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A burst-mode laser system is developed for hybrid femtosecond/picosecond (fs/ps) rotational coherent anti-Stokes Raman scattering (RCARS) at megahertz rates. Using a common fs oscillator, the system simultaneously generates time synchronized 1061 nm, 274 fs and 1064 nm, 15.5 ps pulses with peak powers of 350 MW and 2.5 MW, respectively. The system is demonstrated for two-beam fs/ps RCARS in N 2 at 1 MHz with a signal-to-noise ratio of 176 at room temperature. This repetition rate is an order of magnitude higher than previous CARS using burst-mode ps laser systems and two to three orders of magnitude faster than previous continuously pulsed fs or fs/ps laser systems.

A diode pumped Nd:YAG narrowband spectral amplifier was demonstrated to generate high energy picosecond probe pulses for hybrid femtosecond/picosecond rotational coherent antiStokes Raman scattering thermometry measurements in a calibration flame and rotating detonation engine exhaust.

A burst-mode nitrogen ( N 2 ) picosecond vibrational coherent anti-Stokes Raman scattering (ps-VCARS) system is presented for accurate flame thermometry at 100 kHz repetition rate. A frequency-tripled ps burst-mode laser is used to pump a custom optical parametric generator/amplifier to produce 607 nm broadband Stokes pulses with 120 c m − 1 bandwidth, along with a narrowband 532 nm pump/probe beam. A simultaneous shot-to-shot nonresonant background (NRB) measurement is implemented to account for Stokes spectral profile and beam overlap fluctuations. The 100 kHz ps-VCARS data are benchmarked in a near-adiabatic C H 4 / a i r Hencken calibration flame with an accuracy of 1.5% and precision of 4.7% up to peak flame temperatures. The use of N 2 VCARS and simultaneous NRB measurements enables high-speed thermometry for a wide range of fuels and combustion applications.

Spatio-temporally resolved measurements of temperature using hybrid femtosecond/picosecond rotational coherent anti-Stokes Raman scattering (fs/ps RCARS) are evaluated for characterizing the highly dynamic exhaust flow of a non-premixed hydrogen-air rotating detonation combustor (RDC). The RCARS system utilizes a recently developed kHz-rate probe-pulse amplification system that enables high probe-pulse energies and sufficient sensitivity to track RDC exhaust gas temperatures during the short ~1.5 s run time with a precision of ~2%. Because of the potential for high spatial gradients in temperature and pressure in the RDC exhaust, estimation of bias errors due to spatial averaging in the 700-µm-long RCARS probe volume is conducted by employing the results of a reactive three-dimensional unsteady Reynolds-averaged Navier-Stokes (URANS) model with a structured grid of 48.5 million cells. This results in a potential bias error of ~1.5% due to exhaust temperature gradients and underscores the need for high spatial resolution. The experimental and predicted exhaust temperature histograms show good correspondence with a statistically similar skew-normal distribution relevant to the flow's local dynamical features. By utilizing a high-speed camera synchronized with the RCARS system, it was possible to compare the numerical simulation results with the measured exhaust temperature profile obtained from knowledge of the instantaneous detonation-wave azimuth position. Similar azimuthal spatial variations of ~300 K were observed in the experimental and computed temperatures, indicating a relatively well-mixed exhaust flow. The temperature pattern factors of 0.19 and 0.20 obtained from the experimental and numerical data, respectively, are relatively close to isobaric combustors in modern gas turbine engines. These results illustrate the ability of the fs/ps RCARS and numerical modeling approaches to evaluate characteristics of the RDC exhaust flow for future development in propulsion and power generation systems.

Ignition processes in scramjet pilot cavities are highly transient events that are influenced by factors including freestream Mach number and inlet geometry, turbulence intensity, cavity geometry, ignition source, and fueling composition and flow rate. In particular, the location of the flame kernel and associated propagation rate of the flame front throughout the cavity can significantly influence the end state of the ignition process. In this work formaldehyde (CH2O) was used as a flame marker to track ignition progress in a plane throughout the span-wise width of the cavity, while chemiluminescence imaging provided path-integrated flame location along the span-wise and axial directions. Planar laser-induced fluorescence (PLIF) excitation utilized the 355 nm frequency-tripled output of an Nd:YAG burst-mode laser operating at 50–100 kHz over 10 ms with available pulse energy up to 80 mJ. Simultaneous CH* chemiluminescence imaging from the top of the cavity was obtained with a high-speed complementary metal-oxide semiconductor camera. A freestream Mach number of 2 with ethylene fuel rates from 55–90 standard liters per minute were examined along with two different ignition sources: a spark discharge and pulse detonator. The resulting formaldehyde PLIF and chemiluminescence images indicate a strong correlation between fueling rate and the delay between the onset of ignition and stable combustion. More importantly, the span-wise propagation rate and structure of the flame front is highly dependent on the fueling rate, burning region, and ignition source.

