Rapid intra‐operative diagnosis of kidney cancer by attenuated total reflection infrared spectroscopy of tissue smears

Herein, a technique to analyze air‐dried kidney tissue impression smears by means of attenuated total reflection infrared (ATR‐IR) spectroscopy is presented. Spectral tumor markers—absorption bands of glycogen—are identified in the ATR‐IR spectra of the kidney tissue smear samples. Thin kidney tissue cryo‐sections currently used for IR spectroscopic analysis lack such spectral markers as the sample preparation causes irreversible molecular changes in the tissue. In particular, freeze‐thaw cycle results in degradation of the glycogen and reduction or complete dissolution of its content. Supervised spectral classification was applied to the recorded spectra of the smears and the test spectra were classified with a high accuracy of 92% for normal tissue and 94% for tumor tissue, respectively. For further development, we propose that combination of the method with optical fiber ATR probes could potentially be used for rapid real‐time intra‐operative tissue analysis without interfering with either the established protocols of pathological examination or the ordinary workflow of operating surgeon. Such approach could ensure easier transition of the method to clinical applications where it may complement the results of gold standard histopathology examination and aid in more precise resection of kidney tumors.      

Liver tissue classification of en face images by fractal dimension‐based support vector machine

Full‐field optical coherence tomography (FF‐OCT) has been reported with its label‐free subcellular imaging performance. To realize quantitive cancer detection, the support vector machine model of classifying normal and cancerous human liver tissue is proposed with en face tomographic images. Twenty samples (10 normal and 10 cancerous) were operated from humans and composed of 285 en face tomographic images. Six histogram features and one proposed fractal dimension parameter that reveal the refractive index inhomogeneities of tissue were extracted and made up the training set. The other different 16 samples (8 normal and 8 cancerous) were imaged (190 images) and employed as the test set with the same features. First, a subcellular‐resolution tomographic image library for four histopathological areas in liver tissue was established. Second, the area under the receiver operating characteristics of 0.9378, 0.9858, 0.9391, 0.9517 for prediction of the cancerous hepatic cell, central vein, fibrosis, and portal vein were measured with the test set. The results indicate that the proposed classifier from FF‐OCT images shows promise as a label‐free assessment of quantified tumor detection, suggesting the fractal dimension‐based classifier could aid clinicians in detecting tumor boundaries for resection in surgery in the future.     

Miniaturized all fiber probe for optical coherence tomography and pH detection of biological tissue

We present a novel all-fiber probe with 710-μm outside diameter for combined optical coherence tomography and pH detection. In cancer surgery, a significant challenge is how to completely remove the malignant tumor without cutting too much normal tissue. The difference between cancer tissue and normal tissue not only lies in morphology and structure but also in tissue pH, where malignant tissue has a lower pH. This dual-modality probe combined optical coherence tomography and pH detection of biological tissue, is expected to determine whether the tissue is cancerous quickly and accurately. The probe utilizes a typical three-segment structure (double-clad fiber - no-core fiber - graded-index fiber). We obtained a lateral resolution of ~10.6 μm, a working distance of ~506 μm and a pH measurement accuracy of 0.01 pH unit for the probe. The performance of the all-fiber probe was verified through an ex vivo experiment using the porcine brain specimen.     

Characterization of endogenous fluorescence in nonsmall lung cancerous cells: A comparison with nonmalignant lung normal cells

Monitoring fluorescence properties of endogenous fluorophores such as nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD) in normal and cancerous cells provide substantial information noninvasively on biochemical and biophysical aspects of metabolic dysfunction of cancerous cells. Time‐resolved spectral profiles and fluorescence lifetime images of NADH and FAD were obtained in human lung nonsmall carcinomas (H661 and A549) and normal lung cells (MRC‐5). Both fluorophores show the fast and slowly decaying emission components upon pulsed excitation, and fluorescence spectra of NADH and FAD show blue‐ and red‐shifts, respectively, during their decay. All identified lifetime components of NADH and FAD were found to be shorter in cancerous cells than in normal cells, no matter how they were measured under different extra‐cellular conditions (cells suspended in cuvette and cells attached on glass substrate), indicating that the changes in metabolism likely altered the subcellular milieu and potentially also affected the interaction of NADH and FAD with enzymes to which these cofactors were bound. The intensity ratio of NADH and FAD of cancerous cells was also shown to be larger than that of normal cells.     

