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The aim of this study was to evaluate the application of attenuation coefficients calculated from the cross-polarization optical coherence tomography (CP OCT) data for differentiation of breast cancer from non-tumorous breast tissues. For this purpose, surgically obtained breast specimens from 45 patients were examined using CP OCT in order to construct color-coded en-face OCT maps based on calculation of three optical coefficients (the commonly used rate of attenuation in the co- polarization channel; and, additionally, the attenuation in the cross- polarization channel; and the interchannel attenuation difference). It was shown that the use of these optical coefficients significantly increased the amount of obtained information from the OCT data in comparison with unprocessed images, enabling objective quantification for differentiating non-tumorous and tumorous tissue (adipose tissue, normal stroma, tumor stroma and agglomerates of tumor cells). The attenuation in the cross- polarization channel and the interchannel attenuation difference provided greater contrast for the visualization of the different breast cancer structures compared to the attenuation coefficient in the co- polarization channel. The findings of this study suggest that assessment of CP OCT images of breast cancer based on optical coefficients calculation may in perspective enable real-time feedback about accurate resection margin in patient with breast cancer to the surgeon.

An important factor in assessing tumor status for determining therapeutic and surgical tactics is the study of its morphological structure. In particular, preoperative differentiation/grading of colorectal adenocarcinomas is very important for the adjustment of neoadjuvant therapy, surgical technique choice and survival prediction. To achieve the maximum benefit and to avoid unnecessary side effects, the ability to distinguish high-grade from low-grade colorectal adenocarcinoma, as well as identification of patients with high recurrence risk before the possible initiation of treatment will increase the quality and duration of patients' life. Taking into account the peculiarities of the biology of intestinal cancer, optical coherence elastography, as a method of studying tissue structure at a shallow depth, may be promising in the study of colorectal adenocarcinoma. Possibilities of optical coherence elastography in differentiating low-grade and high-grade types of colorectal adenocarcinoma are demonstrated in this study. The reported results are based on revealed qualitative criteria for assessment of OCE images in combination with quantitative statistically significant differences in characteristic stiffness values for low-grade and high-grade colorectal adenocarcinomas.

The possibility to assess molecular-biological and morphological features of particular breast cancer types can improve the precision of resection margin detection and enable accurate determining of the tumor aggressiveness, which is important for treatment selection. To enable reliable differentiation of breast-cancer subtypes and evaluation of resection margin, without performing conventional histological procedures, here we apply cross-polarization optical coherence tomography (CP-OCT) and compare it with a novel variant of compressional optical coherence elastography (C-OCE) in terms of the diagnostic accuracy (Ac) with histological verification. The study used 70 excised breast cancer specimens with different morphological structure and molecular status (Luminal A, Luminal B, Her2/Neo+, non-luminal and triple-negative cancer). Our first aim was to formulate convenient criteria of visual assessment of CP-OCT and C-OCE images intended (i) to differentiate tumorous and non-tumorous tissues and (ii) to enable more precise differentiation among different malignant states. We identified such criteria based on the presence of heterogeneities and characteristics of signal attenuation in CP-OCT images, as well as the presence of inclusions/mosaic structures combined with visually feasible assessment of several stiffness grades in C-OCE images. Secondly, we performed a blinded reader study of the Ac of C-OCE versus CP-OCT, for delineation of tumorous versus non-tumorous tissues followed by identification of breast cancer subtypes. For tumor detection, C-OCE showed higher specificity than CP-OCT (97.5% versus 93.3%) and higher Ac (96.0 versus 92.4%). For the first time, the Ac of C-OCE and CP-OCT were evaluated for differentiation between non-invasive and invasive breast cancer (90.4% and 82.5%, respectively). Furthermore, for invasive cancers, the difference between invasive but low-aggressive and highly-aggressive subtypes can be detected. For differentiation between non-tumorous tissue and low-aggressive breast-cancer subtypes, Ac was 95.7% for C-OCE and 88.1% for CP-OCT. For differentiation between non-tumorous tissue and highly-aggressive breast cancers, Ac was found to be 98.3% for C-OCE and 97.2% for CP-OCT. In all cases C-OCE showed better diagnostic parameters independently of the tumor type. These findings confirm the high potential of OCT-based examinations for rapid and accurate diagnostics during breast conservation surgery.

