Coherent anti-Stokes Raman Scattering Signal Research Articles

Differential imaging in coherent anti-Stokes Raman scattering microscopy II: a filter-assisted Laguerre-Gaussian beam detection scheme

A new differential imaging technique to obtain contrast improvement in coherent anti-Stokes Raman scattering (CARS) microscopy is proposed through a spatial spiral phase modulation in the collected signal of CARS microscopy. A spiral phase mask makes the CARS signal from the bulk material to be distributed in a ring centered at the detecting pinhole of a confocal microscope resulting in a weak detected CARS signal from a bulk material. When tiny scatters are included in the focal volume of a CARS setup, the ring-shaped distribution of CARS field is distorted, leading to an increase in the detected signal through the pinhole. The sensitivity and the size selectivity of this proposed technique is studied with varying the particle size, and it is found that this method is to be efficient in edge detection. Simulation results obtained by finite difference time domain (FDTD) methods show that the image contrast is enhanced by many times as it is able to highlight the details of the specimen by suppressing the CARS signal from a bulk or a uniform material.

Time-gated single photon counting enables separation of CARS microscopy data from multiphoton-excited tissue autofluorescence

We demonstrate time-gated confocal imaging as a means to separate coherent anti-Stokes Raman scattering (CARS) microscopy data from multi-photon excited endogenous fluorescence in tissue. CARS is a quasi-instantaneous process and its signal decay time is only limited by the system's instrument response function (IRF). Signals due to two-photon-excited (TPE) tissue autofluorescence with excited state lifetimes on the nanosecond scale can be identified and separated from the CARS signal by employing time-gating techniques. We demonstrate this improved contrast on the example of CARS microscopy of intact roots of plant seedlings as well as on rat arterial tissue.

Nonlinear optical interference of two successive coherent anti-Stokes Raman scattering signals for biological imaging applications

The nonlinear optical interference of two successively generated coherent anti-Stokes Raman scattering (CARS) signals from two different samples placed in series is demonstrated for the imaging performance, in which a collinear phase matching geometry is used. The relative phase of two CARS signals is controlled by a phase-shifting unit made of dispersive glass materials of which the thickness can be precisely varied. The clear interference fringes are observed as the thickness of the phase-shifting unit changes. The interference effect is then utilized to achieve a better quality CARS image of a biological tissue taken from a mouse skin. Placing the tissue in the second sample position and performing raster scans of the laser beams on it, we can acquire a CARS image of higher contrast compared to the normal image obtained without interferometric implementation.

Near-field effects on coherent anti-Stokes Raman scattering microscopy imaging

We introduce a numerical method, the finite-difference time-domain (FDTD) method, to study the near-field effects on coherent anti-Stokes Raman Scattering (CARS) microscopy on nanoparticles. Changes of the induced nonlinear polarization, scattering patterns, and polarization properties against different diameters of spherical nanoparticles are calculated and discussed in details. The results show that due to near-field effects, the induced nonlinear polarization is significantly enhanced at the water-particle interface, with 1.5-fold increase in intensity compared to that inside the particles, and the near-field enhancement increases with decreasing diameters of nanoparticles. The enhanced scattering dominates over the scattering contribution from the particles when the nanoparticle size decreases down to the scale of less than a half wavelength of excitation light. Further studies show that near-field effects make the induced perpendicular polarization of CARS signals being strictly confined within the nanoparticles and the particle-water interface, and this perpendicular polarization component could contribute approximately 20% to the backward scattering. The ratio values of the perpendicular polarization component to the total CARS signals from nanoparticles sizing from 75 nm to 300 nm in backward scattering are approximately 3 to 5 times higher than those in forward scattering. Therefore, near-field effects can play an important role in CARS imaging. Employing the perpendicular polarization component of CARS signals can significantly improve the contrast of CARS images, and be particularly useful for revealing the fine structures of bio-materials with nano-scale resolutions.

Adaptive optics for enhanced signal in CARS microscopy

We report the use of adaptive optics with coherent anti-Stokes Raman scattering (CARS) microscopy for label-free deep tissue imaging based on molecular vibrational spectroscopy. The setup employs a deformable membrane mirror and a random search optimization algorithm to improve signal intensity and image quality at large sample depths. We demonstrate the ability to correct for both system and sample-induced aberrations in test samples as well as in muscle tissue in order to enhance the CARS signal. The combined system and sample-induced aberration correction increased the signal by an average factor of approximately 3x for the test samples at a depth of 700 microm and approximately 6x for muscle tissue at a depth of 260 microm. The enhanced signal and higher penetration depth offered by adaptive optics will augment CARS microscopy as an in vivo and in situ biomedical imaging modality.

