Multimodal nonlinear microscope based on a compact fiber-format laser source

Abstract

We present a multimodal non-linear optical (NLO) laser-scanning microscope, based on a compact fiber-format excitation laser and integrating coherent anti-Stokes Raman scattering (CARS), stimulated Raman scattering (SRS) and two-photon-excitation fluorescence (TPEF) on a single platform. We demonstrate its capabilities in simultaneously acquiring CARS and SRS images of a blend of 6-μm poly(methyl methacrylate) beads and 3-μm polystyrene beads. We then apply it to visualize cell walls and chloroplast of an unprocessed fresh leaf of Elodea aquatic plant via SRS and TPEF modalities, respectively. The presented NLO microscope, developed in house using off-the-shelf components, offers full accessibility to the optical path and ensures its easy re-configurability and flexibility.

In-line balanced detection stimulated Raman scattering microscopy

“…The pump beam is modulated at 1 MHz by an acousto-optic modulator (AOM), pump and Stokes are collinearly combined by a dichroic beam splitter (DBS) and sent to a home-built laser-scanning microscope (for a detailed description see ref. 32 ) based on two galvanometric mirrors and a 4-f relay imaging system (not shown in Fig. 1(c) ).…”

“…Initial experiments used a picosecond optical parametric oscillator pumped by a bulk solid-state laser 14 – 17 that, despite its high output power and low intensity noise, is a complex and expensive system. In recent years, research efforts focused on the development of fiber-format excitation laser systems 18 – 32 , leveraging their compactness, cost-effectiveness and insensitivity to misalignment. However, while fiber laser systems can be straightforwardly applied to CARS microscopy, the SRS modality has been much more difficult, and in some cases impossible, to be implemented.…”

High-contrast, fast chemical imaging by coherent Raman scattering using a self-synchronized two-colour fibre laser

“…Different from prior realizations of synchronized pulse generation typically seeded by noise [20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35] , here, the synchronization between the two-colour pulsed laser beams is based on CWG with a passive self-stabilization scheme. Figure 1a depicts the configuration of the selfsynchronized two-colour pulsed fibre laser source; further details are provided in Supplementary Material 2.…”

“…To this end, various two-colour pulsed fibre lasers have recently been proposed and demonstrated for CRS imaging, including fibre-based Ti:sapphire hybrid sources 16,17 and all-fibre lasers based on processes such as active synchronization 18,19 , parametric wavelength conversion [20][21][22][23] , soliton self-frequency shift [24][25][26][27][28][29] and supercontinuum (SC) light generation [30][31][32][33][34][35] . These approaches, however, have specific limitations, either high cost, low power or high intensity noise-the major challenge for directly demodulating a weak SRS signal (on the order of 10 −4 -10 −6 ) 4 ; see Supplementary Material 1.…”

Imaging Collagen in Scar Tissue: Developments in Second Harmonic Generation Microscopy for Biomedical Applications

“…Several groups have been intensively working towards this goal [ 76 , 77 , 78 ]. Crisafi et al have presented the design of a multimodal NLO laser-scanning microscope on a compact fibre-format, integrating three NLO modalities (CARS, SRS, and TPEF) [ 79 ]. Their proposed system offers for the first time the possibility to develop a NLO microscope using off-the-shelf components, providing a cost-effective alternative to commercial systems.…”