[Show abstract][Hide abstract] ABSTRACT: Second Harmonic Generation (SHG) from BiFeO3 nanocrystals is investigated for
the first time to determine their potential as biomarkers for multiphoton
imaging. Nanocrystals are produced by an auto-combustion method with TRIS as a
fuel. Stable colloidal suspensions with mean particle diameters in the range
100-120 nm are then obtained after wet-milling and sonication steps. SHG
properties are determined using two complementary experimental techniques,
Hyper Rayleigh Scattering and nonlinear polarization microscopy. BiFeO3 shows a
very high second harmonic efficiency with an averaged coefficient of 79+-12
pm/V. From the nonlinear polarization response of individual nanocrystals,
relative values of the independent dij coefficients are also determined and
compared with recent theoretical and experimental studies. Additionally, the
particles show a moderate magnetic response, which is attributed to gamma-Fe2O3
impurities. A combination of high nonlinear optical efficiency and magnetic
response within the same particle is of great interest for future bio-imaging
and diagnostic applications.
[Show abstract][Hide abstract] ABSTRACT: A biophotonics approach based on the nonlinear optical process of second harmonic generation is presented and demonstrated on malignant human cell lines labelled by harmonic nanoparticles. The method enables independent imaging and therapeutic action, selecting each modality by simply tuning the excitation laser wavelength from infrared to visible. In particular, the generation of deep ultraviolet radiation at 270 nm allows direct interaction with nuclear DNA in the absence of photosensitizing molecules.
[Show abstract][Hide abstract] ABSTRACT: In this visualized experiment, protocol details are provided for in vitro labeling of human embryonic stem cells (hESC) with second harmonic generation nanoparticles (HNPs). The latter are a new family of probes recently introduced for labeling biological samples for multi-photon imaging. HNPs are capable of doubling the frequency of excitation light by the nonlinear optical process of second harmonic generation with no restriction on the excitation wavelength. Multi-photon based methodologies for hESC differentiation into cardiac clusters (maintained as long term air-liquid cultures) are presented in detail. In particular, evidence on how to maximize the intense second harmonic (SH) emission of isolated HNPs during 3D monitoring of beating cardiac tissue in 3D is shown. The analysis of the resulting images to retrieve 3D displacement patterns is also detailed.
Journal of visualized experiments : JoVE. 01/2014;
[Show abstract][Hide abstract] ABSTRACT: The mammalian circadian timing system consists of a master pacemaker in the suprachiasmatic nucleus (SCN) in the hypothalamus, which is thought to set the phase of slave oscillators in virtually all body cells. However, due to the lack of appropriate in vivo recording technologies, it has been difficult to study how the SCN synchronizes oscillators in peripheral tissues. Here we describe the real-time recording of bioluminescence emitted by hepatocytes expressing circadian luciferase reporter genes in freely moving mice. The technology employs a device dubbed RT-Biolumicorder, which consists of a cylindrical cage with reflecting conical walls that channel photons toward a photomultiplier tube. The monitoring of circadian liver gene expression revealed that hepatocyte oscillators of SCN-lesioned mice synchronized more rapidly to feeding cycles than hepatocyte clocks of intact mice. Hence, the SCN uses signaling pathways that counteract those of feeding rhythms when their phase is in conflict with its own phase.
Genes & development 07/2013; 27(13):1526-36. · 12.08 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A novel bio-photonics approach based on the nonlinear optical process of
second harmonic generation by non-centrosymmetric nanoparticles is presented
and demonstrated on malignant human cell lines. The proposed method allows to
directly interact with DNA in absence of photosensitizing molecules, to enable
independent imaging and therapeutic modalities switching between the two modes
of operation by simply tuning the excitation laser wavelength, and to avoid any
risk of spontaneous activation by any natural or artificial light source.
[Show abstract][Hide abstract] ABSTRACT: A straightforward route is proposed for the multi-gram scale synthesis of heterobifunctional poly(ethylene glycol) (PEG) oligomers containing combination of triethyloxysilane extremity for surface modification of metal oxides and amino or azido active end groups for further functionalization. The suitability of these PEG derivatives to be conjugated to nanomaterials was shown by pegylation of ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles (NPs), followed by functionalization with small peptide ligands for biomedical applications.