Nanosecond electronic-resonance-enhanced coherent anti-Stokes Raman scattering (ERE-CARS) is evaluated for the measurement of formaldehyde ( C H 2 O ) concentrations in reacting and nonreacting conditions. The three-color scheme utilizes a 532 nm pump beam and a scanned Stokes beam near 624 nm for Raman excitation of the C–H symmetric stretch ( ν 1 ) vibrational mode; further, a 342 nm resonant probe is tuned to produce the outgoing CARS signal via the 1 0 1 4 0 3 vibronic transition between the ground ( X ~ 1 A 1 ) and first excited ( A ~ 1 A 2 ) electronic states. This allows detection of C H 2 O at concentrations as low as 9 × 10 14 m o l e c u l e s / c m 3 (55 parts per million) in a calibration cell with C H 2 O and N 2 at 1 bar and 450 K with 3% uncertainty. The measurements show a quadratic dependence of the signal with C H 2 O number density. Pressure scaling experiments up to 11 bar in the calibration cell show an increase in signal up to 8 bar. We study pressure dependence up to 11 bar and further apply the technique to characterize the C H 2 O concentration in an atmospheric premixed dimethyl ether/air McKenna burner flame, with a maximum concentration uncertainty of 11%. This approach demonstrates the feasibility for spatially resolved measurements of minor species such as C H 2 O in reactive environments and shows promise for application in high-pressure combustors.

Optical diagnostics of highly dynamic supersonic and hypersonic flows requires laser sources with a combination of high pulse intensities and fast repetition rates. A burst-mode Nd:YAG laser system is presented for increasing the overall energy of 532 nm pulse trains by ∼ 100 × and the number of high-energy pulses by 30 × for extended duration megahertz-rate flow diagnostics. At a lower repetition rate of 100 kHz, unprecedented energies near 1 J/pulse are achieved at 532 nm over a 1.1 ms burst. The laser performance is characterized and demonstrated for megahertz-rate laser-induced breakdown spectroscopy in a Mach 2 turbulent jet.

The sensitivity of high-repetition-rate hybrid femtosecond/picosecond (fs/ps) rotational coherent anti-Stokes Raman scattering (RCARS) is strongly influenced by the energy available for the ps probe pulse. In this work, a high-energy ps probe pulse that is time-synchronized with the fs pump/Stokes pulse is achieved by using a diode-pumped Nd:YAG amplifier seeded at 1064.4 nm by the output of a fs optical parametric amplifier. Nearly transform-limited, 10 ps pulses with up to 800 µJ/pulse and a bandwidth of 1.9 c m − 1 were generated at the second harmonic 532.2 nm and used for kilohertz-rate fs/ps RCARS thermometry up to 2400 K with accuracies of 1–2%. We furthermore demonstrate the amplification of variable pulsewidths for flexible single-mode (chirp-free) RCARS signal generation.

Quantitative, kiloherz-rate measurement of carbon monoxide mole fractions by femtosecond two-photon, laser-induced fluorescence (TP-LIF) was demonstrated in high-pressure, luminous flames over a range of fuel-air ratios. Femtosecond excitation at 230.1 nm was used to pump CO two-photon rovibrational X1Σ+→B1Σ+ transitions in the Hopfield–Birge system and avoid photolytic interferences with excitation irradiance ∼1.7×1010 W/cm2. The effects of excitation wavelength, detection scheme, and potential sources of de-excitation were also assessed to optimize the signal-to-background and signal-to-noise ratios and achieve excellent agreement with theoretically predicted CO mole fractions at low and high pressure.

High-repetition-rate interferometric-Rayleigh-scattering (IRS) velocimetry is implemented and demonstrated for non-intrusive, high-speed flow-velocity measurements. High temporal resolution is obtained with a quasi-continuous burst-mode laser that is capable of providing bursts of 10-msec duration with pulse widths of 10–100 nsec, pulse energy > 100 mJ at 532 nm, and repetition rates of 10–100 kHz. Coupled with a high-speed camera system, the IRS method is based on imaging the flow field though an etalon with 8-GHz free spectral range and capturing the Doppler shift of the Rayleigh-scattered light from the flow at multiple points having constructive interference. The seed-laser linewidth permits delivery of a laser linewidth of < 150 MHz at 532 nm The technique is demonstrated in a high-speed jet, and high-repetition-rate image sequences are shown.

Laser induced plasmas have found numerous applications including plasma-assisted combustion, combustion diagnostics, laser induced breakdown spectroscopy, light detection and ranging techniques (LIDAR), microwave guiding, reconfigurable plasma antennae etc. Multiphoton ionization (MPI) is a fundamental first step in high-energy laser-matter interaction and is important for understanding of the mechanism of plasma formation. With the discovery of MPI more than 50 years ago, there were numerous attempts to determine basic physical constants of this process in the direct experiments, namely photoionization rates and cross-sections of the MPI, however, no reliable data is available until today and spread in the literature values often reaches 2-3 orders of magnitude. This is due to inability to conduct absolute measurements of plasma electron numbers generated by MPI which leads to uncertainties and, sometimes, contradictions between the MPI cross-section values utilized by different researchers across the field. Here we report first direct measurement of absolute plasma electron numbers generated at MPI of air and subsequently we precisely determine ionization rate and cross-section of eight-photon ionization of oxygen molecule by 800 nm photons ${\sigma}_8=(3.32{\pm}0.3)*10^{-130} W^{-8}m^{16}s^{-1}$. Method is based on the absolute measurement of electron number created by MPI using elastic scattering of microwaves off the plasma volume in Rayleigh regime and establishes a general approach to directly measure and tabulate basic constants of the MPI process for various gases and photon energies.

