Intraoperative Confocal Laser Endomicroscopy Research Articles

292 Intraoperative Confocal Laser Endomicroscopy During 5-ALA Guided Glioma Surgery: Significant Considerations for Resection at the Tumor Margins

INTRODUCTION: Fluorescence-guided surgery using 5-aminolevulinic acid (5-ALA) has proven effective in aiding the detection of glioma marginal zones and enhancing maximal safe resection. FDA-cleared confocal laser endomicroscopy is another imaging modality for intraoperative brain tissue discrimination on cellular level that could improve visualization at the glioma margin. METHODS: 5-ALA wide-field imaging and CLE were used in 9 patients with gliomas (6 enhancing, 3 non-enhancing). CLE imaging was enhanced by the use of a telesurgical pathology software platform to enable real-time conversation between neurosurgeons and pathologists located remotely. CLE was used for imaging tumor regions subjectively regarded as tumor margins under normal visualization with the operative microscope. After CLE imaging, 5-ALA wide-field imaging was performed in the same regions. A tissue was harvested at imaging locations, and interpretations of CLE and 5-ALA wide-field imaging were compared to permanent histological sections. RESULTS: Overall, 20 deep and superficial margin regions of interest (ROI) were imaged with CLE and 5-ALA imaging. Most of the ROIs interpreted by the neuropathologist as infiltrative glioma based on CLE imaging lacked 5-ALA-induced fluorescence. Permanent histological sections from the corresponding regions were concordant with the interpretation of CLE images in 15 of 20 (75%) ROIs and with the interpretation of 5-ALA imaging in 11 of 20 (55%) ROIs. Sensitivity/specificity of CLE and 5-ALA for interpretation of tumor margins were 79%/67%, and 50%/67%, respectively. Positive/negative predictive values for CLE and 5-ALA were 85%/57%, and 78%/37%, respectively. CONCLUSIONS: Conventional intraoperative understanding of tumor margins based on wide-field fluorescence could underestimate the invasiveness of gliomas. The ongoing study demonstrates that CLE shows higher accuracy in detecting regions with infiltrating tumor than intraoperative 5-ALA imaging.

Fluorescein-stained confocal laser endomicroscopy versus conventional frozen section for intraoperative histopathological assessment of intracranial tumors.

The aim of this clinical trial was to compare Fluorescein-stained intraoperative confocal laser endomicroscopy (CLE) of intracranial lesions and evaluation by a neuropathologist with routine intraoperative frozen section (FS) assessment by neuropathology. In this phase II noninferiority, prospective, multicenter, nonrandomized, off-label clinical trial (EudraCT: 2019-004512-58), patients above the age of 18 years with any intracranial lesion scheduled for elective resection were included. The diagnostic accuracies of both CLE and FS referenced with the final histopathological diagnosis were statistically compared in a noninferiority analysis, representing the primary endpoint. Secondary endpoints included the safety of the technique and time expedited for CLE and FS. A total of 210 patients were included by 3 participating sites between November 2020 and June 2022. Most common entities were high-grade gliomas (37.9%), metastases (24.1%), and meningiomas (22.7%). A total of 6 serious adverse events in 4 (2%) patients were recorded. For the primary endpoint, the diagnostic accuracy for CLE was inferior with 0.87 versus 0.91 for FS, resulting in a difference of 0.04 (95% confidence interval -0.10; 0.02; P = .367). The median time expedited until intraoperative diagnosis was 3 minutes for CLE and 27 minutes for FS, with a mean difference of 27.5 minutes (standard deviation 14.5; P < .001). CLE allowed for a safe and time-effective intraoperative histological diagnosis with a diagnostic accuracy of 87% across all intracranial entities included. The technique achieved histological assessments in real time with a 10-fold reduction of processing time compared to FS, which may invariably impact surgical strategy on the fly.

Intraoperative confocal laser endomicroscopy for interpretation of a sellar hemangioblastoma: illustrative case.

