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Introduction: Birefringent crystals such as monosodium-urate (MSU) and cholesterol crystals (CC) likely contribute to the progression of coronary artery disease (CAD) due to their potential to exacerbate inflammation through inflammatory cytokine activation. Here, we present cross-polarized micro-optical coherence tomography (CP-µOCT) for visualizing individual birefringent crystals in human coronary arteries., Methods and Results: Human cadaver coronary arteries with a history of CAD with or without gout were dissected for CP-µOCT imaging. Specimens were processed for histological identification of birefringence under polarization light microscopy (PLM). CP-µOCT visualized needle-crystals that appeared as long projections in orthogonal planes, and PLM confirmed that CP-µOCT-delineated needle-crystals demonstrated negative birefringence. The needle-crystals were dissolved after immersion in uricase ( p  < 0.05), and thus were MSU. CP-µOCT was three-dimensionally volume-rendered for counting MSU and CCs in 79 regions of interest sized [750 ( x ) × 500 ( y ) × 400 ( z ) µm]. Crystal counts were normalized by the total coronary length utilized. The relationship between CP-µOCT-delineated MSU counts and those seen in corresponding histology, and the difference in coronary MSU amongst gout vs. non-gout patients was analyzed. CP-µOCT-delineated MSU counts were significantly correlated with MSU counted by PLM-based histology ( R  = 0.98, p  < 0.01), and with histology-derived intimal thickening ( R  = 0.51, p  < 0.01). MSU and CCs were both significantly greater in gout patients compared with non-gout patients ( p  < 0.05)., Discussion: These results demonstrate a significant increase in CP-µOCT-delineated crystals in gout vs. non-gout patients, suggesting that this technology can be used to improve our understanding of crystal-driven coronary pathogenesis., Competing Interests: GT receives catheter materials from Terumo Corporation. Massachusetts General Brigham has a licensing arrangement with Terumo Corporation. GT has the rights to receive royalties from this licensing arrangement. GT receives sponsored research funding pertaining to coronary OCT from Amgen and, Canon Inc. GT has a financial/fiduciary interest in SpectraWave, a company developing an OCT-Near Infrared Reflectance Spectroscopy (NIRS) intracoronary imaging system and catheter. His financial/fiduciary interest is reviewed and managed by MGB in accordance with their conflict of interest policies. GT also has consulting arrangements with SpectraWave and Novo Nordisk. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2024 Nishimiya, Sharma, Singh, Ahsen, Gardecki and Tearney.)

