Abstract
Multiphoton microscopy (MPM) including two-photon excited fluorescence (TPEF), second harmonic generation (SHG), coherent anti-Stokes Raman scattering (CARS) is an important tool in biology and medicine to explore the dynamics of cells and to investigate tissue structure in living systems and biopsies. However, MPM still suffers from limited applicability due to the integration of highly sophisticated lasers. In this paper, we introduce our triple-modal multicolor MPM platform employing a single direct diode-pumped femtosecond Kerr-lens-mode-locked Ti:sapphire oscillator and acquire simultaneously intrinsic co-registered multicolor TPEF, TPEF, SHG and CARS signals in the backward direction. The complexity and by that the footprint as well the cost of the laser are considerably reduced by the direct diode-pumping scheme. In combination with an Yb fiber amplifier such a laser is suited for biomedical applications allowing for detailed morphologic and molecular spectroscopic in vivo investigation. The optimized emission wavelengths of our Ti:sapphire laser and Yb fiber amplifier provide simultaneous excitation at 805 nm, 1050 nm and 911 nm for multicontrast information from exogenous and endogenous fluorophores. We perform in vivo imaging of C. elegans and visualize metabolic changes in zebrafish larvae to investigate the tumor microenvironment with spectral focusing CARS and multicolor TPEF of green fluorescent protein.
Highlights
U NDERSTANDING the dynamics of metabolism in multicellular organisms is important in unraveling the mechanism behind many biological processes, i.e. tumor development [1]
We introduce an in vivo multimodal multicolor non-linear optical imaging platform solely based on a new direct diode pumped femtosecond Kerr-Lens-modelocked (KLM) Ti:sapphire oscillator and discuss the potential and technical details allowing for optimized excitation of multicolor twophoton excited fluorescence (TPEF), TPEF, second harmonic generation (SHG) and coherent anti-Stokes Raman scattering (CARS) signals
The basic configuration of the proposed multicolor twophoton excited fluorescence microscopy scheme is discussed here.Briefly, our multimodal multicolor non-linear optical imaging shown in Fig. 2(a) uses the direct diode pumped femtosecond Ti:sapphire laser in combination with the inherently synchronized high power Yb fiber amplifier for excitation as described in detail elsewhere [25]
Summary
U NDERSTANDING the dynamics of metabolism in multicellular organisms is important in unraveling the mechanism behind many biological processes, i.e. tumor development [1]. CARS microscopy is a powerful method in biomedical imaging with molecular vibrational contrast for the wide range of biological materials that do not exhibit strong optical transitions to electronic states in the visible and NIR wavelength regime. We demonstrated a multimodal epi-detection non-linear laser-scanning microscope (LSM) with a compact and cost-reduced femtosecond Ti:sapphire laser [25] This system featured simultaneous acquisition of signals arising from freshly excised mouse ear tissue with spectral focusing CARS, TPEF, and SHG, while the laser source still used a water-cooled frequency-doubled distributed Bragg reflectortapered diode laser to pump the Ti:sapphire laser. We introduce an in vivo multimodal multicolor non-linear optical imaging platform solely based on a new direct diode pumped femtosecond Kerr-Lens-modelocked (KLM) Ti:sapphire oscillator and discuss the potential and technical details allowing for optimized excitation of multicolor TPEF, TPEF, SHG and CARS signals. The biomedical imaging performances are demonstrated in living organisms showing the potential for in vivo imaging
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
