Coherent fiber imaging bundles can be used as passive probes for reflectance-mode endomicroscopy providing that the back-reflections from the fiber ends are efficiently rejected. We describe an approach specific to widefield endomicroscopy in which light is injected into a leached fiber bundle near the distal end, thereby avoiding reflections from the proximal face. We use this method to demonstrate the color widefield reflectance endomicroscopy of ex vivo animal tissue.

An intraoperative surgical microscope is an essential tool in a neuro- or ophthalmological surgical environment. Yet, it has an inherent limitation to classify subsurface information because it only provides the surface images. To compensate for and assist in this problem, combining the surgical microscope with optical coherence tomography (OCT) has been adapted. We developed a real-time virtual intraoperative surgical OCT (VISOCT) system by adapting a spectral-domain OCT scanner with a commercial surgical microscope. Thanks to our custom-made beam splitting and image display subsystems, the OCT images and microscopic images are simultaneously visualized through an ocular lens or the eyepiece of the microscope. This improvement helps surgeons to focus on the operation without distraction to view OCT images on another separate display. Moreover, displaying the OCT live images on the eyepiece helps surgeon’s depth perception during the surgeries. Finally, we successfully processed stimulated penetrating keratoplasty in live rabbits. We believe that these technical achievements are crucial to enhance the usability of the VISOCT system in a real surgical operating condition.

In this study, polarization-sensitive optical coherence tomography (PS-OCT) capable of providing polarization contrasts such as phase retardation and degree of polarization uniformity (DOPU) was used for visualizing human meibomian glands (MGs) and investigating morphological characteristics of them. Especially, with the help of the DOPU contrast, MGs were exclusively extracted from the volumetric OCT image. In vivo PS-OCT measurements were performed on the upper eyelids of different age groups. From these measurements, different age-dependent aspects of the MG structure were also observed. Based on these observations, it can be inferred that the PS-OCT system has the potential for clinical diagnosis and investigation of MG-related dry eye diseases like MG dysfunction (MGD) and acinar atrophy.

The expensive fabrication of current opto-microfluidic sensors is a barrier to the successful adoption of these devices in point-of-care testing. This work reports a simple inexpensive opto-microfluidic device incorporating a poly(dimethylsiloxane)-glass hybrid microfluidic chip modified with gold nanoparticles and a high-detectivity, high-stability organic photodetector. The enhancing effect of the gold nanoparticles on horseradish peroxidase-luminol-H 2 O 2 chemiluminescence was exploited in rapid single-analyte immunoassays. The limit of detection for 17-β estradiol was 2.5  pg/ml , which is ∼200 times more sensitive than previously reported chemiluminescent immunosensors employing other organic photodetectors. Detection was also demonstrated in complex media, including natural water and blood serum.

This opinion piece discusses potential funding opportunities by the NIH BRAIN initiative to support the development of deep imaging technologies.

Noninvasive treatments are increasingly being used for the management of basal cell carcinoma (BCC), the predominant type of nonmelanoma skin cancer, making the development of noninvasive diagnostic technologies highly relevant for clinical practice. Laser-induced fluorescence (LIF) spectroscopy emerges as an attractive diagnostic technique for the diagnosis and demarcation of BCC due to its noninvasiveness, high sensitivity, real-time measurements, and user-friendly methodology. LIF relies on the principle of differential fluorescence emission between abnormal and normal skin tissues (ex vivo and in vivo) in response to excitation by a specific wavelength of light. Fluorescence originates either from endogenous fluorophores (autofluorescence) or from exogenously administered fluorophores (photosensitizers). The measured optical properties and fluorophore contributions of normal skin and BCC are significantly different from each other and correlate well with tissue histology. Photodynamic diagnosis (PDD) is based on the visualization of a fluorophore, with the ability to accumulate in tumor tissue, by the use of fluorescence imaging. PDD may be used for detecting subclinical disease, determining surgical margins, and following-up patients for residual tumor or BCC relapse. In this review, we will present the basic principles of LIF and discuss its uses for the diagnosis, management, and follow-up of BCC.

Bone “stress-whitens,” becoming visibly white during mechanical loading, immediately prior to failure. Stress-whitening is known to make materials tougher by dissipating mechanical energy. A greater understanding of stress-whitening, both an optical and mechanical phenomenon, may help explain age-related increases in fracture risk that occur without changes in bone mineralization. In this work, we directly measure the optical properties of demineralized bone as a function of deformation and immersing fluid (with different hydrogen-bonding potentials, water, and ethanol). The change in refractive index of demineralized bone was linear: with deformation and not applied force. Changes in refractive index were likely due to pushing low-refractive-index fluid out of specimens and secondarily due to changes in the refractive index of the collagenous phase. Results were consistent with stress-whitening of demineralized bone previously observed. In ethanol, the refractive index values were lower and less sensitive to deformation compared with deionized water, corroborating the sensitivity to fluid hydration. Differences in refractive index were consistent with structural changes in the collagenous phase such as densification that may also occur under mechanical loading. Understanding bone quality, particularly stress-whitening investigated here, may lead to new therapeutic targets and noninvasive methods to assess bone quality.

