StarBright Violet 570 Dye
StarBright Violet 570 (SBV570) Dye is the latest addition to the StarBright Dye range of fluorescent nanoparticles specifically designed for flow cytometry.
Excitable by the violet 405 nm laser and emitting at 570 nm, it is the perfect replacement for BV570 in your flow cytometry experiments.
Fig. 1. Excitation and emission spectra for SBV570. The dye is excited maximally by the violet laser (dotted line) with minimal excitation by other lasers. The emission profile is narrow to reduce spillover into neighboring filters.
SBV570 is also compatible with spectral flow cytometry and has a unique profile, allowing incorporation into new and existing flow cytometry panels. The full spectral profile is shown below (Figure 2).
Fig. 2. Complete dye signature of SBV570. Full emission spectrum of StarBright Violet 570 Dye showing the emission at all wavelengths. Data was collected on the Cytek Aurora Flow Cytometry System using SpectroFlo Software.
SBV570 Dye has been designed to be bright with narrow excitation and emission, making it a great choice for inclusion in multicolor panels. It is brighter than dyes with similar excitation and emission profiles such as Brilliant Violet 570 (BV570), allowing detection of rare and low antigen density populations, while also giving great separation for more abundantly expressed markers. In addition, as it is not a traditional tandem dye, it does not suffer from inefficient FRET, leading to unwanted signal from other lasers due to excitation of the donor or acceptor.
Although SBV570 can be detected using the ZE5 Cell Analyzer with the 615/24 filter, this is suboptimal and SBV610 is more suitable when using this filter. However, SBV570 can be used on the S3e Cell Sorter fitted with a 405 nm laser as this has a 586/25 or 593/40 filter, depending on the configuration, or any other cell analyzer or sorter with a similar filter. Figure 3 below shows the performance of SBV570 compared to BV570 on the S3e Cell Sorter.
Fig. 3. Brightness comparison. Human peripheral blood mononuclear cells (PBMCs) were stained with A, CD4 BV570 or CD19 BV570 or B, CD4SBV570 (MCA1267SBV570) or CD19SBV570 (MCA1940SBV570) and the stain index calculated. Both antibodies were titrated prior to use to determine the optimal concentration that gave the best resolution between positive and negative.
SBV570 is 2-3 times brighter than BV570, making it easier to select a positive population with SBV570 compared with BV570 during cell sorting, resulting in higher purity and more cell recovery. This can be seen in Figure 4 where CD3FITC+ CD8 T cells stained with either BV570 or SBV570 were sorted from PBMCs. Three times as many cells were recovered with SBV570 compared with BV570 and the purity was 94% compared with 88%.
Fig. 4. Sorting purity and recovery is improved with StarBright Dyes. Human PBMCs were stained with CD3 FITC (MCA463F) and either A, CD8BV570 or B, CD8SBV570 (MCA1226SBV570). The number of cells recovered and the purity can be seen in the statistics below the right hand side plots.
StarBright Violet 570 Dye is compatible with all common fluorophores such as organic fluorophores, protein fluorophores, polymer dyes, and other StarBright Dyes in multicolor panels. SBV570 does not require a special staining buffer, when used with other StarBright Dyes or organic fluorophores, however if it is combined with dyes that do require the use of a special buffer, we recommend staining in that buffer, as it will not harm the StarBright Dye performance.
SBV570 is very stable when kept at 4oC with excellent lot-to-lot reproducibility, meaning you can get the same results time after time. High quality staining can be achieved at 4oC or room temperature, it is not susceptible to photobleaching and the staining does not deteriorate with fixation, giving you flexibility in your experiments. Figure 5 below shows some examples of the staining that can be achieved using SBV570.
Fig. 5. Examples of SBV570 Dye staining. A, mouse peripheral blood was stained with CD122 FITC (MCA2363F) and CD161SBV570 (MCA1266SBV570). Human peripheral blood was stained with B, CD3A647 (MCA463A647) and CD4SBV570 (MCA1267SBV570); and C, CD3SBV515 (MCA463SBV515) and CD4SBV570 (MCA1267SBV570). All staining was performed at 4oC in PBS containing 1% BSA after blocking with either 10% mouse or human serum. Cells were analyzed on the ZE5 Cell Analyzer.
For more information on how the StarBright range can fit into your flow cytometry experiments, please take a look at our StarBright Dyes range.
For more information on fluorophores and immunophenotyping, refer to our cytometry resources.
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