Multiplexed two-photon microscopy of dynamic biological samples with shaped broadband pulses.
Coherent control can be used to selectively enhance or cancel concurrent multiphoton processes, and has been suggested as a means to achieve nonlinear microscopy of multiple signals. Here we report multiplexed two-photon imaging in vivo with fast pixel rates and micrometer resolution. We control broadband laser pulses with a shaping scheme combining diffraction on an optically-addressed spatial light modulator and a scanning mirror allowing to switch between programmable shapes at kiloHertz rates. Using coherent control of the two-photon excited fluorescence, it was possible to perform selective microscopy of GFP and endogenous fluorescence in developing Drosophila embryos. This study establishes that broadband pulse shaping is a viable means for achieving multiplexed nonlinear imaging of biological tissues.
Pillai
Rajesh S
RS
Laboratoire d'Optique et Biosciences, Ecole Polytechnique, CNRS, and INSERM U696, 91128 Palaiseau, France.
Boudoux
Caroline
C
Labroille
Guillaume
G
Olivier
Nicolas
N
Veilleux
Israel
I
Farge
Emmanuel
E
Joffre
Manuel
M
Beaurepaire
Emmanuel
E
D000818:Animals; D015509:Developmental Biology / Q000295:instrumentation; D004330:Drosophila; D004625:Embryo, Nonmammalian / Q000473:pathology*; D004867:Equipment Design; D005583:Fourier Analysis; D049452:Green Fluorescent Proteins / Q000378:metabolism; D007091:Image Processing, Computer-Assisted / Q000379:methods; D007834:Lasers; D036641:Microscopy, Fluorescence, Multiphoton / Q000295:instrumentation* / Q000379:methods*; D015233:Models, Statistical; D055095:Optics and Photonics; D012815:Signal Processing, Computer-Assisted
D049452:Green Fluorescent Proteins