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Significance Raman spectroscopy has been used as a powerful tool for chemical analysis, enabling the noninvasive acquisition of molecular fingerprints from various samples. Raman spectroscopy has proven to be valuable in numerous fields, including pharmaceutical, materials science, and biomedicine. Active research and development efforts are currently underway to bring this analytical instrument into the field, enabling in situ Raman measurements for a wider range of applications. Dispersive Raman spectroscopy using a fixed, narrowband source is a common method for acquiring Raman spectra. However, dispersive Raman spectroscopy requires a bulky spectrometer, which limits its field applicability. Therefore, there has been a tremendous need to develop a portable and sensitive Raman system. Aim We developed a compact swept-source Raman (SS-Raman) spectroscopy system and proposed a signal processing method to mitigate hardware limitations. We demonstrated the capabilities of the SS-Raman spectroscopy by acquiring Raman spectra from both chemical and biological samples. These spectra were then compared with Raman spectra obtained using a conventional dispersive Raman spectroscopy system. Approach The SS-Raman spectroscopy system used a wavelength-swept source laser (822 to 842 nm), a bandpass filter with a bandwidth of 1.5 nm, and a low-noise silicon photoreceiver. Raman spectra were acquired from various chemical samples, including phenylalanine, hydroxyapatite, glucose, and acetaminophen. A comparative analysis with the conventional dispersive Raman spectroscopy was conducted by calculating the correlation coefficients between the spectra from the SS-Raman spectroscopy and those from the conventional system. Furthermore, Raman mapping was obtained from cross-sections of swine tissue, demonstrating the applicability of the SS-Raman spectroscopy in biological samples. Results We developed a compact SS-Raman system and validated its performance by acquiring Raman spectra

Metastatic prostate cancer colonizes the bone to pave the way for bone metastasis, leading to skeletal complications associated with poor prognosis and morbidity. This study demonstrates the feasibility of Raman imaging to differentiate between cancer cells at different stages of tumorigenesis using a nanoclay-based three-dimensional (3D) bone mimetic in vitro model that mimics prostate cancer bone metastasis. A comprehensive study comparing the classification of as received prostate cancer cells in a two-dimensional (2D) model and cancer cells in a 3D bone mimetic environment was performed over various time intervals using principal component analysis (PCA). Our results showed distinctive spectral differences in Raman imaging between prostate cancer cells and the cells cultured in 3D bone mimetic scaffolds, particularly at 1002, 1261, 1444, and 1654 cm−1, which primarily contain proteins and lipids signals. Raman maps capture sub-cellular responses with the progression of tumor cells into metastasis. Raman feature extraction via cluster analysis allows for the identification of specific cellular constituents in the images. For the first time, this work demonstrates a promising potential of Raman imaging, PCA, and cluster analysis to discriminate between cancer cells at different stages of metastatic tumorigenesis.

Cardiovascular diseases pose a serious health risk and have a high mortality rate of 31% worldwide. Digoxin is the most commonly prescribed pharmaceutical preparation to cardiovascular patients particularly in developing countries. The effectiveness of the drug critically depends on its presence in the therapeutic range (0.8–2.0 ng mL−1) in the patient's serum. We fabricated immunoassay chips based on QD photoluminescence (QDs-ELISA) and AuNP Surface Enhanced Raman Scattering (SERS-ELISA) phenomena to detect digoxin in the therapeutic range. Digoxin levels were monitored using digoxin antibodies conjugated to QDs and AuNPs employing the sandwich immunoassay format in both the chips. The limit of detection (LOD) achieved through QDs-ELISA and SERS-ELISA was 0.5 ng mL−1 and 0.4 ng mL−1, respectively. It is demonstrated that the sensitivity of QDs-ELISA was dependent on the charge transfer mechanism from the QDs to the antibody through ionic media, which was further explored using electrochemical impedance spectroscopy. We demonstrate that QDs-ELISA was relatively easy to fabricate compared to SERS-ELISA. The current study envisages replacement of conventional methodologies with small immunoassay chips using QDs and/or SERS-based tags with fast turnaround detection time as compared to conventional ELISA.