Background and Aims: Prior coronary optical coherence tomography (OCT)-near infrared auto-fluorescence (NIRAF) imaging data has shown a correlation between high-risk morphological features and NIRAF signal intensity. This study aims to understand the histopathological origins of NIRAF in human cadaver coronary arteries., Methods: Ex vivo intracoronary OCT-NIRAF imaging was performed on coronary arteries prosected from 23 fresh human cadaver hearts. Arteries with elevated NIRAF were formalin-fixed and paraffin-embedded. Microscopic images of immunostained Glycophorin A (indicating intraplaque hemorrhage) and Sudan Black (indicating ceroid after fixation) stained slides were compared with confocal NIRAF images (ex. 635 nm, em. 655-755 nm) from adjacent unstained slides in each section. Different images from the same section were registered via luminal morphology. Confocal NIRAF-positive 45° sectors were compared to immunohistochemistry and colocalization between NIRAF and intraplaque hemorrhage or ceroid was quantified by Manders' overlap and Dice similarity coefficients., Results: Thirty-one coronary arteries from 14 hearts demonstrated ≥1.5 times higher NIRAF signal than background, and 429 sections were created from them, including 54 sections (12.6%) with high-risk plaques. Within 112 confocal NIRAF-positive 45° sectors, 65 sectors (58.0%) showed both Glycophorin A-positive and Sudan Black-positive, while 7 sectors (6.3%) and 40 sectors (33.6%) only showed Glycophorin A-positive or Sudan black-positive, respectively. A two-tailed McNemar's test showed that Sudan Black more closely corresponded to confocal NIRAF than Glycophorin A (p < 1.0 × 10 -6 ). NIRAF was also found to spatially associate with both Glycophorin A and Sudan Black, with stronger colocalization between Sudan Black and NIRAF (Manders: 0.19 ± 0.15 vs. 0.13 ± 0.14, p < 0.005; Dice: 0.072 ± 0.096 vs. 0.060 ± 0.090, p < 0.01)., Conclusions: As ceroid associates with oxidative stress and intraplaque hemorrhage is implicated in rapid lesion progression, these results suggest that NIRAF provides additional, complementary information to morphologic imaging that may aid in identifying high-risk coronary plaques via translatable intracoronary OCT-NIRAF imaging., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Introduction: Diseases such as celiac disease, environmental enteric dysfunction, infectious gastroenteritis, type II diabetes and inflammatory bowel disease are associated with increased gut permeability. Dual sugar absorption tests, such as the lactulose to rhamnose ratio (L:R) test, are the current standard for measuring gut permeability. Although easy to administer in adults, the L:R test has a number of drawbacks. These include an inability to assess for spatial heterogeneity in gut permeability that may distinguish different disease severity or pathology, additional sample collection for immunoassays, and challenges in carrying out the test in certain populations such as infants and small children. Here, we demonstrate a minimally invasive probe for real-time localized gut permeability evaluation through gut potential difference (GPD) measurement., Materials and Methods: The probe has an outer diameter of 1.2 mm diameter and can be deployed in the gut of unsedated subjects via a transnasal introduction tube (TNIT) that is akin to an intestinal feeding tube. The GPD probe consists of an Ag/AgCl electrode, an optical probe and a perfusion channel all housed within a transparent sheath. Lactated Ringer's (LR) solution is pumped through the perfusion channel to provide ionic contact between the electrodes and the gut lining. The optical probe captures non-scanning (M-mode) OCT images to confirm electrode contact with the gut lining. A separate skin patch probe is placed over an abraded skin area to provide reference for the GPD measurements. Swine studies were conducted to validate the GPD probe. GPD in the duodenum was modulated by perfusing 45 ml of 45 mM glucose., Results: GPD values of -13.1 ± 2.8 mV were measured in the duodenum across four swine studies. The change in GPD in the duodenum with the addition of glucose was -10.5 ± 2.4 mV ( p < 0.001). M-mode OCT images provided electrode-tissue contact information, which was vital in ascertaining the probe's proximity to the gut mucosa., Conclusion: We developed and demonstrated a minimally invasive method for investigating gastrointestinal permeability consisting of an image guided GPD probe that can be used in unsedated subjects., Competing Interests: Conflict of Interest: GT has a financial/fiduciary interest in SpectraWave, a company developing an OCT-NIRS intracoronary imaging system and catheter. His financial/fiduciary interest was reviewed and is managed by the Massachusetts General Hospital and Partners HealthCare in accordance with their conflict of interest policies. GT receives sponsored research from VivoLight, iLumen, Boston Scientific, Vertex, Hamamatsu, AstraZeneca, CN USA Biotech Holdings, Wayvector, and Canon Inc. GT receives catheter materials from Terumo Corporation. GT additional receives royalties from Terumo Corporation, iLumen, and MIT The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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OCT tethered capsule endomicroscopy (TCE) is an emerging noninvasive diagnostic imaging technology for gastrointestinal (GI) tract disorders. OCT measures tissue reflectivity that provides morphologic image contrast, and thus is incapable of ascertaining molecular information that can be useful for improving diagnostic accuracy. Here, we introduce an extension to OCT TCE that includes a fluorescence (FL) imaging channel for attaining complementary, co-registered molecular contrast. We present the development of an OCT-FL TCE capsule and a portable, plug-and-play OCT-FL imaging system. The technology is validated in phantom experiments and feasibility is demonstrated in a methylene blue (MB)-stained swine esophageal injury model, ex vivo and in vivo ., Competing Interests: G.J.T. has a financial/fiduciary interest in SpectraWave, a company developing an OCT-NIRS intracoronary imaging system and catheter. His financial/fiduciary interest was reviewed and is managed by the Massachusetts General Hospital and Partners HealthCare in accordance with their conflict of interest policies. G.J.T. also consults for SpectraWave and receives sponsored research funding from Boston Scientific, Canon Inc., CNUSA Biotech Holdings, AstraZeneca, Translate Bio, Xsphera Biosciences, and WayVector., (© 2021 Optical Society of America under the terms of the OSA Open Access Publishing Agreement.)