Intraoperative frozen sections play a critical role in surgical strategy because of their ability to provide rapid histopathological information. In cases in which intraoperative biopsy carries a significant risk of bleeding, intraoperative confocal laser endomicroscopy (CLE) can assist in decision-making. The authors present a rare case of a large sellar hemangioblastoma. Preoperative radiographic imaging and normal pituitary function suggested a differential diagnosis that included hemangioblastoma. The patient underwent partial preoperative embolization and a right-sided pterional craniotomy for resection of the lesion. Gross intraoperative examination revealed a highly vascular sellar lesion requiring circumferential dissection to minimize blood loss. The serious vascularity precluded intraoperative frozen section analysis, and CLE imaging was performed. CLE imaging provided excellent visualization of the remarkable vascular structure and characteristic histoarchitecture with microvasculature, intracytoplasmic vacuoles, and atypical cells consistent with hemangioblastoma. Resection and decompression of the chiasm was accomplished, and the patient was discharged with improved vision. The final histopathological diagnosis was hemangioblastoma. When the benefits of obtaining intraoperative frozen sections greatly outweigh the associated risks, CLE imaging can aid in decision-making. CLE imaging offers real-time, on-the-fly evaluation of intraoperative tissue without the need to biopsy a vascular lesion.

384 Characterization of Ex Vivo and In Vivo Intraoperative Neurosurgical Confocal Laser Endomicroscopy Imaging

INTRODUCTION: The new FDA-cleared fluorescein sodium (FNa)-based confocal laser endomicroscopy (CLE) imaging system allows for intraoperative on-the-fly cellular level imaging. Two feasibility studies have been completed with intraoperative use of this CLE system in ex vivo and in vivo modalities. METHODS: Images acquired from two prospective CLE clinical studies, one ex vivo and one in vivo, were analyzed quantitatively. Two image quality parameters – brightness and contrast – were measured using Fiji software and compared between ex vivo and in vivo images for imaging timing from FNa dose and in glioma, meningioma, and intracranial metastatic tumor cases. The diagnostic performance of the two studies was compared. RESULTS: Overall, the in vivo images have higher brightness and contrast than the ex vivo images (p &lt; 0.001). A weak negative correlation exists between image quality and timing of imaging after FNa dose for the ex vivo images, but not the in vivo images. In vivo images have higher brightness and contrast than ex vivo images acquired after FNa redosing. In vivo images have higher image quality than ex vivo images (p &lt; 0.001) in glioma, meningioma, and intracranial metastatic tumor cases. In vivo imaging yielded higher sensitivity (90% vs. 72%) and negative predictive value (81% vs 38%) than ex vivo imaging. CONCLUSIONS: In our setting, in vivo CLE optical biopsy outperforms ex vivo CLE by producing higher quality images and less image deterioration, leading to better diagnostic performance. These results support the in vivo modality as the modality of choice for intraoperative CLE imaging.

Intraoperative confocal laser endomicroscopy: prospective in vivo feasibility study of a clinical-grade system for brain tumors.

The authors evaluated the feasibility of using the first clinical-grade confocal laser endomicroscopy (CLE) system using fluorescein sodium for intraoperative in vivo imaging of brain tumors. A CLE system cleared by the FDA was used in 30 prospectively enrolled patients with 31 brain tumors (13 gliomas, 5 meningiomas, 6 other primary tumors, 3 metastases, and 4 reactive brain tissue). A neuropathologist classified CLE images as interpretable or noninterpretable. Images were compared with corresponding frozen and permanent histology sections, with image correlation to biopsy location using neuronavigation. The specificities and sensitivities of CLE images and frozen sections were calculated using permanent histological sections as the standard for comparison. A recently developed surgical telepathology software platform was used in 11 cases to provide real-time intraoperative consultation with a neuropathologist. Overall, 10,713 CLE images from 335 regions of interest were acquired. The mean duration of the use of the CLE system was 7 minutes (range 3-18 minutes). Interpretable CLE images were obtained in all cases. The first interpretable image was acquired within a mean of 6 (SD 10) images and within the first 5 (SD 13) seconds of imaging; 4896 images (46%) were interpretable. Interpretable image acquisition was positively correlated with study progression, number of cases per surgeon, cumulative length of CLE time, and CLE time per case (p ≤ 0.01). The diagnostic accuracy, sensitivity, and specificity of CLE compared with frozen sections were 94%, 94%, and 100%, respectively, and the diagnostic accuracy, sensitivity, and specificity of CLE compared with permanent histological sections were 92%, 90%, and 94%, respectively. No difference was observed between lesion types for the time to first interpretable image (p = 0.35). Deeply located lesions were associated with a higher percentage of interpretable images than superficial lesions (p = 0.02). The study met the primary end points, confirming the safety and feasibility and acquisition of noninvasive digital biopsies in all cases. The study met the secondary end points for the duration of CLE use necessary to obtain interpretable images. A neuropathologist could interpret the CLE images in 29 (97%) of 30 cases. The clinical-grade CLE system allows in vivo, intraoperative, high-resolution cellular visualization of tissue microstructure and identification of lesional tissue patterns in real time, without the need for tissue preparation.

