title: Three-color femtosecond source for simultaneous excitation of three fluorescent proteins in two-photon fluorescence microscopy.
abstract: We demonstrate a fiber-based, three-color femtosecond source for simultaneous imaging of three fluorescent proteins (FPs) using two-photon fluorescence microscopy (2PM). The three excitation wavelengths at 775 nm, 864 nm and 950 nm, are obtained through second harmonic generation (SHG) of the 1550-nm pump laser and the 1728-nm and 1900-nm solitons generated through soliton self-frequency shift (SSFS) in a large-mode-area (LMA) fiber. These energetic pulses are well matched to the two-photon excitation peaks of red, cyan and yellow fluorescent proteins (TagRFPs, TagCFPs, and TagYFPs) for efficient excitation. We demonstrate simultaneous 2PM of human melanoma cells expressing a "rainbow" combination of these three fluorescent proteins.
authors.affiliation: School of Applied and Engineering Physics, Cornell University, Ithaca, NY 14853,  USA ; Each author contributed equally to this work.
authors.forename: Ke
authors.initials: K
authors.lastname: Wang
authors.forename: Tzu-Ming
authors.initials: TM
authors.lastname: Liu
authors.forename: Juwell
authors.initials: J
authors.lastname: Wu
authors.forename: Nicholas G
authors.initials: NG
authors.lastname: Horton
authors.forename: Charles P
authors.initials: CP
authors.lastname: Lin
authors.forename: Chris
authors.initials: C
authors.lastname: Xu