Dominique Van De Sompel

Improved imaging approaches are needed for ovarian cancer screening, diagnosis, staging, and resection guidance. Here, we propose a combined photoacoustic (PA)/Raman approach using gold nanorods (GNRs) as a passively targeted molecular imaging agent. GNRs with three different aspect ratios were studied. Those with an aspect ratio of 3.5 were selected for their highest ex vivo and in vivo PA signal and used to image subcutaneous xenografts of the 2008, HEY, and SKOV3 ovarian cancer cell lines in living mice. Maximum PA signal was observed within 3 h for all three lines tested and increased signal persisted for at least two days postadministration. There was a linear relationship (R(2) = 0.95) between the PA signal and the concentration of injected molecular imaging agent with a calculated limit of detection of 0.40 nM GNRs in the 2008 cell line. The same molecular imaging agent could be used for clear visualization of the margin between tumor and normal tissue and tumor debulking via surface-enhanced Raman spectroscopy (SERS) imaging. Finally, we validated the imaging findings with biodistribution data and elemental analysis. To the best of our knowledge, this is the first report of in vivo imaging of ovarian cancer tumors with a photoacoustic and Raman imaging agent.

Endoscopic imaging is an invaluable diagnostic tool allowing minimally invasive access to tissues deep within the body. It has played a key role in screening colon cancer and is credited with preventing deaths through the detection and removal of precancerous polyps. However, conventional white-light endoscopy offers physicians structural information without the biochemical information that would be advantageous for early detection and is essential for molecular typing. To address this unmet need, we have developed a unique accessory, noncontact, fiber optic-based Raman spectroscopy device that has the potential to provide real-time, multiplexed functional information during routine endoscopy. This device is ideally suited for detection of functionalized surface-enhanced Raman scattering (SERS) nanoparticles as molecular imaging contrast agents. This device was designed for insertion through a clinical endoscope and has the potential to detect and quantify the presence of a multiplexed panel of tumor-targeting SERS nanoparticles. Characterization of the Raman instrument was performed with SERS particles on excised human tissue samples, and it has shown unsurpassed sensitivity and multiplexing capabilities, detecting 326-fM concentrations of SERS nanoparticles and unmixing 10 variations of colocalized SERS nanoparticles. Another unique feature of our noncontact Raman endoscope is that it has been designed for efficient use over a wide range of working distances from 1 to 10 mm. This is necessary to accommodate for imperfect centering during endoscopy and the nonuniform surface topology of human tissue. Using this endoscope as a key part of a multiplexed detection approach could allow endoscopists to distinguish between normal and precancerous tissues rapidly and to identify flat lesions that are otherwise missed.

Raman spectroscopy, amplified by surface enhanced Raman scattering (SERS) nanoparticles, is a molecular imaging modality with ultra-high sensitivity and the unique ability to multiplex readouts from different molecular targets using a single wavelength of excitation. This approach holds exciting prospects for a range of applications in medicine, including identification and characterization of malignancy during endoscopy and intraoperative image guidance of surgical resection. The development of Raman molecular imaging with SERS nanoparticles is presently limited by long acquisition times, poor spatial resolution, small field of view, and difficulty in animal handling with existing Raman spectroscopy instruments. Our goal is to overcome these limitations by designing a bespoke instrument for Raman molecular imaging in small animals. Here, we present a unique and dedicated small-animal Raman imaging instrument that enables rapid, high-spatial resolution, spectroscopic imaging over a wide field of view (> 6 cm(2)), with simplified animal handling. Imaging of SERS nanoparticles in small animals demonstrated that this small animal Raman imaging system can detect multiplexed SERS signals in both superficial and deep tissue locations at least an order of magnitude faster than existing systems without compromising sensitivity.

Photoacoustic imaging combines the high contrast of optical imaging with the spatial resolution and penetration depth of ultrasound. This technique holds tremendous potential for imaging in small animals and importantly, is clinically translatable. At present, there is no accepted standard physical phantom that can be used to provide routine quality control and performance evaluation of photoacoustic imaging instruments. With the growing popularity of the technique and the advent of several commercial small animal imaging systems, it is important to develop a strategy for assessment of such instruments. Here, we developed a protocol for fabrication of physical phantoms for photoacoustic imaging from polyvinyl chloride plastisol (PVCP). Using this material, we designed and constructed a range of phantoms by tuning the optical properties of the background matrix and embedding spherical absorbing targets of the same material at different depths. We created specific designs to enable: routine quality control; the testing of robustness of photoacoustic signals as a function of background; and the evaluation of the maximum imaging depth available. Furthermore, we demonstrated that we could, for the first time, evaluate two small animal photoacoustic imaging systems with distinctly different light delivery, ultrasound imaging geometries and center frequencies, using stable physical phantoms and directly compare the results from both systems.

Topical application and quantification of targeted, surface-enhanced Raman scattering (SERS) nanoparticles offer a new technique that has the potential for early detection of epithelial cancers of hollow organs. Although less toxic than intravenous delivery, the additional washing required to remove unbound nanoparticles cannot necessarily eliminate nonspecific pooling. Therefore, we developed a real-time, ratiometric imaging technique to determine the relative concentrations of at least two spectrally unique nanoparticle types, where one serves as a nontargeted control. This approach improves the specific detection of bound, targeted nanoparticles by adjusting for working distance and for any nonspecific accumulation following washing. We engineered hardware and software to acquire SERS signals and ratios in real time and display them via a graphical user interface. We report quantitative, ratiometric imaging with nanoparticles at pM and sub-pM concentrations and at varying working distances, up to 50 mm. Additionally, we discuss optimization of a Raman endoscope by evaluating the effects of lens material and fiber coating on background noise, and theoretically modeling and simulating collection efficiency at various working distances. This work will enable the development of a clinically translatable, noncontact Raman endoscope capable of rapidly scanning large, topographically complex tissue surfaces for small and otherwise hard to detect lesions.