Increased signals from short-wavelength-excited fluorescent molecules using sub-Ti:Sapphire wavelengths.
We report the use of an all-solid-state ultrashort pulsed source specifically for two-photon microscopy at wavelengths shorter than those of the conventional Ti:Sapphire laser. Our approach involves sum-frequency mixing of the output from an optical parametric oscillator (λ= 1400-1640 nm) synchronously pumped by a Yb-doped fibre laser (λ= 1064 nm), with the residual pump radiation. This generated an fs-pulsed output tunable in the red spectral region (λ= 620-636 nm, ~150 mW, 405 fs, 80 MHz, M(2) ~ 1.3). We demonstrate the performance of our ultrashort pulsed system using fluorescently labelled and autofluorescent tissue, and compare with conventional Ti:Sapphire excitation. We observe a more than 3-fold increase in fluorescence signal intensity using our visible laser source in comparison with the Ti:Sapphire laser for two-photon excitation at equal illumination peak powers of 1.16 kW or less.
Centre for Biophotonics, SIPBS, University of Strathclyde, Glasgow, UK.
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G
Norris
R
R
Amor
J
J
Dempster
W B
WB
Amos
G
G
McConnell
D000818:Animals; D031422:Hosta / Q000166:cytology; D007668:Kidney / Q000166:cytology; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods*; D019441:Plant Epidermis / Q000166:cytology; D018515:Plant Leaves / Q000166:cytology