Observation of CARS signal via maximal atomic coherence prepared by F-STIRAP in a three-level atomic system

We control the atomic coherence and the population transfer among Rb hyperfine atomic levels by the fractional stimulated Raman adiabatic passage (F-STIRAP) in a L-type configuration, and verify the theoretical predictions. Applying this technique, we are able to prepare the atoms with maximal coherence to enhance coherent anti-Stokes Raman scattering (CARS) signal. In our experiment by scanning the frequency of one laser from 794.9839 nm to 794.9844 nm with the 794.9698 nm laser frequency fixed to generate a maximum of the CARS signal we are able to obtain the energy level diagram of the sample.

Differential imaging in coherent anti-Stokes Raman scattering microscopy with Laguerre- Gaussian excitation beams

We propose a new differential imaging technique to visualize the fine structures and the edges of a sample in coherent anti-Stokes Raman Scattering (CARS) microscopy. Both the pump and Stokes excitation fields are modulated simultaneously with a spiral phase mask which transforms them from Gaussian modes into Laguerre-Gaussian modes of LG01 for CARS excitation. With an accurate three dimensional finite-difference time-domain (FDTD) method, the intensity and phase distributions of focused input fields, the scattering pattern of generated CARS signal as well as the formation of differential images are studied detailedly, and by simulating the sensitivity range and reliability of this method, we have verified that it is much suitable for visualizing structures with a scale comparable to the excitation wavelength and has higher reliability in retrieving chemical structural information of the sample compared to common CARS microscopy.

Femtosecond coherent anti-Stokes Raman scattering measurement of gas temperatures from frequency-spread dephasing of the Raman coherence

Gas-phase temperatures and concentrations are measured from the magnitude and decay of the initial Raman coherence in femtosecond coherent anti-Stokes Raman scattering (CARS). A time-delayed probe beam is scattered from the Raman polarization induced by pump and Stokes beams to generate CARS signal; the dephasing rate of this initial coherence is determined by the temperature-sensitive frequency spread of the Raman transitions. Temperature is measured from the CARS signal decrease with increasing probe delay. Concentration is found from the ratio of the CARS and nonresonant background signals. Collision rates do not affect the determination of these quantities.

Coherent anti-Stokes Raman scattering microscopy (CARS): Instrumentation and applications

Recent advances in laser physics have permitted the development of a new kind of microscopy based on stimulated Raman scattering. This new technique known as Coherent anti-Stokes Raman scattering (CARS) microscopy allows vibrational imaging with high sensitivity, high spectral resolution and three-dimensional sectioning capabilities. We review recent advances in CARS microscopy, with applications to chemical and biological systems. We also present an application of CARS microscopy with high optical resolution and spectral selectivity, in resolving structures in surface ex vivo stratum corneum by looking at the CH 2 stretching vibrational band. A strong CARS signal is backscattered from an intense forward generated CARS signal in thick samples. This makes noninvasive imaging of deep structures possible, without labeling or chemical treatments.

Femtosecond coherent anti-Stokes Raman-scattering polarization beat spectroscopy of I2–Xe complex in solid krypton

Time-resolved coherent anti-Stokes Raman-scattering (CARS) measurements are carried out to study the interaction between xenon atom and iodine molecule in a solid krypton matrix. Interference between the CARS polarizations of the "free" and complexed iodine molecules is observed, while the quantum beats of the complex are not detected due to low concentration. Vibrational analysis based on the polarization beats yields accurate molecular constants for the I2-Xe complex. The harmonic frequency of the I2-Xe complex is found to be redshifted by 0.90 cm-1 when compared to the free I2, whereas the anharmonicity is approximately the same. The dephasing rate of the complex is found to be somewhat higher than that of the free iodine molecule in solid Kr, showing that the complexation affects dephasing, although not dramatically. Molecular dynamics simulations are carried out to find the conformation of the complex, and wave packet simulations are used to reproduce the CARS signal to confirm the assignments of the observed beatings as quantum and polarization beats. The results show that the polarization beats are a useful tool for investigating weak interactions in condensed phase.

