With the growing availability of flow cytometers equipped with green (532 nm) and yellow (561 nm) lasers, green light-excitable and yellow light-excitable fluorophores are playing an increasingly critical role in standard multi-parametric flow cytometry. The expansive Alexa Fluor dye, eFluor dye, and NovaFluor dye portfolios support multi-color immunophenotyping by flow cytometry and includes a variety of dyes compatible with the 532 and 561 nm spectral lines of the green and yellow lasers, respectively.
The table below lists the properties of the most commonly used fluorophores for green and yellow lasers.
|Fluorophore||Fluorophore type||Ex/Em (nm)||Relative intensity* (1 = lowest, 5 = highest)||Suggested bandpass filter (nm)||Uses|
|Alexa Fluor 532||Organic dye||532, 532/554||3||555/20||Yellow-fluorescent dye with excitation ideally suited for the 532 nm laser|
|Alexa Fluor 561||Organic dye||561, 558/575||3||555/20||A bright dye optimized for flow cytometers with spectral capabilities which can replace PE to reduce emission spreading into PE tandem dyes|
|NovaFluor Yellow 570||Phiton technology||561, 552/568||2||575/26||Alternative to phycoerythrin (PE) with similar emission spectrum and less cross-laser excitation|
|R-Phycoerythrin (R-PE, PE)||Protein||496, 565/575||5||575/26||Widely used fluorophore with a high quantum yield that is ideal for detection of low-density antigens|
|NovaFluor Yellow 590||Phiton technology||561, 552/590||4||590/40||A unique emission spectrum, which fits in between some PE-tandem dyes, allowing the detection of an additional marker in a complex multi-color panel|
|PE-eFluor 610||Tandem||496, 565/607||3||610/20||Tandem dye, resonance energy transfer from PE molecule to eFluor 610; can be used as a brighter alternative to PE-Texas Red dye|
|NovaFluor Yellow 610||Phiton technology||561, 552/612||3||590/40||Can be used to replace some PE-tandem dyes with less cross-excitation and lower spectral spillover, allowing detection of an additional marker labeled with one of the NovaFluor Blue 610 dyes|
|PE-Texas Red||Tandem||496, 565/615||3||590/40||Tandem dye, resonance energy transfer from PE molecule to Texas Red dye|
|PE-Alexa Fluor 610||Tandem||496, 565/628||3||590/40||Tandem dye, resonance energy transfer from PE molecule to Alexa Fluor 610 dye|
|NovaFluor Yellow 660||Phiton dye||561, 552/663||3||670/14||Can be used to replace some PE tandem dyes with less cross-excitation and ﬂuorescent spillover, allowing for an additional marker to be detected with the blue laser line using NovaFluor Blue 660|
|PE-Cyanine 5 (TRI-COLOR)||Tandem||496, 565/667||3||670/14||Tandem dye, resonance energy transfer from PE molecule to the Cyanine 5 dye|
|NovaFluor Yellow 690||Phiton dye||561, 552/690||3||695/40||Emission spectrum that fits between the emission peaks of PE-Cyanine 5 and PE-Cyanine 7, allowing detection of an additional marker in a complex multi-color panel using a spectral flow cytometer|
|PE-Cyanine 5.5||Tandem||496, 565/694||4||695/40||Tandem dye, resonance energy transfer from PE molecule to the Cyanine 5.5 dye|
|NovaFluor Yellow 700||Phiton dye||561, 552/700||4||695/40||Can be used as an alternative to PE-Cyanine 5.5, with less cross-laser excitation and spectral spillover|
|PE-Alexa Fluor 700||Tandem||496, 565/723||3||695/40||Tandem dye, resonance energy transfer from PE molecule to Alexa Fluor 700 dye|
|NovaFluor Yellow 730||Phiton dye||561, 552/730||3||695/40||Can be used as an alternative to PE-Cyanine 5.5, with far less cross-laser excitation and spectral spillover; NovaFluor Yellow 690, NovaFluor Yellow 700, and NovaFluor 730 dyes cannot be used together with conventional flow cytometry, but their distinct spectral signatures allow them to be successfully unmixed using a spectral flow cytometer|
|NovaFluor Yellow 755||Phiton dye||551/755||3||780/60||Can be used as a non-tandem alternative to PE-Cyanine 7 and cFluor® BYG750 with significantly less cross-laser excitation from the blue (488 nm) laser.|
|PE-Cyanine 7||Tandem||496, 565/767||4||780/60||Tandem dye, resonance energy transfer from PE molecule to Cyanine 7 dye|
|* The relative intensity of any fluorophore is dependent upon cell type being studied, reagent used, instrument, and instrument configuration. The relative intensities provided are to be used as a guideline; observed intensities may vary.|
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