Near-Infrared-Emitting Squaraine Dyes With High 2Pa Cross-Sections For Multiphoton Fluorescence Imaging

Designed to achieve high two-photon absorptivity, new near-infrared (NIR) emitting squaraine dyes, (E)-2-(1-(2-(2-methoxyethoxy)ethyl)-5-(3,4,5- trimethoxystyryl)-1H-pyrrol-2-yl)-4-(1-(2-(2-methoxyethoxy)ethyl)-5-(3,4, 5-trimethoxystyryl)-2H-pyrrolium-2-ylidene)-3-oxocyclobut-1-enolate (1) and (Z)-2-(4-(dibutylamino)-2-hydroxyphenyl)-4-(4-(dibutyliminio) -2-hydroxycyclohexa-2,5-dienylidene)-3-oxocyclobut-1-enolate (2), were synthesized and characterized. Their linear photophysical properties were investigated via UV-visible absorption spectroscopy and fluorescence spectroscopy in various solvents, while their nonlinear photophysical properties were investigated using a combination of two-photon induced fluorescence and open aperture z-scan methods. Squaraine 1 exhibited a high two-photon absorption (2PA) cross-section ( 2PA), ∼20-000 GM at 800 nm, and high photostability with the photochemical decomposition quantum yield one order of magnitude lower than Cy 5, a commercially available pentamethine cyanine NIR dye. The cytotoxicity of the squaraine dyes were evaluated in HCT 116 and COS 7 cell lines to assess the potential of these probes for biomedical imaging. The viability of both cell lines was maintained above 80% at dye concentrations up to 30 μM, indicating good biocompatibility of the probes. Finally, one-photon fluorescence microscopy (1PFM) and two-photon fluorescence microscopy (2PFM) imaging was accomplished after incubation of micelle-encapsulated squaraine probes with HCT 116 and COS 7 cells, demonstrating their potential in 2PFM bioimaging. © 2012 American Chemical Society