Simultaneous 10-kHz OH-PLIF/CH2O-PLIF/PIV (Rayleigh scattering) measurements in DRL-A non-premixed flames were demonstrated using a three-leg burst-mode laser system. High-speed multi-species concentrations, heat-release rate, and flow velocity field (temperature) were measured.

Quantitative measurement of electron number density in laser-induced plasma filament at atmospheric pressure was made using Rayleigh microwave scattering (RMS)1, 2. Plasma filament was produced by focusing 800 nm Ti:Sapphire laser pulses of hundred femtoseconds duration. Electron number densities down to about $10 ^{13}- 10 ^{14}$ cm $^{-3}$ were successfully ascertained using RMS technique. This is about three orders of magnitude below the minimal plasma density measurable by laser interferometry. Traditional laser interferometry for diagnostics of such rarified plasmas is arduous due to very small shifts of the interference fringes. Size of the microplasma filament was determined via imaging through an ICCD camera. The measured electron number density was the average density in the plasma volume and its variation temporally resolved to nanosecond scale. Capability to measure plasma density in rarified plasma filaments paves new ways for their utilization in diagnostic purposes.

Conspectus Traditionally, molecules are analyzed in a test tube. Taking biochemistry as an example, the majority of our knowledge about cellular content comes from analysis of fixed cells or tissue homogenates using tools such as immunoblotting and liquid chromatography–mass spectrometry. These tools can indicate the presence of molecules but do not provide information on their location or interaction with each other in real time, restricting our understanding of the functions of the molecule under study. For real-time imaging of labeled molecules in live cells, fluorescence microscopy is the tool of choice. Fluorescent labels, however, are too bulky for small molecules such as fatty acids, amino acids, and cholesterol. These challenges highlight a critical need for development of chemical imaging platforms that allow in situ or in vivo analysis of molecules. Vibrational spectroscopy based on spontaneous Raman scattering is widely used for label-free analysis of chemical content in cells and tissues. However, the Raman process is a weak effect, limiting its application for fast chemical imaging of a living system. With high imaging speed and 3D spatial resolution, coherent Raman scattering microscopy is enabling a new approach for real-time vibrational imaging of single cells in a living system. In most experiments, coherent Raman processes involve two excitation fields denoted as pump at ωp and Stokes at ωs. When the beating frequency between the pump and Stokes fields (ωp – ωs) is resonant with a Raman-active molecular vibration, four major coherent Raman scattering processes occur simultaneously, namely, coherent anti-Stokes Raman scattering (CARS) at (ωp – ωs) + ωp, coherent Stokes Raman scattering (CSRS) at ωs – (ωp – ωs), stimulated Raman gain (SRG) at ωs, and stimulated Raman loss (SRL) at ωp. In SRG, the Stokes beam experiences a gain in intensity, whereas in SRL, the pump beam experiences a loss. Both SRG and SRL belong to stimulated Raman scattering (SRS), in which the energy difference between the pump and Stokes fields is transferred to the molecule for vibrational excitation. The SRS signal appears at the same wavelengths as the excitation fields and is commonly extracted through a phase-sensitive detection scheme. The detected intensity change because of a Raman transition is proportional to Im[χ(3)]IpIs, where χ(3) represents the third-order nonlinear susceptibility, Ip and Is stand for the intensity of the pump and Stokes fields. In this Account, we discuss the most recent advances in the technical development and enabling applications of SRS microscopy. Compared to CARS, the SRS contrast is free of nonresonant background. Moreover, the SRS intensity is linearly proportional to the density of target molecules in focus. For single-frequency imaging, an SRS microscope offers a speed that is ∼1000 times faster than a line-scan Raman microscope and 10 000 times faster than a point-scan Raman microscope. It is important to emphasize that SRS and spontaneous Raman scattering are complementary to each other. Spontaneous Raman spectroscopy covers the entire window of molecular vibrations, which allows extraction of subtleties via multivariate analysis. SRS offers the speed advantage by focusing on either a single Raman band or a defined spectral window of target molecules. Integrating single-frequency SRS imaging and spontaneous Raman spectroscopy on a single platform allows quantitative compositional analysis of objects inside single live cells.

100-kHz particle image velocimetry (PIV) is demonstrated using a double-pulsed, burst-mode laser with a burst duration up to 100 ms. This enables up to 10,000 time-sequential vector fields for capturing a temporal dynamic range spanning over three orders of magnitude in high-speed turbulent flows. Pulse doublets with inter-pulse spacing of 2 µs and repetition rate of 100 kHz are generated using a fiber-based oscillator and amplified through an all-diode-pumped, burst-mode amplifier. A physics-based model of pulse doublet amplification in the burst-mode amplifier is developed and used to accurately predict oscillator pulse width and pulse intensity inputs required to generate equal-energy pulse doublets at 532 nm for velocity measurements. The effect of PIV particle response and high-speed-detector limitations on the spatial and temporal resolution are estimated in subsonic turbulent jets. An effective spatial resolution of 266–275 µm and temporal resolution of 10 µs are estimated from the 8 × 8 pixel correlation window and inter-doublet time spacing, respectively. This spatiotemporal resolution is sufficient for quantitative assessment of integral time and length scales in highly turbulent jets with Reynolds numbers in the range 15,000–50,000. The temporal dynamic range of the burst-mode PIV measurement is 1200, limited by the 85-ms high-energy portion of the burst and 30-kHz high-frequency noise limit.

