Compressive Raman imaging has emerged as a promising technique to speed up chemical imaging by compressing the data during acquisition. Yet, current scanning imaging speed is fundamentally limited by the sensors pixel dwell times of at best 1µs. Here, we introduce a compressive Raman spectrometer layout equipped with a novel parallelized spatial acquisition using a single-photon avalanche detector array. We show imaging with pixel dwell times of <10µs using the otherwise weak spontaneous Raman effect, thereby unlocking video-rate imaging.
The intricate nature of the brain necessitates the application of advanced probing techniques to comprehensively study and understand its working mechanisms. Neurophotonics offers minimally invasive methods to probe the brain using optics at cellular and even molecular levels. However, multiple challenges persist, especially concerning imaging depth, field of view, speed, and biocompatibility. A major hindrance to solving these challenges in optics is the scattering nature of the brain. This perspective highlights the potential of complex media optics, a specialized area of study focused on light propagation in materials with intricate heterogeneous optical properties, in advancing and improving neuronal readouts for structural imaging and optical recordings of neuronal activity. Key strategies include wavefront shaping techniques and computational imaging and sensing techniques that exploit scattering properties for enhanced performance. We discuss the potential merger of the two fields as well as potential challenges and perspectives toward longer term in vivo applications.
We introduce a phase conjugation method that utilizes multiple incoherent guidestars to control light in complex media. The technique involves the characterization of mutually incoherent scattered fields, followed by their time-reversal. With this approach, we achieve precise light focusing on individual and multiple guidestars, as well as efficient energy delivery to an extended target through scattering media. This method has various potential applications, including optical manipulation, targeted stimulation and deep optical imaging.
In this work, we demonstrate a method that can focus on multiple single guidestar through scattering medium with a spatial light modulator (SLM) and a bucket detector using two-photon fluorescence signals. A gradient descent based multiplexed phase retrieval algorithm is used to non-invasively reconstruct the transmission matrix between the guidestars and the SLM, without any assumptions on the memory effect range. Conversely, if we consider the memory effect, we can reconstruct the image of the sample.
KEYWORDS: Raman spectroscopy, Imaging arrays, Imaging spectroscopy, Data acquisition, Spectroscopy, Sensors, Data conversion, Reconstruction algorithms, Real time imaging, Raman scattering
Compressive Raman imaging has emerged as a promising technique to speed up chemical imaging by compressing the data during acquisition. Yet, current scanning imaging speed is fundamentally limited by the sensors pixel dwell times of at best 1 µs. Here, we introduce a compressive Raman spectrometer layout equipped with a novel parallelized spatial acquisition using a single-photon avalanche detector array. We show imaging with pixel dwell times of <10µs using the otherwise weak spontaneous Raman effect, thereby reaching real-time imaging.
We demonstrate a novel approach to super-resolution in stimulated Raman scattering based on single-pixel detection scheme. We exploit speckle structured illumination, and enable fast super-resolution microscopy using Raman contrasts. The methodology is straightforward to implement and provides label-free super-resolution imaging of optically thick specimens, therefore paving the way for probing complex biological systems when exogenous labelling is challenging.
Light scattering has proven to be a hard limitation in a wide range of sensing applications, such as astronomical or biological imaging. In microscopy systems, the random perturbations introduced to the wavefront limit the achievable spatial resolution and imaging depth. In the past, several methods have been proposed to control how light interacts with the medium, allowing focusing and imaging through multiple scattering media by using wavefront shaping techniques. However, non-invasively imaging objects behind scattering media over large fields of view remains a challenging feat.
Here, we introduce a novel approach that allows to recover fluorescent extended objects behind scattering layers well beyond the optical memory effect (ME) range without the use of neither adaptive optics nor wavefront shaping techniques. To do so, we project a collection of unknown random illumination speckle patterns through the scattering medium by using a simple rotating diffuser. For each position of the rotating diffuser, a different incoherent sum of speckle patterns is recorded by the camera. Even though these images are low-contrast, random, and seem to carry no information at all, they contain the information about the position of the emitters. Here we show that, if enough images are measured, it is possible to use Non-negative Matrix Factorization to demix all the information and to retrieve the relative position of each fluorescent emitter in the sample.
As a proof of the technique, we show experimental results with both sparse and continuous objects, covering fields-of-view of up to three times the optical memory effect range
Deep Raman imaging in complex media has been elusive, due to scattering of the excitation light. Here, we exploit non-invasive wavefront shaping methods using a spectrally resolved speckle variance optimization algorithm in order to enhance the spontaneous Raman signals of a single particle. We demonstrate unprecedented signal enhancement (>5x) in a non-invasive manner for an epi geometry.
The main obstacle for optical imaging deep inside biological tissues is light scattering. Recently, three-photon (3p) microscopy extended the accessible depth ranges of fluorescence imaging due to its enhanced nonlinearity. On a different front, advances in optical wavefront shaping showed that scattering can be compensated for, even in regimes where light entirely lost its directionality. Combining these two approaches, we demonstrate focusing and imaging behind scattering layers, enabled through wavefront shaping guided by 3p fluorescence in situations where no ballistic light reaches the sample. We analyse different sample geometries and compare these results to the case of 2p excitation.