Wild-type p53 cancer therapy-induced senescent cells frequently engulf and degrade neighboring ones inside a massive vacuole in their cytoplasm. After clearance of the internalized cell, the vacuole persists, seemingly empty, for several hours. Despite large vacuoles being associated with cell death, this process is known to confer a survival advantage to cancer engulfing cells, leading to therapy resistance and tumor relapse. Previous attempts to resolve the vacuolar structure and visualize their content using dyes were unsatisfying for lack of known targets and ineffective dye penetration and/or retention. Here, we overcame this problem by applying optical diffraction tomography and Raman spectroscopy to MCF7 doxorubicin-induced engulfing cells. We demonstrated a real ability of cell tomography and Raman to phenotype complex microstructures, such as cell-in-cells and vacuoles, and detect chemical species in extremely low concentrations within live cells in a completely label-free fashion. We show that vacuoles had a density indistinguishable to the medium, but were not empty, instead contained diluted cell-derived macromolecules, and we could discern vacuoles from medium and cells using their Raman fingerprint. Our approach is useful for the noninvasive investigation of senescent engulfing (and other peculiar) cells in unperturbed conditions, crucial for a better understanding of complex biological processes.

Background Margin status is an important prognostic factor for treating colorectal cancer. This study aimed to investigate the usefulness of a multimodal spectroscopic tissue scanner for real-time cancer diagnosis without tissue staining. Patients and methods Diffuse reflectance spectra (DRS) and fluorescence spectra (FS) of < 1-mm-sized paired cancer and normal mucosa tissue were acquired using custom-built spectroscopic tissue scanners. For FS, we analyzed wavelengths and intensities at peaks and highest intensities near (± 1.25 nm) the known fluorescence spectral peaks of collagen (380 nm), reduced nicotinamide adenine dinucleotide (NADH, 460 nm), and flavin adenine dinucleotide (FAD, 550 nm). For DRS, we performed a similar analysis near the peaks of strong absorbers, oxyhemoglobin (oxyHb; 414 nm, 540 nm, and 576 nm) and deoxyhemoglobin (deoxyHb; 432 nm and 556 nm). Logistic regression analysis for these parameters was performed in the testing set. Results We acquired 17,735 spectra of cancer tissues and 9438 of normal tissues from 30 patients. Intensity peaks of representative normal spectra for FS and DRS were higher than those of representative cancer spectra. Logistic regression analysis showed wavelength and intensity at peaks, and the intensities of the peak wavelength of NADH, FAD, deoxyHb, and oxyHb had significant coefficients. The area under the receiver operating characteristic curve was 0.927. The scanner had 100%, 64.3%, and 85.3% sensitivity, specificity, and accuracy, respectively. Conclusions The spectroscopic tissue scanner has high sensitivity and accuracy and provides real-time intraoperative resection margin assessments and should be further investigated as an alternative to frozen section.

We analytically investigate the feasibility of long graded-index (GRIN)-lens-based microendoscopes through wavefront shaping. Following the very well-defined ray trajectories in a GRIN lens, mode-dependent phase delay is first determined. Then, the phase compensation needed for obtaining diffraction limited resolution is derived. Finally, the diffraction pattern of the lens output is computed using the Rayleigh–Sommerfeld diffraction theory. We show that diffraction-limited resolution is obtained for a 0.5 mm diameter lens with a length over 1 m. It is also demonstrated that different imaging working distances (WDs) can be realized by modifying the phase compensation. When a short design WD is used, a large imaging numerical aperture (NA) higher than 0.4 is achievable even when a low NA lens (NA = 0.1) is used. The long- and thin-GRIN-lens-based microendoscope investigated here, which is attractive for biomedical applications, is being prioritized for use in a clinical stage microdevice that measures three-dimensional drug responses inside the body. The advance described in this work may enable superior imaging capabilities in clinical applications in which long and flexible imaging probes are favored., National Institutes of Health (U.S.) (Grants R37CA224144, 41-EB015871), Brigham and Women's Hospital. Department of Radiology. Research Pilot Grant

© 2020 SPIE. The formation and accumulation of advanced glycation end products (AGEs) contribute to diabetic complications such as retinopathy, neuropathy, nephropathy, and cardiovascular diseases. It is clear that the development of effective technique in AGEs detection and the establishing the correlation of measured AGE parameters to diabetic pathogenesis are invaluable in the monitoring of disease progression and drug discovery of anti-AGE compounds. Since some AGE are fluorescent, we propose to investigate the degree of tissue glycation in forming fluorescent AGEs (fAGEs). In this preliminary study, we will investigate the effects of glucose, fructose, and galactose, three of the most abundant dietary simple sugars, in fAGEs production. Excised tissues will be treated in solutions containing the three sugar types; multiphoton autofluorescence imaging will then be performed on the treated tissues to determine their autofluroescence levels.

