Plasmon-enhanced four-wave mixing by nanoholes in thin gold films.

Nonlinear plasmonics opens up for wavelength conversion, reduced interaction/emission volumes, and nonlinear enhancement effects at the nanoscale with many compelling nanophotonic applications foreseen. We investigate nonlinear plasmonic responses of nanoholes in thin gold films by exciting the holes individually with tightly focused laser beams, employing a degenerated pump/probe and Stokes excitation scheme. Excitation of the holes results in efficient generation of both narrowband four-wave mixing (FWM) and broadband multiphoton excited luminescence, blueshifted relative to the excitation beams. Clear enhancements were observed when matching the pump/probe wavelength with the hole plasmon resonance. These observations show that the FWM generation is locally excited by nanoholes and has a resonant behavior primarily governed by the dimensions of the individual holes.

Uptake of gold nanoparticles in healthy and tumor cells visualized by nonlinear optical microscopy.

Understanding the mechanism underlying the interactions between inorganic nanostructures and biological systems is crucial for several rapidly growing fields that rely on nano-bio interactions. In particular, the further development of cell-targeted drug delivery using metallic nanoparticles (NP) requires new tools for understanding the mechanisms triggered by the contact of NPs with membranes in different cells at the subcellular level. Here we present a novel concept of multimodal microscopy, enabling three-dimensional imaging of the distribution of gold NPs in living, unlabeled cells. Our approach combines multiphoton induced luminescence (MIL) with coherent anti-Stokes Raman scattering (CARS) microscopy. Comparison with transmission electron microscopy (TEM) reveals in vivo sensitivity down to the single nanostructure. By monitoring the incorporation of NPs in human healthy epidermal keratinocytes and squamous carcinoma cells (SCC), we address the feasibility of noninvasive delivery of NPs for therapeutic purposes. While neutralizing PEG coating was confirmed to prevent NP integration in SCCs, an unexpectedly efficient integration of NPs into keratinocytes was observed. These results, independently validated using TEM, demonstrate the need for advanced surface modification protocols to obtain tumor selectivity for NP delivery. The CARS/MIL microscopy platform presented here is thus a promising tool for noninvasive study of the interaction between NPs and cell.

Nonlinear microscopy with fiber laser continuum excitation.

A compact high-power fiber-based femtosecond laser system is presented for coherent anti-Stokes Raman scattering/second-harmonic generation (CARS/SHG) microscopy, and quantitatively compared with a conventional picosecond optical parametric oscillator (OPO)-based system. While the broad spectral width of the femtosecond pulses results in 2.5 times lower image contrast and limited spectral selectivity, lipid stores, myosin, and collagen filaments in living cells can clearly be identified at 60 times lower excitation powers compared to the picosecond system. Visually the images contain the same information. Together with simple operation, small footprint, and low cost, the capabilities of this high-power all-fiber-based laser system promise a more general use of nonlinear microscopy within the biosciences.

Nonlinear microscopy of lipid storage and fibrosis in muscle and liver tissues of mice fed high-fat diets.

Hallmarks of high-fat Western diet intake, such as excessive lipid accumulation in skeletal muscle and liver as well as liver fibrosis, are investigated in tissues from mice using nonlinear microscopy, second harmonic generation (SHG), and coherent anti-Stokes Raman scattering (CARS), supported by conventional analysis methods. Two aspects are presented; intake of standard chow versus Western diet, and a comparison between two high-fat Western diets of different polyunsaturated lipid content. CARS microscopy images of intramyocellular lipid droplets in muscle tissue show an increased amount for Western diet compared to standard diet samples. Even stronger diet impact is found for liver samples, where combined CARS and SHG microscopy visualize clear differences in lipid content and collagen fiber development, the latter indicating nonalcoholic fatty liver disease (NAFLD) and steatohepatitis induced at a relatively early stage for Western diet. Characteristic for NAFLD, the fibrous tissue-containing lipids accumulate in larger structures. This is also observed in CARS images of liver samples from two Western-type diets of different polyunsaturated lipid contents. In summary, nonlinear microscopy has strong potential (further promoted by technical advances toward clinical use) for detection and characterization of steatohepatitis already in its early stages.

Raman spectroscopic studies of terthiophenes for molecular electronics.

The effect of thiol and selenol functionalization on the vibrational spectra and photochemical stability of terthiophene based molecular wires was investigated using surface-enhanced Raman scattering (SERS). The molecules were found to exhibit markedly different properties at the silver surface of the SERS substrate, despite having almost identical Raman spectra in solution and in the solid state. In contrast to terthiophene (3T), the bisthiolterthiophene (T3) and biselenol-terthiophene (Se3) molecules were stable against photoinduced structural changes when adsorbed to the metal surface at low concentrations. This indicates that the strong bonds to the silver surface, via S or Se terminal atoms, leads to a rapid decay of photoexcited states. Comparison with ab initio calculations shows that both T3 and Se3 bind with only one of the functional groups to the Ag surface.

Creating hot nanoparticle pairs for surface-enhanced Raman spectroscopy through optical manipulation.

We use optical tweezers to move single silver nanoparticles into near-field contact with immobilized particles, forming isolated surface-enhanced Raman spectroscopy (SERS) active Ag particle dimers. The surface-averaged SERS intensity increases by a factor approximately 20 upon dimerization. Electrodynamics calculations indicate that the final approach between the particles is due to "optical binding". The described methodology may facilitate controlled single molecule SERS analysis.

On the importance of optical forces in surface-enhanced Raman scattering (SERS).

This contribution reports on the combination of optical tweezers with SERS spectroscopy of colloidal silver nanoparticles covered by thiophenol. The experimental design is based on two different laser beams, one used for Raman excitation (lamda = 514.5 nm) and one for optical tweezing (lamda=830 nm). For a fixed Raman excitation power, the SERS signal from thiophenol is found to increase dramatically when the trapping laser is activated. This result is interpreted as a combination of two effects, an accumulation of nanoparticles in the optical trap and an optically induced aggregation of these nanoparticles.