PMON155 Intraoperative, confocal laser endomicroscopy in pituitary surgery

Abstract Introduction Fluorescence imaging, such as confocal laser endomicroscopy (CLE) is a fast modality of image acquisition, which offers the possibility of early intraoperatively diagnosis and may be used to improve surgical decision making. Objective Within a 3-month prospective trail we used the Zeiss CONVIVO CLE in transspehnoidal surgery for different sellar pathologies. CLE is a miniaturized portable, handheld, fiber optic-based, confocal laser scanning microscope used in conjunction with a 488-nm excitable fluorophore. During the trial period we investigated feasibility, diagnostic accuracy and usability in terms of differential diagnosis in transsphenoidal surgeries of various pituitary tumours (15 pituitary adenomas, 2 craniopharyngiomas, 1 sellar metastasis). Method: Patients were randomly selected. Inclusion criteria were a pituitary macroadenoma or a sellar mass lesion exceeding 10 mm in diameter. A transsphenoidal approach was used and fluorescin was injected upon opening the sellar floor. CLE was carried out immediately after dural opening and during tumour resection, whenever possible. Additionally, the microprobe was used ex vivo with the resected tumor material. Histopathologic features of CLE images and H&amp;E-stained permanent sections were assessed and compared postoperatively. Result: We were able to obtain CLE in-vivo images in all cases. Using the microscopic approach, no problems occurred regarding space for the microprobe. To obtain in-vivo images we had to rest the probe within a mount. Wiggling of the surgeon's hand, pulse synchronous movements of the mass and mild intraoperative venous bleeding led to inexact images. To overcome these problems, we used CLE as early as possible at the beginning of tumor resection to avoid bleeding interferences. Additionally we investigated resected tumor specimens ex-vivo within the OR. Since no bleeding and no interference with movements occurred image quality and acquisition time improved. Histopathologic features of CLE images and H&amp;E-stained permanent sections showed, that except for the sellar metastasis, a diagnosis could be made on the basis of the intraoperative CLE images, whereas the ex-vivo images were more precise. Conclusion CLE is a potential imaging modality for transnasal assessment of skull base and pituitary lesions. It may help to establish an early diagnosis, to identify pituitary microadenomas, especially in cushing´s disease and to distinguish the various sellar pathologies. CLE is a fascinating possibility to verify a suspected diagnoses already intraoperatively. Presentation: Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.

P05.01.B Distribution of the staining agent sodium-fluorescein in cerebral neoplasia - a comparison between intraoperative confocal laser endomicroscopy and in vitro fluorescence microscopy