[Show abstract][Hide abstract] ABSTRACT: Optimal Dynamic Discrimination based on the phase-shaping of deep
ultraviolet femtosecond pulses was applied to selectively modulate the
time-resolved fluorescence depletion of pairs of tryptophan-containing
dipeptides. Our results indicate that phase-sensitive excitation allows
their differential identification, beyond the limits of linear and
[Show abstract][Hide abstract] ABSTRACT: We present a novel optical aerosol particle detector based on Xe flash lamp excitation and spectrally resolved fluorescence acquisition. We demonstrate its performances on three natural pollens acquiring in real-time scattering intensity at two wavelengths, sub-microsecond time-resolved scattering traces of the particles' passage in the focus, and UV-excited fluorescence spectra. We show that the device gives access to a rather specific detection of the bioaerosol particles.
The Review of scientific instruments 03/2013; 84(3):033302. · 1.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Nonlinear optical nanocrystals have been recently introduced as a promising alternative to fluorescent probes for multiphoton microscopy. We present for the first time a complete survey of the properties of five nanomaterials (KNbO3, LiNbO3, BaTiO3, KTP, and ZnO), describing their preparation and stabilization and providing quantitative estimations of their nonlinear optical response. In the light of their prospective use as biological and clinical markers, we assess their biocompatibility on human healthy and cancerous cell lines. Finally, we demonstrate the great potential for cell imaging of these inherently nonlinear probes in terms of optical contrast, wavelength flexibility, and signal photostability.
[Show abstract][Hide abstract] ABSTRACT: Label-free selective discrimination of spectrally similar biomolecules, such as peptides and proteins using Optimal Control strategies is a challenge in a variety of practical applications such as label-free fluorescence imaging and protein identification. The principle of Optimal Control is based on the fact that a suitably shaped laser field can differently drive the dynamics of almost identical quantum systems [1, 2].
Most of the biomolecules (including DNA) have absorption bands in the deep UV, a spectral region that is hardly accessible by transmissive devices. MEMS pulse-shapers using Reflective -Electro-Mechanical Systems (MEMS) have proven their broadband applicability for femtosecond pulse shaping [3, 4], even in deep UV and are capable of re-compressing spectrally broadened UV pulses with a closed-loop approach based on a genetic algorithm.
Recent experiments demonstrate that discriminating between nearly identical flavin molecules is possible using a reflective pulse shaping technique. We demonstrate that discrimination is possible between amino-acids, so the Optimal Control of complex systems such as proteins is envisioned as an all-optical method for identification of biomolecules.
[Show abstract][Hide abstract] ABSTRACT: Harmonic Nanoparticles are a new family of exogenous markers for multiphoton imaging exerting optical contrast by second harmonic (SH) generation. In this tutorial, we present the application of Hyper-Rayleigh Scattering (HRS) for a quantitative assessment of the nonlinear optical properties of these particles and discuss the underlying theory and some crucial experimental aspects.
The second harmonic properties of BaTiO3, KNbO3, KiTiOPO4 (KTP), LiNbO3 and ZnO nanocrystals (NCs) are investigated by HRS measurements after careful preparation and characterization of colloidal suspensions.
A detailed analysis of the experimental results is presented with emphasis on the theoretical background and on the influence of some experimental parameters including the accurate determination of the nanocrystal size and concentration. The SH generation efficiency and averaged nonlinear optical coefficients are then derived and compared for six different types of NCs.
After preparation of colloidal NC suspensions and careful examination of their size, concentration and possible aggregation state, HRS appears as a valuable tool to quantitatively assess the SH efficiency of noncentrosymmetric NCs. All the investigated nanomaterials show high SH conversion efficiencies, demonstrating a good potential for bio-labelling applications.
[Show abstract][Hide abstract] ABSTRACT: In the quest for the next generation of imaging bio-markers, successful
probes have to prove to be non toxic, bright, stable against long term
excitation, and able to generate a sharp contrast against background
fluorescence. In all these respects, Harmonic Nanoparticles (HNPs,
"nanodoublers") are receiving an increasing attention as they also open
a series of alternative detection possibilities simply not accessible
with the present generation of fluorescent dyes and quantum dots. In the
first part of the chapter, we report on this novel labelling method with
unprecedented wavelength flexibility, enabled by the non-resonant nature
of the second harmonic process. The possibility of employing infrared
excitation and the consequent deeper tissue penetration is especially
promising for their in vivo applications . The phase-coherent optical
response of HNPs can also be exploited to fully characterize the
excitation laser pulse in the focal spot of a high-NA objective with
nanometric resolution. This proof-of-principle "nano-FROG" experiment
 sets the ground for further phase-sensitive self-referenced
applications, after the recent demonstration of harmonic holography and
heterodyne detection with external references.
