Flavin fluorescence lifetime and autofluorescence optical redox ratio for improved visualization and classification of brain tumors.

Purpose: Modern techniques for improved tumor visualization have the aim to maximize the extent of resection during brain tumor surgery and thus improve patient prognosis. Optical imaging of autofluorescence is a powerful and non-invasive tool to monitor metabolic changes and transformation in brain tumors. Cellular redox ratios can be retrieved from fluorescence emitted by the coenzymes reduced nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) and flavin adenine dinucleotide (FAD). Recent studies point out that the influence of flavin mononucleotide (FMN) has been underestimated. Experimental design: Fluorescence lifetime imaging and fluorescence spectroscopy were performed through a modified surgical microscope. We acquired 361 flavin fluorescence lifetime (500-580 nm) and fluorescence spectra (430-740 nm) data points on freshly excised different brain tumors: low-grade gliomas (N=17), high-grade gliomas (N=42), meningiomas (N=23), metastases (N=26) and specimens from the non-tumorous brain (N=3). Results: Protein-bound FMN fluorescence in brain tumors did increase with a shift toward a more glycolytic metabolism (R=-0.87). This increased the average flavin fluorescence lifetime in tumor entities with respect to the non-tumorous brain. Further, these metrics were characteristic for the different tumor entities and showed promise for machine learning based brain tumor classification. Conclusions: Our results shed light on FMN fluorescence in metabolic imaging and outline the potential for supporting the neurosurgeon in visualizing and classifying brain tumor tissue during surgery.

Physics-aware learning and domain-specific loss design in ophthalmology.

The human cataract, a developing opacification of the human eye lens, currently constitutes the world's most frequent cause for blindness. As a result, cataract surgery has become the most frequently performed ophthalmic surgery in the world. By removing the human lens and replacing it with an artificial intraocular lens (IOL), the optical system of the eye is restored. In order to receive a good refractive result, the IOL specifications, especially the refractive power, have to be determined precisely prior to surgery. In the last years, there has been a body of work to perform this prediction by using biometric information extracted from OCT imaging data, recently also by machine learning (ML) methods. Approaches so far consider only biometric information or physical modelling, but provide no effective combination, while often also neglecting IOL geometry. Additionally, ML on small data sets without sufficient domain coverage can be challenging. To solve these issues, we propose OpticNet, a novel optical refraction network based on an unsupervised, domain-specific loss function that explicitly incorporates physical information into the network. By providing a precise and differentiable light propagation eye model, physical gradients following the eye optics are backpropagated into the network. We further propose a new transfer learning procedure, which allows the unsupervised pre-training on the optical model and fine-tuning of the network on small amounts of surgical patient data. We show that our method outperforms the current state of the art on five OCT-image based data sets, provides better domain coverage within its predictions, and achieves better physical consistency.

Fluorescence Lifetime Imaging and Spectroscopic Co-Validation for Protoporphyrin IX-Guided Tumor Visualization in Neurosurgery.

Maximal safe resection is a key strategy for improving patient prognosis in the management of brain tumors. Intraoperative fluorescence guidance has emerged as a standard in the surgery of high-grade gliomas. The administration of 5-aminolevulinic acid prior to surgery induces tumor-specific accumulation of protoporphyrin IX, which emits red fluorescence under blue-light illumination. The technology, however, is substantially limited for low-grade gliomas and weakly tumor-infiltrated brain, where low protoporphyrin IX concentrations are outweighed by tissue autofluorescence. In this context, fluorescence lifetime imaging has shown promise to distinguish spectrally overlapping fluorophores. We integrated frequency-domain fluorescence lifetime imaging in a surgical microscope and combined it with spatially registered fluorescence spectroscopy, which can be considered a research benchmark for sensitive protoporphyrin IX detection. Fluorescence lifetime maps and spectra were acquired for a representative set of fresh ex-vivo brain tumor specimens (low-grade gliomas n = 15, high-grade gliomas n = 80, meningiomas n = 41, and metastases n = 35). Combining the fluorescence lifetime with fluorescence spectra unveiled how weak protoporphyrin IX accumulations increased the lifetime respective to tissue autofluorescence. Infiltration zones (4.1ns ± 1.8ns, p = 0.017) and core tumor areas (4.8ns ± 1.3ns, p = 0.040) of low-grade gliomas were significantly distinguishable from non-pathologic tissue (1.6ns ± 0.5ns). Similarly, fluorescence lifetimes for infiltrated and reactive tissue as well as necrotic and core tumor areas were increased for high-grade gliomas and metastasis. Meningioma tumor specimens showed strongly increased lifetimes (12.2ns ± 2.5ns, p = 0.005). Our results emphasize the potential of fluorescence lifetime imaging to optimize maximal safe resection in brain tumors in future and highlight its potential toward clinical translation.

Soliton microcomb based spectral domain optical coherence tomography.

Spectral domain optical coherence tomography (OCT) is a widely employed, minimally invasive bio-medical imaging technique, which requires a broadband light source, typically implemented by super-luminescent diodes. Recent advances in soliton based photonic integrated frequency combs (soliton microcombs) have enabled the development of low-noise, broadband chipscale frequency comb sources, whose potential for OCT imaging has not yet been unexplored. Here, we explore the use of dissipative Kerr soliton microcombs in spectral domain OCT and show that, by using photonic chipscale Si3N4 resonators in conjunction with 1300 nm pump lasers, spectral bandwidths exceeding those of commercial OCT sources are possible. We characterized the exceptional noise properties of our source (in comparison to conventional OCT sources) and demonstrate that the soliton states in microresonators exhibit a residual intensity noise floor at high offset frequencies that is ca. 3 dB lower than a traditional OCT source at identical power, and can exhibit significantly lower noise performance for powers at the milli-Watt level. Moreover, we demonstrate that classical amplitude noise of all soliton comb teeth are correlated, i.e., common mode, in contrast to superluminescent diodes or incoherent microcomb states, which opens a new avenue to improve imaging speed and performance beyond the thermal noise limit.

