A Bright, Clean Dye
BD Horizon RealYellow™ 610 (RY610) Reagents leverage an innovative laser-specific fluorochrome, excited primarily by the 561-nm yellow-green laser to offer:
- Minimal cross-laser excitation off the 488-nm blue laser compared to BD Horizon™ PE-CF594 and PE-Dazzle™ 594 Reagents
- A bright fluorochrome to support the detection of markers across a variety of expression levels
- Compatibility with conventional and spectral flow cytometers with the appropriate detectors
RY610 Reagents Can Be Used in Higher Parameter Flow Cytometry Panels to Increase Biological Resolution
| Format | Ex Max | Em Max | Spectral | Conventional | Relative Brightness | Spillover* (1 = low, 4 = high) | Alternative To |
|---|---|---|---|---|---|---|---|
| RY610 | 557 nm | 610 nm | ✓ | ✓ | 1 | PE-CF594, PE-Dazzle™ 594 |
*Value may vary based on instrument configuration and settings. Spillover ranking is based on cross-laser excitation on five-laser spectral instruments and does not take into account spillover into adjacent detectors.
Performance
BD Horizon RealYellow™ 610 Reagents Have a Clean Emission Profile with Minimal Spillover for Flow Cytometers with a 561-nm Yellow-Green Laser
RY610 has minimal cross-laser excitation
Normalized emission profiles of RY610, PE-CF594 and PE-Dazzle™ 594 fluorochromes. Human CD4 SK3 antibody acquired on a BD FACSDiscover™ S8 Cell Sorter.
RY610 reagents have minimal cross-laser excitation into the corresponding UV, violet and blue detectors
Human PBMC were stained with BD Horizon™ RY610 Human CD4 Reagent (clone SK3), acquired and compensated on a BD FACSymphony™ A5 SE Cell Analyzer.
Applications
Mouse splenocytes were stained with PE-CF594 (left) or BD Horizon™ RY610 Reagent (right) Ly-6C (AL-21), co-stained with BUV395 CD8a (53-6.7) and acquired on a BD FACSymphony™ A5 SE Cell Analyzer with compensation.
BD Pharmingen™ HiCK-1 Human Cytokine Positive Control Cells (top) or unstimulated human PBMC (bottom) were permeabilized and then stained with either BD Horizon™ PE-CF594 Reagent or BD Horizon™ RY610 Reagent (clone B27), and matched isotype controls. Samples were acquired on a BD FACSymphony™ A5 SE Cell Analyzer. Histograms are zoomed in to better show IFN-γ-positive population.
BD Horizon™ RY610 Reagents show stable performance with lot-to-lot consistency across made-to-stock reagents and BD OptiBuild™ On-Demand Reagents. Human whole blood was stained with human CD3 (UCHT1, left) or CD4 (SK3, right) RY610 using several different dye lots, followed by lysis with BD FACS™ Lysing Solution. All samples were acquired on a BD FACSymphony™ A5 SE Cell Analyzer. BD Horizon™ and BD OptiBuild™ Reagents were tested in separate experiments.
Multicolor Flow Cytometry
BD Horizon™ RY610 Reagents Have Excellent Resolution as Demonstrated in a 22-color Spectral Flow Cytometry Panel Acquired on the BD FACSDiscover™ S8 Cell Sorter
RY610 and BD Horizon RealBlue™ 613 (RB613) Reagents work well together in a 22-color spectral human lineage panel
Human peripheral fresh whole blood samples were stained with antibodies against cell surface markers prior to bulk red blood cell lysis with BD Pharm Lyse™ Lysing Buffer. Cells were then analyzed on a BD FACSDiscover™ S8 Cell Sorter and analyzed with FlowJo™ Software v10.9. A gating strategy for detection of NK and T cell subsets, after exclusion of doublets and B cells, is shown.
Fluorochrome marker assignment for a 22-color spectral flow cytometry panel
| Specificity | Clone | Fluorochrome | |
|---|---|---|---|
| UV 349 nm | CD56 | R19-760 | |
AutoF | N/A | N/A | |
CD20 | L27 | ||
CD14 | M5E2 | ||
CD127 | HIL-7R-M21 | ||
IgD | IA6-2 | ||
| Violet 405 nm | IgM | G20-127 | |
CD38 | HIT2 | ||
CD8 | RPA-T8 | ||
IgG | G18-145 | ||
CD16 | 3G8 | ||
CD19 | SJ25C1 | ||
| Blue 488 nm | CD45 | HI30 | |
CD24 | ML5 | ||
CD25 | 2A3 | ||
CD123 | 7G3 | ||
TCR γδ | 11F2 | ||
BAFF-R (CD268) | 11C1 | ||
| Yellow-Green 561 nm | CD10 | HI10a | |
CD3 | UCHT1 | ||
| Red 637 nm | CD4 | SK3 | |
CD27 | M-T271 | ||
HLA-DR | L243 |
Fluorochrome marker assignment for a 22-color spectral flow cytometry panel acquired on the BD FACSDiscover™ S8 Cell Sorter.
Buffer Compatibility
BD Horizon RealYellow™ 610 Reagents Are Compatible with a Broad Range of Fixation and Permeabilization Systems
| Buffers | Results |
| BD FACS™ Lysing Solution and BD Pharm Lyse™ Lysing Buffer | Compatible |
| BD Cytofix™ Fixation Buffer | Stable at least 24 hours |
| 1% PFA | Stable at least 24 hours |
| BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution | Compatible with antibody staining before and after fixation |
| BD FACS™ Permeabilizing Solution 2 | Compatible with antibody staining before and after fixation |
| BD Phosflow™ Perm Buffer III | Compatible with antibody staining before and after fixation |
| EDTA and Heparin | Compatible |
| BD Horizon™ Brilliant Stain Buffer (BSB) | Compatible |
For Research Use Only. Not for use in diagnostic or therapeutic procedures.
Refer to manufacturer's instructions for use and related User Manuals and Technical Data Sheets before using this product as described.
Comparisons, where applicable, are made against older BD technology, manual methods or are general performance claims. Comparisons are not made against non-BD technologies, unless otherwise noted.