As was shown earlier in the animal experiments, compression optical coherence elastography (OCE) is a highly sensitive method for studying the morphological features of biological tissues. An attempt was made to study the possibilities of the OCE method for identifying and differentiating from each other various morphological structures in human tissues. Postoperative samples of human breast cancer and human colon cancer were studied. Histological examination identified areas of morphological structures - (I) adipose tissue, (II) stroma and (III) tumor tissue in breast cancer samples; and areas of (I) mucous membrane and (II) tumor tissue in colon cancer samples. For these areas, an accurate comparison was made with OCE images and stiffness ranges for morphological structures were obtained. This observation allows us to make a conclusion that OСE can be a promising method for studying the structure of human tissue in clinics.

Breast cancer is a genetically heterogeneous disease characterized by various biomolecular and morphological features that affect the diagnosis, prognosis, and treatment response. In this study we combined cross-polarization optical coherence tomography (CP OCT) and multiphoton tomography (MPT), based on second harmonic generation (SHG), and two-photon-excited fluorescence (TPEF) to visualize tumor stroma and tumor cells in specimens of a human breast tissue. The data obtained by both techniques were compared with histopathology. The CP OCT and MPT images were quantitatively assessed to distinguish a breast normal tissue from a cancer as well as between a low and a high grade of cancer. Quantitative assessment of the CP OCT image included the calculation of signal attenuation coefficients separately for co- and cross- polarization channels and the formation the color-coded en-face distribution maps of these coefficients. The attenuation coefficient in cross- polarization channel showed better difference between breast cancer of low and high grades and distinguish them from normal tissue. The SHG images were processed using texture analysis techniques to quantify the density of collagen fibers in normal tissue and tumor. Thus, both imaging techniques have great potential to distinguish nontumorous and tumorous human breast tissue of varying degrees of malignancy and could provide identifying breast cancer margins for in surgery.

Compressional optical coherence elastography (OCE) is the emerging tool to evaluate Young modulus of the tissue with resolution close to optical coherence tomography. Recent versions of compressional OCE based on the strain comparison within the tissue and the calibrated reference silicone layer (sensor) located between the OCT probe and tissue. For standard realizations of compressional OCE the evaluation of the strain within the tissue and the sensor performed only at one point of the stress-strain curve. In the same way the applied stress can be easily estimated in a wide range on the base on the stress-strain relation for calibrated sensor strain that allow to evaluate the stress-strain relation for the underlying tissue not in one point but in the wider range. In this report we demonstrate this approach to nonlinear parameter evaluation for wide range of the tissues. We demonstrate the application of this new tool to breast tissue, vaginal wall, bladder, brain, cartilages, coronary arteries walls in various states. The preliminary results shows that several states of the tissues such as vaginal wall state in case of pelvic organ prolapse, benign breast tumor and coronary arteries plague can be determined on the base of elastic nonlinear parameters.

A multimodal optical coherence tomography (MM OCT) combining microstructural cross-polarization (CP) imaging and compression OCT-elastography (OCE) was employed in this study. The research was carried out on the noninvasive and invasive subtypes of human breast cancer. This study demonstrates how CP OCT combined with OCTelastography identifies an example of DCIS cancer that progresses into invasive breast cancer. OCE images more clearly show the cross sections of the ducts filled with tumor cells for DCIS (high-contrast structures with clear boundaries and high stiffness values) surrounded by areas with lower stiffness corresponding to fibrous stroma regions. The regions of invasive breast cancer in OCE-scans look as zones with strongly increased stiffness, which well agrees with the histology. Percentage of pixels with different characteristic stiffness ("stiffness spectrum") in the OCTelastography images was assessed to quantitatively visualize stiffness of tumor. The demonstrated ability of MM OCT imaging for breast cancer subtypes differentiation and breast cancer margin assessment is important for improved managements of patients.

In this study we combined cross-polarization optical coherence tomography (CP OCT), multiphoton tomography (MPT), based on second harmonic generation, and two-photon-excited fluorescence to visualize collagen fibers and tumor cells in the various morphological subtypes of breast cancer. The ability of CP OCT to visualize tissue birefringence and cross-scattering adds new information about the microstructure of such breast cancers, while the MPT provides verification of this microstructure. Mammary glands, both normal and tumorous, were assessed by MPT and CP OCT to establish the relationships between spatial organization features of the cellular component and the intercellular matrix. It was shown, that such multimodal optical imaging has great potential for distinguishing various breast cancer morphological subtypes and could provide useful tools for identifying positive breast cancer margins for surgery.

