Extended Data Fig. 9. Direct comparison of coCATS and rCATS in the same specimen.

a, STED imaging (xy-STED, raw data) in the CA3 region, with stratum radiatum at the top and stratum lucidum in the center in an adult mouse brain after injection of STAR RED-NHS into the lateral ventricle, perfusion fixation and rCATS labeling with WGA-AF594. Note that here, color channels are swapped relative to Fig. 5c, to account for differences in stimulated emission cross section between the fluorophores. The magnified region mainly contains mossy fibers. Both labeling paradigms yield detailed visualization of tissue architecture. b, Lateral and axial sections with near-isotropic STED imaging in a similarly prepared brain in the CA3 stratum lucidum using WGA-AF594 for rCATS and STAR RED-NHS for coCATS. STED imaging was performed at near-isotropic resolution with 80% of STED power with π-tophat phase modulation (z-STED pattern) and 20% of power 4π-helically phase modulated²⁴. N2V was applied to both channels independently. c, Similar measurement but with color channels swapped, using WGA-CF633 for rCATS and AF594-NHS for coCATS. coCATS-rCATS co-labeling was performed in 7 brain sections total from n = 3 animals with different fluorophore combinations.