Abstract Background While conventional frozen sections and other ex vivo microscopy always come along with artefacts, in vivo confocal laser endomicroscopy (CLE) offers less invasive, less manipulated imaging of living tissues. Understanding the distribution of the staining agent sodium-fluorescein (NaFl) plays a key role in establishing in vivo CLE as a new opportunity for real-time transmitted, in vivo imaging. By comparing the distribution of NaFl in in vivo and ex vivo CLE to conventional fluorescence microscopy of NaFl incubated tumor cell cultures we gain a better understanding of the staining mechanism. Material and Methods Sodium-fluorescein is the water-soluble sodium salt of fluorescein and is used as a fluorescent tracer in neurosurgery. The staining agent was applied intravenously at the beginning of the surgical procedure. In vivo CLE of the lesion was performed 30 to 50 minutes later and compared to ex vivo CLE imaging and conventional fluorescence microscopy. In addition, different tumor cell lines derived from malignant gliomas and carcinomas, respectively, were incubated with NaFl in vitro and the uptake of the fluorescent dye was monitored over time. Results From initial results, the intraoperative images showed specific fluorescein distribution depending on the architecture of the tumor entity. In most cases, glial tumors demonstrated higher accumulation of the staining agent in the extracellular tumor matrix, whereas the cells of carcinoma metastases appeared to take up NaFl intracellularly. These results were corroborated by NaFl uptake in cell culture experiments. Compared to ex vivo CLE, in vivo imaging offered a faster assessment of the tissue, brighter images and higher staining levels. Due to movement artefacts and the narrow intraoperative imaging time frame, in vivo CLE images were sometimes impaired by lower image quality. Conclusion The specific distribution of the fluorescent agent NaFl allowed for a discrimination between the different neoplastic entities. Images from in vivo and ex vivo confocal laser endomicroscopy showed NaFl uptake in concordance with the results of NaFl incubated cell cultures. Intraoperative, in vivo confocal laser endomicroscopy shows promising first results in the understanding of brain tumor histomorphology in situ. Being faster and less manipulated by artefacts than ex vivo investigations, it opens up wider opportunities for research and intraoperative diagnostics.

P13.05.A Image annotation guideline for invivo confocal laser endomicroscopy, interrater reliability and how to learn from medical consensus for machine learning algorithms

Abstract Background Intraoperative confocal laser endomicroscopy (CLE) is an in vivo imaging technique increasingly studied in neurosurgery and neuropathology. It can be affected by artifacts introduced by the CLE device or related to the intraoperative setting. We developed and evaluated an image annotation guideline (AGL) to detect and eliminate images bearing no valuable information as a result of such artifacts. Images ware classified into good and bad quality, based on defined technical criteria, which are also considered relevant by clinical experts. Material and Methods Datasets were created from intraoperative CLE in vivo specimens of patients resected for brain tumors. The process from data collection to development of the ML algorithm followed 7 steps: data quality specification, image and metadata collection, AGL development, annotation, data allocation for clinical validation, clinical validation, and, optionally, algorithm development. Final diagnoses were obtained by pathological analysis. Artifacts were grouped into three categories: diminished signal-to-noise-ratio (dSNR), optical distortions (movement/perturbations), and contrast/brightness artifacts. Images were annotated by 4 medical data annotators (T4). For clinical validation, 500 images were excluded from the training data and additionally annotated by 3 board certified neuropathologists (NPs 1-3) with experience in CLE imaging, to determine the medical consensus on good and bad images. All raters (NPs) were compared against each other and against T4; T4 was also compared against the medical consensus. Cohen’s Kappa and overall percentage agreement (OPA) were used to evaluate inter-rater reliability. Positive percent agreement (PPA) and negative percentage agreement (NPA) were also used to evaluate agreement between medical consensus and T4. Results 21,616 CLE images and corresponding clinical metadata were collected from 94 patients and annotated. For each case between 27 and 815 CLE images were acquired over the course of the surgery (mean=175 images per case, SD=170.6). 11% and 13% of images were labeled as dSNR and distortion, respectively, and 34% as class contrast. 42% of the images represented the good quality images. Interrater agreement between the 3 NPs ranged between 0.30 and 0.59. Agreement between T4 and the medical consensus was substantial (Cohen’s Kappa &amp;gt;=0.61). OPA between T4 and the medical consensus was 80.60%, PPA 72.34% and NPA 87.92%. Conclusion Annotations according to a well-structured and expertly curated AGL show higher values for Cohen’s Kappa and Overall Percent Agreement (OPA) with the medical consensus, than that of individual experts among one another. Such an AGL can be considered appropriate and produces on par results with annotations by a group of experts in the field and can be further employed for training machine learning (ML) algorithms.

Characterization of ex vivo and in vivo intraoperative neurosurgical confocal laser endomicroscopy imaging.

