Using a long-Stokes-shift dye for two-photon microscopy
Two colors from one laser: new preprint from my lab about a novel dye application.Motion artifacts and anatomical orientation can pose challenges to two-photon live imaging. A second color channel helps with both problems—but usually requires a second expensive laser. We found another way. The dye ATTO 490LS is a long-Stokes-shift fluorescent dye that’s been around for a decade, but its two-photon properties were unknown. We’ve now found that 490LS works beautifully with a 920 nm laser, the same wavelength used for GFP and GCaMP imaging. Excite with 920 nm, collect both green and red light with two detectors. One laser, two colors.Having a stable red marker like 490LS lets you find a region of interest and distinguish real calcium transients from motion-induced changes. We’re now working toward HaloTag and other conjugates for in vivo chemogenetic labeling, allowing calcium imaging with a stable reference. Please let us know if you’re interested in trying some.Preprint now on bioRxiv:https://www.biorxiv.org/content/10.1101/2025.11.21.689649v2.fullWork was led byKing Yee Cheung, with help fromXianyuan Zhang,Danesha Devini Suresh,Masahiro Fukuda, and the NUS Microscopy Core.