Intralipid is widely used as an optical scattering agent in tissue-mimicking phantoms. Accurate control when using Intralipid is critical to match the optical diffusivity of phantoms to the prescribed value. Currently, most protocols of Intralipid-based hydrogel phantom fabrication focus on factors such as Intralipid brand and concentration. In this note, for the first time to our knowledge, we explore the dependence of the optical reduced scattering coefficient (at 532 nm optical wavelength) on the temperature and the time of mixing Intralipid with gelatin-water solution. The studied samples contained 1% Intralipid and were measured with oblique-incidence reflectometry. It was found that the reduced scattering coefficient increased when the Intralipid-gelatin-water mixture began to solidify at room temperature. For phantoms that had already solidified completely, the diffusivity was shown to be significantly influenced by the temperature and the duration of the mixing course. The dependence of the measured diffusivity on the mixing conditions was confirmed by experimental observations. Moreover, the mechanism behind the dependence behavior is discussed.

Thresholds for microcavitation of bovine and porcine melanosomes were determined using nanosecond laser pulses in the near-infrared (1000 to 1319 nm) wavelength regime. Isolated melanosomes were irradiated by single pulses (10 or 50 ns) using a Q-switched Spectra Physics Nd:YAG laser coupled with an optical parametric oscillator (1000 to 1200 nm) or a continuum laser at 1319 nm. Time-resolved nanosecond strobe photography after the arrival of the irradiation beam allowed imaging of microcavitation events. Average fluence thresholds for microcavitation increased nonlinearly with increasing wavelength from ∼0.5  J/cm 2 at 1000 nm to 2.6  J/cm 2 at 1319 nm. Fluence thresholds were also measured for 10-ns pulses at 532 nm and found to be comparable to visible nanosecond pulse values published in previous reports. Calculated melanosome absorption coefficients decreased from 925  cm −1 at 1000 nm to 176  cm −1 at 1319 nm. This trend was found to be comparable to the decrease in retinal pigmented epithelial layer absorption coefficients reported over the same wavelength region. Estimated corneal total intraocular energy retinal damage threshold values were determined in order to compare to current and proposed maximum permissible exposure (MPE) safe levels. Results from this study support recently proposed changes to the MPE levels.

Photopolymerization is commonly used in a broad range of bioapplications, such as drug delivery, tissue engineering, and surgical implants, where liquid materials are injected and then hardened by means of illumination to create a solid polymer network. However, photopolymerization using a probe, e.g., needle guiding both the liquid and the curing illumination, has not been thoroughly investigated. We present a Monte Carlo model that takes into account the dynamic absorption and scattering parameters as well as solid–liquid boundaries of the photopolymer to yield the shape and volume of minimally invasively injected, photopolymerized hydrogels. In the first part of the article, our model is validated using a set of well-known poly(ethylene glycol) dimethacrylate hydrogels showing an excellent agreement between simulated and experimental volume-growth-rates. In the second part, in situ experimental results and simulations for photopolymerization in tissue cavities are presented. It was found that a cavity with a volume of 152  mm 3 can be photopolymerized from the output of a 0.28-mm 2 fiber by adding scattering lipid particles while only a volume of 38  mm 3 (25%) was achieved without particles. The proposed model provides a simple and robust method to solve complex photopolymerization problems, where the dimension of the light source is much smaller than the volume of the photopolymerizable hydrogel.

Researchers use ultrasound (US) to modulate diffusive light in a highly scattering medium like tissue. This paper analyzes the US–optical interaction in the scattering medium and derives an expression for the US-modulated optical radiance. The diffusion approximation to the radiative transport equation is employed to develop a Green’s function for US-modulated light. The predicted modulated fluence and flux are verified using finite-difference time-domain simulations. The Green’s function is then utilized to illustrate the modulated reflectance as the US–optical interaction increases in depth. The intent of this paper is to focus on high US frequencies necessary for high-resolution imaging because they are of interest for applications such as phase conjugation.

The objectives of this study were to evaluate the effect of low-level laser irradiation (LLLI) on bovine oocyte and granulosa cells metabolism during in vitro maturation (IVM) and further embryo development. Cumulus-oocytes complexes (COCs) were subjected (experimental group) or not (control group) to irradiation with LLLI in a 633-nm wavelength and 1  J/cm 2 fluency. The COCs were evaluated after 30 min, 8, 16, and 24 h of IVM. Cumulus cells were evaluated for cell cycle status, mitochondrial activity, and viability (flow cytometry). Oocytes were assessed for meiotic progression status (nuclear staining), cell cycle genes content [real-time polymerase chain reaction (PCR)], and signal transduction status (western blot). The COCs were also in vitro fertilized, and the cleavage and blastocyst rates were assessed. Comparisons among groups were statistically performed with 5% significance level. For cumulus cells, a significant increase in mitochondrial membrane potential and the number of cells progressing through the cycle could be observed. Significant increases on cyclin B and cyclin-dependent kinase (CDK4) levels were also observed. Concerning the oocytes, a significantly higher amount of total mitogen-activated protein kinase was found after 8 h of irradiation, followed by a decrease in all cell cycle genes transcripts, exception made for the CDK4. However, no differences were observed in meiotic progression or embryo production. In conclusion, LLLI is an efficient tool to modulate the granulosa cells and oocyte metabolism.

