Fluorophores useful for STimulated Emission Depletion (STED) spectroscopy must fulfill strict requirements on depletion efficiency and photostability. These parameters determine the effective resolution of STED imaging. Resolution is typically measured on 30-80 nm spheres heavily decorated with STED bright fluorophores, limiting the possibility to estimate the true resolution achievable on a specific dye. Here we show how single molecule STED microscopy provides an estimate of the fluorophore stimulated emission cross section and of its photostability under STED irradiation. Fluorescein, a green and a yellow mutant of GFP, are tested, and the results are discussed and compared to those obtained with Chromeo488-covered 80 nm spheres on a commercial continuous-wave STED microscope.
Bouzin, M., Chirico, G., D'Alfonso, L., Sironi, L., Soavi, G., Cerullo, G., et al. (2013). Stimulated Emission Properties of Fluorophores by CW-STED Single Molecule Spectroscopy. JOURNAL OF PHYSICAL CHEMISTRY. B, CONDENSED MATTER, MATERIALS, SURFACES, INTERFACES & BIOPHYSICAL, 117(51), 16405-16415 [10.1021/jp409330t].
Stimulated Emission Properties of Fluorophores by CW-STED Single Molecule Spectroscopy
BOUZIN, MARGAUX;CHIRICO, GIUSEPPE;D'ALFONSO, LAURA;SIRONI, LAURA;COLLINI, MADDALENA
2013
Abstract
Fluorophores useful for STimulated Emission Depletion (STED) spectroscopy must fulfill strict requirements on depletion efficiency and photostability. These parameters determine the effective resolution of STED imaging. Resolution is typically measured on 30-80 nm spheres heavily decorated with STED bright fluorophores, limiting the possibility to estimate the true resolution achievable on a specific dye. Here we show how single molecule STED microscopy provides an estimate of the fluorophore stimulated emission cross section and of its photostability under STED irradiation. Fluorescein, a green and a yellow mutant of GFP, are tested, and the results are discussed and compared to those obtained with Chromeo488-covered 80 nm spheres on a commercial continuous-wave STED microscope.
I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