This paper describes a new technology that uses 1-µm-resolution optical coherence tomography (µOCT) to obtain cross-sectional images of intracellular dynamics with dramatically enhanced image contrast. This so-called dynamic µOCT (d-µOCT) is accomplished by acquiring a time series of µOCT images and conducting power frequency analysis of the temporal fluctuations that arise from intracellular motion on a pixel-per-pixel basis. Here, we demonstrate d-µOCT imaging of freshly excised human esophageal and cervical biopsy samples. Depth-resolved d-µOCT images of intact tissue show that intracellular dynamics provides a new contrast mechanism for µOCT that highlights subcellular morphology and activity in epithelial surface maturation patterns., Competing Interests: G.J.T. receives sponsored research for µOCT research from CN USA Biotech Holdings and Wayvector., (© 2020 Optical Society of America under the terms of the OSA Open Access Publishing Agreement.)

Cross-sectional visualisation of the cellular and subcellular structures of human atherosclerosis in vivo is significant, as this disease is fundamentally caused by abnormal processes that occur at this scale in a depth-dependent manner. However, due to the inherent resolution-depth of focus tradeoff of conventional focusing optics, today's highest-resolution intravascular imaging technique, namely, optical coherence tomography (OCT), is unable to provide cross-sectional images at this resolution through a coronary catheter. Here, we introduce an intravascular imaging system and catheter based on few-mode interferometry, which overcomes the depth of focus limitation of conventional high-numerical-aperture objectives and enables three-dimensional cellular-resolution intravascular imaging in vivo by a submillimetre diameter, flexible catheter. Images of diseased cadaver human coronary arteries and living rabbit arteries were acquired with this device, showing clearly resolved cellular and subcellular structures within the artery wall, such as individual crystals, smooth muscle cells, and inflammatory cells. The capability of this technology to enable cellular-resolution, cross-sectional intravascular imaging will make it possible to study and diagnose human coronary disease with much greater precision in the future., Competing Interests: Conflict of interestB.Y. and G.J.T. have patent applications on the invention reported in this paper. Massachusetts General Hospital has a licensing agreement with Terumo Corporation and G.J.T. has the rights to receive royalties from this relationship. G.J.T. has a financial/fiduciary interest in Spectrawave, a company developing an OCT-NIRS intracoronary imaging system and catheter. His financial/fiduciary interest was reviewed and is managed by the Massachusetts General Hospital and Partners HealthCare in accordance with their conflict of interest policies. G.J.T.’s laboratory receives sponsored research funding from Canon Inc., Vivolight, and CN USA Biotech Holdings., (© The Author(s) 2019.)

Context.—: The accuracy of needle biopsy for the detection of prostate cancer is limited by well-known sampling errors. Thus, there is an unmet need for a microscopic screening tool that can screen large regions of the prostate comprehensively for cancer. Previous prostate imaging by optical coherence tomography (OCT) has had insufficient resolution for imaging cellular features related to prostate cancer. We have recently developed micro-optical coherence tomography (μOCT) that generates depth-resolved tissue images at a high frame rate with an isotropic resolution of 1 μm., Objective.—: To demonstrate that optical images obtained with μOCT provide cellular-level contrast in prostate specimens that will enable differentiation and diagnosis of prostate pathologies., Design.—: Fresh prostate specimens obtained from surgical resections were scanned with μOCT ex vivo. Histologic features in the μOCT images were correlated to the corresponding conventional histology., Results.—: Findings indicate that μOCT is capable of resolving many of the architectural and cellular features associated with benign and neoplastic prostate., Conclusions.—: Because μOCT can be implemented in a small-diameter flexible probe, this study suggests that high-resolution μOCT imaging may be a useful tool for needle-based virtual biopsy of the prostate gland.

