Manojit Pramanik

Sentinel Lymph Nodes in the Rat: Noninvasive Photoacoustic and US Imaging with a Clinical US System

PURPOSEnTo evaluate in vivo sentinel lymph node (SLN) mapping by using photoacoustic and ultrasonographic (US) imaging with a modified clinical US imaging system.nnnMATERIALS AND METHODSnAnimal protocols were approved by the Animal Studies Committee. Methylene blue dye accumulation in axillary lymph nodes of seven healthy Sprague-Dawley rats was imaged by using a photoacoustic imaging system adapted from a clinical US imaging system. To investigate clinical translation, the imaging depth was extended up to 2.5 cm by adding chicken or turkey breast on top of the rat skin surface. Three-dimensional photoacoustic images were acquired by mechanically scanning the US transducer and light delivery fiber bundle along the elevational direction.nnnRESULTSnPhotoacoustic images of rat SLNs clearly help visualization of methylene blue accumulation, whereas coregistered photoacoustic/US images depict lymph node positions relative to surrounding anatomy. Twenty minutes following methylene blue injection, photoacoustic signals from SLN regions increased nearly 33-fold from baseline signals in preinjection images, and mean contrast between SLNs and background tissue was 76.0 +/- 23.7 (standard deviation). Methylene blue accumulation in SLNs was confirmed photoacoustically by using the optical absorption spectrum of the dye. Three-dimensional photoacoustic images demonstrate dynamic accumulation of methylene blue in SLNs after traveling through lymph vessels.nnnCONCLUSIONnIn vivo photoacoustic and US mapping of SLNs was successfully demonstrated with a modified clinical US scanner. These results raise confidence that photoacoustic and US imaging can be used clinically for accurate, noninvasive imaging of SLNs for axillary lymph node staging in breast cancer patients.

Design and evaluation of a novel breast cancer detection system combining both thermoacoustic (TA) and photoacoustic (PA) tomography

We have developed a novel scanner for breast cancer detection, integrating both thermoacoustic and photoacoustic techniques to achieve dual contrast (microwave and light absorption) imaging. This scanner is nonionizing, low cost, and can potentially provide high-resolution, dual modality three-dimensional images of the breast. The scanner uses front instead of side breast compression and dry instead of gel ultrasonic coupling. Here we present the design of the breast scanner along with initial tissue phantom study results as a precursor to an actual patient study.

Thermoacoustic and photoacoustic sensing of temperature

We present a novel temperature-sensing technique using thermoacoustic and photoacoustic measurements. This noninvasive method has been demonstrated using a tissue phantom to have high temporal resolution and temperature sensitivity. Because both photoacoustic and thermoacoustic signal amplitudes depend on the temperature of the source object, the signal amplitudes can be used to monitor the temperature. A temperature sensitivity of 0.15 degrees C was obtained at a temporal resolution as short as 2 s, taking the average of 20 signals. The deep-tissue imaging capability of this technique can potentially lead us to in vivo temperature monitoring in thermal or cryogenic applications.

Single-walled carbon nanotubes as a multimodal-thermoacoustic and photoacoustic-contrast agent

We have developed a novel carbon nanotube-based contrast agent for both thermoacoustic and photoacoustic tomography. In comparison to deionized water, single-walled carbon nanotubes exhibited more than twofold signal enhancement for thermoacoustic tomography at 3 GHz. In comparison to blood, they exhibited more than sixfold signal enhancement for photoacoustic tomography at 1064 nm wavelength. The large contrast enhancement of single-walled carbon nanotubes was further corroborated by tissue phantom imaging studies.

