Much Brighter Equivalent for DyLight 680

Results Summary

WT mouse splenocytes were stained with anti-CD45 clone 30-F11 at a range of dilutions. With splenocytes, greater than 90% of the cells are expected to stain brightly with anti-CD45, and I had been disappointed with the Dylight 680 conjugate as it only produced dim staining when using a very large amount of antibody. In contrast, the Spark NIR 685 conjugate stained brightly, and exhibited adequate staining in the range of 1:5000-1:1000. This is similar to what is required when using relatively bright fluorophores, suggesting that Spark NIR 685 is likely to be brighter than Alexa Fluor 700. The emission profile is similar to APC-Cy5.5Spark NIR exhibited less spillover into other far red channels on the Aurora. Thus, users may expect less interference.Attempts to unmix Spark NIR 685 in the context of a 40+ color panel containing Alexa Fluor 647, APC, Alexa Fluor 700, APC-Cy5.5 and eFluor780 were not very successful. However, it would likely work well to remove APC-Cy5.5 and replace it with Spark NIR 685, as this would be a cleaner option and not have issues due to tandem degradation.