There is an increasing use of high-power fiber lasers in manufacturing and telecommunications industries operating in the infrared spectrum between 1000 and 2000 nm, which are advertised to provide as much as 10 kW continuous output power at 1070 nm. Safety standards have traditionally been based on experimental and modeling investigations with scant data available for these wavelengths. A series of studies using 1070-nm infrared lasers to determine the minimum visible lesion damage thresholds in skin using the Yucatan miniature pig (Sus scrofa domestica) for a range of beam diameters (0.6, 1.1, 1.9, 2.4, 4.7, and 9.5 cm) and a range of exposure durations (10 ms to 10 s) is presented. Experimental peak temperatures associated with each damage threshold were measured using thermal imaging. Peak temperatures at damage threshold for the 10-s exposures were ∼10°C lower than those at shorter exposures. The lowest and highest experimental minimum visible lesion damage thresholds were found to have peak radiant exposures of 19 and 432  J/cm 2 for the beam diameter-exposure duration pairs of 2.4 cm, 25 ms and 0.6 cm, 10 s, respectively. Thresholds for beam diameters >2.5  cm had a weak to no effect on threshold radiant exposure levels for exposure times ≤0.25  s , but may have a larger effect on thresholds for exposures ≥10  s .

In vivo harmonic generation microscopy (HGM) has been applied successfully in healthy human skin and can achieve a submicron resolution, similar to histopathologic examination, even at a penetration depth up to 270 μm. This study aims to investigate the clinical applicability of HGM imaging for differential diagnosis of nonmelanoma pigmented skin lesions. A total of 42 pigmented skin tumors, including pigmented basal cell carcinoma, melanocytic nevus, and seborrheic keratosis were evaluated by HGM ex vivo or in vivo. Based on the standard histopathologic characteristics, we established the corresponding HGM imaging criteria for each pigmented tumor. Diagnostic performance of HGM for differentiating nonmelanoma pigmented skin tumors was evaluated through the observers’ direct general assessment (overall evaluation) or the presence of two imaging criteria with the highest sensitivity and specificity (major criteria evaluation). Our results show that, based on the direct general assessment, the sensitivity is 92% [95% confidence interval (CI): 67 to 97%] and the specificity is 96% (95% CI: 83 to 99%); by major criteria evaluation, 94% sensitivity (95% CI: 70 to 99%) and 100% specificity (95% CI: 87 to 100%) are achieved. Our study indicates that HGM serves as a promising histopathological examination tool for noninvasive differential diagnostics of nonmelanoma pigmented skin tumors.

The invention of green fluorescent protein and other molecular fluorescent probes has promoted applications of confocal and two-photon fluorescence microscopy in biology and medicine. However, exogenous fluorescence contrast agents may affect cellular structure and function, and fluorescence microscopy cannot image nonfluorescent chromophores. We overcome this limitation by integrating optical-resolution photoacoustic microscopy into a modern Olympus IX81 confocal, two-photon, fluorescence microscope setup to provide complementary, label-free, optical absorption contrast. Automatically coregistered images can be generated from the same sample. Imaging applications in ophthalmology, developmental biology, and plant science are demonstrated. For the first time, in a familiar microscopic fluorescence imaging setting, this trimodality microscope provides a platform for future biological and medical discoveries.

Photoacoustic microscopy (PAM) is becoming a vital tool for various biomedical studies, including functional and molecular imaging of cancer. However, due to the use of a focused ultrasonic transducer for photoacoustic detection, the image quality of conventional PAM degrades rapidly away from the ultrasonic focal zone. To improve the image quality of PAM for out-of-focus regions, we have developed compressed sensing based virtual-detector photoacoustic microscopy (CS-PAM). Through phantom and in vivo experiments, it has been demonstrated that CS-PAM can effectively extend the depth of focus of PAM, and thus may greatly expand its potential biomedical applications.

Quantification of cell proliferation and monitoring its kinetics are essential in fields of research such as developmental biology, oncology, etc. Although several proliferation assays exist, monitoring cell proliferation kinetics remains challenging. We present a novel cell proliferation assay based on real-time monitoring of cell culture inside a standard incubator using a lensfree video-microscope, combined with automated detection of single cell divisions over a population of several thousand cells. Since the method is based on direct visualization of dividing cells, it is label-free, continuous, and not sample destructive. Kinetics of cell proliferation can be monitored from a few hours to several days. We compare our method to a standard assay, the EdU proliferation assay, and as proof of principle, we demonstrate concentration-dependent and time-dependent effect of actinomycin D—a cell proliferation inhibitor.

