Bright and stable fluorophores for multiparameter cell analysis

  • Bright mean fluorescence intensity for excellent signals
  • High stain index for clear population discrimination
  • Hassle-free multicolor experiments due to low spillover

New fluorophores for the VioDye portfolio

VioDyes family

Vio® and Vio® Bright Dyes represent a family of fluorochromes for flow cytometry and fluorescence microscopy. They are characterized by high fluorescence intensities and low spillover, making them an ideal choice for multicolor applications.

Combined with traditional fluorochromes, such as FITC, PE, PerCP, and APC, they expand available dye options, giving you more flexibility in setting up multiparameter cell analysis.

What is the excitation and emission data of the fluorophores?

Whether you perform flow cytometry, flow sorting, or microscopy, our portfolio of fluorophores – covering wavelengths from 400 nm to 800 nm – enables you to do comprehensive multicolor analysis.

Our Vio Dye family of fluorophores includes both single fluorochromes and tandem dyes. In addition various traditional fluorophores are available to allow for fluorescence analysis over a wide range of the visual spectrum. 

Fluorescent dye

Excitation (nm)

Exmax (nm)

Emmax (nm)

Violet laser fluorochromes
VioBlue® 405400452
VioGreenTM405388520
Vio® Bright V423405399420
Blue laser tandem dyes
Vio® B515488488514
Vio® Bright B515488488514
Vio® Bright FITC488496522
FITC488495520
Blue and yellow-green laser dyes
PE-Vio® 615488 or 561565619
PE-Vio® 770488 or 561565775
PE488 or 561565578
PerCP-Vio® 700488482704
PerCP488482675
Red laser fluorochromes
Vio® R667561 or 635 or 640645668
Vio® Bright R667561 or 635 or 640645668
Vio® R720640695720
Vio® Bright R720640695720
APC-Vio® 770561 or 635 or 640652775
APC561 or 635652660

What are the characteristics and applications of the different fluorophores?

To help you plan your experiment properly, the table below summarizes important characteristics of all Vio Dye fluorophores that are available for Miltenyi Biotec antibodies. 

Characteristics of fluorescent dyes

Fluorophore

Brightness

Fixation stability

Photostability

MACSQuant Analyzer Detection filter (nm)

Microscopy

Violet laser fluorescent dyes
VioBlue3PFA, ethanol, methanol, and acetoneMedium450/50Not tested
VioGreen1PFA, ethanol, methanol, and acetoneHigh525/50Not tested
Vio Bright V4235PFAHigh450/50Not tested
Blue laser fluorochromes
Vio B5154PFA, ethanol, methanol, and acetoneVery high525/50Recommended
Vio Bright B5155PFA, ethanol, methanol, and acetoneMedium525/50Suitable
Vio Bright FITC4PFA, ethanol, methanol, and acetoneLow525/50Suitable
Blue and yellow-green laser tandem dyes
PE-Vio 6155PFANot tested655-730 or 615/20Suitable
PE-Vio 7704PFANot tested750 LP or 667/30Not tested
PerCP-Vio 7003PFANot tested655-730 or 615/20 
Red laser fluorescent dyes
APC-Vio 7703PFANot tested750 LP or 725/40Not tested
Vio R667      5PFA, ethanol, methanol, and acetoneVery high655-730 or 661/20 or 667/30Recommended
Vio Bright R667  5+PFA, ethanol, methanol, and acetoneMedium655-730 or 661/20 or 667/30Suitable
Vio R7203PFAHigh725/40Not tested
Vio Bright R7204PFAHigh725/40Not tested

Violet laser (405 nm)

Vio Bright V423 (Emmax 420 nm)

Vio Bright V423 is Miltenyi Biotec’s brightest dye, excited at 405 nm. In addition to high brightness, this dye shows a significantly improved signal-to-noise ratio, minimal intra-laser compensation, and a superior detection of rare antigens. 
Vio Bright V423 is a synthetic dye which is 5× brighter than VioBlue Dye, making it an excellent choice for multiparameter cell analysis. 
 

