A real-time spectral mapper as an emerging diagnostic technology in biomedical sciences.

Real time spectral imaging and mapping at video rates can have tremendous impact not only on diagnostic sciences but also on fundamental physiological problems. We report the first real-time spectral mapper based on the combination of snap-shot spectral imaging and spectral estimation algorithms. Performance evaluation revealed that six band imaging combined with the Wiener algorithm provided high estimation accuracy, with error levels lying within the experimental noise. High accuracy is accompanied with much faster, by 3 orders of magnitude, spectral mapping, as compared with scanning spectral systems. This new technology is intended to enable spectral mapping at nearly video rates in all kinds of dynamic bio-optical effects as well as in applications where the target-probe relative position is randomly and fast changing.

Dynamic contrast enhanced optical imaging of cervix, in vivo: a paradigm for mapping neoplasia-related parameters.

We present a novel biophotonic method and imaging modality for estimating and mapping neoplasia-specific functional and structural parameters of the cervical precancerous epithelium. Estimations were based on experimental data obtained from dynamic contrast-enhanced optical imaging of cervix, in vivo. We have developed a pharmacokinetic, in silico, model of the optical tracer's uptake by the epithelium. We have identified that the kinetic parameters of the model correlate well with pathologic alterations in both metabolic and structural characteristics of the tissue, associated with the neoplasia progress. Global sensitivity analysis and global optimization methods were employed for identifying the key determinant set of biological parameters that dictate the model's output. Particularly, the shuffled complex evolution algorithm converged to a set of four parameters that can be estimated with an error of 7%, indicating a good accuracy and precision. These results are unique in the sense that for the first time functional and microstructural parameter maps can be estimated and displayed together, thus maximizing the diagnostic information. The quantity and the quality of this information are unattainable by other invasive and non invasive methods.

A novel endoscopic spectral imaging platform integrating k-means clustering for early and non-invasive diagnosis of endometrial pathology.

We report a multimodal endoscopic system capable of performing both color and fast multispectral imaging in the spectral range 400-1000 nm. The system is based on a computer controllable tunable light source, which can be coupled with all types of endoscopes. Performance evaluation showed about 60% flat transmittance in almost all the operating wavelengths, at about 13 nm bandwidth per tuning step. With this system adapted to a thin hysteroscope, we also report, for the first time, spectral analysis of the endometrium and unsupervised/objective clustering of the spectra. We have implemented a method combining the k-means algorithm with the silhouette criterion for estimating the number of the distinguishable spectral classes that may correspond to different medical conditions of the tissue. It was found that there are five-well defined clusters of spectra, while preliminary clinical data seem to correlate well with the tissue pathology.

Estimation of neoplasia-related biological parameters through modeling and sensitivity analysis of optical molecular imaging data.

We combine system's biology approaches with in vivo optical molecular imaging of epithelial neoplasia for estimating disease-specific biological parameters. Molecular imaging measures and maps the dynamic optical effects, generated by the topical application of acetic acid diluted solution. The dynamic characteristics of the in vivo measured optical signal are governed by the epithelial transport effects of the biomarker. Nine biological parameters, both structural and functional, have been identified to be potentially correlated with the neoplasia growth and to be manifested to the measured data in a convoluted manner. A compartmental model of the cervical neoplastic epithelium has been developed, which predicts the dynamic optical effects in all possible parameter value combinations. We have performed global sensitivity analysis for the purpose of identifying the subset of the input parameters that are the key determinants of the model's output. Finally, we have for the first time shown that it is possible to estimate, from in vivo measured dynamic optical data, the following neoplasia related parameters: number of neoplastic layers, intracellular and extracellular space dimensions, functionality of tight junctions, and extracellular pH. These findings have been (in part) validated with optical data and biopsies obtained from 30 women with cervical neoplasia.

In vivo dynamic imaging, in silico modeling and global sensitivity analysis for the study and the diagnosis of epithelial neoplasia.

