BD Biosciences Fluorochrome Reference Chart PDF

BD Biosciences Fluorochrome Reference Chart Visit bdbiosciences.com/colors for detailed information about our newest fluorochromes and instrumentation. To select your optimal combination of fluorochromes, visit bdbiosciences.com/spectra to use an interactive fluorescence spectrum tool....

BD Biosciences Fluorochrome Reference Chart Visit bdbiosciences.com/colors for detailed information about our newest fluorochromes and instrumentation. To select your optimal combination of fluorochromes, visit bdbiosciences.com/spectra to use an interactive fluorescence spectrum tool. Excitation Laser Fluorescence Stain index of various fluorochrome Instrument Laser Line (nm) Channel Fluorochromes provided by BD Biosciences conjugates on a BD™ LSR II BD FACSArray™ Green Diode 532 Yellow PE Far Red PerCP-Cy5.5 PE-Cy7 Stain bioanalyzer Reagent Clone Filter Index Red Diode 635 Red APC Alexa Fluor® 647 Infrared BD APC-H7 APC-Cy7 PE RPA-T4 575/26 305 *BD FACSCalibur™ Argon 488 FL1 Green FITC Alexa Fluor® 488 flow cytometry system FL2 Yellow PE APC1 RPA-T4 660/20 263 FL3 Red PE-Cy5a PerCP PerCP-Cy5.5 PE-Cy7 Red Diode 635 FL4 Red APCa Alexa Fluor® 647 PE-Cy™52 RPA-T4 695/40 198 *BD FACSCanto™ Solid State 488 Green FITC Alexa Fluor® 488 Alexa Fluor® 6471 RPA-T4 660/20 184 flow cytometry system Yellow PE Red PerCP PerCP-Cy5.5 PE-Cy™7 RPA-T4 780/60 122 Infrared PE-Cy7 HeNe 633 Red APC Alexa Fluor® 647 PerCP-Cy™5.52 RPA-T4 695/40 99 Infrared BD APC-H7 APC-Cy7 *†BD FACSCanto™ II Solid State 488 Green FITC Alexa Fluor® 488 Alexa Fluor® 4883 RPA-T4 530/30 68 flow cytometry system Yellow PE Orange PE-Texas Red®b BD Horizon™ V4505 RPA-T4 450/50 65 Red PerCP PerCP-Cy5.5 Infrared PE-Cy7 Alexa Fluor® 700 RPA-T4 720/40 64 HeNe 633 Red APC Alexa Fluor® 647 Far Red Alexa Fluor® 700b Pacific Blue™,5 RPA-T4 450/50 63 Infrared BD APC-H7 APC-Cy7 Solid Stateb 405 Green BD Horizon™ 500b AmCyanb FITC3 RPA-T4 530/30 43 Blue BD Horizon™ V450b Pacific Blue™,b Preconfigured BD™ LSR II Solid State 488 Green FITC Alexa Fluor® 488 AmCyan6 RPA-T4 525/50 37 (typical setup)d Yellow PE Orange PE-Texas Red® APC-Cy74 RPA-T4 780/60 36 Red PerCP PE-Cy5 a PerCP-Cy5.5 PerCP2 RPA-T4 695/40 30 Infrared PE-Cy7 Solid State 640 Red APCa Alexa Fluor® 647 BD Horizon™ V5006 RPA-T4 525/50 27 Far Red Alexa Fluor® 700 Infrared BD APC-H7 APC-Cy7 BD APC-H74 RPA-T4 780/60 25 Solid State 405 Green BD Horizon V500 AmCyan Blue BD Horizon V450 Pacific Blue™ Freshly isolated lymphocytes, stained with anti-human CD4 antibodies conjugated with Special Order BD™ LSR II Solid State 488 Green FITC Alexa Fluor® 488 various fluorochromes run on a BD™ LSR II flow cytometer. This chart is meant as a guideline of relative stain indices of various fluorochromes. Observed relative stain indices Special Order Yellow PE may vary depending on instrument configurations and reagents used. BD LSRFortessa™ Orange PE-Texas Red® 1, 2, 3, 4, 5, 6 F luorochromes listed with the same superscript number are read in the same (typical setup)d Red PerCP PE-Cy5a PerCP-Cy5.5 detector, and thus would not normally be used in combination. Infrared PE-Cy7 Solid State 532 or 561 Yellow PE Orange PE-Texas Red® Red PE-Cy5a Infrared PE-Cy7 W1 Solid State 640 Red APCa Alexa Fluor® 647 Far Red Alexa Fluor® 700 Infrared BD APC-H7 APC-Cy7 W2 Solid State 405 Green BD Horizon V500 AmCyan Blue BD Horizon V450 Pacific Blue™ BD FACSAria™ cell sorter Solid State 488 Green FITC Alexa Fluor® 488 familyc Yellow PE (typical setup)d Orange PE-Texas Red® Red PerCP PE-Cy5a PerCP-Cy5.5 Infrared PE-Cy7 Solid Stateb 561 Yellow PE D Orange PE-Texas Red® Red PE-Cy5a Infrared PE-Cy7 HeNe 640 Red APCa Alexa Fluor® 647 Far Red Alexa Fluor® 700 Stain Index = D/W Infrared BD APC-H7 APC-Cy7 Resolution sensitivity (the ability to resolve a dim positive signal from Solid Stateb 405 Green BD Horizon V500 AmCyan background) depends upon the difference between positive and Blue BD Horizon V450 Pacific Blue™ background peak means (D) and