This paper considers valuable visual assessment criteria for distinguishing between tumorous and non-tumorous tissues, intraoperatively, using cross-polarization OCT (CP OCT) – OCT with a functional extension, that enables detection of the polarization properties of the tissues in addition to their conventional light scattering. Materials and methods. The study was performed on 176 ex vivo human specimens obtained from 30 glioma patients. To measure the degree to which the typical parameters of CP OCT images can be matched to the actual histology, 100 images of tumors and white matter were selected for visual analysis to be undertaken by three “single-blinded” investigators. An evaluation of the inter-rater reliability between the investigators was performed. Application of the identified visual CP OCT criteria for intraoperative use was performed during brain tumor resection in 17 patients. Results. The CP OCT image parameters that can typically be used for visual assessment were separated: (1) signal intensity; (2) homogeneity of intensity; (3) attenuation rate; (4) uniformity of attenuation. The degree of match between the CP OCT images and the histology of the specimens was significant for the parameters “signal intensity” in both polarizations, and “homogeneity of intensity” as well as the “uniformity of attenuation” in co-polarization. A test based on the identified criteria showed a diagnostic accuracy of 87-88%. Intraoperative in vivo CP OCT images of white matter and tumors have similar signals to ex vivo ones, whereas the cortex in vivo is characterized by indicative vertical striations arising from the “shadows” of the blood vessels; these are not seen in ex vivo images or in the case of tumor invasion. Conclusion. Visual assessment of CP OCT images enables tumorous and non-tumorous tissues to be distinguished. The most powerful aspect of CP OCT images that can be used as a criterion for differentiation between tumorous tissue and white matter is the signal intensity. In distinguishing white matter from tumors the diagnostic accuracy using the identified visual CP OCT criteria was 87-88%. As the CP OCT data is easily associated with intraoperative neurophysiological and neuronavigation findings this can provide valuable supplementary information for the neurosurgeon during tumor resection.

In this report we discuss some practical obstacles/pitfalls arising in realization of quantitative compressional OCE, as well as discuss possible ways of their resolution. More specifically we consider (i) complications in quantification of the Young modulus of tissues related to influence of partial adhesion between the OCT probe and pre-calibrated reference layers - "compliant sensors", (ii) distorting influence of surface curvature/corrugation on strain distribution in the tissue bulk, (iii) ways of enhancement of effective SNR in OCT-based strain mapping without periodic averaging, and (iv) potentially significant influence of nonlinearity of the elastic response of biological tissues on quantification of their stiffness.

Optical coherence tomography (OCT) is a promising method of glial tumors borders diagnostics. Nowadays it is possible to use hand-held and microscope mounted OCT devices during tumor removal. But still there are no clearly visual assessment criteria of OCT images, on the basis of which good differentiation between glioma tissue and white matter can be performed. This paper presents such criteria for crosspolarization OCT (CP OCT), which can detect both the scattering and polarization properties of tissues.

Soft biological tissues manifest strongly pronounced nonlinear elastic behavior. Namely, the Young modulus for some tissues strongly depends on the applied stress. This property can be evaluated by compressional OCT elastography. We demonstrate the evaluation of nonlinear elastic properties on the samples of coronary arteries, breast cancer and cornea. The developed technique can be used for further investigation of the nonlinear properties of healthy and pathological tissues.

Early response of cancer cells to chemotherapeutic drugs were studied using novel fluorescence tools and microscopy techniques. We applied two-photon fluorescence lifetime imaging (microscopy and in vivo imaging) to assess metabolic and oxygen changes in cancer cells in different tumor models. The dynamics of energy metabolism, intracellular pH, oxygen under chemotherapy were evaluated. In patients with skin neoplasia using of optical imaging, including FLIM, we have showed possibilities to distinguish benign and malignant pigmented lesions.

OCT-based local strain relaxation/creep evaluation is an emerging tool for tissue viscoelasticity characterization. We present a tool for 2D visualization of local strain relaxation and creep time/rate inside the tissue.

Multimodal OCT is a promising tool for monitoring of individual tumor response to antitumor therapies. The changes of tumor cells, connective tissue, microcirculation and stiffness can be estimated simultaneously in real time with high resolution.

We discuss the application of compressional OCT-based elastography for measuring elastic response of biological tissues in the strain range from fractions of one per cent up to strains over ~10% and greater. Such fairly large strains are not typical of phase-sensitive OCT-based elastographic methods and can be measurable due to application of interframe strain-estimation method in which interframe phase gradients are estimated without the necessity of preliminary measuring total displacements of scattering particles in combination with summation of interframe strains. For estimating applied stresses, intervenient soft layer of translucent silicone is used a a reference fairly linear material. The obtained results based on comparison of strains in the reference layer and tissue demonstrate highly nonlinear character of elasticity of biological tissues even for rather moderate strains on order of several per cent.