Extracellular matrix (ECM) remodeling contributes to the pathogenic changes in chronic obstructive pulmonary disease (COPD) and is both complex and not well understood. Collagen I, a component of the ECM altered in COPD airways, has second harmonic generation (SHG) properties. The SHG signal is coherent, propagating both forward (F) (primarily organized/mature collagen fibrils) and backward (B) (primarily disorganized/immature collagen fibrils) parallel to the incident light. The F/B SHG ratio was used to determine the proportion of organized to disorganized collagen, with lower variation in F/B ratio between sampling regions within the same patient and between patients in the same disease group compared with analyzing F and B data alone. The F/B ratio was independent of laser power drift, regions analyzed within a tissue and tissue orientation during analysis. Using this method, we identified a significant difference in collagen organization in airway tissue between COPD and nondiseased. We have developed a robust optimization and calibration methodology that will allow direct comparison of data obtained at different times and from multiple microscopes, which is directly adaptable for use with other tissue types. We report a powerful new tool for advancing our understanding of pathological ECM remodeling that may uncover new therapeutic targets in the future.

Recent studies indicate that the type 2 cannabinoid receptors (CB 2 R ) have become an attractive target for treating a variety of pathologies, including cancers, neurodegenerative diseases, inflammation, pain, osteoporosis, immunological disorders and drug abuse. In addition, it appears that many of these diseases have up-regulated CB 2 R expression. However, the precise role of CB 2 R in the regulation of diseases remains unclear. The ability to specifically image CB 2 R would contribute to develop reliable CB 2 R -based therapeutic approaches with a better understanding of the mechanism of CB 2 R action in these diseases. We developed a CB 2 R -targeted zwitterionic near-infrared (NIR) fluorescent probe, ZW760-mbc94. When compared with a previously reported CB 2 R probe (NIR760-mbc94) with the same targeting moiety but a charged NIR fluorescent dye, ZW760-mbc94 showed improved binding specificity in vitro and ex vivo. Overall, ZW760-mbc94 appears to have great potential as a CB 2 R -targeted contrast agent.

We propose a two-dimensional (2-D) space-scale analysis of fringe patterns collected from a diffraction phase microscope based on the 2-D Morlet wavelet transform. We show that the adaptation of a ridge detection method with anisotropic 2-D Morlet mother wavelets is more efficient for analyzing cellular and high refractive index contrast objects than Fourier filtering methods since it can separate phase from intensity modulations. We compare the performance of this ridge detection method on theoretical and experimental images of polymer microbeads and experimental images collected from living myoblasts.

A Monte Carlo simulation of light propagation through the retina has been developed to understand the path-length distributions within the retinal vessel. For full-field illumination, the path-length distribution within the vessel comprises directly backscattered light and light that has passed once or twice through the vessel. The origins of these light path-length distributions can be better understood by investigating different combinations of single-point illumination and detection positions. Perhaps the most significant observation is that illumination at the edges of the vessel, rather than over the whole field of view, and detection directly above the vessel capture only the light that has taken a single pass through the vessel. This path-length distribution is tightly constrained around the diameter of the vessel and can potentially provide enhancements for oxygen saturation imaging. The method could be practically implemented using an offset-pinhole confocal imaging system or structured light illumination.

We describe a combination of fabrication techniques and a general process to construct a three-dimensional (3-D) phantom that mimics the size, macroscale structure, microscale surface topology, subsurface microstructure, optical properties, and functional characteristics of a cancerous bladder. The phantom also includes features that are recognizable in white light (i.e., the visual appearance of blood vessels), making it suitable to emulate the bladder for emerging white light+optical coherence tomography (OCT) cystoscopies and other endoscopic procedures of large, irregularly shaped organs. The fabrication process has broad applicability and can be generalized to OCT phantoms for other tissue types or phantoms for other imaging modalities. To this end, we also enumerate the nuances of applying known fabrication techniques (e.g., spin coating) to contexts (e.g., nonplanar, 3-D shapes) that are essential to establish their generalizability and limitations. We anticipate that this phantom will be immediately useful to evaluate innovative OCT systems and software being developed for longitudinal bladder surveillance and early cancer detection.

Optical microangiography based on optical coherence tomography (OCT) is prone to noise that arises from a static tissue region. Here, we propose a method that can significantly reduce this noise. The method is developed based on an approach that uses the magnitude information of OCT signals to produce tissue microangiograms, especially suitable for the case where a swept-source OCT system is deployed. By combined use of two existing OCT microangiography methods—ultrahigh-sensitive optical microangiography (UHS-OMAG) and correlation mapping OCT (cmOCT)—the final tissue microangiogram is generated by masking UHS-OMAG image using the binary representation of cmOCT image. We find that this process masks the residual static artifacts while preserving the vessel structures. The noise rejection capability of the masked approach (termed as mOMAG) is tested on a tissue-like flow phantom as well as an in vivo human skin tissue. Compared to UHS-OMAG and cmOCT, we demonstrate that the proposed method is capable of achieving improved signal-to-noise ratio in providing microcirculation images. Finally, we show its clinical potential by quantitatively assessing the vascular difference between a burn scar and a normal skin of human subject in vivo.