In order to enhance spontaneous Raman signals, we exploit non-invasive wavefront shaping methods using a spectrally resolved speckle variance optimization algorithm. We demonstrate unprecedented signal enhancement > 5x in an epi-geometry.
We present a simulation of stimulated Raman scattering that could make deep super resolution imaging possible with chemical selectivity and relatively low power levels. For this, we directly use speckles as a structured illumination pattern in a coherent Raman scattering processes, in particular Stimulated Raman Scattering. Using off-the-shelf conjugate gradient-based algorithm, we demonstrate that the method enables super-resolution better than conventional raster scanning techniques.
Raman imaging is recognized as a powerful label-free approach to provide contrasts based on chemical selectivity. Nevertheless, Raman-based microspectroscopy has drawbacks mostly due to its inherent overwhelming data size, which slows imaging speeds, and non-trivial post-processing, in particular for non-specialists in vibrational spectroscopy. In parallel, compressive sensing has shown a paradigm shift approach where one can computationally reconstruct accurate information from highly undersampled data. Following the compressive sensing spirit, compressive Raman microspectroscopy has emerged as a potential approach to speed up the imaging and concomitantly simplify the post-processing analysis. In this contribution, I will discuss the concepts and assumptions in compressive Raman, in particular focusing on current challenges for fast and high-sensitivity imaging of challenging biological specimens.
Raman imaging is recognized as a powerful label-free approach to provide contrasts based on chemical selectivity. Nevertheless, Raman-based microspectroscopy still have drawbacks. The first issue is the inherent high data throughput in Raman microspectroscopy, posing challenges for dynamic and large-scale imaging, and its subsequent data storage. The second issue is data presentation: often, Raman microspectroscopy acquires overwhelming data sets, which information is then post-processed to a more useful and straightforward presentation (typically limited to the number of different target chemicals in a system). In parallel, compressive sensing has shown a paradigm shift approach where one can obtain accurate information from fewer samples than assumed by Nyquist-Shannon sampling theorem. A key concept in compressive sensing is to recognize that data sparsity can be exploited to reconstruct data that has been considerably undersampled. Following the compressive sensing spirit, various approaches were proposed to demonstrate compressive Raman microspectroscopy in different flavors, exploiting the fact that both the vibrational spectrum and the chemical components are often sparse. In this contribution, I will discuss different ways of performing compressive Raman, in particular focusing on the challenges that precludes fast imaging of biological specimens, and how we recently tackled them. With these outcomes, compressive Raman imaging soon may be routinely used by non-specialists in vibrational spectroscopy in a “blind” manner.
The high data bandwidth of Raman imaging precludes high-speed spectroscopic imaging. Conversely, emerging compressive sensing hyperspectroscopy techniques could, in principle, address this issue by using undersampling methodologies with computational reconstructions. However, compressive spectrometer layouts have prohibitive losses for low-light levels applications, such as in the spontaneous Raman imaging of dynamic biological specimens. These losses are due to the fact that high-sensitivity light detectors (photo-counters) have too small active area (typically 100 um) compared to the size of digital micromirror devices (DMD) (~10 mm) used in most compressive layouts. Inspired by pulse shaping techniques of ultrafast spectroscopy, we present a new programmable spectrometer layout with high-throughput and large spectral coupling bandwidths. Exploiting amplitude spectral modulation with DMD allows conventional and compressive Raman imaging and spectroscopy acquisitions with shot-noise-limited sensitivity. With this spectrometer, we demonstrate compressed hyperspectroscopy at faster speeds and at lower costs than traditional cameras used in Raman imaging applications. We showcase imaging of biological specimens at high spatial resolution (250 nm).
Spatial and temporal properties of an ultrashort pulse of light are naturally scrambled upon propagation in thick scattering media. Significant progresses have been realized over the last decade to manipulate light propagation in scattering media, mostly using monochromatic light. However, applications that require a broadband ultrashort pulse of light remain limited, as the pulse gets temporally broadened because of scattering effects. A monochromatic optical transmission matrix does not allow temporal control of broadband light. Although measuring multiple transmission matrices with spectral resolution allows fine temporal control, it requires lengthy measurements, as well as stability of the medium.
In this work, we show that a single linear operator that we named Broadband Transmission Matrix, can be straightforwardly measured for a broadband pulse with a co-propagating reference. We exploit this operator for focusing purposes, and we analyze its phase conjugation properties. While the operator naturally allows for spatial focusing, unexpectedly, the focus duration is on average shorter than the natural temporal broadening due to the medium. More precisely, we observe a two-fold temporal recompression at the focus that we fully explain theoretically. We also explore the spectral content at the focus, and demonstrate a narrowing of the spectrum.
These results are particularly relevant for non-linear imaging techniques in biological tissues, at depth where an ultrashort excitation pulse is broadened.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.