OBJECTIVEIn patients with suspected diffusely infiltrating low-grade gliomas (LGG), the prognosis is dependent especially on extent of resection and precision of tissue sampling. Unfortunately, visible 5-aminolevulinic acid (5-ALA) fluorescence is usually only present in high-grade gliomas (HGGs), and most LGGs cannot be visualized. Recently, spectroscopic probes were introduced allowing in vivo quantitative analysis of intratumoral 5-ALA–induced protoporphyrin IX (PpIX) accumulation. The aim of this study was to intraoperatively investigate the value of visible 5-ALA fluorescence and quantitative PpIX analysis in suspected diffusely infiltrating LGG.METHODSPatients with radiologically suspected diffusely infiltrating LGG were prospectively recruited, and 5-ALA was preoperatively administered. During resection, visual fluorescence and absolute tissue PpIX concentration (CPpIX) measured by a spectroscopic handheld probe were determined in different intratumoral areas. Subsequently, corresponding tissue samples were safely collected for histopathological analysis. Tumor diagnosis was established according to the World Health Organization 2016 criteria. Additionally, the tumor grade and percentage of tumor cells were investigated in each sample.RESULTSAll together, 69 samples were collected from 22 patients with histopathologically confirmed diffusely infiltrating glioma. Visible fluorescence was detected in focal areas in most HGGs (79%), but in none of the 8 LGGs. The mean CPpIX was significantly higher in fluorescing samples than in nonfluorescing samples (0.693 μg/ml and 0.008 μg/ml, respectively; p < 0.001). A significantly higher mean percentage of tumor cells was found in samples with visible fluorescence compared to samples with no fluorescence (62% and 34%, respectively; p = 0.005), and significant correlation of CPpIX and percentage of tumor cells was found (r = 0.362, p = 0.002). Moreover, high-grade histology was significantly more common in fluorescing samples than in nonfluorescing samples (p = 0.001), whereas no statistically significant difference in mean CPpIX was noted between HGG and LGG samples. Correlation between maximum CPpIX and overall tumor grade was highly significant (p = 0.005). Finally, 14 (40%) of 35 tumor samples with no visible fluorescence and 16 (50%) of 32 LGG samples showed significantly increased CPpIX (cutoff value: 0.005 μg/ml).CONCLUSIONSVisible 5-ALA fluorescence is able to detect focal intratumoral areas of malignant transformation, and additional quantitative PpIX analysis is especially useful to visualize mainly LGG tissue that usually remains undetected by conventional fluorescence. Thus, both techniques will support the neurosurgeon in achieving maximal safe resection and increased precision of tissue sampling during surgery for suspected LGG.Clinical trial registration no.: NCT01116661 (clinicaltrials.gov)

In fluorescence guided surgery, data visualization represents a critical step between signal capture and display needed for clinical decisions informed by that signal. The diversity of methods for displaying surgical images are reviewed, and a particular focus is placed on electronically detected and visualized signals, as required for near-infrared or low concentration tracers. Factors driving the choices such as human perception, the need for rapid decision making in a surgical environment, and biases induced by display choices are outlined. Five practical suggestions are outlined for optimal display orientation, color map, transparency/alpha function, dynamic range compression, and color perception check.

OBJECTIVE The objective of this study was to detect 5-aminolevulinic acid (ALA)-induced tumor fluorescence from glioma below the surface of the surgical field by using red-light illumination. METHODS To overcome the shallow tissue penetration of blue light, which maximally excites the ALA-induced fluorophore protoporphyrin IX (PpIX) but is also strongly absorbed by hemoglobin and oxyhemoglobin, a system was developed to illuminate the surgical field with red light (620-640 nm) matching a secondary, smaller absorption peak of PpIX and detecting the fluorescence emission through a 650-nm longpass filter. This wide-field spectroscopic imaging system was used in conjunction with conventional blue-light fluorescence for comparison in 29 patients undergoing craniotomy for resection of high-grade glioma, low-grade glioma, meningioma, or metastasis. RESULTS Although, as expected, red-light excitation is less sensitive to PpIX in exposed tumor, it did reveal tumor at a depth up to 5 mm below the resection bed in 22 of 24 patients who also exhibited PpIX fluorescence under blue-light excitation during the course of surgery. CONCLUSIONS Red-light excitation of tumor-associated PpIX fluorescence below the surface of the surgical field can be achieved intraoperatively and enables detection of subsurface tumor that is not visualized under conventional blue-light excitation. Clinical trial registration no.: NCT02191488 (clinicaltrials.gov).

