This PDF file contains the front matter associated with SPIE Proceedings Volume 11657, including the Title Page, Copyright information, and Table of Contents.

Angular scattering can be used to obtain morphological information from biological specimen, such as the mean size of organelles. We will investigate the limitations that arise when fitting cell scattering to a Mie theory model and extracting organelle size estimates. Using simulation studies of the scattering of two organelle populations in a cell (mitochondria and lysosomes), we will determine under what conditions multiple size distribution parameters can be estimated. Additionally, the analysis method that will allow for the most accurate and smallest organelle size changes to be measured will be investigated computationally.

While fluorescence labeling remains a powerful tool for many studies, it also poses fundamental limitations, which have motivated many groups to develop fluorescence-free measurement methods. Scattering offers many interesting opportunities for these efforts, but lack of spectral specificity makes the detection of small objects via scattering challenging. Nearly two decades ago, we showed that, nevertheless, single gold nanoparticles as small as 5 nm could be detected via interferometric detection, giving birth to a new detection scheme that was later coined iSCAT. Since then, it has been shown that small unlabeled proteins could be detected and counted, and transmembrane proteins tracked on living cells using gold nanoparticles as scattering label. In this presentation, I discuss the technical subtleties of iSCAT, its principal advantages and several new studies that have become possible by it.

Quantitative phase imaging (QPI) and 3D refractive index (RI) tomography are important tools in biomedicine that yield label-free access to cellular and subcellular structures with unprecedented clarity. However, implementation of these technologies involves a transmission-based geometry which requires thin samples. Here we describe quantitative oblique back illumination microscopy (qOBM), which overcomes this significant limitation and achieves epiillumination quantitative phase imaging and 3D RI tomography in thick samples, including intact thick tissues. In this presentation, we will describe the method in detail and show results from thick samples including intact whole mouse brains and organoids. qOBM opens the door to many exciting new applications in biomedicine.

Oesophageal cancer and colon cancer have five year survival rates of 15% and 63% respectively. These low survival rates are due in part to poor early detection during endoscopic screening, with conventional endoscopes providing insufficient information about tissue properties to spot a wide range of potential tumours. Improving early detection of gastrointestinal cancers would dramatically increase their five year survival rates. Spatial Frequency Domain Imaging (SFDI) is a low-cost imaging technique that can measure absorption, scattering and shape as potential indicators of cancer. Specific absorption and scattering properties are known to be linked to malignancy in the oesophagus, and shape is an important indicator in colon cancer. Though a range of research and commercial SFDI systems have been developed, adapting these for in vivo clinical application is challenging due to constraints imposed by miniaturisation, sample geometry and illumination conditions. To facilitate design of novel SFDI systems under such constraints, we have developed a model of an SFDI imaging system built on the open-source 3D modelling software Blender. Using Blender’s Cycles ray-tracing engine, we are able to simulate a range of different scattering and absorption coefficients for a number of different imaging configurations, sample geometries and illumination patterns. Using established processing algorithms, we show we can recover maps of absorption, scattering and shape in a range of simulated ex vivo and in vivo imaging geometries with relevance to clinical detection of tumours. Our system enables accessible exploration of different optical configurations and realistic illumination conditions that will inform future design of compact, low-cost instruments.

We present a method for large volume imaging of highly scattering tissues using a dual-axis optical coherence tomography system (DA-OCT) at 1.3 μm featuring a dynamic focus-tracking method to create an enhanced depth of focus. Our approach is validated for skin imaging, using an in vivo rat skin model. A quantitative discussion of imaging performance in highly scattering tissue for both DA-OCT and conventional OCT at 1.3 μm is presented.

Multiple scattering and angle-dependent scattering anisotropy confound interrogation of tissues with OCT and are generally considered noise. Here we characterize a new localization-diverse OCT system that measures the scattering through a pair of neighboring locations. By varying the offset and direction between the locations, we could distinguish single- from multiple-scattering in tissue-mimicking scattering phantoms. This system has the potential to detect previously unobserved tissue anisotropy by leveraging localization diversity in OCT.

Traditional optical imaging systems are designed with the assumption that light travels along straight paths inside the sample. Light scattering in biological tissue leads to high background and blurry images in traditional imaging methods. We report technique advancements in structured illumination with light sheet imaging (SIM-LS) in deep tissue. SIM-LS is capable of removing image background generated by scattering and restoring optical resolution in deep tissue imaging. With two-photon Bessel beam excitation, SIM-LS supports up to 500 micron wide field of view 3D imaging in embryos and brain tissue at subcellular resolution.

We will overview the technological development and emerging applications of Brillouin scattering microscopy

We present results from depth-resolved light scattering measurements of triple transgenic mouse retinas for Alzheimer’s Disease (AD) using a multimodal coherent imaging system. Use of a co-registered angle-resolved low-coherence interferometry (a/LCI) and optical coherence tomography (OCT) system allows unique analysis that is otherwise unavailable using a single modality to provide complementary information on tissue structural changes associated with AD. This abstract summarizes the light scattering parameters drawn using this system at selective retinal layers guided by OCT image segmentation. Future developments of this combined system for human retinal imaging, which involve a low-cost OCT engine, are also discussed.