Vio Bright V423 at a glance:

  • Is the brightest Vio Dye for the violet laser.
  • Provides a bright alternative for the confident detection of rare cells, as well as dim and uncharacterized markers.
  • Shows minimal spillover to other detection channels within the violet laser, thereby minimizing compensation effort.
  • Is available in an extensive portfolio conjugated to recombinantly engineered REAfinity™ Antibodies for higher reproducibility.

Excitation and emission maxima of fluorochromes related to Vio Bright V423.

FluorochromeExcitation max. (nm)Emission max. (nm)
Vio Bright V423399 420
Brilliant Violet 421 (BD)408425
Brilliant Violet 421 (Biolegend)408424
Super Bright 436415434

VioBlue Dye (Emmax 452 nm)

Designed to maximize the potential of a flow cytometer’s violet laser, the VioBlue Dye shows superior performance compared to many other fluorophores excited at 405 nm, including significant advances in brightness, signal-to-noise ratios, and intralaser compensation requirements. In addition, VioBlue exhibits minimal photo-induced degradation, and can consequently be used for many different applications, such as fluorescence microscopy.
 

VioBlue Dye at a glance:

  • Coumarin-based dye with excitation and emission wavelengths of 400 nm and 455 nm, respectively.
  • Brighter alternative to Pacific Blue™, Alexa Fluor® 405, or BD Horizon™ V450.
  • Multiplexing of VioBlue with other fluorochromes is easily possible, adding to the variety of marker combination for multiparameter flow cytometry.
  • Combined use with the VioGreen Dye.

For detailed information including stability, spillover, and brightness data, please visit the MACS Handbook.
 

Excitation and emission maxima of fluorochromes related to VioBlue.

FluorochromeExcitation max. (nm)Emission max. (nm)
VioBlue    400455
Pacific Blue405455
Cascade Blue(375); 401423
Alexa Fluor 405405421
eFluor 405405450
BD Horizon V450404448

VioGreen Dye (Emmax 520 nm)

Designed to maximize the potential of a flow cytometer’s violet laser, the VioGreen Dye shows superior performance compared with many other fluorophores excited at 405 nm, including significant advances in brightness, signal-to-noise ratios, and intralaser compensation requirements. In addition, VioGreen exhibits minimal levels of photo-induced degradation, and can consequently be used for many different applications, such as fluorescence microscopy.

VioGreen Dye at a glance:

  • Large Stokes shift fluorophore, emitting strong fluorescence at 520 nm upon excitation at 405 nm.
  • Significantly increased mean fluorescence intensities and higher stain indices than Pacific Orange™, Krome Orange™, and BD Horizon™ V500.
  • Non-protein fluorophore.
  • Combined use with the VioBlue Dye.
  • Perfectly suited for multiparameter flow cytometry using all current flow cytometers.

For detailed information including stability, spillover, and brightness data, please visit the MACS Handbook.
 

Excitation and emission maxima of fluorochromes related to VioGreen.

FluorochromeExcitation max. (nm)Emission max. (nm)
VioGreen388520
Pacific Orange400551
Krome Orange398528
AmCyan458489
HorizonV500415500

Blue laser (488nm) 

VioBright B515 and Vio B515 (Emmax 514 nm)

Vio Bright B515 and Vio B515 are extremely bright dyes for the B1/FITC-channel. The proprietary Vio Bright multimerization technology results in a 4-fold increase in brightness over the traditional FITC dye. With a 25% reduced spill-over into the B2/PE-channel and high photo stability, these dyes are excellent choices for immunofluorescence microscopy applications.

Vio Bright B515 at a glance:

  • Brightest Vio Dye for the blue laser.
  • Excellent choice for detection of surface markers via flow cytometry, in particular for low-expressed antigens.

Vio B515 at a glance:

  • Brighter alternative to FITC.
  • Perfectly suited for detection of intracellular markers via flow cytometry, such as cytokines and transcription factors.
  • Optimal for use in fluorescence microscopy applications with higher photostability compared to Alexa Fluor 488.

For detailed information including stability, spillover, and brightness data, please visit the MACS Handbook.
 