the spread of the background peak BD Influx™ cell sorter Solid State 488 Green FITC Alexa Fluor® 488 (W). W1 and W2 represent background peaks with different spreads. The stain index is a metric that captures both of these factors. Yellow PE Orange PE-Texas Red® Red PE-Cy5 PerCP-Cy5.5 Infrared PE-Cy7 * For In Vitro Diagnostic Use. Solid State 532 or 561 Yellow PE † Seven- and eight-color assays on this device are for Research Use Only. Orange PE-Texas Red® Unless otherwise specified, all products are for Research Use Only. Red PE-Cy5 Class I (1) laser product Infrared PE-Cy7 APC-Cy7: US patent 5,714,386 Solid State 640 Red APC Alexa Fluor® 647 Cy™ is a trademark of Amersham Biosciences Corp. Far Red Alexa Fluor®700 Pacific Blue is a trademark and Alexa Fluor and Texas Red are registered trademarks of Molecular Probes, Inc. Alexa Fluor®, Texas Red®, and Pacific Blue™ dye antibody conjugates Infrared BD APC-H7 APC-Cy7 are sold under license from Molecular Probes, Inc., for research use only, excluding use in Solid State 405 Green BD Horizon V500 AmCyan combination with microarrays, or as analyte specific reagents. The Alexa Fluor® dyes (except for Alexa Fluor® 430) and Pacific Blue™ dye are covered by pending and issued patents. Blue BD Horizon V450 Pacific Blue™ BD, BD Logo and all other trademarks are property of Becton, Dickinson and Company. © 2009 BD a APC and PE-Cy5 may be used together on instruments with cross-beam compensation. Available through laser and/or detector options. b c BD FACSAria™ and BD FACSAria™ II 23-9582-04 d More laser and detector options are available through the Special Order Research Products (SORP) program. Choose a winning combination - Guidelines for selecting reagents for multicolor flow cytometry 1 The basics: Know your instrument Reagent selection starts with 2 Fluorochromes: Go for the bright Rank available dyes according 3 Minimize spillover As soon as cells are stained with multiple reagents, spectral overlap 4 Colors and specificities: Define winning combinations Once the fluorochromes to be used 5 Tandem dyes APC-Cy7, and to a lesser extent, PE-Cy7, can degrade in the presence 6 Validation Use controls (such as fluorescence-minus-one, or FMO) your instrument configuration. to their intrinsic brightness on (or spillover) becomes an issue. The have been defined, you can begin of light, fixative, and elevated to validate your selected multicolor The lasers and detectors in your a particular instrument (when more colors you attempt to resolve to match antibody specificities to temperatures so that they emit in the reagent cocktail. FMO controls help configuration dictate how well your configured with a specified set of on any particular cell, the more particular fluorochromes. Generally, parent dye detector (APC or PE). By define the contribution of spillover cytometer can excite and measure lasers and filters). spillover impacts sensitivity. We reserve the brightest fluorochromes minimizing the exposure of samples to the background in a given a given fluorochrome, and whether use compensation, an adjustment for dim antigens, and vice versa, to light, heat, and formaldehyde- detector, and are therefore useful you have enough detectors to applied to all colors, to correct but avoid spillover from bright based fixatives, this problem can in gauging the sensitivity of that read out a given combination of for spillover. For example, a cell cell populations into detectors be largely avoided. For more stable detector in the context of a certain fluorochromes. population fluorescing only in FITC requiring high sensitivity for those tandem dyes, BD now offers reagent cocktail. will show no PE fluorescence, on populations. BD APC-H7 conjugated antibodies. average, but will likely exhibit more spread in the PE detector after compensation than completely For additional guidelines, visit bdbiosciences.com/colors to download the unstained cells. Application Note “Selecting Reagents for Multicolor Flow Cytometry.”