BackgroundThe new US Food and Drug Administration-cleared fluorescein sodium (FNa)-based confocal laser endomicroscopy (CLE) imaging system allows for intraoperative on-the-fly cellular level imaging. Two feasibility studies have been completed with intraoperative use of this CLE system in ex vivo and in vivo modalities. This study quantitatively compares the image quality and diagnostic performance of ex vivo and in vivo CLE imaging.MethodsImages acquired from two prospective CLE clinical studies, one ex vivo and one in vivo, were analyzed quantitatively. Two image quality parameters – brightness and contrast – were measured using Fiji software and compared between ex vivo and in vivo images for imaging timing from FNa dose and in glioma, meningioma, and intracranial metastatic tumor cases. The diagnostic performance of the two studies was compared.ResultsOverall, the in vivo images have higher brightness and contrast than the ex vivo images (p < 0.001). A weak negative correlation exists between image quality and timing of imaging after FNa dose for the ex vivo images, but not the in vivo images. In vivo images have higher image quality than ex vivo images (p < 0.001) in glioma, meningioma, and intracranial metastatic tumor cases. In vivo imaging yielded higher sensitivity and negative predictive value than ex vivo imaging.ConclusionsIn our setting, in vivo CLE optical biopsy outperforms ex vivo CLE by producing higher quality images and less image deterioration, leading to better diagnostic performance. These results support the in vivo modality as the modality of choice for intraoperative CLE imaging.

Intraoperative confocal laser endomicroscopy for brain tumors - potential and challenges from a neuropathological perspective.

Confocal laser endomicroscopy (CLE) represents a new non-invasive in vivo imaging technique that holds considerable promise in neurosurgery and neuropathology. CLE is based on the principle of optical sectioning which uses pinholes placed in the light path to selectively image photons of a specific focal plane by filtering out photons above and below the focal plane. Potential indications of CLE in neurosurgery and neuropathology include intraoperative tumor diagnosis and staging as well as assessment of tumor resection margins notably in the case of diffusely infiltrating gliomas. CLE-based tumor analysis in near-real time may also have a significant impact on future tumor resection strategies. We here discuss the technical features of CLE, its potential for wide-field imaging, its role in comparison to established histological techniques for intraoperative tumor assessment and its position in digital pathology and telepathology. Based on our group's experience with a commercially available confocal laser endomicroscope (ZEISS CONVIVO), we critically address the current state of intraoperative CLE in brain tumor surgery, the applicability of classical histological criteria and the strategies required to further improve the diagnostic accuracy of CLE. We finally discuss how a widespread use of CLE in neurosurgery may modify the role of neuropathologists in intraoperative consultation, generating both new opportunities and new challenges.

High-Dose Fluorescein Reveals Unusual Confocal Endomicroscope Imaging of Low-Grade Glioma.

Background: Fluorescence-guided brain tumor surgery using fluorescein sodium (FNa) for contrast is effective in high-grade gliomas. However, the effectiveness of this technique for visualizing noncontrast-enhancing and low-grade gliomas is unknown. This report is the first documented case of the concurrent use of wide-field fluorescence-guided surgery and confocal laser endomicroscopy (CLE) with high-dose FNa (40 mg/kg) for intraoperative visualization of tumor tissue cellularity in a nonenhancing glioma.Case Description: A patient underwent fluorescence-guided surgery for a left frontal lobe mass without contrast enhancement on magnetic resonance imaging. The patient received 40 mg/kg FNa intravenously at the induction of anesthesia. Surgery was performed under visualization with a Yellow 560 filter and white-light wide-field imaging. Intraoperative CLE produced high-quality images of the lesion 1.5 h after FNa injection. Frozen-section analysis demonstrated findings comparable to those of intraoperative CLE visualization and consistent with World Health Organization (WHO) glioma grades II–III. The patient recovered without complications. Analysis of the permanent histologic sections identified the tumor as an anaplastic oligodendroglioma, IDH-mutant, 1p/19q co-deleted, consistent with WHO grade III because of discrete foci of hypercellularity and increased mitotic figures, but large regions of the lesion were low grade.Conclusions: The use of high-dose FNa in this patient with a nonenhancing borderline low-grade/high-grade glioma produced actionable wide-field fluorescence imaging using the operating microscope and improved CLE visualization of tumor cellularity. Higher doses of FNa for intraoperative CLE imaging and possible simultaneous wide-field fluorescence surgical guidance in nonenhancing gliomas merit further investigation.

