Cytek Aurora Fluorochrome Selection Guidelines · 2019-04-01 · Zombie Red. Blue Laser Excitable...
Transcript of Cytek Aurora Fluorochrome Selection Guidelines · 2019-04-01 · Zombie Red. Blue Laser Excitable...
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Cytek AuroraFluorochrome Selection Guidelines
Last Updated: October, 2018
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Fluorochrome SignaturesDyes can be used in combination as long as they have a distinct spectrum signature.
In designing a multicolor panel, you should look for dyes with unique spectra and consider their impacts on other dyes (see slide 20)
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Violet Laser Excitable Dyes
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Violet Laser Excitable Dyes with Similar or Identical SignatureseFluor450 –Pacific Blue –vioBlue – Live/Dead Violet
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Super Bright 436, Zombie Violet
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Violet Laser Excitable Dyes with Similar or Identical SignaturesBV510 – VioGreen– Zombie Aqua –Live/Dead Yellow – eFluor 506
e506 is slightly different
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Brilliant Violet vs Super Bright (SB): almost identical
BV421vs
SB 436
BV605vs
SB 600
BV650vs
SB 645
BV711vs
SB 702
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Qdot655
Violet Laser Excitable Dyes Unique SignaturesBV421
SB436, Zombie Violet
SB 600-BV605
Qdot605
SB 645-BV650
Qdot705
SB702, BV711
BV750
BV785
Qdot 800
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Live/Dead Blue
Pacific Blue, eFluor450, vioBlue, Live/Dead Violet
Zombie Yellow
BV570
Live/Dead Red
BV480
efluor506
BV510, VioGreen, Zombie Aqua, Live/Dead Yellow
Zombie Red
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Blue Laser Excitable Dyes
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Blue Laser Excitable Dyes with Identical Signatures (1)BB515- sVio515- Vio515 AF 488, FITC, vioBright FITC, Zombie Green
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PE Dazzle594, PE eFluor 610, PE Texas Red PerCP vio700, PerCP eFluor 710
PE Vio770, PE Cy7
Blue Laser Excitable Dyes with Identical Signatures (2)
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Blue Laser Excitable Dyes Unique Signatures
BB515- sVio515- Vio515
AF 488, FITC, vioBright FITC, Zombie Green
PE
Alexa Fluor 532
PE Cy5
PE Cy55
PerCP
BB700
PerCP vio700, PerCP eFluor 710
PE Vio770, PE Cy7
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Live/Dead Green
PE Dazzle594, PE CF594, PE eFluor 610, PE Tx Red
PerCP Cy55
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Red Laser Excitable Dyes
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Red Laser Excitable Dyes with Identical SignaturesAlexa 647, Vio 667, Live/Dead Far Red
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APC-Cy7, APC-H7, APC Fire 750 , APC Vio770, Live/Dead Near-IR
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Red Laser Excitable Dyes with Unique Signatures
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APC
Alexa Fluor 647, Vio667, Live/Dead Far Red
APC Cy55
Alexa Fluor 700
APC H7, APC Fire750, APC Cy7, APCvio770
Zombie NIR
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Example of 24 dyes that can be used in combination(CAREFUL PANEL DESIGN IS NEEDED)
Fluorophore Fluorophore Fluorophore
BB515 APC BV421
Alexa Fluor 488 or FITC Alexa Fluor 647 Super Bright 436
Alexa Fluor 532 APC-R700 or AF700 eFluor450 or equivalent
PEAPC-Fire 750 or
equivalentBV480
PE/Dazzle 594 or equivalent
BV510
PE-Cy5 BV570
PerCPCy55 BV605
PerCP-eFluor710 BV650
PE-Cy7 BV711
BV750
BV785
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Stain IndexesData generated using CD4 staining in human whole blood
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600
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800
900
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Stain Index 53 dyes
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400
450
Stain Index Violet Excitable dyes
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600
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Stain Index Blue Excitable dyes
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600
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800
900
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Stain Index Red Excitable dyes
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Spread MatrixDyes used in combination need to have a unique spectrum AND also need to be assessed in terms of spread that they introduce to other dyes.
For example PerCP Cy5.5 and PE Cy5.5 have distinct signatures, but because of emission in the same wavelength range and significant spread introduced by PE Cy5.5, you will use one or the other.
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To read this table: fluor in the row impacts the one in the column. Red means the fluor in that row has significant spread into the dye in the column (for example PE into BV570). Areas in bright pink and red is where more attention to panel design is needed.
Spread matrix for 24 Fluors that can be use in combination
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In house unique spectra
Live/dead
DAPI
Zombie NIR
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UV excited dyes
BUV496
BUV563
BUV661
BUV737
BUV805
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Violet excited fluorescent protein
eBFP
eCFP
eCerulean3
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Blue excited fluorescent protein
eGFP
eYFP
mVenus
ZsGreen
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Red excited fluorescent protein Not optimally excited, test it before you use or wait until the 561 laser is installed
DsRed
mCherry
mPlum
mStrawberry
mRuby