Two-dimensional gas-phase coherent anti-Stokes Raman scattering (2D-CARS) thermometry is demonstrated at 1 kHz in a heated jet. A hybrid femtosecond/picosecond CARS configuration is used in a two-beam phase-matching arrangement with a 100-femtosecond pump/Stokes pulse and a 107-picosecond probe pulse. The femtosecond pulse is generated using a mode-locked oscillator and regenerative amplifier that is synchronized to a separate picosecond oscillator and burst-mode amplifier. The CARS signal is spectrally dispersed in a custom imaging spectrometer and detected using a high-speed camera with image intensifier. 1-kHz, single-shot planar measurements at room temperature exhibit error of 2.6% and shot-to-shot variations of 2.6%. The spatial variation in measured temperature is 9.4%. 2D-CARS temperature measurements are demonstrated in a heated Osub2/subjet to capture the spatiotemporal evolution of the temperature field.

Simultaneous high-speed quantitative imaging of mixture fraction and velocity is demonstrated using the fourth- and second-harmonic outputs, respectively, of a burst-mode Nd:YAG laser. A tenfold increase in the record length and 16-fold increase in per-pulse energy are achieved compared with previous measurements of mixture fraction using burst-mode and continuously pulsed diode-pumped solid-state lasers, respectively. The high output energy is used for quantitative, high-speed mixture-fraction imaging with acetone planar laser-induced fluorescence, which also enables simultaneous particle-based velocimetry without interference from particle scattering. A semiquantitative model is used to determine the limitations on fourth-harmonic output energy due to the effects of transient absorption and thermal phase mismatch over a range of repetition rates. Data are presented for mixing within a turbulent jet (Reynolds number of 15,000) and are validated by comparisons with known turbulent mixing laws and previously published data.

Gout is a disease process where the nucleation and growth of crystals in the synovial fluid of joints elicit painful arthritis-like symptoms. Raman spectroscopy is evolving as a potential diagnostic tool in identifying such crystals; however, attainment of sufficient Raman signal while overcoming the background fluorescence remains as a major challenge. The current study focused on assessing whether excitation in 532-700 nm range will provide greater signal intensity than the standard 785 nm while not being impeded by background fluorescence. We characterized the fluorescence spectra, absorption spectra and Raman spectra of synovial fluid from patients who presented "gout-like symptoms" (symptomatic) and controls (asymptomatic). A digestion and filtration method was developed to isolate crystals from synovial fluid while reducing the organic burden. Spectral profile and photobleaching dynamics during Raman spectroscopy were observed under an excitation wavelength range spanning 532 to 785 nm. Absorbance and fluorescence profiles indicated the digestion and filtration worked effectively to extract crystals from symptomatic synovial fluid without introducing additional fluorescence. Raman spectral analyses at 532 nm, 660 nm, 690 nm and 785 nm indicated that both asymptomatic and symptomatic samples had significant levels of fluorescence at excitation wavelengths below 700 nm, which either hindered the collection of Raman signal or necessitated prolonged durations of photobleaching. Raman-based diagnostics were more feasible at the longest excitation wavelength of 785 nm without employing photobleaching. This study further demonstrated that a near-infrared OEM based lower-cost Raman system at 785 nm excitation has sufficient sensitivity to identify crystals isolated from the synovial fluid. In conclusion, while lower excitation wavelengths provide greater signal, the fluorescence necessitates near-infrared wavelengths for Raman analysis of crystal species observed in synovial aspirates.

Nitrogen oxide clusters (NO)n have been studied in He droplets via infrared depletion spectroscopy and by quantum chemical calculations. The ν1 and ν5 bands of cis-ON-NO dimer have been observed at 1868.2 and 1786.5 cm(-1), respectively. Furthermore, spectral bands of the trimer and tetramer have been located in the vicinity of the corresponding dimer bands in accord with computed frequencies that place NO-stretch bands of dimer, trimer, and tetramer within a few wavenumbers of each other. In addition, a new line at 1878.1 cm(-1) close to the band origin of single molecules was assigned to van der Waals bound dimers of (NO)2, which are stabilized due to the rapid cooling in He droplets. Spectra of larger clusters (n > 5), have broad unresolved features in the vicinity of the dimer bands. Experiments and calculations indicate that trimers consist of a dimer and a loosely bound third molecule, whereas the tetramer consists of two weakly bound dimers.

We demonstrated high-speed flame temperature measurements with high temporal and spatial resolution based on coherent anti-Stokes Raman scattering (CARS) spectroscopy at a rate of 100 kHz employing a picosecond burst-mode laser.