Intraoperative Confocal Laser Endomicroscopy Ex Vivo Examination of Tissue Microstructure During Fluorescence-Guided Brain Tumor Surgery.

BackgroundNoninvasive intraoperative optical biopsy that provides real-time imaging of histoarchitectural (cell resolution) features of brain tumors, especially at the margin of invasive tumors, would be of great value. To assess clinical-grade confocal laser endomicroscopy (CLE) and to prepare for its use intraoperatively in vivo, we performed an assessment of CLE ex vivo imaging in brain lesions.MethodsTissue samples from patients who underwent intracranial surgeries with fluorescein sodium (FNa)–based wide-field fluorescence guidance were acquired for immediate intraoperative ex vivo optical biopsies with CLE. Hematoxylin-eosin–stained frozen section analysis of the same specimens served as the gold standard for blinded neuropathology comparison. FNa 2 to 5 mg/kg was administered upon induction of anesthesia, and FNa 5 mg/kg was injected for CLE contrast improvement. Histologic features were identified, and the diagnostic accuracy of CLE was assessed.ResultsOf 77 eligible patients, 47 patients with 122 biopsies were enrolled, including 32 patients with gliomas and 15 patients with other intracranial lesions. The positive predictive value of CLE optical biopsies was 97% for all specimens and 98% for gliomas. The specificity of CLE was 90% for all specimens and 94% for gliomas. The second FNa injection in seven patients, a mean of 2.6 h after the first injection, improved image quality and increased the percentage of accurately diagnosed images from 67% to 93%. Diagnostic CLE features of lesional glioma biopsies and normal brain were identified. Seventeen histologic features were identified.ConclusionsResults demonstrated high specificity and positive predictive value of ex vivo intraoperative CLE optical biopsies and justify an in vivo intraoperative trial. This new portable, noninvasive intraoperative imaging technique provides diagnostic features to discriminate lesional tissue with high specificity and is feasible for incorporation into the fluorescence-guided surgery workflow, particularly for patients with invasive brain tumors.

Diagnostic value of probe-based confocal laser endomicroscopy versus conventional endoscopic biopsies of non-muscle invasive bladder tumors: a pilot study

Purpose The standard procedure for diagnostics and follow-up for non-muscle invasive bladder cancer (NMIBC) is cystoscopy in the outpatient clinic. Suspicious lesions are biopsied for histopathological assessment. This pilot study aimed to evaluate the ability of Confocal Laser Endomicroscopy (CLE) to rule out High Grade Urothelial Carcinoma (HGUC) to select patients for in-office fulguration. Materials and methods We performed a prospective non-randomized, single surgeon study. Intraoperative CLE was performed independently by the surgeon and a blinded on-site uropathologist. Following the procedure, a CLE evaluation was performed by another blinded urologist. Lesions were classified as normal/inflammatory, Low Grade Urothelial Carcinoma (LGUC) or HGUC. With the histological evaluations as the gold standard we calculated sensitivity, specificity, PPV and NPV for HGUC and the accuracy for each CLE assessor. The primary outcome was the NPV for HGUC for the surgeon. Results Twelve patients with a total of 34 lesions were included. Six lesions were flat and 28 were exophytic. On histopathology, 25 lesions were classified as normal/inflammatory or LGUC, while nine were classified as HGUC. For the surgeon, the uropathologist and the second urologist, the sensitivity was 44%, 78% and 22%, respectively. Specificities for the three observers were 84%, 68% and 96%. This corresponded to PPVs for HGUC of 50%, 47% and 67% and NPV for HGUC of 81%, 89% and 77%. Conclusions In our hands the NPV of CLE is not high enough for it to be considered an alternative to histopathological assessment of bladder lesions.