An in vivo near-infrared fluorescence (NIRF) imaging technique is described for therapy monitoring of ankle joints affected by collagen-induced arthritis, a model of human rheumatoid arthritis. Arthritis was induced in rats by intradermal injections of collagen and Freund’s incomplete adjuvant. For in vivo imaging, the nonspecific NIR dye tetrasulfocyanine (TSC) was used. Prior to and after treatment with a nonsteroidal anti-inflammatory drug, meloxicam, or analgesic drug, tramadol hydrochloride (which served as no-therapy control), normalized fluorescence intensities of each ankle joint were measured. Additionally, each ankle joint was characterized by clinical arthritis scoring and histopathology. Over a 3-week treatment period, a significant difference in disease progression between animals treated with meloxicam and tramadol hydrochloride was detected. A statistically significant improvement in ankle joint pathology from high- or moderate-grade to moderate- or low-grade upon meloxicam therapy, as determined by clinical evaluation, translated into a significant decrease in fluorescence intensity. In contrast, all arthritic joints of the no-therapy control group deteriorated to high-grade arthritis with high-fluorescence intensities in NIRF imaging.

A new approach of the criterion assignment for registration of erythrocyte agglutinates to instrumentally determine blood group type is suggested. The criterion is based on comparison of R and G components of RGB decomposition of microscopy digital image taken for the blood-serum mixture sample. For the chosen experimental conditions, the minimal size (area) of RBC agglutinate to be registered by the criterion suggested is estimated theoretically. The proposed method was tested experimentally on the example of monitoring agglutinates in flow. The encouraging experimental results were obtained for improvement of the resolving power of the method; the optimal experimental conditions were revealed for maximum resolution. Though the suggested method was realized for dynamic (flow) blood group determination, it could also be applied for diagnostics in a stationary environment. This approach increases the reliability of RBC agglutinates registration and, hence, blood group typing. The results may be used to develop the apparatus for automated determination of human blood group.

Field carcinogenesis is the initial stage of cancer progression. Understanding field carcinogenesis is valuable for both cancer biology and clinical medicine. Here, we used inverse spectroscopic optical coherence tomography to study colorectal cancer (CRC) and pancreatic cancer (PC) field carcinogenesis. Depth-resolved optical and ultrastructural properties of the mucosa were quantified from histologically normal rectal biopsies from patients with and without colon adenomas (n=85 ) as well as from histologically normal peri-ampullary duodenal biopsies from patients with and without PC (n=22 ). Changes in the epithelium and stroma in CRC field carcinogenesis were separately quantified. In both compartments, optical and ultra-structural alterations were consistent. Optical alterations included lower backscattering (μ b ) and reduced scattering (μ ′ s ) coefficients and higher anisotropy factor g . Ultrastructurally pronounced alterations were observed at length scales up to ∼450  nm , with the shape of the mass density correlation function having a higher shape factor D , thus implying a shift to larger length scales. Similar alterations were found in the PC field carcinogenesis despite the difference in genetic pathways and etiologies. We further verified that the chromatin clumping in epithelial cells and collagen cross-linking caused D to increase in vitro and could be among the mechanisms responsible for the observed changes in epithelium and stroma, respectively.

Multicomponent tissue models are viable tools to better understand cell responses in complex environments, but present challenges when investigated with live cell microscopy noninvasively. In this study, integrated nonlinear optical microscopy-optical coherence microscopy (NLOM-OCM) was used to characterize cell interactions within three-dimensional (3-D), multicomponent extracellular matrices. In fibrin-collagen mixtures, 3T3 fibroblasts were observed to recruit both fibrin and collagen fibers while remodeling matrices. Also, NLOM-OCM was used to observe collagen deposition by neonatal human dermal fibroblasts within originally fibrin matrices over an extended time. It was observed that preferentially aligned collagen deposition could be achieved with aligned fibroblasts but that cell alignment could be achieved without aligning the extant extracellular matrix. In summary, this multimodel imaging system has potential for both real-time and longitudinal imaging of living 3-D cultures, which is particularly important for evaluating cell microenvironments in composite scaffolds or serial characterization of engineered tissue constructs during culture.