We present a method for detecting and studying neoplasia-specific functional and structural features through the combination of in vivo dynamic imaging, in silico modeling and global sensitivity analysis. We particularly present the case of cervical epithelium interacting with acetic acid solution, which is employed as an optical biomarker. The in vivo measured dynamic scattering characteristics are strongly correlated with the output of the biomarker's pharmacokinetic model that we have developed. Model global sensitivity analysis has shown that the measured/modeled bio-optical processes can be used for probing, in vivo, the number of neoplastic layers, the extracellular pH, the intracellular buffering efficiency and the size of the extracellular space.

Dynamic spectral imaging: improving colposcopy.

PURPOSE: Colposcopy occupies a key role in the prevention of cervical cancer by identifying preinvasive or invasive lesions. However, colposcopy is subjective and is responsible for 52% of screening failures. Dynamic spectral imaging (DSI) is based on the objective, quantitative assessment of the acetowhitening effect. This study compared DSI with colposcopy. EXPERIMENTAL DESIGN: Women referred for colposcopy were examined simultaneously with colposcopy and DSI using a precommercial DySIS model (FPC-03) in an international, multicenter trial. The colposcopy impression and DySIS values were compared with consensus histology reports of biopsies. Subjects were recruited to a training group and subsequently to a test group. Measures were taken to avoid verification bias. RESULTS: The training and test groups comprised 82 and 308 eligible women, respectively. A cutoff value to identify high-grade disease was selected from the results of the training group and data from previous work. Receiver operator curve analysis of the test data showed an area under the curve of 0.844. DySIS detected 62.9% more high-grade cases than colposcopy (57 versus 35, P=0.0001). DySIS exceeded end points approved by the Food and Drug Administration for similar studies, with increments in the true positive rate of 22/308 (7.1%; lower 95% CL, 4.5% versus 2%) and in the false positive rate of 32/308 (10.4%; upper 95% CL, 14.7% versus 15%). CONCLUSIONS: DySIS is more sensitive than colposcopy in detecting high-grade lesions and can provide improved guidance for biopsy. The results are obtained in a user-independent fashion, making it suitable for use by nursing personnel.

Spectral characteristics of acute lymphoblastic leukemia in childhood.

Although the differentiation and classification of acute leukemia are based upon cytochemical features as well as immunologic, cytogenetic, and molecular characteristics, in many cases the morphological distinction of normal lymphocytes from lymphoblasts of acute lymphoblastic leukemia (ALL) is difficult using light microscopy. In this study the distinction between normal lymphocytes and lymphoblasts of childhood ALL is proposed using their spectral characteristics. The method has been based upon the analysis and classification of optical absorption characteristics of the bone marrow cells. Spectral microscopy system is capable of capturing a great number of narrow-band images, in the wide spectral range of the optical spectrum. The analysis showed statistically significant difference (P < 0.0001) between normal lymphocytes and lymphoblasts as far as it concerns the detection, identification and mapping of their spectral absorption characteristics. Our results suggest the potential of spectral imaging as a new method for the distinction of lymphocytes from lymphoblasts in cases that with the light microscope, the morphologic differences are not visible in the bone marrow smears at diagnosis or the follow up of the children with ALL.

A novel spectral microscope system: application in quantitative pathology.

In this paper, a novel spectral microscope system is presented together with a method for the quantitative assessment of the uptake by histologic samples of stains used in pathology to label tissue features of diagnostic importance. The critical component of the microscope is a variable interference filter-based monochromator. The system is capable of performing real-time spectral imaging in a plurality of spectral bands and micro-spectroscopy in any image pixel, in the spectral range 400-1000 nm. The wavelength-tuning step is 2.4-2.6 nm, while the full-width at half maximum in each step is about 1.5% of the operating central wavelength. The developed system integrates algorithms and calibration procedures for the calculation of the stain-uptake by the tissue. The acquired spectra from both stained tissue and calibration stain solutions enable the calculation of the concentration maps of the stains, even if the latter are multiple and overlap spatially and spectrally. The system was used for the quantitative mapping of the expression of estrogen and progesterone receptors in breast cancer cells. In this particular case, model validation shows that although two stains are employed, capturing of their transmittance at more than ten wavelengths is required in order to obtain an acceptable accuracy. These findings highlight the need for the development and implementation of spectral microscopy in pathology and its potential to introduce novel more reliable diagnostic criteria.