The main goal of the study was a vasculature targeted PDT. A new approach named M-mode-like OCT (MML OCT) was applied to monitor early response to PDT. Due to the chosen filtering parameters (96 Hz filter threshold), this approach visualizes only vessels with flowing blood. Without flowing blood even filled vessels are not visualized because flow-less blood causes speckle variations with significantly lower frequencies (

In the case of infiltrative brain tumors the surgeon faces difficulties in determining their boundaries to achieve total resection. The aim of the investigation was to evaluate the performance of multimodal OCT (MM OCT) for differential diagnostics of normal brain tissue and glioma using an experimental model of glioblastoma. The spectral domain OCT device that was used for the study provides simultaneously two modes: cross-polarization and microangiographic OCT. The comparative analysis of the both OCT modalities images from tumorous and normal brain tissue areas concurrently with histologic correlation shows certain difference between when accordingly to morphological and microvascular tissue features.

The combined use of fluorescence cystoscopy and cross-polarization optical coherence tomography (CP OCT) with quantitative estimation of the OCT signal was assessed in 92 bladder zones. It demonstrated the diagnostic accuracy in detecting superficial bladder cancer of 93.6%, sensitivity 96.4%, specificity 92.1%, positive predictive value 87% and negative predictive value 97.9%. Quantitative estimation of OCT signal standard deviation in cross-polarization (CP OCT SD index) makes the visual criteria of CP OCT image assessment more objective. The level of CP OCT SD index for diagnosing superficial bladder cancer, including cancer in situ, was 4.32 dB and lower. When tumor is located on a postoperative scar, CP OCT SD index may be higher than the threshold level of 4.32 dB due to strong scattering and depolarization in scar fibrous tissue. A high inverse correlation was found between CP OCT SD index and the level expressed by p63, Ki-67, p53, CD44v6 markers.

A combination of approaches to the image analysis in cross-polarization optical coherence tomography (CP OCT) and high-resolution imaging by nonlinear microscopy and atomic force microscopy (AFM) at the different stages of atherosclerotic plaque development is studied. This combination allowed us to qualitatively and quantitatively assess the disorganization of collagen in the atherosclerotic arterial tissue (reduction and increase of CP backscatter), at the fiber (change of the geometric distribution of fibers in the second-harmonic generation microscopy images) and fibrillar (violation of packing and different nature of a basket-weave network of fibrils in the AFM images) organization levels. The calculated CP channel-related parameters are shown to have a statistically significant difference between stable and unstable (also called vulnerable) plaques, and hence, CP OCT could be a potentially powerful, minimally invasive method for vulnerable plaques detection.

In this study we demonstrate the capability of cross-polarization optical coherence tomography (CP OCT) to assess collagen and elastin fibers condition in atherosclerotic plaques basing on ratio of the OCT signal levels in cross- and со- polarizations. We consider the depolarization factor (DF) and the effective birefringence (Δn) as quantitative characteristics of CP OCT images. We revealed that calculation of both DF and Δn in the region of interest (fibrous cap) yields a statistically significant difference between stable and unstable plaques (0.46±0.21 vs 0.09±0.04 for IDF; (4.7±1.0)•10–4 vs (2.5±0.7)•10–4 for Δn; p

Multimodal optical coherence tomography (OCT) is an emerging tool for tissue state characterization. Optical coherence elastography (OCE) is an approach to mapping mechanical properties of tissue based on OCT. One of challenging problems in OCE is elimination of the influence of residual local tissue relaxation that complicates obtaining information on elastic properties of the tissue. Alternatively, parameters of local relaxation itself can be used as an additional informative characteristic for distinguishing the tissue in normal and pathological states over the OCT image area. Here we briefly present an OCT-based approach to evaluation of local relaxation processes in the tissue bulk after sudden unloading of its initial pre-compression. For extracting the local relaxation rate we evaluate temporal dependence of local strains that are mapped using our recently developed hybrid phase resolved/displacement-tracking (HPRDT) approach. This approach allows one to subtract the contribution of global displacements of scatterers in OCT scans and separate the temporal evolution of local strains. Using a sample excised from of a coronary arteria, we demonstrate that the observed relaxation of local strains can be reasonably fitted by an exponential law, which opens the possibility to characterize the tissue by a single relaxation time. The estimated local relaxation times are assumed to be related to local biologically-relevant processes inside the tissue, such as diffusion, leaking/draining of the fluids, local folding/unfolding of the fibers, etc. In general, studies of evolution of such features can provide new metrics for biologically-relevant changes in tissue, e.g., in the problems of treatment monitoring.