Molecular photoacoustic imaging of angiogenesis with integrin-targeted gold nanobeacons

Photoacoustic tomography (PAT) combines optical and acoustic imaging to generate highresolution images of microvasculature. Inherent sensitivity to hemoglobin permits PAT to image blood vessels but precludes discriminating neovascular from maturing microvasculature. αvβ3‐Gold nanobeacons (αvβ3‐GNBs) for neovascular molecular PAT were developed, characterized, and demonstrated in vivo using a mouse Matrigel‐plug model of angiogenesis. PAT results were microscopically corroborated with fluorescent αvβ3‐GNB localization and supporting immunohistology in RαgïtmlMom Tg(Tie‐2‐lacZ)182‐Sato mice. αvβ3‐GNBs (154 nm) had 10‐fold greater contrast than blood on an equivolume basis when imaged at 740 nm to 810 nm in blood. The lowest detectable concentration in buffer was 290 nM at 780 nm. Noninvasive PAT of angiogenesis using a 10‐MHz ultrasound receiver with αvβ3‐GNBs produced a 600% increase in signal in a Matrigel‐plug mouse model relative to the inherent hemoglobin contrast pretreatment. In addition to increasing the contrast of neovessels detected at baseline, αvβ3‐GNBs allowed visualization of numerous angiogenic sprouts and bridges that were undetectable before contrast injection. Competitive inhibition of αvβ3‐GNBs with αvβ3‐NBs (no gold particles) almost completely blocked contrast enhancement to pretreatment levels, similar to the signal from animals receiving saline only. Consistent with other studies, nontargeted GNBs passively accumulated in the tortuous neovascular but provided less than half of the contrast enhancement of the targeted agent. Microscopic studies revealed that the vascular constrained, rhodamine‐labeled αvβ3‐GNBs homed specifically to immature neovasculature (PECAM+, Tie‐2) along the immediate tumor periphery, but not to nearby mature microvasculature (PECAM+, Tie‐2+). The combination of PAT and αvβ3‐GNBs offered sensitive and specific discrimination and quantification of angiogenesis in vivo, which may be clinically applicable to a variety of highly prevalent diseases, including cancer and cardiovascular disease.—Pan, D., Pramanik, M., Senpan, A., Allen, J. S., Zhang, H., Wickline, S. A., Wang, L. V., Lanza, G. M. Molecular photoacoustic imaging of angiogenesis with integrin‐targeted gold nanobeacons. FASEB J. 25, 875–882 (2011). www.fasebj.org

Near infrared photoacoustic detection of sentinel lymph nodes with gold nanobeacons.

Detection of sentinel lymph node (SLN) using photoacoustic imaging is an emerging technique for noninvasive axillary staging of breast cancer. Due to the absence of intrinsic contrast inside the lymph nodes, exogenous contrast agents are used for photoacoustic detection. In this work, we have demonstrated near infrared detection of SLN with gold nanobeacons (GNBs) providing the photoacoustic contrast in a rodent model. We found that size dictates the in vivo characteristics of these nanoparticles in SLN imaging. Larger nanobeacons with high payloads of gold were not as efficient as smaller size nanobeacons with lower payloads for this purpose. Colloidal GNBs were designed as a nanomedicine platform with soft nature that is amenable to bio-elimination, an essential feature for in vivo efficacy and safety. The GNBs were synthesized as lipid- or polymer-encapsulated colloidal particles incorporating tiny gold nanoparticles (2-4 nm) in three tunable sizes (90 nm, 150 nm and 290 nm). Smaller GNBs were noted trafficking through the lymphatic system and accumulating more efficiently in the lymph nodes in comparison to the bigger nanoagents. At 20 min, the GNBs reached the SLN and were no longer observed within the draining lymphatic vessel. Within 1 h post-injection, the contrast ratio of the lymph nodes with the surrounding blood vessels was 9:1. These findings were also supported by analytical measurements of the ex vivo tissue samples. Results indicate that cumulative nanoparticle deposition in lymph nodes is size dependent and that high payloads of gold, although offering greater contrast in vitro, may yield nanoagents with poor intradermal migration and lymphatic transport characteristics.