Because each nonlinear optical (NLO) imaging modality is sensitive to specific molecules or structures, multimodal NLO imaging capitalizes the potential of NLO microscopy for studies of complex biological tissues. The coupling of multiphoton fluorescence, second harmonic generation, and coherent anti-Stokes Raman scattering (CARS) has allowed investigation of a broad range of biological questions concerning lipid metabolism, cancer development, cardiovascular disease, and skin biology. Moreover, recent research shows the great potential of using CARS microscope as a platform to develop more advanced NLO modalities such as electronic-resonance-enhanced four-wave mixing, stimulated Raman scattering, and pump-probe microscopy. This article reviews the various approaches developed for realization of multimodal NLO imaging as well as developments of new NLO modalities on a CARS microscope. Applications to various aspects of biological and biomedical research are discussed.

Femtosecond laser electronic excitation tagging (FLEET) velocimetry is characterized for the first time at high-pressure, low-temperature conditions. FLEET signal intensity and signal lifetime data are examined for their thermodynamic dependences; temperatures range from 89 K to 275 K while pressures are varied from 85 kPa to 400 kPa. The FLEET signal intensity is found to scale linearly with the flow density. An inverse density dependence is observed in the FLEET signal lifetime data, with little independent sensitivity to the other thermodynamic conditions apparent. FLEET velocimetry is demonstrated in the NASA Langley 0.3-m Transonic Cryogenic Tunnel. Velocity measurements are made over the entire operational envelope: Mach numbers from 0.2 to 0.75, total (stagnation) temperatures from 100 K to 280 K, and total pressures from 100 kPa to 400 kPa. The velocity measurement accuracy is assessed over this domain of conditions. Measurement errors below 1.15 percent are typical, with slightly decreasing accuracy as temperatures are decreased. Assessment of the measurement precision finds a zero-velocity precision of 0.4 m/s. The precision is observed to have a weak temperature dependence as well, likely a result of the shorter lifetimes experienced at higher densities. The velocity dynamic range is found to have a nominal value of 650. Finally the spatial resolution of the measurements is found to be a dominated by the physical size of the FLEET signal and advective motion. The transverse spatial resolution is found to be 1 mm, while the streamwise spatial resolution is dependent on velocity with a minimum of 2 mm and a maximum of 3.3 mm.

Interference-free hybrid femtosecond/picosecond vibrational coherent anti-Stokes Raman scattering (CARS) of nitrogen is reported for temperature measurements of 1300–2300 K in high-pressure, laminar H2–air and CH4–air diffusion flames up to 10 bar. Following coherent Raman excitation by 100 fs duration pump and Stokes pulses, a time-asymmetric probe pulse is used for the detection of spectrally resolved N2 CARS signals at probe delays as early as ∼200–300 fs. This allows for full rejection of nonresonant contributions while being independent of collisions for single-shot precision of ±2% at elevated pressures. The effects of collisions at longer probe-pulse delays are also investigated to determine the feasibility of varying the detection timing from 200 fs to 100 ps.

Cells store excess energy in the form of cytoplasmic lipid droplets. At present, it is unclear how different types of fatty acids contribute to the formation of lipid droplets. We describe a compound Raman microscope capable of both high-speed chemical imaging and quantitative spectral analysis on the same platform. We used a picosecond laser source to perform coherent Raman scattering imaging of a biological sample and confocal Raman spectral analysis at points of interest. The potential of the compound Raman microscope was evaluated on lipid bodies of cultured cells and live animals. Our data indicate that the in vivo fat contains much more unsaturated fatty acids (FAs) than the fat formed via de novo synthesis in 3T3-L1 cells. Furthermore, in vivo analysis of subcutaneous adipocytes and glands revealed a dramatic difference not only in the unsaturation level but also in the thermodynamic state of FAs inside their lipid bodies. Additionally, the compound Raman microscope allows tracking of the cellular uptake of a specific fatty acid and its abundance in nascent cytoplasmic lipid droplets. The high-speed vibrational imaging and spectral analysis capability renders compound Raman microscopy an indispensible analytical tool for the study of lipid-droplet biology.

Temperature measurements based on gas-phase coherent anti-Stokes Raman scattering (CARS) spectroscopy are demonstrated in reacting flows at a rate of 100 kHz employing a burst-mode laser with a pulse duration of ∼100 ps. The recently developed picosecond-duration, high-energy burst-mode laser is used to pump an optical parametric generator/optical parametric amplifier that produces broadband light centered at ∼680 nm to provide the Stokes beams for excitation of the rovibrational Raman transitions of H(2). The 532-nm output of the picosecond burst-mode laser is then utilized as a pump beam for the CARS process that generates 100 single-shot spectra at a rate of 100 kHz during the 1-ms duration burst. Coherent spectroscopy-based temperature measurements at 100 kHz will significantly aid the understanding of transient and unsteady flow phenomena related to turbulent combustion, transonic and hypersonic flows, high-enthalpy flows, and the dynamics of energetic materials.

Picosecond-laser electronic-excitation tagging (PLEET), a seedless picosecond-laser-based velocimetry technique, is demonstrated in non-reactive flows at a repetition rate of 100 kHz with a 1064 nm, 100 ps burst-mode laser. The fluorescence lifetime of the PLEET signal was measured in nitrogen, and the laser heating effects were analyzed. PLEET experiments with a free jet of nitrogen show the ability to measure multi-point flow velocity fluctuations at a 100 kHz detection rate or higher. Both spectral and dynamic mode decomposition analyses of velocity on a Ma=0.8 free jet show two dominant Strouhal numbers around 0.24 and 0.48, respectively, well within the shear-layer flapping frequencies of the free jets. This technique increases the laser-tagging repetition rate for velocimetry to hundreds of kilohertz. PLEET is suitable for subsonic through supersonic laminar- and turbulent-flow velocity measurements.

