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Regional lymph node metastasis is the strongest prognostic factor in patients with melanoma. Published reports that used lymphoscintigraphy with radioactive colloids and blue dye demonstrated accurate sentinel lymph node (SLN) identification in inguinal nodes and axillary nodes, but decreased accuracy in cervical, popliteal, epitrochlear, and parascapular nodes. Near-infrared imaging (NIR) may utilize indocyanine green (ICG) to improve SLN identification. The safety, feasibility and optimal dose of albumin-bound ICG (ICG:HSA) was assessed by NIR to improve SLN mapping in patients with melanoma. Twenty-five consecutive patients with biopsy-proven melanoma underwent standard SLN mapping with preoperatively administered technetium-99 m nanocolloid (Tc-99 m). Intraoperative NIR fluorescence imaging was performed after injection of 1.0 ml of 100, 250 or 500 μM of ICG:HSA in four quadrants around the primary lesion. NIR fluorescent imaging demonstrated accuracy of 98 % when compared with radioactive colloid. A total of 65 lymph nodes were identified (65 with Tc-99 m, 64 with ICG:HSA). Overall, successful mapping that used either technique was 96 % as one patient failed to map with either modality. As the dose of ICG was increased, the signal-to-background ratio increased from a median of 3.1 to 8.4 to 10.9 over the range of 100, 250, and 500 μM, respectively. SLN mapping with ICG:HSA is feasible and accurate in melanoma. ICG has the added advantage of a low cost and an intraoperative technique that does not alter the surgical field, thus allowing for easy identification of SLNs.

Invisible NIR fluorescent light can provide high sensitivity, high-resolution, and real-time image-guidance during oncologic surgery, but imaging systems that are presently available do not display this invisible light in the context of surgical anatomy. The FLARE imaging system overcomes this major obstacle.Color video was acquired simultaneously, and in real-time, along with two independent channels of NIR fluorescence. Grayscale NIR fluorescence images were converted to visible "pseudo-colors" and overlaid onto the color video image. Yorkshire pigs weighing 35 kg (n = 5) were used for final preclinical validation of the imaging system. A six-patient pilot study was conducted in women undergoing sentinel lymph node (SLN) mapping for breast cancer. Subjects received (99m)Tc-sulfur colloid lymphoscintigraphy. In addition, 12.5 microg of indocyanine green (ICG) diluted in human serum albumin (HSA) was used as an NIR fluorescent lymphatic tracer.The FLARE system permitted facile positioning in the operating room. NIR light did not change the look of the surgical field. Simultaneous pan-lymphatic and SLN mapping was demonstrated in swine using clinically available NIR fluorophores and the dual NIR capabilities of the system. In the pilot clinical trial, a total of nine SLNs were identified by (99m)Tc- lymphoscintigraphy and nine SLNs were identified by NIR fluorescence, although results differed in two patients. No adverse events were encountered.We describe the successful clinical translation of a new NIR fluorescence imaging system for image-guided oncologic surgery.

Neuroendocrine tumors of the pancreas, such as insulinoma, are difficult to localize, and complete resection is essential for cure. Our hypothesis is that a near-infrared (NIR) fluorophore exhibiting uptake in insulinoma could provide high-sensitivity detection intraoperatively.The optical properties of methylene blue (MB) were measured in vitro in 100% serum at 37 degrees C and in vivo after tissue uptake. MB was injected as a rapid intravenous bolus at doses ranging from 0.25 to 2 mg/kg into wildtype rats and pigs, and into insulinoma-bearing transgenic mice. The FLARE imaging system was used to acquire color video and NIR fluorescence images simultaneously and in real-time. The signal-to-background ratios (SBR) of tissues and tumors were quantified using FLARE software.When appropriately diluted, MB exhibits moderate NIR fluorescence emission peaking at 688 nm. At dosesor =1 mg/kg, certain normal tissues, such as pancreas, accumulate MB and remain NIR fluorescent for up to 1 h with an SBRor = 1.6. MB spectral properties are maintained after uptake into tissue. Interestingly, insulinoma exhibits even higher MB signal than normal pancreas, resulting in insulinoma-to-pancreas ratios of 3.7 and insulinoma-to-muscle ratios of 16.2. MB permitted high-sensitivity, real-time localization of primary, multicentric, and metastatic insulinoma and permitted differentiation among tumor, normal pancreas, and other abdominal structures.A single intravenous injection of a clinically available, commonly used NIR fluorophore provides prolonged intraoperative localization of normal pancreas and insulinoma using invisible NIR fluorescent light.

Because many gastrointestinal (GI) tumors spread by way of lymphatics, histological assessment of the first draining lymph nodes has both prognostic and therapeutic significance. However, sentinel lymph node mapping of the GI tract by using available techniques is limited by unpredictable drainage patterns, high background signal, and the inability to image lymphatic tracers relative to surgical anatomy in real time. Our goal was to develop a method for patient-specific intraoperative sentinel lymph node mapping of the GI tract by using invisible near-infrared light. We developed an intraoperative near-infrared fluorescence imaging system that simultaneously displays surgical anatomy and otherwise invisible near-infrared fluorescence images of the surgical field. Near-infrared fluorescent quantum dots were injected intraparenchymally into the stomach, small bowel, and colon, and draining lymphatic channels and sentinel lymph nodes were visualized. Dissection was performed under real-time image guidance. In 10 adult pigs, we demonstrated that 200 pmol of quantum dots quickly and accurately map lymphatic drainage and sentinel lymph nodes. Injection into the mid jejunum and colon results in fluorescence of a single lymph node at the root of the bowel mesentery. Injection into the stomach resulted in identification of a retrogastric node. Histological analysis in all cases confirmed the presence of nodal tissue. We report the use of invisible near-infrared light for intraoperative sentinel lymph node mapping of the GI tract. This technology overcomes the limitations of currently available methods, permits patient-specific imaging of lymphatic flow and sentinel nodes, and provides highly sensitive, real-time image-guided dissection.