This study examines the application of backscattered ultrasound (US) and photoacoustic (PA) signals for assessment of bone structure and density variations. Both methods were applied in the frequency-domain, employing linear frequency modulation chirps. A near-IR laser (800 nm) was used for inducing the PA signal. The backscattered pressure waves were detected with a 2.2-MHz US transducer. Experiments were focused on detection and evaluation of PA and US signals from in-vitro animal and human bones with cortical and trabecular sublayers. It was shown that PA signals can be detected as deep as a few millimeters below trabecular and cortical layers. The occurrence of multiple scattering was demonstrated in PA detected signals from cancellous bone. Osteoporotic changes in the bone were simulated by using a very mild demineralization ethylenediaminetetraacetic acid solution. Changes in the time-domain signals as well as integrated backscattering spectra were compared for the samples before and after demineralization. The results demonstrated the sensitivity of PA to variations in bone minerals. In comparison to PA, US was capable of generating detectable signals from deeper bone sublayers (few centimeters). However, while US signal variations with changes in the cortical layer were insignificant, PA proved to be sensitive even to minor variations of the cortical bone density.

A new approach for generating high-speed multispectral confocal images has been developed. The central concept is that spectra can be acquired for each pixel in a confocal spatial scan by using a fast spectrometer based on optical fiber delay lines. This approach merges fast spectroscopy with standard spatial scanning to create datacubes in real time. The spectrometer is based on a serial array of reflecting spectral elements, delay lines between these elements, and a single element detector. The spatial, spectral, and temporal resolution of the instrument is described and illustrated by multispectral images of laser-induced autofluorescence in biological tissues.

An optimal measurement selection strategy based on incoherence among rows (corresponding to measurements) of the sensitivity (or weight) matrix for the near infrared diffuse optical tomography is proposed. As incoherence among the measurements can be seen as providing maximum independent information into the estimation of optical properties, this provides high level of optimization required for knowing the independency of a particular measurement on its counterparts. The proposed method was compared with the recently established data-resolution matrix-based approach for optimal choice of independent measurements and shown, using simulated and experimental gelatin phantom data sets, to be superior as it does not require an optimal regularization parameter for providing the same information.

We report on the feasibility of using long-range swept-source optical coherence tomography (OCT) to detect airway changes following smoke inhalation in a sheep model. The long-range OCT system (with axial imaging range of 25 mm) and probe are capable of rapidly obtaining a series of high-resolution full cross-sectional images and three-dimensional reconstructions covering 20-cm length of tracheal and bronchial airways with airway diameter up to 25 mm, regardless of the position of the probe within the airway lumen. Measurements of airway thickness were performed at baseline and postinjury to show mucosal thickness changes following smoke inhalation.

Quantitative analysis of optical clearing effects (OCE) induced by hyperosmotic agents is very important to optical tissue clearing applications in biomedical diagnostic imaging and therapeutics. This study aims at investigating the effect of glycerol concentration on the laser-scanning optical-resolution photoacoustic microscopy (LSOR-PAM) imaging contrast and light penetration depth. The photoacoustic (PA) signal amplitude changes are evaluated as a function of the concentration of glycerol. The results reveal that the PA signal amplitudes are enhanced with the glycerol concentration increasing, and also show that higher concentration of glycerol produces better light penetration and OCE on a phantom. The PA signal amplitude increases only 8.1% for 20% glycerol, but for higher concentrations, the increases are 76% and 165% for 40% and 60% glycerol, respectively. This preliminary study demonstrates that application of glycerol as an optical contrast agent reduces the tissue scattering and is beneficial to PAM imaging and optical diagnosis in clinical dermatology.

Ovarian cancer is the most deadly gynecologic cancer, a fact which is attributable to poor early detection and survival once the disease has reached advanced stages. Intraoperative laparoscopic volume holographic imaging has the potential to provide simultaneous visualization of surface and subsurface structures in ovarian tissues for improved assessment of developing ovarian cancer. In this ex vivo ovarian tissue study, we assembled a benchtop volume holographic imaging system (VHIS) to characterize the microarchitecture of 78 normal and 40 abnormal tissue specimens derived from ovarian, fallopian tube, uterine, and peritoneal tissues, collected from 26 patients aged 22 to 73 undergoing bilateral salpingo-oophorectomy, hysterectomy with bilateral salpingo-oophorectomy, or abdominal cytoreductive surgery. All tissues were successfully imaged with the VHIS in both reflectance- and fluorescence-modes revealing morphological features which can be used to distinguish between normal, benign abnormalities, and cancerous tissues. We present the development and successful application of VHIS for imaging human ovarian tissue. Comparison of VHIS images with corresponding histopathology allowed for qualitatively distinguishing microstructural features unique to the studied tissue type and disease state. These results motivate the development of a laparoscopic VHIS for evaluating the surface and subsurface morphological alterations in ovarian cancer pathogenesis.