Refractive effects in coherent anti-Stokes Raman scattering microscopy

Coherent anti-Stokes Raman scattering (CARS) microscopy with high sensitivity and high three- dimensional resolution has been developed for the vibrational imaging of chemical species. Due to the coherent nature of the CARS emission, it has been reported that the detection of epi-CARS and forward-CARS (F-CARS) signals depends on the size and shape of the sample. We investigate theoretically and experimentally the effects on the CARS signal of refractive index mismatches between the sample and its surroundings. Backward-CARS and F-CARS signals are measured for different polystyrene bead diameters embedded in different refractive index solvents. We show that index mismatches result in a backward-reflected F-CARS signal that generally dominates the experimentally backward-detected signal. Simulations based on geometrical and wave optics comparing forward- and backward-detected signals for polystyrene beads embedded in different index solvents confirm our findings. Furthermore, we demonstrate that the maxima of forward- and backward-detected signals are generated at different positions along the optical axis in the sample if refractive index mismatches are present between the sample and its surroundings.

Homogeneous Raman gain saturation at high pump and Stokes powers

A Raman gain spectral profile has been measured in a phosphosilicate fiber at high pump and Stokes (signal) wave powers by a coherent anti-Stokes Raman scattering (CARS) technique. It has been shown that the profile saturates homogeneously. The main saturation mechanism is proved to be the pump depletion; i.e., the Raman gain coefficient gR does not depend on the pump and signal wave power of several watts. The possible influence of stimulated Brillouin scattering and four-wave mixing is discussed. In addition, a transient regime of the CARS signal has been experimentally observed and theoretically explained.

Development of simultaneous frequency- and time-resolved coherent anti-Stokes Raman scattering for ultrafast detection of molecular Raman spectra

The development of a time-resolved coherent anti-Stokes Raman scattering (CARS) variant for use as a probe of excited electronic state Raman-active modes following excitation with an ultrafast pump pulse is detailed. Application of this technique involves a combination of broadband fs-time scale pulses and a narrowband pulse of ps duration that allows multiplexed detection of the CARS signal, permitting direct observation of molecular Raman frequencies and intensities with time resolution dictated by the broadband pulses. Thus, this nonlinear optical probe, designated fs/ps CARS, is suitable for observation of Raman spectral evolution following excitation with a pump pulse. Because of the spatial separation of the CARS output signal relative to the three input beams inherent in a folded BOXCARS arrangement, this technique is particularly amenable to probing low-frequency vibrational modes, which play a significant role in accepting vibrational energy during intramolecular vibrational energy redistribution within electronically excited states. Additionally, this spatial separation allows discrimination against strong fluorescence signal, as demonstrated in the case of rhodamine 6G.

Evidence for mesoscopic local structures in ionic liquids: CARS signal spatial distribution of C nmim[PF 6] ( n = 4, 6, 8)

We measure the spatial distribution of the coherent anti-Stokes Raman scattering (CARS) signal from three room-temperature ionic liquids, 1-alkyl-3-methylimidazolium hexafluorophosphates, where alkyl is butyl, hexyl, and octyl. The spatial distribution pattern becomes narrower as the alkyl-chain length increases. This result suggests that, although these ionic liquids are uniform and transparent macroscopically, specific local structures are formed with their size increasing with the alkyl-chain length. From the broadening mechanism of the CARS spatial distribution, it is most likely that the size of local structures in these ionic liquids is in the range of several tens of nanometers.

Single-pulse coherent anti-Stokes Raman scattering microscopy employing an octave spanning pulse

We demonstrate two complementary types of microscopy using an identical setup for single-pulse coherent anti-Stokes Raman scattering (CARS) imaging, which employs an ultrabroadband laser pulse with a spectral bandwidth of 4800 cm(-1) and enables the suppression of nonresonant CARS signals. One is a novel type of microscopy that uses spectral phase modulation for the selective excitation of a single Raman mode. The selective excitation is achieved by the modulated pulse focusing its difference-frequency spectrum into a narrow spectral region. Another type is Fourier-transform CARS (FT-CARS) microspectroscopy based on the measurement of the CARS spectrum obtained from the Fourier-transform of the interferometric autocorrelation (IAC) signal. Vibrational spectral imaging of chemical and biological samples is demonstrated using the two types of microscopy.