Background and Objectives: Endoscopic optical coherence tomography probes suffer from various artifacts due to dispersion imbalance and polarization mismatch between reference and sample arm light. Such artifacts can be minimized using a common path approach. In this work, we demonstrate a miniaturized common path probe for optical coherence tomography using an inline fiber mirror., Materials and Methods: A common path optical fiber probe suitable for performing high-resolution endoscopic optical coherence tomography imaging was developed. To achieve common path functionality, an inline fiber mirror was fabricated using a thin gold layer. A commercially available swept source engine was used to test the designed probe in a cadaver human coronary artery ex vivo., Results: We achieved a sensitivity of 104 dB for this probe using a swept source optical coherence tomography system. To test the probe, images of a cadaver human coronary artery were obtained, demonstrating the quality that is comparable to those obtained by OCT systems with separate reference arms. Additionally, we demonstrate recovery of ranging depth by use of a Michelson interferometer in the detection path., Conclusion: We developed a miniaturized monolithic inline fiber mirror-based common path probe for optical coherence tomography. Owing to its simplicity, our design will be helpful in endoscopic applications that require high-resolution probes in a compact form factor while reducing system complexity. Lasers Surg. Med. 50:230-235, 2018. © 2017 Wiley Periodicals, Inc., (© 2017 Wiley Periodicals, Inc.)

Improving lateral resolution for cross-sectional optical coherence tomography (OCT) imaging is difficult due to the rapid divergence of light once it is focused to a small spot. To overcome this obstacle, we introduce a fiber optics system that generates a coaxially focused multimode (CAFM) beam for depth of focus (DOF) extension. We fabricated a CAFM beam OCT probe and show that the DOF is more than fivefold that of a conventional Gaussian beam, enabling cross-sectional imaging of biological tissues with clearly resolved cellular and subcellular structures over more than a 400 μm depth range. The compact and straightforward design and high-resolution extended DOF imaging capabilities of this technique suggests that it will be very useful for endoscopic cross-sectional imaging of human internal organs in vivo .

We report the design and fabrication of a flexible, longitudinally scanning high-resolution micro-optical coherence tomography (μOCT) endobronchial probe, optimized for micro-anatomical imaging in airways. The 2.4 mm diameter and flexibility of the probe allows it to be inserted into the instrument channel of a standard bronchoscope, enabling real-time video guidance of probe placement. To generate a depth-of-focus enhancing annular beam, we utilized a new fabrication method, whereby a hollow glass ferrule was angle-polished and gold-coated to produce an elongated annular reflector. We present validation data that verifies the preservation of linear scanning, despite the use of flexible materials. When utilized on excised, cultured mouse trachea, the probe acquired images of comparable quality to those obtained by a benchtop μOCT system.

Objectives: The authors present the clinical imaging of human coronary arteries in vivo using a multimodality optical coherence tomography (OCT) and near-infrared autofluorescence (NIRAF) intravascular imaging system and catheter., Background: Although intravascular OCT is capable of providing microstructural images of coronary atherosclerotic lesions, it is limited in its capability to ascertain the compositional/molecular features of plaque. A recent study in cadaver coronary plaque showed that endogenous NIRAF is elevated in necrotic core lesions. The combination of these 2 technologies in 1 device may therefore provide synergistic data to aid in the diagnosis of coronary pathology in vivo., Methods: We developed a dual-modality intravascular imaging system and 2.6-F catheter that can simultaneously acquire OCT and NIRAF data from the same location on the artery wall. This technology was used to obtain volumetric OCT-NIRAF images from 12 patients with coronary artery disease undergoing percutaneous coronary intervention. Images were acquired during a brief, nonocclusive 3- to 4-ml/s contrast purge at a speed of 100 frames/s and a pullback rate of 20 or 40 mm/s. OCT-NIRAF data were analyzed to determine the distribution of the NIRAF signal with respect to OCT-delineated plaque morphological features., Results: High-quality intracoronary OCT and NIRAF image data (>50-mm pullback length) were successfully acquired without complication in all patients (17 coronary arteries). The maximum NIRAF signal intensity of each plaque was compared with OCT-defined type, showing a statistically significant difference between plaque types (1-way analysis of variance, p < 0.0001). Interestingly, coronary arterial NIRAF intensity was elevated only focally in plaques with a high-risk morphological phenotype (p < 0.05), including OCT fibroatheroma, plaque rupture, and fibroatheroma associated with in-stent restenosis., Conclusions: This OCT-NIRAF study demonstrates that dual-modality microstructural and fluorescence intracoronary imaging can be safely and effectively conducted in human patients. Our findings show that NIRAF is associated with a high-risk morphological plaque phenotype. The focal distribution of NIRAF in these lesions furthermore suggests that this endogenous imaging biomarker may provide complementary information to that obtained by structural imaging alone., (Copyright © 2016 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.)

