ABSTRACT: Spectrally resolved photoacoustic imaging is promising for label-free imaging in optically scattering materials. However, this technique often requires acquisition of a separate image at each wavelength of interest. This reduces imaging speeds and causes errors if the sample changes in time between images acquired at different wavelengths. We demonstrate a solution to this problem by using dual-comb spectroscopy for photoacoustic measurements. This approach enables a photoacoustic measurement at thousands of wavelengths simultaneously. In this technique, two optical-frequency combs are interfered on a sample and the resulting pressure wave is measured with an ultrasound transducer. This acoustic signal is processed in the frequency-domain to obtain an optical absorption spectrum. For a proof-of-concept demonstration, we measure photoacoustic signals from polymer films. The absorption spectra obtained from these measurements agree with those measured using a spectrophotometer. Improving the signal-to-noise ratio of the dual-comb photoacoustic spectrometer could enable high-speed spectrally resolved photoacoustic imaging.
Project description:Optical frequency combs have developed into powerful tools for distance metrology. In this paper we demonstrate absolute long distance measurement using a single femtosecond frequency comb laser as a multi-wavelength source. By applying a high-resolution spectrometer based on a virtually imaged phased array, the frequency comb modes are resolved spectrally to the level of an individual mode. Having the frequency comb stabilized against an atomic clock, thousands of accurately known wavelengths are available for interferometry. From the spectrally resolved output of a Michelson interferometer a distance is derived. The presented measurement method combines spectral interferometry, white light interferometry and multi-wavelength interferometry in a single scheme. Comparison with a fringe counting laser interferometer shows an agreement within <10(-8) for a distance of 50 m.
Project description:Dual-comb spectroscopy is a powerful technique for real-time, broadband optical sampling of molecular spectra, which requires no moving components. Recent developments with microresonator-based platforms have enabled frequency combs at the chip scale. However, the need to precisely match the resonance wavelengths of distinct high quality-factor microcavities has hindered the development of on-chip dual combs. We report the simultaneous generation of two microresonator combs on the same chip from a single laser, drastically reducing experimental complexity. We demonstrate broadband optical spectra spanning 51 THz and low-noise operation of both combs by deterministically tuning into soliton mode-locked states using integrated microheaters, resulting in narrow (<10 kHz) microwave beat notes. We further use one comb as a reference to probe the formation dynamics of the other comb, thus introducing a technique to investigate comb evolution without auxiliary lasers or microwave oscillators. We demonstrate high signal-to-noise ratio absorption spectroscopy spanning 170 nm using the dual-comb source over a 20-?s acquisition time. Our device paves the way for compact and robust spectrometers at nanosecond time scales enabled by large beat-note spacings (>1 GHz).
Project description:Non-invasive imaging of biological processes in vivo is invaluable in advancing biology. Photoacoustic tomography is a scalable imaging technique that provides higher resolution at greater depths in tissue than achievable by purely optical methods. Here we report the application of two spectrally distinct near-infrared fluorescent proteins, iRFP670 and iRFP720, engineered from bacterial phytochromes, as photoacoustic contrast agents. iRFPs provide tissue-specific contrast without the need for delivery of any additional substances. Compared to conventional GFP-like red-shifted fluorescent proteins, iRFP670 and iRFP720 demonstrate stronger photoacoustic signals at longer wavelengths, and can be spectrally resolved from each other and hemoglobin. We simultaneously visualized two differently labeled tumors, one with iRFP670 and the other with iRFP720, as well as blood vessels. We acquired images of a mouse as 2D sections of a whole animal, and as localized 3D volumetric images with high contrast and sub-millimeter resolution at depths up to 8?mm. Our results suggest iRFPs are genetically-encoded probes of choice for simultaneous photoacoustic imaging of several tissues or processes in vivo.
Project description:Photo-acoustic spectroscopy (PAS) is one of the most sensitive non-destructive analysis techniques for gases, fluids and solids. It can operate background-free at any wavelength and is applicable to microscopic and even non-transparent samples. Extension of PAS to broadband wavelength coverage is a powerful tool, though challenging to implement without sacrifice of wavelength resolution and acquisition speed. Here we show that dual-frequency comb spectroscopy (DCS) and its potential for unmatched precision, speed and wavelength coverage can be combined with the advantages of photo-acoustic detection. Acoustic wave interferograms are generated in the sample by dual-comb absorption and detected by a microphone. As an example, weak gas absorption features are precisely and rapidly sampled; long-term coherent averaging further increases the sensitivity. This novel approach of dual-frequency comb photo-acoustic spectroscopy (DCPAS) generates unprecedented opportunities for rapid and sensitive multi-species molecular analysis across all wavelengths of light.
Project description:Optical-resolution photoacoustic microscopy (OR-PAM) is an imaging modality with superb penetration depth and excellent absorption contrast. Here we demonstrate, for the first time, that this technique can advance quantitative analysis of conventional chromogenic histochemistry. Because OR-PAM can quantify the absorption contrast at different wavelengths, it is feasible to spectrally resolve the specific biomolecules involved in a staining color. Furthermore, the tomographic capability of OR-PAM allows for noninvasive volumetric imaging of a thick sample without microtoming it. By immunostaining the sample with different chromogenic agents, we further demonstrated the ability of OR-PAM to resolve different types of cells in a coculture sample with imaging depths up to 1?mm. Taken together, the integration of OR-PAM with (immuno)histochemistry offers a simple and versatile technique with broad applications in cell biology, pathology, tissue engineering, and related biomedical studies.