[Show abstract][Hide abstract] ABSTRACT: We show that the coherent manipulation of molecular wavepackets in the excited states of trp-containing dipeptides allows efficient discrimination among them. Optimal dynamic discrimination fails, however, for some dipeptide couples. When considering the limited spectral resources at play (3 nm bandwidth at 266 nm), we discuss the concept of discriminability, which appears uncorrelated to both static spectra and relaxation lifetimes.
[Show abstract][Hide abstract] ABSTRACT: We demonstrate direct amplitude shaping of high harmonics (HHs) using a reflective micromirror array based on micro-electromechanical-system (MEMS) technology. We show independent control over the intensity of each HH in the observed range (14 - 36 eV). These results are used to calculate the control achieved over the temporal structure of the attosecond pulses in the train.
[Show abstract][Hide abstract] ABSTRACT: Potassium niobate nonlinear nanoparticles are used for the first time to monitor the evolution of embryonic stem cells (ESC) by second harmonic microscopy. These particles feature the complete absence of photo-bleaching and unlimited excitation wavelength flexibility. The potential of this approach is made evident for tissue-regeneration studies and applications, by capturing a high-speed movie of ESC-derived cardiomyocytes autonomously beating within a cluster. Time-resolved data are analyzed to retrieve 3D information of the contraction pattern at the cellular level.
[Show abstract][Hide abstract] ABSTRACT: We propose a DAST/SiO(2) multilayer structure for efficient generation of near-single-cycle THz transients with average frequency around 6 THz via collinear optical rectification of 800 nm femtosecond laser pulses. The use of such a composite material allows compensation for the phase mismatch that accompanies THz generation in bulk DAST crystals. The presented calculations indicate a strong increase in the THz generation efficiency in the DAST/SiO(2) structure in comparison to the case of bulk DAST crystal.
[Show abstract][Hide abstract] ABSTRACT: Coherent manipulation of molecular wavepackets in biomolecules might contribute to the quest towards label-free cellular imaging and protein identification. We report the use of optimally tailored UV laser pulses in pump-probe depletion experiments that selectively enhance or decrease fluorescence between two aromatic amino acids: tryptophan (Trp) and tyrosine (Tyr). Selective fluorescence modulation is achieved with a contrast of ~35%. A neat modification of the time-dependent fluorescence depletion signal of Trp is observed, while the Tyr transient trace remains unchanged. The mechanism invoked for explaining the change of the depletion of Trp is a less efficient coupling between the fluorescing state and the higher non-radiative excited states by the optimally shaped pulse, than by the reference pulse.
Physical Chemistry Chemical Physics 03/2012; 14(26):9317-22. · 3.83 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We present a novel concept for optical spectroscopy called nonlinear correlation spectroscopy (NLCS). NLCS analyses coherent field fluctuations of the second and third harmonic light generated by diffusing nanoparticles. Particles based on noncentrosymmetric nonlinear materials such as KNbO(3) show a strong second as well as third harmonic response. The method and the theory are introduced and experimental NLCS results in fetal calf serum are presented showing the promising selectivity of this technique for measurement in complex biological environments.
[Show abstract][Hide abstract] ABSTRACT: Nonlinear optical nanocrystals have been recently introduced as a promising alternative to fluorescent probes for multiphoton microscopy. We present for the first time a complete survey of the properties of five nanomaterials (KNbO(3), LiNbO(3), BaTiO(3), KTP, and ZnO), describing their preparation and stabilization and providing quantitative estimations of their nonlinear optical response. In the light of their prospective use as biological and clinical markers, we assess their biocompatibility on human healthy and cancerous cell lines. Finally, we demonstrate the great potential for cell imaging of these inherently nonlinear probes in terms of optical contrast, wavelength flexibility, and signal photostability.