The hybrid femtosecond/picosecond coherent anti-Stokes Raman scattering (fs/ps CARS) technique presents a promising alternative to either fs time-resolved or ps frequency-resolved CARS in both gas-phase thermometry and condensed-phase excited-state dynamics applications. A theoretical description of time-dependent CARS is used to examine this recently developed probe technique, and quantitative comparisons of the full time–frequency evolution show excellent accuracy in predicting the experimental vibrational CARS spectra obtained for two model systems. The interrelated time- and frequency-domain spectral signatures of gas-phase species produced by hybrid fs/ps CARS are explored with a focus on gas-phase N2 vibrational CARS, which is commonly used as a thermometric diagnostic of combusting flows. In particular, we discuss the merits of the simple top-hat spectral filter typically used to generate the ps-duration hybrid fs/ps CARS probe pulse, including strong discrimination against non-resonant background that often contaminates CARS signal. It is further demonstrated, via comparison with vibrational CARS results on a time-evolving solvated organic chromophore, that this top-hat probe-pulse configuration can provide improved spectral resolution, although the degree of improvement depends on the dephasing timescales of the observed molecular modes and the duration and timing of the narrowband final pulse. Additionally, we discuss the virtues of a frequency-domain Lorentzian probe-pulse lineshape and its potential for improving the hybrid fs/ps CARS technique as a diagnostic in high-pressure gas-phase thermometry applications.

We demonstrate a hyperspectral stimulated Raman scattering (SRS) microscope through spectral-transformed excitation. The 1064-nm Stokes pulse was from a synchronized time-lens source, generated through time-domain phase modulation of a continuous wave (CW) laser. The tunable pump pulse was from linear spectral filtering of a femtosecond laser output with an intra-pulse spectral scanning pulse shaper. By electronically modulating the time-lens source at 2.29 MHz, hyperspectral stimulated Raman loss (SRL) images were obtained on a laser-scanning microscope. Using this microscope, DMSO in aqueous solution with a concentration down to 28 mM could be detected at 2 μs time constant. Hyper-spectral SRL images of prostate cancer cells were obtained. Multivariate curve resolution analysis was further applied to decompose the SRL images into concentration maps of CH2 and CH3 bonds. This method offers exciting potential in label-free imaging of live cells using fingerprint Raman bands.

The stimulated Raman scattering signal is often accompanied by unwanted background arising from other pump-probe modalities. We demonstrate an approach to overcome this challenge based on spectral domain modulation, enabled by a compact, cost-effective angle-to-wavelength pulse shaper. The pulse shaper switches between two spectrally narrow windows, which are cut out of a broadband femtosecond pulse and selected for on- and off- Raman resonance excitation, at 2.1 MHz frequency for detection of stimulated Raman scattering signal. Such spectral modulation reduced the unwanted pump-probe signals by up to 20 times and enabled stimulated Raman scattering imaging of molecules in a pigmented environment.

We demonstrate a low-cost-stimulated Raman scattering (SRS) microscope using continuous-wave (cw) lasers as excitation sources. A dual modulation scheme is used to remove the electronic background. The cw-SRS imaging of lipids in fatty liver is demonstrated by excitation of C─H stretch vibration.

Photoacoustic imaging employing molecular overtone vibration as contrast mechanism opens a new avenue for deep tissue imaging with chemical bond selectivity. Here, we demonstrate vibration-based photoacoustic tomography with an imaging depth on the centimeter scale. To provide sufficient pulse energy at the overtone transition wavelengths, we constructed a compact, barium nitrite crystal-based Raman laser for excitation of 2nd overtone of C-H bond. Using a 5-ns Nd:YAG laser as pumping source, up to 105 mJ pulse energy at 1197 nm was generated. Vibrational photoacoutic spectroscopy and tomography of phantom (polyethylene tube) immersed in whole milk was performed. With a pulse energy of 47 mJ on the milk surface, up to 2.5 cm penetration depth was reached with a signal-to-noise ratio of 12.

Though single-color coherent Raman microscopy has been widely used for vibrational imaging of isolated Raman bands, it is still challenging to visualize molecules having overlapping Raman bands. We address this issue by developing a spectroscopic SRS microscope with a time-lens laser source synchronized to a femtosecond laser. The time-lens source provides 2-ps pulse at the wavelength of 1064 nm. A pulse shaper is installed for intra-pulse spectral scanning of the femtosecond laser output. By electronically modulating the time-lens source at MHz frequency, spectroscopic stimulated Raman loss (SRL) images were obtained on a laser-scanning microscope. Using this microscope, we have been able to detect 0.2% DMSO in aqueous solution. Spectroscopic SRL images of prostate cancer cells were obtained. Multivariate curve resolution analysis was further applied to decompose the SRL images into concentration maps of proteins and lipids. With high sensitivity and high spectral resolution, this method offers exciting potential in label-free imaging of live cells using fingerprint Raman bands.