Optical biopsy of penile cancer with in vivo confocal laser endomicroscopy

IntroductionSurgical management of penile cancer depends on accurate margin assessment and staging. Advanced optical imaging technologies may improve penile biopsy and organ-sparing treatment. We evaluated the feasibility of confocal laser endomicroscopy for intraoperative assessment of benign and malignant penile tissue. Patients and methodsWith institutional review board approval, 11 patients were recruited, 9 with suspected penile cancer, and 2 healthy controls. Confocal laser endomicroscopy using a 2.6-mm fiber-optic probe was performed at 1 or 2 procedures on all subjects, for 13 imaging procedures. Fluorescein was administered intravenously approximately 3 minutes prior to imaging for contrast. Video sequences from in vivo (n = 12) and ex vivo (n = 6) imaging were obtained of normal glans, suspicious lesions, and surgical margins. Images were processed, annotated, characterized, and correlated with standard hematoxylin and eosin histopathology. ResultsNo adverse events related to imaging were reported. Distinguishing features of benign and malignant penile tissue could be identified by confocal laser endomicroscopy. Normal skin had cells of uniform size and shape, with distinct cytoplasmic membranes consistent with squamous epithelium. Malignant lesions were characterized by disorganized, crowded cells of various size and shape, lack of distinct cytoplasmic membranes, and hazy, moth-eaten appearance. The transition from normal to abnormal squamous epithelium could be identified. ConclusionsWe report the initial feasibility of intraoperative confocal laser endomicroscopy for penile cancer optical biopsy. Pending further evaluation, confocal laser endomicroscopy could serve as an adjunct or replacement to conventional frozen section pathology for management of penile cancer.

Confocal-Assisted Fluorescence Resection of Low-Grade Gliomas

Abstract INTRODUCTION Low-grade gliomas (LGG) are the most common intra-axial brain tumors. Intraoperatively it's not possible to distinguish optically between the tumor and the normal tissue. Furthermore, it can not be detected with fluorescent agents like 5-aminolevulinacid (5-ALA). Those reasons often lead to inaccurate resection of the tumor. Confocal laser endomicroscopy (CLE) is a cellular imaging modality used for optical biopsies. The aim of this study was to use intraoperative CLE to differentiate tumor from normal tissue and to define borders during resection of LGG on a cellular level. METHODS We investigated the use of intraoperative CLE on 6 patients with LGG. We used a confocal system that is able to detect fluorescence on the infrared spectrum and we applied indocyanine green as a detecting fluorescent agent. We analyzed the accuracy of the intraoperative cellular CLE optical biopsies in comparison with the hematoxylin and eosin stains of the biopsies taken from the tumor and normal tissue borders area. RESULTS With CLE we were able intraoperatively to diagnose the tumor entity, normal brain structures, and vascularization modalities and to control the resection zone on a cellular level. Furthermore, CLE pictures showed exactly the cellular borders between tumor and normal tissue. The correlation of the CLE pictures with the H&amp;E staining was 100%. CONCLUSION CLE, on one hand, allows intraoperative detection and differentiation of single tumor cells, while, on the other hand, it allows defining borders between tumor and normal tissue on a cellular level, dramatically improving the accuracy of surgical resection of LGG. The application and implementation of CLE-assisted surgery increases not only the options for real-tumor identification but also the therapeutic options by extending the resection borders of LGG on a cellular level and, more importantly, by protecting the functionality of normal tissue on the adjacent areas of the human brain.

Utilization of intraoperative confocal laser endomicroscopy in brain tumor surgery.

Precise identification of tumor margins is of the utmost importance in neuro-oncology. Confocal microscopy is capable of rapid imaging of fresh tissues at cellular resolution and has been miniaturized into handheld probe-based systems suitable for use in the operating room. We aimed to perform a literature review to provide an update on the current status of confocal laser endomicroscopy (CLE) technology for brain tumor surgery. Aside from benchtop confocal microscopes used in ex vivo fashion, there are four CLE systems that have been investigated for potential application in the workflow of brain tumor surgery. Preclinical studies on animal tumor models and clinical studies on human brain tumors have assessed in vivo and ex vivo imaging approaches, suggesting that confocal microscopy holds promise for rapid identification of the characteristic (diagnostic) histological features of tumor and normal brain tissues. However, there are few studies assessing diagnostic accuracy sufficient to provide a definitive determination of the clinical and economical value of CLE in brain tumor surgery. Intraoperative real-time, high-resolution tissue imaging has significant clinical potential in the field of neuro-oncology. CLE is an emerging imaging technology that shows promise for improving brain tumor surgery workflow in in vivo and ex vivo studies. Future clinical studies are necessary to demonstrate clinical and economic benefit of CLE.

