title: Time-resolved multifocal multiphoton microscope for high speed FRET imaging in vivo.
abstract: Imaging the spatiotemporal interaction of proteins in vivo is essential to understanding the complexities of biological systems. The highest accuracy monitoring of protein-protein interactions is achieved using Förster resonance energy transfer (FRET) measured by fluorescence lifetime imaging, with measurements taking minutes to acquire a single frame, limiting their use in dynamic live cell systems. We present a diffraction limited, massively parallel, time-resolved multifocal multiphoton microscope capable of producing fluorescence lifetime images with 55 ps time-resolution, giving improvements in acquisition speed of a factor of 64. We present demonstrations with FRET imaging in a model cell system and demonstrate in vivo FLIM using a GTPase biosensor in the zebrafish embryo.
authors.forename: Simon P
authors.initials: SP
authors.lastname: Poland
authors.forename: Nikola
authors.initials: N
authors.lastname: Krstajić
authors.forename: Simao
authors.initials: S
authors.lastname: Coelho
authors.forename: David
authors.initials: D
authors.lastname: Tyndall
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Walker
authors.forename: Viviane
authors.initials: V
authors.lastname: Devauges
authors.forename: Penny E
authors.initials: PE
authors.lastname: Morton
authors.forename: Nicole S
authors.initials: NS
authors.lastname: Nicholas
authors.forename: Justin
authors.initials: J
authors.lastname: Richardson
authors.forename: David Day-Uei
authors.initials: DD
authors.lastname: Li
authors.forename: Klaus
authors.initials: K
authors.lastname: Suhling
authors.forename: Claire M
authors.initials: CM
authors.lastname: Wells
authors.forename: Maddy
authors.initials: M
authors.lastname: Parsons
authors.forename: Robert K
authors.initials: RK
authors.lastname: Henderson
authors.forename: Simon M
authors.initials: SM
authors.lastname: Ameer-Beg
mesh_terms: D000818:Animals; D031541:Fluorescence Resonance Energy Transfer; D061986:MCF-7 Cells; D036641:Microscopy, Fluorescence, Multiphoton / Q000379:methods*; D013997:Time Factors; D015027:Zebrafish