Spectroscopic imaging has been an increasingly critical approach for unveiling specific molecules in biological environments. Towards this goal, we demonstrate hyperspectral stimulated Raman loss (SRL) imaging by intra-pulse spectral scanning through a femtosecond pulse shaper. The hyperspectral stack of SRL images is further analyzed by a multivariate curve resolution (MCR) method to reconstruct quantitative concentration images for each individual component and retrieve the corresponding vibrational Raman spectra. Using these methods, we demonstrate quantitative mapping of dimethyl sulfoxide concentration in aqueous solutions and in fat tissue. Moreover, MCR is performed on SRL images of breast cancer cells to generate maps of principal chemical components along with their respective vibrational spectra. These results show the great capability and potential of hyperspectral SRL microscopy for quantitative imaging of complicated biomolecule mixtures through resolving overlapped Raman bands.

We report a compact, cost-effective tuned amplifier for frequency-selective amplification of the modulated signal in heterodyne detected nonlinear optical microscopy. Our method improved the signal to noise ratio by an order of magnitude compared to conventional lock-in detection, as demonstrated through stimulated Raman scattering imaging of live cells and tissues at the speed of 2 μsec/pixel. Application of the tuned amplifier to transient absorption microscopy is also demonstrated. The increased signal to noise ratio allowed epi-detected in vivo imaging of myelin and blood in rat spinal cord with high spatial resolution.

High-repetition-rate, single-laser-shot measurements are important for the investigation of unsteady flows where temperature and species concentrations can vary significantly. Here, we demonstrate single-shot, pure-rotational, hybrid femtosecond/picosecond coherent anti-Stokes Raman scattering (fs/ps RCARS) thermometry based on a kHz-rate fs laser source. Interferences that can affect nanosecond (ns) and ps CARS, such as nonresonant background and collisional dephasing, are eliminated by selecting an appropriate time delay between the 100-fs pump/Stokes pulses and the pulse-shaped 8.4-ps probe. A time- and frequency-domain theoretical model is introduced to account for rotational-level dependent collisional dephasing and indicates that the optimal probe-pulse time delay is 13.5 ps to 30 ps. This time delay allows for uncorrected best-fit N2-RCARS temperature measurements with ~1% accuracy. Hence, the hybrid fs/ps RCARS approach can be performed with kHz-rate laser sources while avoiding corrections that can be difficult to predict in unsteady flows.

Hybrid femtosecond/picosecond coherent anti-Stokes Raman scattering (fs/ps CARS) offers accurate thermometry at kHz rates for combustion diagnostics. In high-temperature flames, selection of probe-pulse characteristics is key to simultaneously optimizing signal-to-nonresonant-background ratio, signal strength, and spectral resolution. We demonstrate a simple method for enhancing signal-to-nonresonant-background ratio by using a narrowband Lorentzian filter to generate a time-asymmetric probe pulse with full-width-half-maximum (FWHM) pulse width of only 240 fs. This allows detection within just 310 fs after the Raman excitation for eliminating nonresonant background while retaining 45% of the resonant signal at 2000 K. The narrow linewidth is comparable to that of a time-symmetric sinc2 probe pulse with a pulse width of ~2.4 ps generated with a conventional 4-f pulse shaper. This allows nonresonant-background-free, frequency-domain vibrational spectroscopy at high temperature, as verified using comparisons to a time-dependent theoretical fs/ps CARS model.

We demonstrate nonlinear vibrational imaging of isolated Raman bands by detecting femtosecond pulse stimulated Raman loss. Femtosecond pulse excitation produces a stimulated Raman loss signal that is 12 times larger than what picosecond pulse excitation produces. The strong signal allowe d real-time, bond-selective im aging of deuterated palmitic acid-d 31 inside live cells, and 3D sectioning of fat storage in live C. elegans. With the high peak power provided by femtosecond pulses, this system is highly compatible with other nonlinear optical modalities such as two-photon excited fluorescence. With most of the excitation power contributed by the Stokes beam in the 1.0 - 1.2 µm wavelength range, photodamage of biological samples was not observed. Keywords: stimulated Raman scattering (SRS), nonlinear microscopy, multimodal imaging, femtosecond laser, coherent Raman scattering, lipid INTRODUCTION Label-free chemically selective imaging based on inherent molecular vibrations is an attractive alternative to fluorescence imaging. Vibrational microscopes based on spontaneous Raman scattering and infrared (IR) absorption have been routinely used for chemical imaging of unstained samples. Nevertheless, the application of IR microscopy to live cell imaging has been hindered by the poor spatial resolution and IR absorption of water. Raman microscopy avoids these problems because water is a weak Raman scatterer. Moreover, a higher spatial resolution can be achieved by visible light excitation and confocal detection. However, Raman scattering has an extremely small cross section (~ 10

This work expands on previous studies to utilize hybrid femtosecond/picosecond coherent anti-Stokes Raman scattering (fs/ps CARS) in high-temperature flames for background-free thermometry. The goal of the current work is to quantify the precision and accuracy of the temperature measurements over an expanded temperature range from 1250 – 2400 K, while also quantifying the influence of nonresonant background on the measurements. Temporal suppression of the nonresonant background by three orders of magnitude while retaining up to 20% of the time-zero signal is demonstrated while the background-free single-shot spectra show a signal-to-noise ratio of ~ 200:1 at ~2400 K. The accuracy and precision of the singleshot temperature measurements are investigated over an expanded range of 1250 – 2400 K to address hybrid fs/ps CARS applicability to lower temperature flames. The resulting accuracy and precision of the measurements is better than 10% and 3.5%, respectively, at all conditions. Finally, the effect of the nonresonant background on the accuracy and precision of the temperature measurement is quantified at ~2400 K, and the need for nonresonant-free detection is discussed.