While optical coherence tomography (OCT) has been shown to be capable of imaging coronary plaque microstructure, additional chemical/molecular information may be needed in order to determine which lesions are at risk of causing an acute coronary event. In this study, we used a recently developed imaging system and double-clad fiber (DCF) catheter capable of simultaneously acquiring both OCT and red excited near-infrared autofluorescence (NIRAF) images (excitation: 633 nm, emission: 680nm to 900nm). We found that NIRAF is elevated in lesions that contain necrotic core - a feature that is critical for vulnerable plaque diagnosis and that is not readily discriminated by OCT alone. We first utilized a DCF ball lens probe and a bench top setup to acquire en face NIRAF images of aortic plaques ex vivo (n = 20). In addition, we used the OCT-NIRAF system and fully assembled catheters to acquire multimodality images from human coronary arteries (n = 15) prosected from human cadaver hearts (n = 5). Comparison of these images with corresponding histology demonstrated that necrotic core plaques exhibited significantly higher NIRAF intensity than other plaque types. These results suggest that multimodality intracoronary OCT-NIRAF imaging technology may be used in the future to provide improved characterization of coronary artery disease in human patients.

Mucociliary clearance, characterized by mucus secretion and its conveyance by ciliary action, is a fundamental physiological process that plays an important role in host defense. Although it is known that ciliary activity changes with chemical and mechanical stimuli, the autoregulatory mechanisms that govern ciliary activity and mucus transport in response to normal and pathophysiological variations in mucus are not clear. We have developed a high-speed, 1-μm-resolution, cross-sectional imaging modality, termed micro-optical coherence tomography (μOCT), which provides the first integrated view of the functional microanatomy of the epithelial surface. We monitored invasion of the periciliary liquid (PCL) layer by mucus in fully differentiated human bronchial epithelial cultures and full thickness swine trachea using μOCT. We further monitored mucociliary transport (MCT) and intracellular calcium concentration simultaneously during invasion of the PCL layer by mucus using colocalized μOCT and confocal fluorescence microscopy in cell cultures. Ciliary beating and mucus transport are up-regulated via a calcium-dependent pathway when mucus causes a reduction in the PCL layer and cilia height. When the load exceeds a physiological limit of approximately 2 μm, this gravity-independent autoregulatory mechanism can no longer compensate, resulting in diminished ciliary motion and abrogation of stimulated MCT. A fundamental integrated mechanism with specific operating limits governs MCT in the lung and fails when periciliary layer compression and mucus viscosity exceeds normal physiologic limits.

The presence of cholesterol crystals is a hallmark of atherosclerosis, but until recently, such crystals have been considered to be passive components of necrotic plaque cores. Recent studies have demonstrated that phagocytosis of cholesterol crystals by macrophages may actively precipitate plaque progression via an inflammatory pathway, emphasizing the need for methods to study the interaction between macrophages and crystalline cholesterol. In this study, we demonstrate the feasibility of detecting cholesterol in macrophages in situ using Micro-Optical Coherence Tomography (µOCT), an imaging modality we have recently developed with 1-µm resolution. Macrophages containing cholesterol crystals frequently demonstrated highly scattering constituents in their cytoplasm on µOCT imaging, and µOCT was able to evaluate cholesterol crystals in cultured macrophage cells. Our results suggest that µOCT may be useful for the detection and characterization of inflammatory activity associated with cholesterol crystals in the coronary artery.

Background: Biopsy surveillance protocols for the assessment of Barrett's esophagus can be subject to sampling errors, resulting in diagnostic uncertainty. Optical coherence tomography is a cross-sectional imaging technique that can be used to conduct volumetric laser endomicroscopy (VLE) of the entire distal esophagus. We have developed a biopsy guidance platform that places endoscopically visible marks at VLE-determined biopsy sites., Objective: The objective of this study was to demonstrate in human participants the safety and feasibility of VLE-guided biopsy in vivo., Design: A pilot feasibility study., Setting: Massachusetts General Hospital., Patients: A total of 22 participants were enrolled from January 2011 to June 2012 with a prior diagnosis of Barrett's esophagus. Twelve participants were used to optimize the laser marking parameters and the system platform. A total of 30 target sites were selected and marked in real-time by using the VLE-guided biopsy platform in the remaining 10 participants., Intervention: Volumetric laser endomicroscopy., Main Outcome Measurements: Endoscopic and VLE visibility, and accuracy of VLE diagnosis of the tissue between the laser cautery marks., Results: There were no adverse events of VLE and laser marking. The optimal laser marking parameters were determined to be 2 seconds at 410 mW, with a mark separation of 6 mm. All marks made with these parameters were visible on endoscopy and VLE. The accuracies for diagnosing tissue in between the laser cautery marks by independent blinded readers for endoscopy were 67% (95% confidence interval [CI], 47%-83%), for VLE intent-to-biopsy images 93% (95% CI, 78%-99%), and for corrected VLE post-marking images 100% when compared with histopathology interpretations., Limitations: This is a single-center feasibility study with a limited number of patients., Conclusion: Our results demonstrate that VLE-guided biopsy of the esophagus is safe and can be used to guide biopsy site selection based on the acquired volumetric optical coherence tomography imaging data. (, Clinical Trial Registration Number: NCT01439633.)., (Copyright © 2014 American Society for Gastrointestinal Endoscopy. Published by Mosby, Inc. All rights reserved.)