Angular optimization for cancer identification with circularly polarized light

Depolarization of circularly polarized light scattered from biological tissues depends on structural changes in cell nuclei, which can provide valuable information for differentiating cancer tissues concealed in healthy tissues. In this study, we experimentally verified the possibility of cancer identification using scattering of circularly polarized light. We investigated the polarization of light scattered from a sliced biological tissue with various optical configurations. A significant difference between circular polarizations of light scattered from cancerous and healthy tissues is observed, which is sufficient to distinguish a cancerous region. The line-scanning experiments along a region incorporating healthy and cancerous parts indicate step-like behaviors in the degree of circular polarization corresponding to the state of tissues, whether cancerous or normal. An oblique and perpendicular incidence induces different resolutions for identifying cancerous tissues, which indicates that the optical arrangement can be selected according to the priority of resolution.     

Fourier transform infrared microspectroscopy as a diagnostic tool for distinguishing between normal and malignant human gastric tissue

Fourier transform infrared (FTIR) microspectroscopy can be considered to be a fast and non-invasive tool for distinguishing between normal and cancerous cells and tissues without the need for laborious and invasive sampling procedures. Gastric samples from four patients (age, 65±2 years) were analysed. Samples were obtained from the organs removed during gastrectomy and then classified as normal or cancerous. Classification was based on histopathological examinations at our institution. Formalin-fixed sections of gastric tissue were analysed by FTIR- microspectroscopy. To characterize differences between sections of normal and cancerous tissue, specific regions of the spectra were analysed to study variations in the levels of metabolites. To distinguish between two conditions (normal and cancerous), changes in the relative intensity of bands in the range 600–4000 cm⁻¹ were analysed. A FTIR spectral map of the bands in the region 2800–3100 cm^–1 and 900–1800 cm^–1 were created to analyse pathological changes in tissues. The limited data available showed that normal gastric tissue had stronger absorption than cancerous tissue over a wide region in the four patients. There was a significant decrease in total biomolecular components for cancerous tissue compared with normal tissue.     

Rapid intraoperative diagnosis of gynecological cancer by ATR‐FTIR spectroscopy of fresh tissue biopsy

A rapid and reliable intraoperative diagnostic technique to support clinical decisions was developed using Fourier‐transform infrared (FTIR) spectroscopy. Twenty‐six fresh tissue samples were collected intraoperatively from patients undergoing gynecological surgeries. Frozen section (FS) histopathology aimed to discriminate between malignant and benign tumors was performed, and attenuated total reflection (ATR) FTIR spectra were collected from these samples. Digital dehydration and principal component analysis and linear discriminant analysis (PCA‐LDA) models were developed to classify samples into malignant and benign groups. Two validation schemes were employed: k‐fold and “leave one out.” FTIR absorption spectrum of a fresh tissue sample was obtained in less than 5 minutes. The fingerprint spectral region of malignant tumors was consistently different from that of benign tumors. The PCA‐LDA discrimination model correctly classified the samples into malignant and benign groups with accuracies of 96% and 93% for the k‐fold and “leave one out” validation schemes, respectively. We showed that a simple tissue preparation followed by ATR‐FTIR spectroscopy provides accurate means for very rapid tumor classification into malignant and benign gynecological tumors. With further development, the proposed method has high potential to be used as an adjunct to the intraoperative FS histopathology technique.     

In vivo sun protection factor and UVA protection factor determination using (hybrid) diffuse reflectance spectroscopy and a multi-lambda-LED light source

The sun protection factor (SPF) values are currently determined using an invasive procedure, in which the volunteers are irradiated with ultraviolet (UV) light. Non-invasive approaches based on hybrid diffuse reflectance spectroscopy (HDRS) have shown a good correlation with conventional SPF testing. Here, we present a novel compact and adjustable DRS test system. The in vivo measurements were performed using a multi-lambda-LED light source and an 84-channel imaging spectrograph with a fiber optic probe for detection. A transmission spectrum was calculated based on the reflectance measured with sunscreen and the reflectance measured without sunscreen. The preexposure in vitro spectrum was fitted to the in vivo spectrum. Each of the 11 test products was investigated on 10 volunteers. The SPF and UVA-PF values obtained by this new approach were compared with in vivo SPF results determined by certified test institutes. A correlation coefficient R² = 0.86 for SPF, and R² = 0.92 for UVA-PF were calculated. Having examined various approaches to apply the HDRS principle, the method we present was found to produce valid and reproducible results, suggesting that the multi-lambda-LED device is suitable for in-vivo SPF testing based on the HDRS principle as well as for in-vivo UVA-PF measurements.     