In vivo carbon nanotube-enhanced non-invasive photoacoustic mapping of the sentinel lymph node

Sentinel lymph node biopsy (SLNB), a less invasive alternative to axillary lymph node dissection (ALND), has become the standard of care for patients with clinically node-negative breast cancer. In SLNB, lymphatic mapping with radio-labeled sulfur colloid and/or blue dye helps identify the sentinel lymph node (SLN), which is most likely to contain metastatic breast cancer. Even though SLNB, using both methylene blue and radioactive tracers, has a high identification rate, it still relies on an invasive surgical procedure, with associated morbidity. In this study, we have demonstrated a non-invasive single-walled carbon nanotube (SWNT)-enhanced photoacoustic (PA) identification of SLN in a rat model. We have successfully imaged the SLN in vivo by PA imaging (793 nm laser source, 5 MHz ultrasonic detector) with high contrast-to-noise ratio (=89) and good resolution ( approximately 500 microm). The SWNTs also show a wideband optical absorption, generating PA signals over an excitation wavelength range of 740-820 nm. Thus, by varying the incident light wavelength to the near infrared region, where biological tissues (hemoglobin, tissue pigments, lipids and water) show low light absorption, the imaging depth is maximized. In the future, functionalization of the SWNTs with targeting groups should allow the molecular imaging of breast cancer.

Molecular Photoacoustic Tomography with Colloidal Nanobeacons

Spotting clots: Vascularly constrained colloidal gold nanobeacons (GNBs; see picture) can be used as exogenous photoacoustic contrast agents for the targeted detection of fibrin, a major biochemical feature of thrombus. Fibrin-targeted GNBs provide a more than tenfold signal enhancement in photoacoustic tomography in the near-IR wavelength window, indicating their potential for diagnostic imaging.

Recent advances in colloidal gold nanobeacons for molecular photoacoustic imaging

Photoacoustic imaging (PAI) represents a hybrid, nonionizing modality, which has been of particular interest because of its satisfactory spatial resolution and high soft tissue contrast. PAI has the potential to provide both functional and molecular imaging in vivo since optical absorption is sensitive to physiological parameters. In this review we summarize our effort to advance molecular PAI with colloidal gold nanobeacons (GNB). GNB represents a robust nanoparticle platform that entraps multiple copies of tiny gold nanoparticles (2-4u2009nm) within a larger colloidal particle encapsulated by biocompatible synthetic or natural amphilines. The utilization of numerous small gold particles greatly amplifies the signal without exceeding the renal elimination threshold size. With fibrin-targeted GNB, the robust detection of microthrombus formed over a ruptured atherosclerotic plaque has been achieved, which offers an important opportunity to recognize patients with moderate lumen stenosis but high risk of stroke. With the use of second-generation smaller GNBs, the potential to improve sentinel lymph node assessment and biopsy was advanced with respect to rapidity and sensitivity of detection in mice. Finally, for angiogenesis, an essential microanatomical biomarker of tumor and cardiovascular disease progression, integrin-targeted GNBs allowed visualization of numerous angiogenic sprouts and bridges that were otherwise undetectable from inherent blood signal alone, offering sensitive and specific discrimination and quantification of angiogenesis in vivo.

Photoacoustic tomography of foreign bodies in soft biological tissue

In detecting small foreign bodies in soft biological tissue, ultrasound imaging suffers from poor sensitivity (52.6%) and specificity (47.2%). Hence, alternative imaging methods are needed. Photoacoustic (PA) imaging takes advantage of strong optical absorption contrast and high ultrasonic resolution. A PA imaging system is employed to detect foreign bodies in biological tissues. To achieve deep penetration, we use near-infrared light ranging from 750 to 800 nm and a 5-MHz spherically focused ultrasonic transducer. PA images were obtained from various targets including glass, wood, cloth, plastic, and metal embedded more than 1 cm deep in chicken tissue. The locations and sizes of the targets from the PA images agreed well with those of the actual samples. Spectroscopic PA imaging was also performed on the objects. These results suggest that PA imaging can potentially be a useful intraoperative imaging tool to identify foreign bodies.