Project description:Removing the comb artifact introduced by imaging fibre bundles, or 'fibrescopes', for example in medical endoscopy, is essential to provide high quality images to the observer. Multispectral imaging (MSI) is an emerging method that combines morphological (spatial) and chemical (spectral) information in a single data 'cube'. When a fibrescope is coupled to a spectrally resolved detector array (SRDA) to perform MSI, comb removal is complicated by the demosaicking step required to reconstruct the multispectral data cube. To understand the potential for using SRDAs as multispectral imaging sensors in medical endoscopy, we assessed five comb correction methods with respect to five performance metrics relevant to biomedical imaging applications: processing time, resolution, smoothness, signal and the accuracy of spectral reconstruction. By assigning weights to each metric, which are determined by the particular imaging application, our results can be used to select the correction method to achieve best overall performance. In most cases, interpolation gave the best compromise between the different performance metrics when imaging using an SRDA.
Project description:Multispectral photoacoustic microscopy uses a wavelength-dependent absorption difference as a contrast mechanism to image the target molecule. In this paper, we present a novel multispectral pulsed fiber laser source, which selectively alternates the excitation wavelengths between green and red colors based on the stimulated Raman scattering (SRS) effect for imaging. This laser has a high pulse repetition rate (PRR) of 300 kHz and high pulse energy of more than 200 nJ meeting the real-time requirements of optical-resolution photoacoustic microscopy imaging. By switching the polarization state of the pump light and optical paths of the pump light, the operating wavelengths of the light source can be selectively alternated at the same fast PRR for any two SRS peak wavelengths between 545 and 655 nm. At 545 nm excitation wavelength, molecular photoacoustic signals from both blood vessels and gold nanorods were obtained simultaneously. However, at 655 nm, the photoacoustic signals of gold nanorods were dominant because the absorption of light by the blood vessels decreased drastically in the spectral region over 600 nm. Thus the multispectral photoacoustic system designed using the novel laser source implemented here could simultaneously monitor the time-dependent fast movement of two molecules independently, having different wavelength-dependent absorption properties at a high repetition rate of 0.49 frames per second (fps).
Project description:Optical frequency combs are innovative tools for broadband spectroscopy because a series of comb modes can serve as frequency markers that are traceable to a microwave frequency standard. However, a mode distribution that is too discrete limits the spectral sampling interval to the mode frequency spacing even though individual mode linewidth is sufficiently narrow. Here, using a combination of a spectral interleaving and dual-comb spectroscopy in the terahertz (THz) region, we achieved a spectral sampling interval equal to the mode linewidth rather than the mode spacing. The spectrally interleaved THz comb was realized by sweeping the laser repetition frequency and interleaving additional frequency marks. In low-pressure gas spectroscopy, we achieved an improved spectral sampling density of 2.5 MHz and enhanced spectral accuracy of 8.39 × 10(-7) in the THz region. The proposed method is a powerful tool for simultaneously achieving high resolution, high accuracy, and broad spectral coverage in THz spectroscopy.
Project description:Optical frequency combs, consisting of well-controlled equidistant frequency lines, have been widely used in precision spectroscopy and metrology. Terahertz combs have been realized in quantum cascade lasers (QCLs) by employing either an active mode-locking or phase seeding technique, or a dispersion compensator mirror. However, it remains a challenge to achieve the passive comb formation in terahertz semiconductor lasers due to the insufficient nonlinearities of conventional saturable absorbers. Here, a passive terahertz frequency comb is demonstrated by coupling a multilayer graphene sample into a QCL compound cavity. The terahertz modes are self-stabilized with intermode beat note linewidths down to a record of 700 Hz and the comb operation of graphene-coupled QCLs is validated by on-chip dual-comb measurements. Furthermore, the optical pulse emitted from the graphene-coupled QCL is directly measured employing a terahertz pump-probe technique. The enhanced passive frequency comb operation is attributed to the saturable absorption behavior of the graphene-integrated saturable absorber mirror, as well as the dispersion compensation introduced by the graphene sample. The results provide a conceptually different graphene-based approach for passive comb formation in terahertz QCLs, opening up intriguing opportunities for fast and high-precision terahertz spectroscopy and nonlinear photonics.
Project description:Spectroscopic ellipsometry is a means of investigating optical and dielectric material responses. Conventional spectroscopic ellipsometry is subject to trade-offs between spectral accuracy, resolution, and measurement time. Polarization modulation has afforded poor performance because of its sensitivity to mechanical vibrational noise, thermal instability, and polarization-wavelength dependency. We combine spectroscopic ellipsometry with dual-comb spectroscopy, namely, dual-comb spectroscopic ellipsometry. Dual-comb spectroscopic ellipsometry (DCSE). DCSE directly and simultaneously obtains the ellipsometric parameters of the amplitude ratio and phase difference between s-polarized and p-polarized light signals with ultra-high spectral resolution and no polarization modulation, beyond the conventional limit. Ellipsometric evaluation without polarization modulation also enhances the stability and robustness of the system. In this study, we construct a polarization-modulation-free DCSE system with a spectral resolution of up to 1.2?×?10-5?nm throughout the spectral range of 1514-1595?nm and achieved an accuracy of 38.4?nm and a precision of 3.3?nm in the measurement of thin-film samples.Spectroscopic ellipsometry is an established technique to characterize the optical properties of a material. Here, Minamikawa et al. combine the method with dual-comb spectroscopy, which allows them to obtain ellipsometric parameters including the phase difference between s-polarized and p-polarized light.