© COPYRIGHT SPIE. Downloading of the abstract is permitted for personal use only. Cardiovascular complication is a major health concern for diabetic patients. Elevated blood sugar levels lead to the formation of advanced glycation end products (AGEs), which are implicated in diabetic pathogenesis. In this study, we investigate the effect of prolonged exposure to elevated sugar level by studying the combined effect of diffusion and glycation rate in arteries from different commonly-consumed simple sugars. Since some AGEs are autofluorescent, we will perform multiphoton autofluorescence imaging to quantify the rates of fluorescent AGEs formation in elastic fibers and collagen fibers. Croß-section imaging of arteries and spatial and temporal profiles of autofluorescence of the blood veßel will be investigated.

We analytically investigate the feasibility of long graded-index (GRIN)-lens-based microendoscopes through wavefront shaping. Following the very well-defined ray trajectories in a GRIN lens, mode-dependent phase delay is first determined. Then, the phase compensation needed for obtaining diffraction limited resolution is derived. Finally, the diffraction pattern of the lens output is computed using the Rayleigh–Sommerfeld diffraction theory. We show that diffraction-limited resolution is obtained for a 0.5 mm diameter lens with a length over 1 m. It is also demonstrated that different imaging working distances (WDs) can be realized by modifying the phase compensation. When a short design WD is used, a large imaging numerical aperture (NA) higher than 0.4 is achievable even when a low NA lens (NA = 0.1) is used. The long- and thin-GRIN-lens-based microendoscope investigated here, which is attractive for biomedical applications, is being prioritized for use in a clinical stage microdevice that measures three-dimensional drug responses inside the body. The advance described in this work may enable superior imaging capabilities in clinical applications in which long and flexible imaging probes are favored.

Label-free live cell imaging was performed using a custom-built high-speed confocal Raman microscopy system. For various cell types, cell-intrinsic Raman bands were monitored. The high-resolution temporal Raman images clearly delineated the intracellular distribution of biologically important molecules such as protein, lipid, and DNA. Furthermore, optical phase delay measured using quantitative phase microscopy shows similarity with the image reconstructed from the protein Raman peak. This reported work demonstrates that Raman imaging is a powerful label-free technique for studying various biomedical problems in vitro with minimal sample preparation and external perturbation to the cellular system.

© COPYRIGHT SPIE. Downloading of the abstract is permitted for personal use only. Cardiovascular complication is a major health concern for diabetic patients. Elevated blood sugar levels lead to the formation of advanced glycation end products (AGEs), which are implicated in diabetic pathogenesis. In this study, we investigate the effect of prolonged exposure to elevated sugar level by studying the combined effect of diffusion and glycation rate in arteries from different commonly-consumed simple sugars. Since some AGEs are autofluorescent, we will perform multiphoton autofluorescence imaging to quantify the rates of fluorescent AGEs formation in elastic fibers and collagen fibers. Croß-section imaging of arteries and spatial and temporal profiles of autofluorescence of the blood veßel will be investigated.

Advances in the intratumor measurement of drug responses have included a pioneering biomedical microdevice for high throughput drug screening in vivo, which was further advanced by integrating a graded-index lens based two-dimensional fluorescence micro-endoscope to monitor tissue responses in situ across time. While the previous system provided a bulk measurement of both drug delivery and tissue response from a given region of the tumor, it was incapable of visualizing drug distribution and tissue responses in a three-dimensional (3D) way, thus missing the critical relationship between drug concentration and effect. Here we demonstrate a next-generation system that couples multiplexed intratumor drug release with continuous 3D spatial imaging of the tumor microenvironment via the integration of a miniaturized two-photon micro-endoscope. This enables optical sectioning within the live tissue microenvironment to effectively profile the entire tumor region adjacent to the microdevice across time. Using this novel microimaging-microdevice (MI-MD) system, we successfully demonstrated the four-dimensional imaging (3 spatial dimensions plus time) of local drug delivery in tissue phantom and tumors. Future studies include the use of the MI-MD system for monitoring of localized intra-tissue drug release and concurrent measurement of tissue responses in live organisms, with applications to study drug resistance due to nonuniform drug distribution in tumors, or immune cell responses to anti-cancer agents.

Label-free live cell imaging was performed using a custom-built high-speed confocal Raman microscopy system. For various cell types, cell-intrinsic Raman bands were monitored. The high-resolution temporal Raman images clearly delineated the intracellular distribution of biologically important molecules such as protein, lipid, and DNA. Furthermore, optical phase delay measured using quantitative phase microscopy shows similarity with the image reconstructed from the protein Raman peak. This reported work demonstrates that Raman imaging is a powerful label-free technique for studying various biomedical problems in vitro with minimal sample preparation and external perturbation to the cellular system.