Owing to its superior resolution, intravascular optical coherence tomography (IVOCT) is a promising tool for imaging the microstructure of coronary artery walls. However, IVOCT does not identify chemicals and molecules in the tissue, which is required for a more complete understanding and accurate diagnosis of coronary disease. Here we present a dual-modality imaging system and catheter that uniquely combines IVOCT with diffuse near-infrared spectroscopy (NIRS) in a single dual-modality imaging device for simultaneous acquisition of microstructural and compositional information. As a proof-of-concept demonstration, the device has been used to visualize co-incident microstructural and spectroscopic information obtained from a diseased cadaver human coronary artery.

Optical frequency domain imaging (OFDI) can identify key components related to plaque vulnerability but can suffer from artifacts that could prevent accurate identification of lipid rich regions. In this paper, we present a model of depth resolved spectral analysis of OFDI data for improved detection of lipid. A quadratic Discriminant analysis model was developed based on phantom compositions known chemical mixtures and applied to a tissue phantom of a lipid-rich plaque. We demonstrate that a combined spectral and attenuation model can be used to predict the presence of lipid in OFDI images.

Background: Several high-risk morphological features (HRM) of plaques, especially in combination, are associated with an increased risk of a clinical event. Although plaque inflammation is also associated with atherothrombosis, the relationship between inflammation and number of HRM is not well understood., Methods and Results: Thirty-four patients underwent (18)flurodeoxyglucose positron emission tomography (FDG-PET) imaging, and carotid atherosclerotic inflammation was assessed (target-to-, Background: =0.0003) and increased with the number of HRM observed (P<0.001 for trend). Similarly, inflammation within atherosclerotic specimens (% CD68 staining) was higher in plaques with (versus without) HRM (median [interquartile range]: 10 [0, 19.85] versus 0 [0, 1.55], P=0.01) and increased with the number of HRM observed (P<0.001 for trend)., Conclusions: Inflammation, as assessed by both FDG uptake and histology, is increased in plaques containing HRM and increases with increasing number of HRM. These data support the concept that inflammation accumulates relative to the burden of morphological abnormalities.

Progress in understanding, diagnosis, and treatment of coronary artery disease (CAD) has been hindered by our inability to observe cells and extracellular components associated with human coronary atherosclerosis in situ. The current standards for microstructural investigation, histology and electron microscopy are destructive and prone to artifacts. The highest-resolution intracoronary imaging modality, optical coherence tomography (OCT), has a resolution of ~10 μm, which is too coarse for visualizing most cells. Here we report a new form of OCT, termed micro-optical coherence tomography (μOCT), whose resolution is improved by an order of magnitude. We show that μOCT images of cadaver coronary arteries provide clear pictures of cellular and subcellular features associated with atherogenesis, thrombosis and responses to interventional therapy. These results suggest that μOCT can complement existing diagnostic techniques for investigating atherosclerotic specimens, and that μOCT may eventually become a useful tool for cellular and subcellular characterization of the human coronary wall in vivo.