Vio Bright FITC (Emmax 522 nm)

Vio Bright FITC is a blue laser–excited (488 nm) revolutionary dye, which allows binding of an increased number of FITC molecules per antibody, as compared to conventional FITC conjugation. With brightness similar to PE, Vio Bright FITC expands the dimensions of multicolor flow analysis. In addition, it provides a bright alternative for confident detection of rare cells, as well as dim and uncharacterized markers.

VioBright FITC at a glance:

  • Brightness similar to PE for confident detection of low-expressed and rare markers.
  • Excellent bright alternative to PE for flexible multicolor panel design.
  • Signal detectable in standard FITC channel with peak excitation and emission wavelength of 496 nm and 522 nm, respectively.
  • Works with your standard staining protocol.

For detailed information including stability, spillover, and brightness data, please visit the MACS Handbook.
 

Excitation and emission peaks of fluorochromes compared to Vio Bright B515, Vio B515, and Vio Bright FITC.
 

FluorochromeExcitation max. (nm)Emission max. (nm)
Vio Bright B515488514
Vio B515488514
VioBright FITC496522
FITC495520
Alexa Fluor 488495519
BD Horizon Brilliant Blue 515490515

PerCP-Vio 700 (Emmax 704 nm)

The PerCP-Vio 700 Dye is a tandem conjugate that combines the peridinin chlorophyll protein (PerCP) and the new Vio 700 Dye to emit a strong fluorescence at 655–730 nm upon blue laser excitation at 488 nm.  This dye is suited perfectly for the B3 channel of the MACSQuant Analyzer.
 

For detailed information including stability, spillover, and brightness data, please visit the MACS Handbook.
 

Excitation and emission maximums of fluorochromes comparable to PerCP-Vio 700.
 

FluorochromeExcitation max. (nm)Emission max. (nm)
PerCP482675
PerCP-Vio 700482704
PerCP-Cy5.5490695

Blue laser (488 nm) / yellow laser (561 nm) 

PE-Vio 770 (Emmax 770 nm)

The PE-Vio 770 Dye is a tandem conjugate, like PE-Cy™7, that exploits the principle of fluorescence-resonance-energy-transfer (FRET) allowing for large Stokes shifts between the absorbed energy of a fluorescence donor (PE) and the emission wavelength of a suitable acceptor (Vio 770) dye.

PE-Vio 770 Dye at a glance:

  • Excitation with the blue (488 nm) or yellow (561 nm) laser, emission in the near-infrared region at 775 nm.
  • The combination of Vio 770 as the acceptor dye and an optimized chemistry have furnished a tandem dye that is characterized by a high fluorescence intensity, minimal spillover to adjacent detection channels, and low non-specific binding to non-target cells to meet the complexity of multiparameter flow cytometry.
  • Higher MFI and stain index values, in addition to lower compensation settings when compared to PE-Cy7

For detailed information including stability, spillover, and brightness data, please visit the MACS Handbook.
 

Excitation and emission maximums of fluorochromes comparable to PE-Vio 770.
 

FluorochromeExcitation max. (nm)Emission max. (nm)
PE-Vio 770565775
PE-Cy7496774
PE-Alexa Fluor 750496775

PE-Vio 615 (Emmax 619 nm)

PE-Vio 615 is a tandem dye with PE as the donor dye and Vio 615 as the acceptor dye. This tandem dye is optimized for efficient donor-to-acceptor dye energy transfer, high fluorescent intensity, and low spillover into the donor dye detection channel. Designed to be a superior alternative to ECD, PE-Texas Red®, PE-efluor® 610, PE-CF594 and PE/Dazzle™ 594, this Vio Dye expands the options for flexible multicolor panel design and provides a bright dye for confident detection of dim and rare markers.

PE-Vio 615 at a glance:

  • Optimal excitation with blue (488 nm), green (532 nm), and yellow green (561 nm) laser lines for maximum flexibility.
  • Excellent brightness for confident detection of dim, rare, and uncharacterized markers.
  • Extensive portfolio of PE-Vio 615 dye conjugated to recombinantly engineered REAfinity Antibodies for higher reproducibility.