Optoacoustic (photoacoustic) imaging has already showcased the capacity to offer high-resolution small animal visualization in vivo in a variety of cancer, cardiovascular, or neuroimaging applications. In particular, multispectral optoacoustic tomography (MSOT) has shown imaging along the spectral and the time dimensions, enabling sensing of multiple molecules over time and, more recently, in real time. Furthermore, cross-sectional imaging of at least 20 mm diameter has been showcased in vivo in animals and humans using 64-element curved transducers placed along a single curved line. Herein, we investigated the imaging improvements gained by utilizing a larger number of detectors and inquired whether more detectors will result in measurable image quality improvements. For this reason, we implemented MSOT using 64-, 128-, and 256-element transducers and imaged the same phantoms and animals under similar conditions. Further, corroborated by numerical simulation analysis, our findings quantify the improvements in resolution and overall image quality for the increasing number of detectors used pointing to significant improvements in image quality for the 256 detector array, over 64 or 128 detectors.

Autofluorescence (AF) imaging can provide valuable information about the structural and metabolic state of tissue that can be useful for elucidating physiological and pathological processes. Optical coherence tomography (OCT) provides high resolution detailed information about tissue morphology. We present coregistered AF-OCT imaging of human lung sections. Adjacent hematoxylin and eosin stained histological sections are used to identify tissue structures observed in the OCT images. Segmentation of these structures in the OCT images allowed determination of relative AF intensities of human lung components. Since the AF imaging was performed on tissue sections perpendicular to the airway axis, the results show the AF signal originating from the airway wall components free from the effects of scattering and absorption by overlying layers as is the case during endoscopic imaging. Cartilage and dense connective tissue (DCT) are found to be the dominant fluorescing components with the average cartilage AF intensity about four times greater than that of DCT. The epithelium, lamina propria, and loose connective tissue near basement membrane generate an order of magnitude smaller AF signal than the cartilage fluorescence.

Currently, laser fluence calibration is typically required for quantitative measurement of particle concentration in photoacoustic imaging. Here, we present a calibration-free method to quantify the absolute particle concentration by statistically analyzing photoacoustic signals. The proposed method is based on the fact that Brownian motion induces particle count fluctuation in the detection volume. If the count of particles in the detection volume is assumed to follow the Poisson distribution, its expected value can be calculated by the photoacoustic signal mean and variance. We first derived a theoretical model for photoacoustic signals. Then, we applied our method to quantitative measurement of different concentrations of various particles, including red blood cells. Finally, we performed in vivo experiments to demonstrate the potential of our method in biological applications. The experimental results agreed well with the predictions from the theoretical model suggesting that our method can be used for noninvasive measurement of absolute particle concentrations in deep tissue without fluence calibration.

Review of the existing studies on the contact pressure–induced changes in the optical properties of biological tissues showed that the reported changes in transmittance, reflectance, absorption, and scattering coefficient are vastly inconsistent. In order to gain more insight into the contact pressure–induced changes observed in biomedical applications involving common probe-spectrometer diffuse reflectance measurement setups and provide a set of practical guidelines minimizing the influence of the changes on the analysis of acquired spectra, we conducted a series of in vivo measurements, where the contact pressure was precisely controlled, and the spectral and contact pressure information were acquired simultaneously. Classification of three measurement sites on a human hand, representing the natural variability in the perfusion and structure of the underlying tissue, was assessed by training and evaluating classifiers at different contact pressure levels and for different probe operators. Based on the results, three practical guidelines have been proposed to avoid classification performance degradation. First, the most suitable pressure level should be identified. Second, the pressure level should be kept in a narrow range during the acquisition of spectra. Third, applications utilizing probes equipped with a calibrated spring can use several classifiers trained at different contact pressure levels to improve classification performance.

Steady-state and time-resolved fluorescence spectroscopy were employed in the discrimination of cervical cancer patients from healthy subjects using urine samples. Fluorescence emission at 390 and 440 nm was considered to monitor the fluorescence of indoxyl sulfate and neopterin. Significant spectral differences were observed between healthy and cancer subjects. Different ratio parameters were calculated from the spectral intensity at 280- and 350-nm excitation and were subjected to stepwise linear discriminant analysis. In total, 84.0% of samples were correctly classified at 280 nm and 96.4% were correctly classified at 350 nm. The fluorescence decay kinetics of urine samples at 390-nm emission was best described by bi- exponential fits, whereas the decay characteristics at 440 nm of urine samples was best explained by bi-exponential fits and, in some cases, by tri-exponential fits. However, the decay kinetics of both indoxyl sulfate and neopterin standards was well described by bi-exponential decays. Based on the fluorescence emission characteristics and statistical analysis, the fluorophores indoxyl sulfate, neopterin, and riboflavin may be considered as potential biomarkers for cervical cancer diagnosis.

Tissue vasculature is altered when cancer develops. Consequently, noninvasive methods of monitoring blood vessel size, density, and oxygenation would be valuable. Simple spectroscopy employing fiber optic probes to measure backscattering can potentially determine hemoglobin parameters. However, heterogeneity of blood distribution, the dependence of the tissue-volume-sampled on scattering and absorption, and the potential compression of tissue all hinder the accurate determination of hemoglobin parameters. We address each of these issues. A simple derivation of a correction factor for the absorption coefficient, μ a , is presented. This correction factor depends not only on the vessel size, as others have shown, but also on the density of blood vessels. Monte Carlo simulations were used to determine the dependence of an effective pathlength of light through tissue which is parameterized as a ninth-order polynomial function of μ a . The hemoglobin bands of backscattering spectra of cervical tissue are fit using these expressions to obtain effective blood vessel size and density, tissue hemoglobin concentration, and oxygenation. Hemoglobin concentration and vessel density were found to depend on the pressure applied during in vivo acquisition of the spectra. It is also shown that determined vessel size depends on the blood hemoglobin concentration used.