Monitoring the renal cortical microperfusion (RCM) can aid in the determination of an adequate treatment plan to eventually improve transplantation outcome and improve patient care. In this paper we report on a feasibility study on the use of LSCI to monitor RCM in human sized porcine kidneys using an isolated perfusion model and a Lapvas-Imaging speckle imager. Our data shows that there is a discrepancy between overall- and RCM perfusion indicating that the implementation of LSCI during transplant surgery could help with the establishment of an appropriate treatment plan possibly decreasing the chance of renal allograft rejection.

We examine the relative merits of wide-field, sub-diffuse light scatter imaging and micro-computed tomography for visualizing breast tumor heterogeneity. A large dataset of breast tumor slices was imaged using both modalities, and image data were spatially co-registered with histopathological images to facilitate direct comparisons in feature visualization between tissue types, including normal, benign, and malignant tissues, and microcalcifications. Mechanisms of scatter for the two modalities are discussed and related to the features detected by each modality.

Three-dimensional (3D) cell culture models are developed as a promising platform to screen anticancer therapeutics and treatments. However, current imaging techniques cannot provide 3D structures of tumor spheroids in situ. In this study, we employed label-free and noninvasive optical coherence tomography (OCT) for imaging and quantifying the 3D structures of tumor spheroids. We imaged ovarian cancer spheroids with OVCAR-8 cell line over a period of 10 days with 5,000 and 50,000 initial cell numbers. We successfully reconstructed the 3D necrotic regions via label-free intrinsic scattering attenuation contrast and evaluated the effect of Cisplatin treatment on tumor spheroids.

We present a machine learning method for the detection and staging of cervical dysplasia tissue using a convolutional neural network (CNN)-based architecture. Depth-resolved angular scattering measurements collected from two clinical trials consisting of 6660 and 1600 depth scans were used as training and validation sets separately. Our results demonstrated high sensitivity and specificity for classifying cervical dysplasia at a hundredfold faster processing time compared with the traditional Mie-theory inverse light scattering analysis (ILSA) method, offering a promising approach for a/LCI in the clinic for assessing cervical dysplasia.

We assess a novel means of measuring cerebrovascular reactivity (CVR) in awake mice using intraperitoneal injection of acetazolamide combined with continuous monitoring of cerebral blood flow with a minimally invasive diffuse correlation spectroscopy (DCS).

Near-infrared spectroscopy has been widely employed in biophotonics to study and quantify the optical properties of biological tissues. Unlike steady-state approaches, time-resolved spectroscopic techniques enable optical absorption and scattering properties of the medium to be separated, allowing for quantitation of depth-dependent absolute tissue optical properties. However, robust analysis of time-resolved signals requires careful consideration of calibration techniques and computational models. Here, we consider the effect of the time window employed when fitting a diffusion theory model to Monte-Carlo simulations. Next, we describe the impact of the temporal position of the instrument response function (IRF) in recovery of the optical properties. Finally, we discuss a technique to analyze time-resolved measurements without knowledge of the timescale of the IRF or the time-resolved measurement by fitting the relative shape of the photons’ distribution time-of-flight (DTOF).

Deciphering molecular changes (such as structure conformation) in complex systems can be challenging. If these conformation changes could be monitored in real time and modeled, it would open up new opportunities to gain a deeper understanding of signal pathways in biological systems. Through the use of confocal Raman spectroscopy, which captures the molecular fingerprint with high precision, we monitored the evolution of these changes over time. The key was to identify the spectral regions within the Raman spectrum. We employed an adaptive principal component analysis (PCA) technique to study Raman spectra and modeled strain conditions in this molecular network. Experiments were completed according to a full factorial design of experiment (DOE) approach with variable parameters including laser power density and stage temperature over the spectral range of 50-4000 /cm. Thermal effects were also introduced through the controllable micro-stage heater. We implemented this adaptive PCA technique on both individual and blended amino acids in order to highlight vibrational modes within complex samples. We examined three structurally similar branched chain amino acids to study similarities and identified specific vibrational modes that indicate molecular bending, rocking, and wagging. Results demonstrate that adaptive PCA is capable of highlighting subtle changes in molecular networks due to environmental and compositional variations. With an understanding of which data (spectral band) is more important, this speeds up computation and provides real-time analysis for monitoring conformational changes.

In this work, Surface-enhanced Raman Scattering had been applied to detect L-Asparagine. The presence of this amino acid in tissues and fluids helps to the prevalence of acute lymphoblastic leukemia cells and promotes the proliferation of metastasis for some other malignant tumors; depletion of L-Asparagine is up to date the best treatment for leukemia and could improve the prognosis for other cancers. We have carried out the preparation and characterization of two types of metallic surfaces, one with nanoparticles and the other one with nanostructures; we obtained and compared the SERS spectra and enhancement factors between them to emphasize the advantages in each one. The principal aim of this work is to establish the basis for a new reliable biosensor capable of being used in clinical applications.