High-sensitivity vibrational imaging with frequency modulation coherent anti-Stokes Raman scattering (FM CARS) microscopy

We demonstrate a new approach to coherent anti-Stokes Raman scattering (CARS) microscopy that significantly increases the detection sensitivity. CARS signals are generated by collinearly overlapped, tightly focused, and raster scanned pump and Stokes laser beams, whose difference frequency is rapidly modulated. The resulting amplitude modulation of the CARS signal is detected through a lock-in amplifier. This scheme efficiently suppresses the nonresonant background and allows for the detection of far fewer vibrational oscillators than possible through existing CARS microscopy methods.

CARS microscopy in a wide‐field geometry with nanosecond pulses

AbstractWe demonstrate coherent anti‐Stokes Raman scattering (CARS) microscopy in a wide‐field geometry and discuss the reasons why this approach is capable of becoming a complementary method to confocal scanning CARS microscopy. By means of nanosecond laser pulses and a non‐confocal excitation geometry, the CARS signal is generated over a sample area of approximately 50 µm in diameter. As an example, we recorded images of oil drops within finely sliced sunflower seeds by tuning the laser frequency to the strong Raman transition of linoleic acid around 2870 cm−1. Meaningful images with spectral selectivity could already be recorded using only a few nanosecond pulses. Copyright © 2006 John Wiley & Sons, Ltd.

Optical nonlinearities of nanocomposite constituents selectively addressed by polarization‐controlled coherent anti‐Stokes Raman scattering

AbstractPolarization coherent anti‐Stokes Raman scattering (CARS) is shown to provide a means to selectively and independently address optical nonlinearities of materials constituting a multicomponent nanostructure, thus suggesting the way to tailor the nonlinear‐optical response of nanocomposite materials. Polarization properties of CARS in porous glass infiltrated with a toluene solution are experimentally studied. Rotation of the polarization analyzer with respect to the polarization vectors of the linearly polarized pump fields is shown to radically modify CARS line profiles, indicating changes in the ratio of CARS signal components related to toluene molecules and the porous glass host. Copyright © 2006 John Wiley & Sons, Ltd.

Dynamical Symmetry Breaking Induced by Ultrashort Laser Pulses in KTaO3

Raman selection rules were observed to be violated dynamically under resonant and coherent pumping of the Raman-active two-phonon mode in the cubic perovskite-type oxide KTaO 3 . In this study, two intense femtosecond laser pulses having near infrared frequencies were used to excite the crystal with a relative time delay within ±200 fs. The frequency difference of the pulses was set to be almost equal to the frequency of one of the Raman-active two-phonon modes, with both phonons being at the Brillouin-zone (BZ) edge point. As a result, not only the conventional multistep coherent anti-Stokes Raman scattering (CARS) signals, but a new series of multistep signals with the frequency spacing of originally Raman-inactive single phonons were observed in the wavelength range from near infrared to the whole visible region. Importantly, the observed frequency spacing of the multistep signals does not correspond to the single-phonon frequency at the Γ-point but that at the BZ-edge point. In addition, the angle spacing of the observed signals is almost constant; it is also quite different from the angle distribution of the conventional CARS signals. A model mechanism of BZ folding from the BZ edge to the Γ-point due to coherent accumulation of two phonons at the BZ edge is presented to explain all aspects of this new phenomenon. Two phonons at the BZ edge and those at the Γ-point are anharmonically coupled within a decay time of coherent phonons, and those originally at the BZ edge become Raman-active. This novel phenomenon should provide a new tool to observe phonon modes at the BZ edge other than inelastic neutron scattering.

Fourier-transform coherent anti-Stokes Raman scattering microscopy

We report a novel Fourier-transform-based implementation of coherent anti-Stokes Raman scattering (CARS) microscopy. The method employs a single femtosecond laser source and a Michelson interferometer to create two pulse replicas that are fed into a scanning multiphoton microscope. By varying the time delay between the pulses, we time-resolve the CARS signal, permitting easy removal of the nonresonant background while providing high resolution, spectrally resolved images of CARS modes over the laser bandwidth (approximately 1500 cm(-1)). We demonstrate the method by imaging polystyrene beads in solvent.