Hybrid fs/ps CARS is employed for time- and frequency-domain spectroscopy in reacting flows. Advantages include nonresonant background suppression, chemical specificity, detection at high pressures, potential for kHz-rate thermometry, and ability to resolve energy transfer phenomena.

We report on a femtosecond-OPO based multimodal nonlinear optical microscope that permits multiphoton fluorescence, SHG, THG, CARS, and stimulated Raman scattering imaging on the same platform.

We detail the development of an optical probe technique based on time-resolved Raman-induced Kerr effect polarization spectroscopy (tr-RIKES). This technique, termed fs/ps RIKES, combines an ultrafast pump pulse with a narrowband probe that directly allows spectral resolution of low-frequency (0-600 cm(-1)) modes typically observable via RIKES. The narrowband probe pulse alleviates the need to scan the time delay between pump and probe pulses to observe molecular coherences, thus making this multiplexed technique a convenient probe for studying low-frequency molecular dynamics. An important distinguishing characteristic of this polarization-sensitive technique arises from the fact that the delay between the impulsive pump pulse and the picosecond-duration probe pulse is optimized to maximize suppression of nonresonant background signal. Model systems, including the rotational spectrum of gas-phase hydrogen and the low-frequency vibrational spectrum of neat bromoform, are used to compare fs/ps RIKES with the conventional time-resolved RIKES technique.

Real-time vibrational spectroscopic imaging is desired for monitoring cellular states and cellular processes in a label-free manner. Raman spectroscopic imaging of highly dynamic systems is inhibited by relatively slow spectral acquisition on millisecond to second scale. Here, we report microsecond scale vibrational spectroscopic imaging by lock-in free parallel detection of spectrally dispersed stimulated Raman scattering signal. Using a homebuilt tuned amplifier array, our method enables Raman spectral acquisition, within the window defined by the broadband pulse, at the speed of 32 µs and with close to shot-noise limited detection sensitivity. Incorporated with multivariate curve resolution analysis, our platform allows compositional mapping of lipid droplets in single live cells, observation of intracellular retinoid metabolism, discrimination of fat droplets from protein-rich organelles in Caenorhabditis elegans, spectral detection of fast flowing tumor cells and monitoring drug diffusion through skin tissue in vivo. The reported technique opens new opportunities for compositional analysis of cellular compartment in a microscope setting and high-throughput spectral profiling of single cells in a flow cytometer setting. Vibrational spectroscopic imaging on a microsecond time scale has been realized using multiplex Raman microscopy. Real-time vibrational spectroscopic imaging holds out the promise of enabling cellular processes to be monitored without labeling, but this has been difficult to achieve because of the long acquisition times needed. Now, researchers at Purdue University in the USA have realized vibrational spectroscopic imaging by parallel detection of a spectrally dispersed stimulated Raman signal. Using a homemade system, they acquired Raman spectra in 32 microseconds with a detection sensitivity that is nearly shot-noise limited. The scientists illustrate the potential of the technique by using it to obtain compositional maps of lipid droplets in living single cells, observe intracellular retinoid metabolism, and discriminate between fat droplets and protein-rich organelles in a nematode.

Extended abstract of a paper presented at Microscopy and Microanalysis 2009 in Richmond, Virginia, USA, July 26 – July 30, 2009

Photoacoustic imaging employing molecular overtone vibration as a contrast mechanism opens a new avenue for bond-selective imaging of deep tissues. Broad use of this modality is, however, hampered by the extremely low conversion efficiency of optical parametric oscillators at the overtone transition wavelengths. To overcome such a barrier, we demonstrate the construction and use of a compact, barium nitrite crystal-based Raman laser for photoacoustic imaging of C–H overtone vibrations. Using a 5-ns Nd∶YAG laser as the pumping source, up to 21.4 mJ pulse energy at 1197 nm was generated, corresponding to a conversion efficiency of 34.8%. Using the 1197 nm pulses, three-dimensional photoacoustic imaging of intramuscular fat was demonstrated.

We report on multimodal coherent anti-Stokes Raman scattering (CARS) imaging with a source composed of a femtosecond fiber laser and a photonic crystal fiber (PCF)-based optical parametric oscillator (FOPO). By switching between two PCFs with different zero dispersion wavelengths, a tunable signal beam from the FOPO covering the range from 840 to 930 nm was produced. By combining the femtosecond fiber laser and the FOPO output, simultaneous CARS imaging of a myelin sheath and two-photon excitation fluorescence imaging of a labeled axons in rat spinal cord have been demonstrated at the speed of 20 μs per pixel.