Understanding the spatial and temporal drainage patterns of the pleural space could have profound impact on the treatment of lung cancer and mesothelioma. The purpose of this study was to identify the in vivo pattern of drainage from the pleural space to prognostic lymph node stations.Fifty-six rats underwent pleural space injection of a novel lymph tracer composed of recombinant human serum albumin (HSA) covalently conjugated to the near-infrared (NIR) fluorophore IRDye78 via an amide bond (HSA-78). Nodal uptake was imaged at 10, 20, 30, and 60 minutes and 4, 12, and 24 hours after injection with a custom system that simultaneously acquires color video, NIR fluorescence of HSA-78, and a merged picture of the two. Six pigs underwent the same procedure with imaging at 30 minutes, 1 hour, and 24 hours.In both the rat model and the pig model, HSA-78 drained from the pleural space to superior mediastinal lymph nodes first, followed by other intrathoracic and then extrathoracic lymph nodes over the course of 24 hours.NIR fluorescence imaging in two species shows that the superior mediastinal lymph nodes are the first to drain the pleural space. Over the course of 24 hours, the pleural space also communicates with other intrathoracic and then extrathoracic lymph nodes. This study also demonstrates an intraoperative method for identifying nodes communicating with the pleural space, with potential utility in the staging and/or resection of lung cancer and mesothelioma.

BACKGROUND: Sentinel lymph node (SLN) mapping in colorectal cancer may have prognostic and therapeutic significance; however, currently available techniques are not optimal. We hypothesized that the combination of invisible near-infrared (NIR) fluorescent light and ex vivo injection could solve remaining problems of SLN mapping in colorectal cancer. METHODS: The FLARE imaging system was used for real-time identification of SLNs after injection of the NIR lymphatic tracer HSA800 in the colon and rectum of (n = 4) pigs. A total of 32 SLN mappings were performed in vivo and ex vivo after oncologic resection using an identical injection technique. Guided by these results, SLN mappings were performed in ex vivo tissue specimens of 24 consecutive colorectal cancer patients undergoing resection. RESULTS: Lymph flow could be followed in real-time from the injection site to the SLN using NIR fluorescence. In pigs, the SLN was identified in 32 of 32 (100%) of SLN mappings under both in vivo and ex vivo conditions. Clinically, SLNs were identified in all patients (n = 24) using the ex vivo strategy within 5 min after injection of fluorescent tracer. Also, 9 patients showed lymph node involvement (N1 disease). In 1 patient, a 3-mm mesenteric metastasis was found adjacent to a tumor-negative SLN. CONCLUSIONS: The current pilot study shows proof of principle that ex vivo NIR fluorescence-guided SLN mapping can provide high-sensitivity, rapid, and accurate identification of SLNs in colon and rectum. This creates an experimental platform to test optimized, non-FDA-approved NIR fluorescent lymphatic tracers in a clinical setting.

Background Near-infrared (NIR) fluorescent sentinel lymph node (SLN) mapping in breast cancer requires optimized imaging systems and lymphatic tracers. Materials and Methods A small, portable version of the FLARE imaging system, termed Mini-FLARE, was developed for capturing color video and two semi-independent channels of NIR fluorescence (700 and 800 nm) in real time. Initial optimization of lymphatic tracer dose was performed using 35-kg Yorkshire pigs and a 6-patient pilot clinical trial. More refined optimization was performed in 24 consecutive breast cancer patients. All patients received the standard of care using 99mTechnetium-nanocolloid and patent blue. In addition, 1.6 ml of indocyanine green adsorbed to human serum albumin (ICG:HSA) was injected directly after patent blue at the same location. Patients were allocated to 1 of 8 escalating ICG:HSA concentration groups from 50 to 1000 μM. Results The Mini-FLARE system was positioned easily in the operating room and could be used up to 13 in. from the patient. Mini-FLARE enabled visualization of lymphatic channels and SLNs in all patients. A total of 35 SLNs (mean = 1.45, range 1–3) were detected: 35 radioactive (100%), 30 blue (86%), and 35 NIR fluorescent (100%). Contrast agent quenching at the injection site and dilution within lymphatic channels were major contributors to signal strength of the SLN. Optimal injection dose of ICG:HSA ranged between 400 and 800 μM. No adverse reactions were observed. Conclusions We describe the clinical translation of a new NIR fluorescence imaging system and define the optimal ICG:HSA dose range for SLN mapping in breast cancer. Electronic supplementary material The online version of this article (doi:10.1245/s10434-011-1566-x) contains supplementary material, which is available to authorized users.