Tryptophan is investigated as the key native marker in cells to determine the level of metastasis competence in breast cell lines using native fluorescence spectroscopy. The ratio of fluorescence intensity at 340 nm to intensity at 460 nm is associated with aggressiveness of the cancer cells. We found that the fluorescence of aggressive breast cancer cell has a much higher contribution from tryptophan compared with that from the normal cells and nonaggressive breast cancer cell.

The purpose of the present study was to measure the intradiscal pressure signal of an anesthetized sheep under spontaneous breathing. An ultra-miniature fiber optic high-pressure sensor was implanted into the nucleus pulposus of the fifth lumbar intervertebral using a dorsolateral transforaminal approach. Results suggested the periodicity of the intradiscal pressure signal was similar to the mean respiratory rate of the animal. The average resting intradiscal pressure was also calculated and compared to available data.

Low-level light therapy has been shown to improve in vitro wound healing. However, well-defined parameters of different light sources for this therapy are lacking. The goal of this study was (1) to determine if the wavelengths tested are effective for in vitro wound healing and (2) to compare a laser and a light-emitting diode (LED) source at similar wavelengths. We show four wavelengths, delivered by either a laser or LED array, improved in vitro wound healing in A549, U2OS, and PtK2 cells. Improved wound healing occurred through increased cell migration demonstrated through scratch wound and transwell assays. Cell proliferation was tested by the (3-(4,5-dimethylthiazol-2-yl)-5-(3-car-boxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) (MTS) assay and was found generally not to be involved in the wound healing process. The laser and LED sources were found to be comparable when equal doses of light were applied. The biological response measured was similar in most cases. We conclude that the laser at 652 (5.57  mW/cm 2 , 10.02  J/cm 2 ) and 806 nm (1.30  mW/cm 2 , 2.334  J/cm 2 ) (full bandwidth 5 nm), and LED at 637 (5.57  mW/cm 2 , 10.02  J/cm 2 ) and 901 nm (1.30  mW/cm 2 , 2.334  J/cm 2 ) (full bandwidth 17 and 69 nm respectively) induce comparable levels of cell migration and wound closure.

Suture ligation with subsequent cutting of blood vessels to maintain hemostasis during surgery is time consuming and skill intensive. Energy-based electrosurgical and ultrasonic devices are often used to replace sutures and mechanical clips to provide rapid hemostasis and decrease surgery time. Some of these devices may create undesirably large collateral zones of thermal damage and tissue necrosis, or require separate mechanical blades for cutting. Infrared lasers are currently being explored as alternative energy sources for vessel sealing applications. In a previous study, a 1470-nm laser was used to seal vessels 1 to 6 mm in diameter in 5 s, yielding burst pressures of ∼500  mmHg . The purpose of this study was to provide vessel sealing times comparable with current energy-based devices, incorporate transection of sealed vessels, and demonstrate high vessel burst pressures to provide a safety margin for future clinical use. A 110-W, 1470-nm laser beam was transmitted through a fiber and beam shaping optics, producing a 90-W linear beam 3.0 by 9.5 mm for sealing (400  W/cm 2 ), and 1.1 by 9.6 mm for cutting (1080  W/cm 2 ). A two-step process sealed and then transected ex vivo porcine renal vessels (1.5 to 8.5 mm diameter) in a bench top setup. Seal and cut times were 1.0 s each. A burst pressure system measured seal strength, and histologic measurements of lateral thermal spread were also recorded. All blood vessels tested (n=55 seal samples) were sealed and cut, with total irradiation times of 2.0 s and mean burst pressures of 1305±783  mmHg . Additional unburst vessels were processed for histological analysis, showing a lateral thermal spread of 0.94±0.48  mm (n=14 seal samples). This study demonstrated that an optical-based system is capable of precisely sealing and cutting a wide range of porcine renal vessel sizes and, with further development, may provide an alternative to radiofrequency- and ultrasonic-based vessel sealing devices.

Lasers are part of our everyday life. They are used from manufacturing to entertainment, warfare (laser-guided weapons), construction, do-it-yourself (DIY) projects, communication, education, medicine, robotics, automotive, aviation, space, and more. Since their invention over five decades ago, the field of medicine has been a major beneficiary of laser technology. The book Lasers for Medical Applications: Diagnostics, Therapy and Surgery describes this subject in detail. This book is edited by Prof. Helena Jelinkova of the Czech Technical University of the Czech Republic in Eastern Europe.