For detailed information including stability, spillover, and brightness data, please visit the MACS Handbook.
 

Excitation and emission maximums of fluorochromes comparable to PE-Vio 615.
 

FluorochromeExcitation max. (nm)Emission max. (nm)
PE-Vio 615565619
ECD, PE-Texas Red565613
PE-efluor 610565606
PE-CF594565614
PE/Dazzle 594566612

Red laser (635 nm) 

APC-Vio 770 (Emmax 770 nm)

The APC-Vio 770 Dye is a tandem conjugate, like APC-Cy™7 or APC-H7, that exploits the principle of fluorescence-resonance-energy transfer (FRET) allowing for large Stokes shifts between the absorbed energy of a fluorescence donor (APC) and the emission wavelength of a suitable acceptor (Vio 770) dye.

APC-Vio 770 Dye at a glance:

  • Excitation with the yellow (561 nm) or red (635 nm) laser, emission in the near-infrared region at 775 nm.
  • The combination of Vio 770 as the acceptor dye and an optimized chemistry have furnished a tandem dye that is characterized by a high fluorescence intensity, minimal spillover to adjacent detection channels, and low non-specific binding to non-target cells to meet the complexity of multiparameter flow cytometry.
  • Higher MFI and stain index values, in addition to lower compensation settings when compared to APC-Cy7.

For detailed information including stability, spillover, and brightness data, please visit the MACS Handbook.
 

Excitation and emission maxima of fluorochromes related to APC-Vio 770.
 

FluorochromeExcitation max. (nm)Emission max. (nm)
APC-Vio 770625775
APC-Cy7650774
APC-Alexa Fluor750775

Vio Bright R667 and Vio R667 (Emmax 668 nm)

Vio Bright R667 and Vio R667 are extremely bright dyes for the R1/APC-channel. The proprietary Vio Bright multimerization technology results in a two-fold increase in brightness over the traditional APC dye. 
 

Vio Bright R667 at a glance:

  • Brighter alternative to APC.
  • Brightest Vio Dye for the red laser.
  • Excellent choice for surface markers, in particular for low-expressed antigens.

Vio R667 at a glance:

  • Small molecule alternative to APC.
  • Perfectly suited for detection of intracellular markers via flow cytometry, such as cytokines and transcription factors.
  • Optimal for use in fluorescence microscopy applications with higher photostability compared to Alexa Fluor 647.

For detailed information including stability, spillover, and brightness data, please visit the MACS Handbook.
 

Excitation and emission maxima of fluorochromes related to APC.

FluorochromeExcitation max. (nm)Emission max. (nm)
Vio Bright R667645668
Vio R667645668
APC650660

Vio Bright R720 and Vio R720 (Emmax 720nm)

Vio Bright R720 and Vio R720 are organic dyes that can be detected in Alexa Fluor 700 channel (R2 channel of MACSQuant® Analyzer 16). The proprietary multimerization technology underlying the Vio Bright R720 dye typically allows for a more than 2-fold increase in brightness over the non-multimerized Vio R720. This makes Vio Bright R720 an excellent choice for multiparameter cell analysis.

 

Vio Bright R720 at a glance:

  • Enhanced brightness with significant reduction in background noise.
  • Excellent choice for surface markers, in particular for low-expressed antigens.

Vio R720 at a glance:

  • Increased specificity by reducing background noise in conjugation to REAfinity Recombinant Antibodies.
  • Perfectly suited for intracellular markers, such as cytokines and transcription factors.

Excitation and emission maxima of fluorochromes related to APC.

FluorochromeExcitation max. (nm)Emission max. (nm)
Vio Bright R720695720
Vio R720695720
Alexa Fluor 700 (BD)697722
APC-R700653713

Resources for VioDyes

Download PDF

Brochure
Vio® and Vio®Bright Dyes - Bright and stable fluorochromes for multicolor flow cytometry

Download our brochure to find out more about Vio and Vio Bright Dyes.
 

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