Thermo-optic measurements and their inter-dependencies for delineating cancerous breast biopsy tissue from adjacent normal

The histopathological diagnosis of cancer is the current gold standard to differentiate normal from cancerous tissues. We propose a portable platform prototype to characterize the tissue's thermal and optical properties, and their inter-dependencies to potentially aid the pathologist in making an informed decision. The measurements were performed on 10 samples from five subjects, where the cancerous and adjacent normal were extracted from the same patient. It was observed that thermal conductivity (k) and reduced-scattering-coefficient (μ'_s) for both the cancerous and normal tissues reduced with the rise in tissue temperature. Comparing cancerous and adjacent normal tissue, the difference in k and μ'_s (at 940 nm) were statistically significant (p = 7.94e-3), while combining k and μ'_s achieved the highest statistical significance (6.74e-4). These preliminary results promise and support testing on a large number of samples for rapidly differentiating cancerous from adjacent normal tissues.     

SERS-based sensor for direct L-selectin level determination in plasma samples as alternative method of tumor detection

Selectin ligands are present on the surface of tumor cells, for this reason lowering the L-selectin level in the blood and lymph can indicate presence of the tumor. Therefore the selectin level in the plasma are potential targets for anticancer therapy. We demonstrate the surface enhanced Raman spectroscopy (SERS)-based sensor for the determination of L-selectin level in biological samples that can be used in medical diagnosis. The combination of SERS with the method of multivariate analysis as principle component analysis (PCA) allows to strengthen the presented data analysis. The loadings of PCA permit to indicate those vibration modes, that are the most important for the assumed identification (bands at 1574, 1450, 1292 cm⁻¹). Two bands at 1286 and 1580 cm⁻¹ were selected for the determination of the calibration curve (bands intensities I₁₂₈₆/I₁₅₈₀ ratio). The L-selectin level of biological samples can be read, directly from the calibration curve. The presented sensor is as a sensitive tool with good specificity and selectivity of L-selectin, even in the case of coexistence of P- and E-selectin.     

Raman spectral discrimination in human liquid biopsies of oesophageal transformation to adenocarcinoma

The aim of this study was to determine whether Raman spectroscopy combined with chemometric analysis can be applied to interrogate biofluids (plasma, serum, saliva and urine) towards detecting oesophageal stages through to oesophageal adenocarcinoma [normal/squamous epithelium, inflammatory, Barrett's, low‐grade dysplasia, high‐grade dysplasia and oesophageal adenocarcinoma (OAC)]. The chemometric analysis of the spectral data was performed using principal component analysis, successive projections algorithm or genetic algorithm (GA) followed by quadratic discriminant analysis (QDA). The genetic algorithm quadratic discriminant analysis (GA‐QDA) model using a few selected wavenumbers for saliva and urine samples achieved 100% classification for all classes. For plasma and serum, the GA‐QDA model achieved excellent accuracy in all oesophageal stages (>90%). The main GA‐QDA features responsible for sample discrimination were: 1012 cm^?1 (C─O stretching of ribose), 1336 cm^?1 (Amide III and CH₂ wagging vibrations from glycine backbone), 1450 cm^?1 (methylene deformation) and 1660 cm^?1 (Amide I). The results of this study are promising and support the concept that Raman on biofluids may become a useful and objective diagnostic tool to identify oesophageal disease stages from squamous epithelium to OAC.     

Optical fan for single‐cell screening

The single‐cell screening has attracted great attentions in advanced biomedicine and tissue biology, especially for the early disease diagnosis and treatment monitoring. In this work, by using a specific‐designed fiber probe with a flat facet, we propose an “optical fan” strategy to screen K562 cells at the single‐cell level from a populations of RBCs. After the 980‐nm laser beam injected into the fiber probe, the RBCs were blown away but holding target K562 cells in place. Further, multiple leukemic cells can be screened from hundreds of red blood cells, providing an efficient approach for the cell screening. The experimental results were interpreted by the numerical simulation, and the stiffness of optical fan was also discussed.     