Fluorescence guided surgery (FGS) using aminolevulinic-acid (ALA) induced protoporphyrin IX (PpIX) provides intraoperative visual contrast between normal and malignant tissue during resection of high grade gliomas. However, maps of the PpIX biodistribution within the surgical field based on either visual perception or the raw fluorescence emissions can be masked by background signals or distorted by variations in tissue optical properties. This study evaluates the impact of algorithmic processing of hyperspectral imaging acquisitions on the sensitivity and contrast of PpIX maps. Measurements in tissue-simulating phantoms showed that (I) spectral fitting enhanced PpIX sensitivity compared with visible or integrated fluorescence, (II) confidence-filtering automatically determined the lower limit of detection based on the strength of the PpIX spectral signature in the collected emission spectrum (0.014–0.041 μg/ml in phantoms), and (III) optical-property corrected PpIX estimates were more highly correlated with independent probe measurements (r = 0.98) than with spectral fitting alone (r = 0.91) or integrated fluorescence (r = 0.82). Application to in vivo case examples from clinical neurosurgeries revealed changes to the localization and contrast of PpIX maps, making concentrations accessible that were not visually apparent. Adoption of these methods has the potential to maintain sensitive and accurate visualization of PpIX contrast over the course of surgery.

We have developed an image-updating system to compensate for brain deformation during resection using a computational model with data assimilation of displacements measured with intraoperative stereovision imaging that maintains TREs less than 2 mm on average.

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A 55-year-old man presented to the emergency department with a 1-week history of severe headache. Magnetic resonance imaging of the brain with and without gadolinium-based contrast agent revealed a rim-enhancing mass (4.5 cm in diameter) in the right temporal lobe. At the time of surgery for tumor resection, fluorescein was administered intravenously (Video); Panel A shows the baseline condition before injection. The intact blood-brain barrier of normal brain parenchyma prevents the uptake of fluorescein; however, a tumor can disrupt the blood-brain barrier and allow for the accumulation of the fluorophore. In this patient, within the first 35 seconds after the injection, fluorescence was first evident in arterial vessels (Panel B), then in the smaller veins, and then in the larger veins (Panel C). By 35 seconds after the injection, fluorescein had begun to accumulate in the tumor tissue (Panel C). At 86 seconds, the fluorescein within the cortical veins had begun to fade, providing a sharp contrast between the tumor (Panel D, area inside the dashed line) and normal surrounding tissue; a small area of blood obscures some tumor tissue. Pathological studies revealed a glioblastoma, and postoperative magnetic resonance imaging confirmed gross total resection of the tumor. Four weeks after surgery, the patient began to receive external-beam radiation therapy and temozolomide chemotherapy. At follow-up 2 months after surgery, he had no symptoms of headache and did not have any neurologic deficits.

Mapping the optical absorption and scattering properties of tissues using spatial frequency-domain imaging (SFDI) enhances quantitative fluorescence imaging of protoporphyrin IX (PpIX) in gliomas in the preclinical setting. The feasibility of using SFDI in the operating room was investigated here. A benchtop SFDI system was modified to mount directly to a commercial operating microscope. A digital light processing module imposed a selectable spatial light pattern from a broad-band xenon arc lamp to illuminate the surgical field. White light excitation and a liquid crystal-tunable filter allowed the diffuse reflectance images to be recorded at discrete wavelengths from 450 to 720 nm on a sCMOS camera. The performance was first tested in tissue-simulating phantoms, and data were then acquired intraoperatively during brain tumor resection surgery. The optical absorption and transport scattering coefficients could be estimated with average errors of 3.2% and 4.5% for the benchtop and clinical systems, respectively, with spatial resolution of better than 0.7 mm. These findings suggest that SFDI can be implemented in a clinically relevant configuration to achieve accurate mapping of the optical properties in the surgical field that can then be applied to achieve quantitative imaging of the fluorophore.