Introduction
Fluorescent dyes absorb light at certain wavelengths and in turn emit their fluorescence energy at a higher wavelength. Each dye has a distinct emission spectrum, which can be exploited for multicolor analysis. eBioscience antibodies are available conjugated to a wide variety of fluorochromes.
The tables below lists the properties of the most commonly used fluorochromes in flow cytometry. The eBioscience product number is the prefix used in our catalog numbering system to indicate the corresponding formats of our antibodies. We are continually developing new fluorochrome conjugated antibodies for both flow cytometry and microscopy. Please feel free to contact us at tech@ebioscience.com for more information!
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Fluorescent Dyes for Flow Cytometric Analysis - 488nm Excitation
|
| eBioscience Product Prefix # |
Name |
Excitation laser (nm) |
Peak Emission (nm) |
Relative Intensity (1=lowest, 5=highest) |
Uses |
| 11
|
FITC (Fluorescein isothiocyanate)
|
488
|
518
|
3
|
Detected in FL1 channel on most instruments; also a good choice for fluorescence microscopy
|
| 53
|
Alexa FluorŽ 488
|
488
|
519
|
3
|
Detected in FL1 channel on most instruments; photostable; an excellent choice for fluorescence microscopy
|
| 12
|
R-PE
(R-Phycoerythrin)
|
488
|
575
|
5
|
Detected in FL2 channel on most instruments; very high quantum yield (ideal for detection of low density antigens); subject to photobleaching.
|
| -
|
PE-Texas Red
|
488
|
615
|
3
|
Detected in FL3 on single laser instruments
|
| -
|
PE-Alexa FluorŽ 610
|
488
|
628
|
3
|
This alternative to PE-Texas Red has less overlap with PE due to its 628nm emission wavelength and is less subject to Fc mediated binding than PE-Cy5.
|
| 15
|
PE-Cy5
|
488
|
670
|
4
|
Detected in FL3 or FL4 on most instruments; tandem dye, resonance energy transfer from PE molecule to the cyanine dye Cy5; when used with APC on dual laser machines, needs a cytometer capable of interlaser compensation. See PE-Cy5.5 for dual laser instruments
|
| 35
|
PE-Cy5.5
|
488
|
690
|
3
|
Detected in FL3 or FL4 on most instruments; tandem dye, resonance energy transfer from PE molecule to cyanine dye Cy5.5; when used with APC on dual laser machines, needs a cytometer capable of interlaser compensation. Better choice for dual laser instruments than PE-Cy5.
|
| 45
|
PerCP-Cy5.5
|
488
|
690
|
3
|
Detected in FL3 or FL4 on most instruments; tandem dye, resonance energy transfer from PerCP molecule to cyanine dye Cy5.5; when used with APC on dual laser machines, needs a cytometer capable of inter-laser compensation. Better choice for dual laser instruments than PE-tandems as a result of its large stokes shift.
|
| 46
|
PerCP-eFluorŽ 710
|
488
|
710
|
4
|
Detected in FL3 on most instruments; tandem dye, resonance energy transfer from PerCP molecule to eFluorŽ 710; when used with APC on dual laser machines, needs a cytometer capable of inter-laser compensation. Better choice for dual laser instruments than PE-tandems as a result of its large stokes shift. 2-3 fold brighter than PerCP-Cy5.5.
|
| 25
|
PE-Cy7
|
488
|
760
|
3
|
Detected in appropriate channel based on filter set arrangement of the instrument; tandem dye, resonance energy transfer from PE molecule to cyanine dye Cy7; minimal cross-beam compensation required when used simultaneously
|
| -
|
7-AAD
|
488
|
647
|
|
Detected in FL3 channel on most instruments; binds selectively to GC regions of DNA; used as a viability dye to discriminate between viable and non-viable cells
|
| -
|
Propidium Iodide (PI)
|
488
|
617
|
|
Detected in FL2 and FL3 channels on most instruments; a DNA intercalating dye; used as a viability dye to discriminate between viable and nonviable cells
|
|
Fluorescent Dyes for Flow Cytometric Analysis - 633nm Excitation
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| eBioscience Product Prefix # |
Name |
Excitation laser (nm) |
Peak Emission (nm) |
Relative Intensity (1=lowest, 5=highest) |
Uses |
| 17
|
Allophycocyanin (APC)
|
633
|
660
|
4
|
Detected in FL4 on dual laser instruments; can be used interchangeably with Cy5 or Alexa FluorŽ 647
|
| 50
|
eFluorŽ 660
|
633
|
660
|
4
|
eFluorŽ 660 is a replacement for Alexa FluorŽ 647. Detected in FL4 on dual laser instruments; can be used interchangeably with APC; photostable, making it a great choice for microscopy, in addition to flow cytometry.