We present the first prospective test of Raman spectroscopy in diagnosing normal, benign, and malignant human breast tissues. Prospective testing of spectral diagnostic algorithms allows clinicians to accurately assess the diagnostic information contained in, and any bias of, the spectroscopic measurement. In previous work, we developed an accurate, internally validated algorithm for breast cancer diagnosis based on analysis of Raman spectra acquired from fresh-frozen in vitro tissue samples. We currently evaluate the performance of this algorithm prospectively on a large ex vivo clinical data set that closely mimics the in vivo environment. Spectroscopic data were collected from freshly excised surgical specimens, and 129 tissue sites from 21 patients were examined. Prospective application of the algorithm to the clinical data set resulted in a sensitivity of 83%, a specificity of 93%, a positive predictive value of 36%, and a negative predictive value of 99% for distinguishing cancerous from normal and benign tissues. The performance of the algorithm in different patient populations is discussed. Sources of bias in the in vitro calibration and ex vivo prospective data sets, including disease prevalence and disease spectrum, are examined and analytical methods for comparison provided.

We demonstrate a fluorescence spectrometer that utilizes principles of Fourier transform spectroscopy to measure excitation emission matrices (EEM) rapidly and with high spectral resolution. For this EEM fluorometer, incoherent excitation light is first input into a differential-delay scanning Michelson interferometer. Light from the output port excites sample fluorescence. The fluorescence remitted from the sample is directed to a second Michelson interferometer, whose differential-delay scanning is synchronized with the first interferometer. The EEM is obtained by two-dimensional Fourier analysis of the detected signal from the output port of the second interferometer. EEM results from the system are verified by comparing with results from a standard spectrometer. The system provides a wide spectral range, adjustable spectral resolution, and fast EEM acquisition speed, which allows EEM's to be acquired in 40 seconds at a spectral resolution of 81-cm-1.

We present the first demonstration of in vivo collection of Raman spectra of breast tissue. Raman spectroscopy, which analyzes molecular vibrations, is a promising new technique for the diagnosis of breast cancer. We have collected 31 Raman spectra from nine patients undergoing partial mastectomy procedures to show the feasibility of in vivo Raman spectroscopy for intraoperative margin assessment. The data was fit with an established model, resulting in spectral-based tissue characterization in only 1 second. Application of our previously developed diagnostic algorithm resulted in perfect sensitivity and specificity for distinguishing cancerous from normal and benign tissues in our small data set. Significantly, we have detected a grossly invisible cancer that, upon pathologic review, required the patient to undergo a second surgical procedure. Had Raman spectroscopy been used in a real-time fashion to guide tissue excision during the procedure, the additional reexcision surgery might have been avoided. These preliminary findings suggest that Raman spectroscopy has the potential to lessen the need for reexcision surgeries resulting from positive margins and thereby reduce the recurrence rate of breast cancer following partial mastectomy surgeries.

Vulnerable plaques, which are responsible for most acute ischemic events, are presently invisible to x-ray angiography. Their primary morphological features include a thin or ulcerated fibrous cap, a large necrotic core, superficial foam cells, and intraplaque hemorrhage. We present evidence that multimodal spectroscopy (MMS), a novel method that combines diffuse reflectance spectroscopy (DRS), intrinsic fluorescence spectroscopy (IFS), and Raman spectroscopy (RS), can detect these markers of plaque vulnerability. To test this concept, we perform an MMS feasibility study on 17 human carotid artery specimens. Following the acquisition of spectra, each specimen is histologically evaluated. Two parameters from DRS, hemoglobin concentration and a scattering parameter, are used to detect intraplaque hemorrhage and foam cells; an IFS parameter that relates to the amount of collagen in the topmost layers of the tissue is used to detect the presence of a thin fibrous cap; and an RS parameter related to the amount of cholesterol and necrotic material is used to detect necrotic core. Taken together, these spectral parameters can generally identify the vulnerable plaques. The results indicate that MMS provides depth-sensitive and complementary morphological information about plaque composition. A prospective in vivo study will be conducted to validate these findings.

In vitro experiments have demonstrated the ability of Raman spectroscopy to diagnose a wide variety of diseases. Recent in vivo investigations performed with optical fiber probes were promising but generally limited to easily accessible organs, often requiring relatively long collection times. We have implemented an optical design strategy to utilize system throughput fully by characterizing the Raman distribution from tissue. This scheme optimizes collection efficiency, minimizes noise, and has resulted in small-diameter, highly efficient Raman probes that are capable of collecting high-quality data in 1 s. Performance has been tested through simulations and experiments with tissue models and several in vitro tissue types, demonstrating that this new design can advance Raman spectroscopy as a clinically practical technique.