Diagnosing colorectal abnormalities using scattering coefficient maps acquired from optical coherence tomography

Optical coherence tomography (OCT) has shown potential in differentiating normal colonic mucosa from neoplasia. In this study of 33 fresh human colon specimens, we report the first use of texture features and computer vision-based imaging features acquired from en face scattering coefficient maps to characterize colorectal tissue. En face scattering coefficient maps were generated automatically using a new fast integral imaging algorithm. From these maps, a gray-level cooccurrence matrix algorithm was used to extract texture features, and a scale-invariant feature transform algorithm was used to derive novel computer vision-based features. In total, 25 features were obtained, and the importance of each feature in diagnosis was evaluated using a random forest model. Two classifiers were assessed on two different classification tasks. A support vector machine model was found to be optimal for distinguishing normal from abnormal tissue, with 94.7% sensitivity and 94.0% specificity, while a random forest model performed optimally in further differentiating abnormal tissues (i.e., cancerous tissue and adenomatous polyp) with 86.9% sensitivity and 85.0% specificity. These results demonstrated the potential of using OCT to aid the diagnosis of human colorectal disease.     

Raman spectroscopy of human skin for kidney failure detection

The object of this paper is in vivo study of skin spectral-characteristics in patients with kidney failure by conventional Raman spectroscopy in near infrared region. The experimental dataset was subjected to discriminant analysis with the projection on latent structures (PLS-DA). Application of Raman spectroscopy to investigate the forearm skin in 85 adult patients with kidney failure (90 spectra) and 40 healthy adult volunteers (80 spectra) has yielded the accuracy of 0.96, sensitivity of 0.94 and specificity of 0.99 in terms of identifying the target subjects with kidney failure. The autofluorescence analysis in the near infrared region identified the patients with kidney failure among healthy volunteers of the same age group with specificity, sensitivity, and accuracy of 0.91, 0.84, and 0.88, respectively. When classifying subjects by the presence of kidney failure using the PLS-DA method, the most informative Raman spectral bands are 1315 to 1330, 1450 to 1460, 1700 to 1800 cm⁻¹. In general, the performed study demonstrates that for in vivo skin analysis, the conventional Raman spectroscopy can provide the basis for cost-effective and accurate detection of kidney failure and associated metabolic changes in the skin.     

Real‐time augmented reality for delineation of surgical margins during neurosurgery using autofluorescence lifetime contrast

Current clinical brain imaging techniques used for surgical planning of tumor resection lack intraoperative and real‐time feedback; hence surgeons ultimately rely on subjective evaluation to identify tumor areas and margins. We report a fluorescence lifetime imaging (FLIm) instrument (excitation: 355 nm; emission spectral bands: 390/40 nm, 470/28 nm, 542/50 nm and 629/53 nm) that integrates with surgical microscopes to provide real‐time intraoperative augmentation of the surgical field of view with fluorescent derived parameters encoding diagnostic information. We show the functionality and safety features of this instrument during neurosurgical procedures in patients undergoing craniotomy for the resection of brain tumors and/or tissue with radiation damage. We demonstrate in three case studies the ability of this instrument to resolve distinct tissue types and pathology including cortex, white matter, tumor and radiation‐induced necrosis. In particular, two patients with effects of radiation‐induced necrosis exhibited longer fluorescence lifetimes and increased optical redox ratio on the necrotic tissue with respect to non‐affected cortex, and an oligodendroglioma resected from a third patient reported shorter fluorescence lifetime and a decrease in optical redox ratio than the surrounding white matter. These results encourage the use of FLIm as a label‐free and non‐invasive intraoperative tool for neurosurgical guidance.     

Pilot feasibility study of in vivo intraoperative quantitative optical coherence tomography of human brain tissue during glioma resection

This study investigates the feasibility of in vivo quantitative optical coherence tomography (OCT) of human brain tissue during glioma resection surgery in six patients. High‐resolution detection of glioma tissue may allow precise and thorough tumor resection while preserving functional brain areas, and improving overall survival. In this study, in vivo 3D OCT datasets were collected during standard surgical procedure, before and after partial resection of the tumor, both from glioma tissue and normal parenchyma. Subsequently, the attenuation coefficient was extracted from the OCT datasets using an automated and validated algorithm. The cortical measurements yield a mean attenuation coefficient of 3.8 ± 1.2 mm^?1 for normal brain tissue and 3.6 ± 1.1 mm^?1 for glioma tissue. The subcortical measurements yield a mean attenuation coefficient of 5.7 ± 2.1 and 4.5 ± 1.6 mm^?1 for, respectively, normal brain tissue and glioma. Although the results are inconclusive with respect to trends in attenuation coefficient between normal and glioma tissue due to the small sample size, the results are in the range of previously reported values. Therefore, we conclude that the proposed method for quantitative in vivo OCT of human brain tissue is feasible during glioma resection surgery.     