Towards real-time wide-field fluorescence lifetime imaging of 5-ALA labeled brain tumors with multi-tap CMOS cameras.

Fluorescence guided neurosurgery based on 5-aminolevulinic acid (5-ALA) has significantly increased maximal safe resections. Fluorescence lifetime imaging (FLIM) of 5-ALA could further boost this development by its increased sensitivity. However, neurosurgeons require real-time visual feedback which was so far limited in dual-tap CMOS camera based FLIM. By optimizing the number of phase frames required for reconstruction, we here demonstrate real-time 5-ALA FLIM of human high- and low-grade glioma with up to 12 Hz imaging rate over a wide field of view (11.0 x 11.0 mm). Compared to conventional fluorescence imaging, real-time FLIM offers enhanced contrast of weakly fluorescent tissue.

Surgical microscope with integrated fluorescence lifetime imaging for 5-aminolevulinic acid fluorescence-guided neurosurgery.

SIGNIFICANCE: 5-Aminolevulinic acid (5-ALA)-based fluorescence guidance in conventional neurosurgical microscopes is limited to strongly fluorescent tumor tissue. Therefore, more sensitive, intrasurgical 5-ALA fluorescence visualization is needed. AIM: Macroscopic fluorescence lifetime imaging (FLIM) was performed ex vivo on 5-ALA-labeled human glioma tissue through a surgical microscope to evaluate its feasibility and to compare it to fluorescence intensity imaging. APPROACH: Frequency-domain FLIM was integrated into a surgical microscope, which enabled parallel wide-field white-light and fluorescence imaging. We first characterized our system and performed imaging of two samples of suspected low-grade glioma, which were compared to histopathology. RESULTS: Our imaging system enabled macroscopic FLIM of a 6.5 × 6.5 mm2 field of view at spatial resolutions <20 µm. A frame of 512 × 512 pixels with a lifetime accuracy <1 ns was obtained in 65 s. Compared to conventional fluorescence imaging, FLIM considerably highlighted areas with weak 5-ALA fluorescence, which was in good agreement with histopathology. CONCLUSIONS: Integration of macroscopic FLIM into a surgical microscope is feasible and a promising method for improved tumor delineation.

Widefield fluorescence lifetime imaging of protoporphyrin IX for fluorescence-guided neurosurgery: An ex vivo feasibility study.

Achieving a maximal safe extent of resection during brain tumor surgery is the goal for improved patient prognosis. Fluorescence-guided neurosurgery using 5-aminolevulinic acid (5-ALA) induced protoporphyrin IX has thereby become a valuable tool enabling a high frequency of complete resections and a prolonged progression-free survival in glioblastoma patients. We present a widefield fluorescence lifetime imaging device with 250 mm working distance, working under similar conditions such as surgical microscopes based on a time-of-flight dual tap CMOS camera. In contrast to intensity-based fluorescence imaging, our method is invariant to light scattering and absorption while being sensitive to the molecular composition of the tissue. We evaluate the feasibility of lifetime imaging of protoporphyrin IX using our system to analyze brain tumor phantoms and fresh 5-ALA-labeled human tissue samples. The results demonstrate the potential of our lifetime sensing device to go beyond the limitation of current intensity-based fluorescence-guided neurosurgery.

Feasibility of intrasurgical spectral-domain optical coherence tomography.

PURPOSE: To evaluate the feasibility of intrasurgical spectral-domain optical coherence tomography in a pilot study. METHODS: Using a Carl Zeiss Meditec Cirrus HD-OCT system adapted to the optical pathway of a Zeiss OPMI VISU 200 surgical microscope, 512 × 128 macular cube scans were performed during various steps of microsurgical procedures in 25 cases. The acquired volume data were postprocessed and visualized using a ray-traced three-dimensional display system. RESULTS: The surgical procedures included pars plana vitrectomies for epiretinal membranes (n = 8), macular holes (n = 4), primary rhegmatogenous retinal detachment (n = 1), proliferative diabetic retinopathy (n = 3), silicone oil removal (n = 2), and cataract surgery only (n = 7). It was possible to acquire intraretinal scans with sufficient quality from all patients. Decisions for additional membrane peeling, knowledge about the behavior of the macular hole and the foveal depression during and after membrane removal, information about clinically invisible fluid accumulation under silicone oil or in a clinically diagnosed "macula-on" retinal detachment, and the condition of the fovea immediately after cataract removal could be gained. CONCLUSION: Intrasurgical spectral-domain optical coherence tomography evaluation is feasible using the tested system and may positively influence surgical decisions and techniques resulting in an improved patient outcome.

Interferometer for optical coherence tomography.

We describe a new interferometer setup for optical coherence tomography (OCT). The interferometer is based on a fiber arrangement similar to Young's two-pinhole interference experiment with spatial coherent and temporal incoherent light. Depth gating is achieved detection of the interference signal on a linear CCD array. Therefore no reference optical delay scanning is needed. The interference signal, the modulation of the signal, the axial resolution, and the depth range are derived theoretically and compared with experiments. The dynamic range of the setup is compared with OCT sensors in the time domain. To our knowledge, the first images of porcine brain and heart tissue and human skin are presented.