intracellular targets
|
• Cannot be used in combination with APC-eFluor® 780 or APC-Alexa Fluor® 750
|
65-0865
|
Calcein AM
|
495 nm
|
515 nm
|
Labels live cells
|
• Can be used to identify live cells by flow cytometry
• Non-fluorescent compound freely enters live cells and intracellular esterases converts it to a fluorescent dye
• Measures intracellular esterase activity in live cells
• Is not retained in dead cells with compromised cell membranes
|
• Will not be retained in cells after fixation or permeabilization and therefore, is not useful for intracellular staining protocols
• High concentrations of dye will require greater compensation out of PE, PerCP-Cy5.5, and PE-Cy7 channels
• May require co-staining with Annexin, 7-AAD, or propidium iodide to optimally separate live/dead cells
• Cannot be used in combination with FITC, Alexa Fluor® 488, or GFP
|
65-0853
|
Calcein Violet AM
|
408 nm
|
450 nm
|
Labels live cells
|
• Can be used to identify live cells by flow cytometry
• Excited by the violet laser line
• Non-fluorescent compound freely enters live cells and intracellular esterases converts it to a fluorescent dye
• Measures intracellular esterase activity in live cells
• Is not retained in dead cells with compromised cell membranes
|
• Will not be retained in cells after fixation or permeabilization and therefore, is not useful for intracellular staining protocols
• Cannot be used in combination with Pacific Blue® or eFluor® 450
|
65-0854
|
Calcein Blue AM
|
360 nm
|
445 nm
|
Labels live cells
|
• Can be used to identify live cells by flow cytometry
• Excited by the UV laser line
• Non-fluorescent compound freely enters live cells and intracellular esterases converts it to a fluorescent dye
• Measures intracellular esterase activity in live cells
• Is not retained in dead cells with compromised cell membranes
|
• Will not be retained in cells after fixation or permeabilization and therefore, is not useful for intracellular staining protocols
|
65-0855
|
Propidium Iodide
|
488 nm
|
617 nm
|
Labels dead cells
|
• Allows exclusion of dead cells from flow cytometric analysis
• Enters cells with compromised cell membranes and binds dsDNA/RNA
• Cannot pass through intact cell membranes
• May also be used for DNA content/cell cycle analysis (see DNA content, Cell cycle analysis, Identification of nucleated cells)
|
• Will not be retained in cells after fixation or permeabilization and therefore, is not useful for intracellular staining protocols
|
00-6990
|
7-AAD
|
548 nm
|
647 nm
|
Labels dead cells
|
• Allows exclusion of dead cells from flow cytometric analysis
• Enters cells with compromised cell membranes and binds dsDNA
• Cannot pass through intact cell membranes
• May also be used for DNA content/cell cycle analysis (see DNA content, Cell cycle analysis, Identification of nucleated cells)
|
• Will not be retained in cells after fixation or permeabilization and therefore, is not useful for intracellular staining protocols
|
00-6993
|
DNA Content, Cell Cycle Analysis, and Identification of Nucleated Cells
|
Dye |
Excitation |
Emission |
Function |
Benefits |
Limitations |
Links |
Nuclear RED (DRAQ5™)
|
488-647 nm
|
>670 nm
|
Labels dsDNA
|
• Binds DNA stoichiometrically for analysis of cell cycle and ploidy
• Does not require fixation/ permeabilization
• Can be used as a nuclear counterstain for microscopy
• Can be used to distinguish nucleated/nonnucleated cells for flow cytometry
|
• Broad excitation / emission spectra may limit use in multi-color flow cytometry & may not be compatible with PerCP-eFluor® 710/PerCP-Cy5.5, PE-Cy7, APC/Alexa Fluor® 647, Alexa Fluor® 700, or APC-eFluor® 780/APC-Alexa Fluor® 750
• For cell cycle analysis, emission should be collected with far-red filters (780/60) to obtain optimal CVs
• For cell cycle analysis, it may be necessary to label cells for a longer period of time and with a higher concentration to obtain optimal CVs
|
65-0880
|
Nuclear ORANGE (CyTRAK Orange™)
|
488-550 nm
|
610 nm
|
Labels dsDNA
|
• Binds DNA
• Does not require fixation/ permeabilization
• Can be used to distinguish nucleated and nonnucleated cells for flow cytometry
|
• Background staining of the cytoplasm may limit its use as a nuclear counterstain in fluorescent microscopy
|
65-0881
|
Propidium Iodide
|
488 nm
|
617 nm
|
Labels dsDNA/RNA
|
• Binds DNA/RNA stoichiometrically for analysis of cell cycle and ploidy
• May also be used for live/dead discrimination in flow cytometry
|
• Must fix/permeabilize cells
• Requires degradation of cellular RNA for DNA/cell cycle analysis
• Has a broad emission spectra making it difficult to use in multi-color flow cytometric applications
|
00-6990
|
7-AAD
|
548 nm
|
647 nm
|
Labels dsDNA
|
• Specifically binds DNA (does not bind to RNA) stoichiometrically for analysis of cell cycle and ploidy
• May also be used for live/dead discrimination in flow cytometry
|
• Must fix/permeabilize cells
|
00-6993
|
Calcium Flux
|
Dye |
Excitation |
Emission |
Function |
Benefits |
Limitations |
Links |
eFluor® 514 Calcium Sensor Dye
|
490 nm
|
514 nm
|
Fluorescence intensity increases upon binding free Ca2+
|
• Allows qualitative/semi-quantitative measurements of intracellular free Ca2+
• Compatible with flow cytometry, fluorescence microscopy, microplate readers, and spectrophotometry
• Better cellular uptake and brighter than Fluo-3 or Fluo-4
|
• Only semi-quantitatve
|
65-0859
|
Indo-1 AM
|
346 nm
|
410 nm (bound) 485 nm (unbound)
|
Peak emission decreases upon binding free Ca2+
|
• Allows quantitative measurement of free Ca2+
• Compatible with flow cytometry, fluorescence microscopy, microplate readers, and spectrophotometry
• Ratiometric measurements possible
|
• Compensation between bound/ unbound emission may be difficult
• Not compatible with eFluor® 450, 605NC, 625NC, 650 NC, or other violet laser-excitable dyes
|
65-0856 65-0857
|
Fura-2 AM
|
300 nm (bound)
400 nm (unbound)
|
510 nm
|
Peak excitation decreases upon binding free Ca2+
|
• Ideal for measuring low to medium concentrations of Ca2+ by spectrophotometry or microscopy (when specific excitiation wavelengths can be used)
• Compatible with ratiometric imaging microscopy
|
• Not for use with flow cytometry
• Kd is highly sensitive to changes in pH, temperature, ionic strength and viscosity of the cytosol
|
65-0858
|
Mitochondrial Membrane Potential
|
Dye |
Excitation |
Emission |
Function |
Benefits |
Limitations |
Links |
JC-1
|
515 nm
|
530 nm
(monomer/ depolarized)
590 nm
(aggregates/ polarized)
|
Measures polarization/ depolarization of mitochondrial membrane
|
• Selectively enters mitochondria & can be used to indicate the initiation of apoptosis
• Changes in fluorescence is reversible
• Compatible with flow cytometry and fluorescent spectrophotometry
• Provides qualitative and quantitative measurements
|
• Compensation between the monomer and aggregate emission can be difficult
|
65-0851
|
|