Colon cancer detection by using Poincaré sphere and 2D polarimetric mapping of ex vivo colon samples

This work is dedicated to the diagnosis and grading of colon cancer by a combined use of Poincaré sphere and 2D Stokes vector polarimetry mapping approaches. The major challenge consists in exploring the applicability of polarized light for noninvasive screening of the histological abnormalities within the samples of biological tissues. Experimental studies were conducted in ex vivo colon sample, excised after surgical procedure for colon tumor removal of G2‐adenocarcinoma lesion. Polarimetric measurements in linear and circular regime were carried via personally developed polarimetric, optical set‐up, using supercontinuous fiber laser with irradiation fixed at 635 nm. We apply the Poincaré sphere and two‐dimensional Stokes vector scanning approach for screening the corresponding tissue samples. A comparison between linear and circular polarization states is made both for quantitative and qualitative evaluations. It is shown that circular polarization has better diagnostic capabilities than linear polarization, with higher dynamic ranges of the polarimetric parameters and better values of the diagnostic quantities. In addition to the standard polarimetry parameters, utilized as essential diagnostic markers, we apply statistical analysis to obtain more detailed information in frame of the applied diagnostic approach.     

Immunotargeting of urothelial cell carcinoma with intravesically administered Tc-99m labeled HMFG1 monoclonal antibody

Ten patients with transitional cell carcinoma (TCC) of the bladder received 3–6 mCi of HMFG1 monoclonal antibody (MAb) intravesically. The antibody was labeled with Tc-99m using the 2-Iminothiolane method. All patients underwent transurethral resection of the bladder tumor within 12–20 h following intravesical administration of^99m-Tc-HMFG1. The presence of the radiolabeled MAb in the circulation was studied by measuring the radioactivity in the serum for a period up to 20 h. Three of 10 patients underwent immunoscintigraphy (SPECT) 2–3 h postadministration and cancerous areas could be easily localized. Biopsies were taken from the tumor sites as well as from normal bladder mucosa. Absolute uptake of the administered MAb expressed as percent administered dose/kg of tissue could be evaluated only in eight patients. Multiple specimen taken from various tumor sites in every patient gave a wide range of uptake values ranging from 0 to 9.29% adm. dose/kg, whereas normal tissue uptake values ranged from 0 to 1.63, respectively.     

Blood plasma resonance Raman spectroscopy combined with multivariate analysis for esophageal cancer detection

We herein report a novel, reliable and inexpensive method for detecting esophageal cancer using blood plasma resonance Raman spectroscopy combined with multivariate analysis methods. The blood plasma samples were divided into late stage cancer group (n = 164), early stage cancer group (n = 35) and normal group (n = 135) based on clinical pathological diagnosis. Using a specially designed quartz capillary tube as sample holder, we obtained higher quality resonance Raman spectra of blood plasma than existing method. The study demonstrated that the carotenoids levels in blood plasma were reduced in esophageal cancer patients. The area under the receiver operating characteristic curve (and 95% confidence interval) calculated by wavenumber selection and principal component analysis combined with linear discriminant analysis (PC-LDA) algorithm were 0.894 (0.858-0.929), 0.901 (0.841-0.960) and 0.871 (0.799-0.942) for differentiating late cancer from normal, late cancer from early cancer, and early cancer from normal respectively. The contribution from the two carotenoids wavenumber regions of 1155 and 1515 cm⁻¹ were more than 84.2%. The results show that the plasma carotenoids could be a potential biomarker for screening esophageal cancer using resonance Raman spectroscopy combined with wavenumber selection and PC-LDA algorithms.      

Optoacoustic monitoring of water content in tissue phantoms and human skin

Skin water content monitoring is important for diagnostics and management of edema, dehydration, and other skin conditions as well as for cosmetic applications. Because optoacoustic (OA) technique has high (optical) contrast and (ultrasound) resolution and significant probing depth, it may be suitable for accurate, noninvasive water content monitoring in the skin. In this work we studied OA response from skin tissue phantoms and human wrist skin in the wavelength range from 1370 nm to 1650 nm using a novel, tunable OPO OA system. We identified optimal wavelengths for OA water content monitoring in different skin layers. The results of our study suggest that the OA technique may become a valuable, quantitative tool for accurate, high-resolution water content monitoring in the skin and other tissues and may find wide applications in dermatology, cosmetology, and tissue trauma management.