Popular fluorophores for dSTORM imaging Imaging guide Legend OS: Oxygen Scavenger | MEA: Mercaptoethanol | BME: ß-Mercaptoethanol PBS: Phosphate Buffer Solution | STORM: Stochastic Optical Reconstruction Microscopy Fluorophore Laser line (nm) Excitation max (nm) Emission max (nm) Buffer composition Compatible with ONI buffer Comments Fluorescent labels CF® 488 488 490 515 MEA/OS Yes A bright, photostable dye in the green range. Compatible with super-resolution dSTORM using the 488 nm laser. ATTO 488 488 501 523 MEA/OS Yes (ONI’s top recommended fluorophore) One of the highest performing dSTORM dyes using 488 nm excitation. This is recommended to use over Alexa Fluor® 488. Alexa Fluor® 555 532 555 580 MEA/OS Yes An good dSTORM dye in the red range. Alexa Fluor® 555 emits a high number of photons per switching cycle. Cy3B 561 559 570 BME/OS or MEA/ OS Yes Considered to be the best dye for dSTORM imaging in the red range. CF® 568 561 562 583 MEA/OS Yes (ONI’s top recommended fluorophore) Brighter and more photostable than Alexa Fluor® 568. Compatible with multi- color dSTORM microscopy. Dylight 550 561 562 576 BME/OS or MEA/ OS Yes This dye is recommended for dSTORM imaging, however Alexa Fluor® 568 or CF®568 are better alternatives for this range.. Alexa Fluor® 568 561 578 603 MEA/OS Yes Considered a good dye for dSTORM microscopy. CF®568 is still recommended to use over Alexa Fluor® 568. Cy5 640 649 670 BME/OS or MEA/ OS Yes One of the highest performing dSTORM dyes. Considered one of the best dyes for dSTORM imaging CF® 647 640 650 665 MEA/OS Yes Excellent brightness and photostability. Compatible with super-resolution dSTORM microscopy. Alexa Fluor® 647 640 650 665 BME/OS or MEA/ OS Yes (ONI’s top recommended fluorophore) This is regarded as the best dye for dSTORM imaging due to its ability to emit thousands of photons per switching cycle. Nuclear dyes SYTO13 488 481 509 BME/OS or MEA/ OS Further testing needed Nucleic acid stain that is cell permeant. This dye is not DNA specific and can bind to RNA. This dye cannot be activated with a 405 nm laser. YOYO-1 488 491 509 BME/OS or MEA/ OS Further testing needed A high affinity nucleic acid stain which is cell impermeant. This dye is not DNA specific and can bind to RNA.YOYO-1 is often chosen over other dyes for super-resolution imaging of DNA. SYTOX™ Green 488 504 523 MEA/ OS Yes Nucleic acid stain that is cell impermeant. A good quality dSTORM dye for the green range. This dye is not DNA specific and can bind to RNA. Other dyes Tetramethyl Rhodamine (TMR) 532 546 570 MEA/OS Yes This organic fluorophore can be conjugated to genetically coded self labeling tag proteins, ie HaloTag® or SNAP-Tag®. Janelia Fluor® 549 532 549 571 PBS or MEA Yes Janelia Fluor® dyes can be conjugated to genetically coded self labeling tag proteins, ie HaloTag® or SNAP-Tag®. These dyes are reported as compatible with dSTORM microscopy in fixed cells. CellMask™ Orange Plasma Membrane Stain 561 554 567 BME/OS or MEA/ OS Needs further testing A good stain for the labeling the plasma membrane. This dye can survive cell fixation but not cell permabilisation. This dye has been investigated for dSTORM microscopy, but needs further testing before recommending as a suitable dye. MitoTracker™ Deep Red 640 644 665 BME/OS or MEA/ OS Needs further testing Specifically stains the mitochondria of cells. This dye has been investigated for dSTORM microscopy, but needs further testing before recommending as a suitable dye. CellMask™ Deep Red Plasma Membrane Stain 640 649 666 BME/OS or MEA/ OS Needs further testing A good stain for the labeling the plasma membrane. This dye can survive cell fixation but not cell permabilisation. This dye has been investigated for dSTORM microscopy, but further testing is needed before recommending as a suitable dye. Janelia Fluor® 646 640 646 664 PBS or MEA Yes Janelia Fluor® dyes can be conjugated to genetically coded self labeling tag proteins, ie HaloTag® or SNAP-Tag®. These dyes are reported as compatible with dSTORM microscopy in fixed cells. Read more about dSTORM imaging: www. oni.bio/storm-microscopy/