Recently proposed in vivo label-free optical coherence angiography techniques based on phase and amplitude speckle variability often require additional signal pre- and post processing operations to enhance vessel-contrast. We observe here 1) contrast enhancement by optimizing the signal normalization/weighing before processing; 2) algorithm based on Kasai estimator for phase compensation between processed A-scans to reduce masking role of motion artifacts; and 3) image projection through the imaging depth for en face plotting. We demonstrate the efficiency of proposed additional algorithms as for the microcirculation imaging of hamsters cheek in vivo as for the preliminary microcirculation imaging of patients after radiotherapy. This technical framework complete in details our recent publications on M-Mode like OCT algorithms and its implementation.

We combined cross-polarization optical coherence tomography (CP OCT) and non-linear microscopy based on second harmonic generation (SHG) and two-photon-excited fluorescence (2PEF) to assess collagen and elastin fibers in the development of the atherosclerotic plaque (AP). The study shows potential of CP OCT for the assessment of collagen and elastin fibers condition in atherosclerotic arteries. Specifically, the additional information afforded by CP OCT, related to birefringence and cross-scattering properties of arterial tissues, may improve the robustness and accuracy of assessment about the microstructure and composition of the plaque for different stages of atherosclerosis.

We combined cross-polarization optical coherence tomography (CP OCT) and non-linear microscopy based on second harmonic generation (SHG) and two-photon-excited fluorescence (2PEF) to assess collagen and elastin fibers and other vascular structures in the development of atherosclerosis, including identification of vulnerable plaques, which remains an important clinical problem and imaging application. CP OCT's ability to visualize tissue birefringence and cross-scattering adds new information about the microstructure and composition of the plaque. However its interpretation can be ambiguous, because backscattering contrast may have a similar appearance to the birefringence related fringes. Our results represent a step towards minimally invasive characterization and monitoring of different stages of atherosclerosis, including vulnerable plaques. CP OCT image of intimal thickening in the human coronary artery. The dark stripe in the cross-polarization channel (arrow) is a polarization fringe related to the phase retardation between two eigen polarization states. It is histologically located in the area of the lipid pool, however this stripe is a polarization artifact, rather than direct visualization of the lipid pool.

In this paper, we point out some practical obstacles arising in realization of compressional optical coherence elastography (OCE) that have not attracted sufficient attention previously. Specifically, we discuss (i) complications in quantification of the Young modulus of tissues related to partial adhesion between the OCE probe and soft intervening reference layer sensor, (ii) distorting influence of tissue surface curvature/corrugation on the subsurface strain distribution mapping, (iii) ways of signal-to-noise ratio (SNR) enhancement in OCE strain mapping when periodic averaging is not realized, and (iv) potentially significant influence of tissue elastic nonlinearity on quantification of its stiffness. Potential practical approaches to mitigate the effects of these complications are also described.

The goal of the research was analysis of the effect of collagen condition in formation of cross-polarized CP OCT images. We used of the CP OCT technique for studying collagen condition on an example of oral mucosa. Special histologic picrosirius red (PSR) staining of cheek mucosa specimens was used with subsequent assessing of the result of collagen staining in polarized light. High correlation (r = 0.692, p = 0.0001) between OCT signal standard deviation (SD) in cross-polarized images and brightness of PSR stained collagen fibers in cheek mucosa specimens was demonstrated in patients with inflammatory intestine and oral mucosa diseases. We have found that the OCT signal SD in cross-polarized images reflects two boundary conditions of collagen disorganization, namely, loss of fiber properties at active inflammation which attenuates the signal and fibrosis that occurs due to synthesis of a new remodeled collagen which amplifies the OCT signal.

Quantitative image analysis and parameter extraction using a specific implementation of polarization-sensitive optical coherence tomography (OCT) provides differential diagnosis of mucosal pathologies in in-vivo human bladders. We introduce a cross-polarization (CP) OCT image metric called Integral Depolarization Factor (IDF) to enable automatic diagnosis of bladder conditions (assessment the functional state of collagen fibers). IDF-based diagnostic accuracy of identification of the severe fibrosis of normal bladder mucosa is 79%; recurrence of carcinoma on the post-operative scar is 97%; and differentiation between neoplasia and acute inflammation is 75%. The promising potential of CP OCT combined with image analysis in human urology is thus demonstrated in vivo.