Sachin V. Patwardhan

Time-dependent whole-body fluorescence tomography of probe bio-distributions in mice

We present a fast scanning fluorescence optical tomography system for imaging the kinetics of probe distributions through out the whole body of small animals. Configured in a plane parallel geometry, the system scans a source laser using a galvanometer mirror pair (tauswitch~1ms) over flexible source patterns, and detects excitation and emission light using a high frame rate low noise, 5 MHz electron multiplied charge-coupled device (EMCCD) camera. Phantom studies were used to evaluate resolution, linearity, and sensitivity. Time dependent (deltat=2.2 min.) in vivo imaging of mice was performed following injections of a fluorescing probe (indocyanine green). The capability to detect differences in probe delivery route was demonstrated by comparing an intravenous injection, versus an injection into a fat pocket (retro orbital injection). Feasibility of imaging the distribution of tumor-targeted molecular probes was demonstrated by imaging a breast tumor-specific near infrared polypeptide in MDA MB 361 tumor bearing nude mice. A tomography scan, at 24 hour post injection, revealed preferential uptake in the tumor relative to surrounding tissue.

In vivo fluorescence lifetime tomography

Local molecular and physiological processes can be imaged in vivo through perturbations in the fluorescence lifetime (FLT) of optical imaging agents. In addition to providing functional information, FLT methods can quantify specific molecular events and multiplex diagnostic and prognostic information. We have developed a fluorescence lifetime diffuse optical tomography (DOT) system for in vivo preclinical imaging. Data is captured using a time-resolved intensified charge coupled device (ICCD) system to measure fluorescence excitation and emission in the time domain. Data is then converted to the frequency domain, and we simultaneously reconstruct images of yield and lifetime using an extension to the normalized Born approach. By using differential phase measurements, we demonstrate DOT imaging of short lifetimes (from 350 ps) with high precision (+/-5 ps). Furthermore, this system retains the efficiency, speed, and flexibility of transmission geometry DOT. We demonstrate feasibility of FLT-DOT through a progressive series of experiments. Lifetime range and repeatability are first measured in phantoms. Imaging of subcutaneous implants then verifies the FLT-DOT approach in vivo in the presence of inhomogeneous optical properties. Use in a common research scenario is ultimately demonstrated by imaging accumulation of a targeted near-infrared (NIR) fluorescent-labeled peptide probe (cypate-RGD) in a mouse with a subcutaneous tumor.

Classification of melanoma using tree structured wavelet transforms.

This paper presents a wavelet transform based tree structure model developed and evaluated for the classification of skin lesion images into melanoma and dysplastic nevus. The tree structure model utilizes a semantic representation of the spatial-frequency information contained in the skin lesion images including textural information. Results show that the presented method is effective in discriminating melanoma from dysplastic nevus. The results are also compared with those obtained using another method of developing tree structures utilizing the maximum channel energy criteria with a fixed energy ratio threshold.

Monte Carlo simulation of light-tissue interaction: three-dimensional simulation for trans-illumination-based imaging of skin lesions

Three-dimensional, voxel-based, and wavelength-dependent skin lesion models are developed and simulated using Monte Carlo techniques. The optical geometry of the Nevoscope with trans-illumination is used in the simulations for characterizing the lesion thickness. Based on the correlation analysis between the lesion thickness and the diffuse reflectance, optical wavelengths are selected for multispectral imaging of skin lesions using the Nevoscope. Tissue optical properties reported by various researchers are compiled together to form a voxel library. Tissue models used in the simulations are developed using the voxel library which offers flexibility in updating the optical properties and adding new media types into the models independent of the Monte Carlo simulation code.

Quantitative diffuse optical tomography for small animals using an ultrafast gated image intensifier

The quantitative accuracy of fluorescence and bioluminescence imaging of small animals can be improved by knowledge of the in situ optical properties of each animal. Obtaining in situ optical property maps is challenging, however, due to short propagation distances, requirements for high dynamic range, and the need for dense spatial, temporal, and spectral sampling. Using an ultrafast gated image intensifier and a pulsed laser source, we have developed a small animal diffuse optical tomography system with multiple synthetic modulation frequencies up to >1 GHz. We show that amplitude and phase measurements with useful contrast-to-noise ratios can be obtained for modulation frequencies over the range of approximately 250 to 1250 MHz. Experiments with tissue simulating phantoms demonstrate the feasibility of reconstructing the absorption and scattering optical properties in a small animal imaging system.

Multispectral optical imaging of skin-lesions for detection of malignant melanomas

Optical imaging of skin-lesions for early detection and management of the most fatal skin-cancer malignant melanoma is of significant interest in mass screening of skin-lesions with high-risk population. Surface illumination based optical imaging methods such as epiluminescence light microscopy (ELM) through “Dermascopy” has shown a significant potential in improving early diagnosis of malignant melanomas. Limitations of surface reflectance based imaging systems have been realized in analyzing images for important vascular and depth dependent information. We have developed a novel optical imaging system, the Nevoscope, that uses multispectral transillumination as to provide images of skin-lesions showing sub-surface pigmentation as well as vascular architecture based blood volume information. This paper presents multispectral Nevoscope transillumination method to compare and analyze ratiometric measurements to epiluminescence imaging for its ability to discriminate malignant melanomas from dysplastic nevi and other normal skin-lesions.

Classification of melanoma using wavelet-transform-based optimal feature set

The features used in the ABCD rule for characterization of skin lesions suggest that the spatial and frequency information in the nevi changes at various stages of melanoma development. To analyze these changes wavelet transform based features have been reported. The classification of melanoma using these features has produced varying results. In this work, all the reported wavelet transform based features are combined to form a single feature set. This feature set is then optimized by removing redundancies using principal component analysis. A feed forward neural network trained with the back propagation algorithm is then used in the classification process to obtain better classification results.

Quantitative small animal fluorescence tomography using an ultra- fast gated image intensifier

Optical approaches to small animal in vivo molecular imaging provide high sensitivity, stable non-radioactive probes, and an extensive array of functional reporting strategies. However, quantitative whole body assays remain illusive. The quantitative accuracy of optical imaging is affected by the depth of the buried target and the heterogeneity of tissue optical properties. Tomography approaches, to obtaining in-vivo optical property maps, and whole body distributions of fluorescing probes, provide a strategy for improving the quality and quantitative accuracy of small animal optical imaging. Here we present a time-resolved, charged coupled device (CCD) based system for quantitative small animal fluorescence tomography

Wavelength selection for multi-spectral imaging of skin lesions using Nevoscope

Wavelength dependent, voxel-based skin lesion models with different depths are simulated using the Monte Carlo technique with the Nevoscope optical geometry. A correlation analysis between the lesion depth and the diffuse reflectance is used for selecting optimal wavelengths for skin lesion imaging.

Fluorescence Lifetime Tomography for Whole Body Small Animal Imaging

Fluorescent lifetime provides a rich approach to quantitative molecular in vivo imaging. Here we report in vivo FLT-tomography using full spatial sampling for 3-D localization, and time-resolved (up to >1 GHz) analysis of lifetime contrasts.