|
| 51
|
Alexa FluorŽ 647
|
633
|
668
|
4
|
Detected in FL4 on dual laser instruments; can be used interchangeably with Cy5 or APC; photostable, making it a great choice for microscopy, in addition to flow cytometry
|
| 19
|
Cy5
|
633
|
670
|
3
|
Detected in FL4 on dual laser instruments (used interchangeably with APC or Alexa FluorŽ 647); good choice for intracellular labeling and microscopy
|
| -
|
APC-Cy5.5
|
633
|
690
|
3
|
Detected in appropriate channel based on filter set arrangement; tandem dye, resonance energy transfer from APC molecule to Cy5.5
|
| 56
|
Alexa FluorŽ 700
|
633
|
723
|
2
|
Although not optimally excited by the He-Ne laser, it does produce a bright signal that can be used as a third dye off this laser; please note that emission wavelength requires a distinct set of filters; most instruments will require a 685 LP mirror and 710/20 filter.
|
| 47
|
APC-eFluorŽ 780
|
633
|
780
|
2
|
Appropriate channel for detection based on filter set arrangement of the instrument; more photostable than APC-Cy7; tandem dye, resonance energy transfer from APC molecule to eFluorŽ 780, APC-Cy7, APC-Alexa FluorŽ 750 and APC-eFluorŽ 780 may be used interchangeably.
|
| 27
|
APC-Alexa FluorŽ 750
|
633
|
775
|
2
|
Appropriate channel for detection based on filter set arrangement of the instrument; more photostable than APC-Cy7; tandem dye, resonance energy transfer from APC molecule to Alexa FluorŽ 750. APC-Cy7, APC-Alexa FluorŽ 750 and APC-eFluorŽ 780 may be used interchangeably.
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|
Fluorescent Dyes for Flow Cytometric Analysis - 405nm Excitation
|
| eBioscience Product Prefix # |
Name |
Excitation laser (nm) |
Peak Emission (nm) |
Relative Intensity (1=lowest, 5=highest) |
Uses |
| 93
|
eFluorŽ 605NC
|
405
|
605
|
2
|
Excited by the violet laser; eFluorŽ 605NC can be used in combination with eFluorŽ 450, please make sure to use this conjugate for your brightest antigen/proteins of interest
|
| 94
|
eFluorŽ 625NC
|
405
|
625
|
2
|
Excited by the violet laser; eFluorŽ 625NC can be used in combination with eFluorŽ 450, please make sure to use this conjugate for your brightest antigen/proteins of interest
|
| 95
|
eFluorŽ 650NC
|
405
|
650
|
3
|
Excited by the violet laser; eFluorŽ 650NC can be used in combination with eFluorŽ 450 and eFluorŽ 605NC, please make sure to use this conjugate for your brightest antigen/proteins of interest.
|
| 48
|
eFluorŽ 450
|
405
|
450
|
2
|
Excited by the violet laser; eFluorŽ 450 can be used interchangeably with Pacific BlueŽ, can be used in combination with eFluorŽ 605NC or eFluorŽ 625NC; please make sure to use this conjugate for your brightest antigen/proteins of interest
|
| -
|
Alexa FluorŽ 405
|
405
|
421
|
1
|
Extremely dim dye excited by the violet laser; suitable for only the brightest antigens
|
| 57
|
Pacific BlueŽ
|
405
|
455
|
2
|
Excited by the violet laser; Pacific BlueŽ can be used interchangeably with eFluorŽ 450, typically a dimmer fluorochrome; please make sure to use this conjugate for your brightest antigen/proteins of interest
|
| -
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Alexa FluorŽ 430
|
405
|
541
|
1
|
Can be used as second dye excited by the violet laser
|
| -
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Pacific OrangeŽ
|
405
|
551
|
1
|
Dim and only suitable for densely expressed targets, due to its 551nm emission wavelength the Pacific OrangeŽ dye can be used with Pacific BlueŽ for multicolor analysis with the violet laser, however eFluorŽ 605NC is a better alternative
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Cy-ChromeTM is a registered trademark of Becton, Dickinson and Company. Tri-ColorTM is a registered trademark of Caltag Laboratories. Alexa FluorŽ and Pacific BlueŽ are registered trademarks of and licensed under patents assigned to Molecular Probes, Inc., for research use only.
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