Prospective evaluation of the utility of intraoperative confocal laser endomicroscopy in patients with brain neoplasms using fluorescein sodium: experience with 74 cases.

This study evaluated the utility, specificity, and sensitivity of intraoperative confocal laser endomicroscopy (CLE) to provide diagnostic information during resection of human brain tumors. CLE imaging was used in the resection of intracranial neoplasms in 74 consecutive patients (31 male; mean age 47.5 years; sequential 10-month study period). Intraoperative in vivo and ex vivo CLE was performed after intravenous injection of fluorescein sodium (FNa). Tissue samples from CLE imaging-matched areas were acquired for comparison with routine histological analysis (frozen and permanent sections). CLE images were classified as diagnostic or nondiagnostic. The specificities and sensitivities of CLE and frozen sections for gliomas and meningiomas were calculated using permanent histological sections as the standard. CLE images were obtained for each patient. The mean duration of intraoperative CLE system use was 15.7 minutes (range 3-73 minutes). A total of 20,734 CLE images were correlated with 267 biopsy specimens (mean number of images/biopsy location, in vivo 84, ex vivo 70). CLE images were diagnostic for 45.98% in vivo and 52.97% ex vivo specimens. After initiation of CLE, an average of 14 in vivo images and 7 ex vivo images were acquired before identification of a first diagnostic image. CLE specificity and sensitivity were, respectively, 94% and 91% for gliomas and 93% and 97% for meningiomas. CLE with FNa provided intraoperative histological information during brain tumor removal. Specificities and sensitivities of CLE for gliomas and meningiomas were comparable to those for frozen sections. These data suggest that CLE could allow the interactive identification of tumor areas, substantially improving intraoperative decisions during the resection of brain tumors.

Intraoperative Optical Biopsy during Robotic Assisted Radical Prostatectomy Using Confocal Endomicroscopy

Intraoperative optical biopsy technologies may aid in the identification of important anatomical landmarks and improve surgical outcomes of robotic assisted radical prostatectomy. We evaluate the feasibility of confocal laser endomicroscopy during robotic assisted radical prostatectomy. A total of 21 patients with biopsy proven prostate cancer scheduled for robotic assisted radical prostatectomy were recruited. After intravenous administration of fluorescein 15 patients underwent invivo intraoperative confocal laser endomicroscopy of prostatic and periprostatic structures using a 2.6 or 0.85 mm imaging probe. Standard robotic instruments were used to grasp and maneuver the confocal laser endomicroscopy probes for image acquisition. Confocal laser endomicroscopy imaging was performed exvivo on fresh prostate specimens from 20 patients. Confocal video sequences acquired invivo and exvivo were reviewed and analyzed, with additional image processing using a mosaicing algorithm. Processed confocal images were compared with standard hematoxylin and eosin analysis of imaged regions. Confocal laser endomicroscopy was successfully integrated with robotic surgery, including co-registration of confocal video sequences with white light and probe handling with standard robotic instrumentation. Intraoperative confocal laser endomicroscopy imaging of the neurovascular bundle before and after nerve sparing dissection revealed characteristic features including dynamic vascular flow and intact axon fibers. Exvivo confocal imaging of the prostatic parenchyma demonstrated normal prostate glands, stroma and prostatic carcinoma. We report the initial feasibility of optical biopsy of prostatic and periprostatic tissue during robotic assisted radical prostatectomy. Image guidance and tissue interrogation using confocal laser endomicroscopy offer a new intraoperative imaging method that has the potential to improve the functional and oncologic outcomes of prostate cancer surgery.

758 Intraoperative confocal laser endomicroscopy during robotic-assisted radical prostatectomy: Initial feasibility study

758 Intraoperative confocal laser endomicroscopy during robotic-assisted radical prostatectomy: Initial feasibility study