Color Processing in the Early Visual System of Drosophila.

Color vision extracts spectral information by comparing signals from photoreceptors with different visual pigments. Such comparisons are encoded by color-opponent neurons that are excited at one wavelength and inhibited at another. Here, we examine the circuit implementation of color-opponent processing in the Drosophila visual system by combining two-photon calcium imaging with genetic dissection of visual circuits. We report that color-opponent processing of UVshort/blue and UVlong/green is already implemented in R7/R8 inner photoreceptor terminals of "pale" and "yellow" ommatidia, respectively. R7 and R8 photoreceptors of the same type of ommatidia mutually inhibit each other directly via HisCl1 histamine receptors and receive additional feedback inhibition that requires the second histamine receptor Ort. Color-opponent processing at the first visual synapse represents an unexpected commonality between Drosophila and vertebrates; however, the differences in the molecular and cellular implementation suggest that the same principles evolved independently.

Molecular imaging in oncology: Current impact and future directions.

The authors define molecular imaging, according to the Society of Nuclear Medicine and Molecular Imaging, as the visualization, characterization, and measurement of biological processes at the molecular and cellular levels in humans and other living systems. Although practiced for many years clinically in nuclear medicine, expansion to other imaging modalities began roughly 25 years ago and has accelerated since. That acceleration derives from the continual appearance of new and highly relevant animal models of human disease, increasingly sensitive imaging devices, high-throughput methods to discover and optimize affinity agents to key cellular targets, new ways to manipulate genetic material, and expanded use of cloud computing. Greater interest by scientists in allied fields, such as chemistry, biomedical engineering, and immunology, as well as increased attention by the pharmaceutical industry, have likewise contributed to the boom in activity in recent years. Whereas researchers and clinicians have applied molecular imaging to a variety of physiologic processes and disease states, here, the authors focus on oncology, arguably where it has made its greatest impact. The main purpose of imaging in oncology is early detection to enable interception if not prevention of full-blown disease, such as the appearance of metastases. Because biochemical changes occur before changes in anatomy, molecular imaging-particularly when combined with liquid biopsy for screening purposes-promises especially early localization of disease for optimum management. Here, the authors introduce the ways and indications in which molecular imaging can be undertaken, the tools used and under development, and near-term challenges and opportunities in oncology.

Optical in situ deciphering of the surface reconstruction-assistant multielectron transfer event of single Co3O4 nanoparticles.

Surface reconstruction determines the fate of catalytic sites on the near-surface during the oxygen evolution reaction. However, deciphering the conversion mechanism of various intermediate-states during surface reconstruction remains a challenge. Herein, we employed an optical imaging technique to draw the landscape of dynamic surface reconstruction on individual Co3O4 nanoparticles. By regulating the surface states of Co3O4 nanoparticles, we explored dynamic growth of the CoOx(OH)y sublayer on single Co3O4 nanoparticles and directly identified the conversion between two dynamics. Rich oxygen vacancies induced more active sites on the surface and prolonged surface reconstruction, which enhanced electrochemical redox and oxygen evolution. These results were further verified by in situ electrochemical extinction spectroscopy of single Co3O4 nanoparticles. We elucidate the heterogeneous evolution of surface reconstruction on individual Co3O4 nanoparticles and present a unique perspective to understand the fate of catalytic species on the nanosurface, which is of enduring significance for investigating the heterogeneity of multielectron-transfer events.

In vivo three-dimensional brain imaging with chemiluminescence probes in Alzheimer's disease models.

Optical three-dimensional (3D) molecular imaging is highly desirable for providing precise distribution of the target-of-interest in disease models. However, such 3D imaging is still far from wide applications in biomedical research; 3D brain optical molecular imaging, in particular, has rarely been reported. In this report, we designed chemiluminescence probes with high quantum yields, relatively long emission wavelengths, and high signal-to-noise ratios to fulfill the requirements for 3D brain imaging in vivo. With assistance from density-function theory (DFT) computation, we designed ADLumin-Xs by locking up the rotation of the double bond via fusing the furan ring to the phenyl ring. Our results showed that ADLumin-5 had a high quantum yield of chemiluminescence and could bind to amyloid beta (Aß). Remarkably, ADLumin-5's radiance intensity in brain areas could reach 4 × 107 photon/s/cm2/sr, which is probably 100-fold higher than most chemiluminescence probes for in vivo imaging. Because of its strong emission, we demonstrated that ADLumin-5 could be used for in vivo 3D brain imaging in transgenic mouse models of Alzheimer's disease.

Speckle-correlation imaging through a kaleidoscopic multimode fiber.

Speckle-correlation imaging techniques are widely used for noninvasive imaging through complex scattering media. While light propagation through multimode fibers and scattering media share many analogies, reconstructing images through multimode fibers from speckle correlations remains an unsolved challenge. Here, we exploit a kaleidoscopic memory effect emerging in square-core multimode fibers and demonstrate fluorescence imaging with no prior knowledge on the fiber. Experimentally, our approach simply requires to translate random speckle patterns at the input of a square-core fiber and to measure the resulting fluorescence intensity with a bucket detector. The image of the fluorescent object is then reconstructed from the autocorrelation of the measured signal by solving an inverse problem. This strategy does not require the knowledge of the fragile deterministic relation between input and output fields, which makes it promising for the development of flexible minimally invasive endoscopes.

Cortical Networks Relating to Arousal Are Differentially Coupled to Neural Activity and Hemodynamics.

Even in the absence of specific sensory input or a behavioral task, the brain produces structured patterns of activity. This organized activity is modulated by changes in arousal. Here, we use wide-field voltage imaging to establish how arousal relates to cortical network voltage and hemodynamic activity in spontaneously behaving head-fixed male and female mice expressing the voltage-sensitive fluorescent FRET sensor Butterfly 1.2. We find that global voltage and hemodynamic signals are both positively correlated with changes in arousal with a maximum correlation of 0.5 and 0.25, respectively, at a time lag of 0 s. We next show that arousal influences distinct cortical regions for both voltage and hemodynamic signals. These include a broad positive correlation across most sensory-motor cortices extending posteriorly to the primary visual cortex observed in both signals. In contrast, activity in the prefrontal cortex is positively correlated to changes in arousal for the voltage signal while it is a slight net negative correlation observed in the hemodynamic signal. Additionally, we show that coherence between voltage and hemodynamic signals relative to arousal is strongest for slow frequencies below 0.15 Hz and is near zero for frequencies >1 Hz. We finally show that coupling patterns are dependent on the behavioral state of the animal with correlations being driven by periods of increased orofacial movement. Our results indicate that while hemodynamic signals show strong relations to behavior and arousal, these relations are distinct from those observed by voltage activity.

Ultraliser: a framework for creating multiscale, high-fidelity and geometrically realistic 3D models for in silico neuroscience.

Ultraliser is a neuroscience-specific software framework capable of creating accurate and biologically realistic 3D models of complex neuroscientific structures at intracellular (e.g. mitochondria and endoplasmic reticula), cellular (e.g. neurons and glia) and even multicellular scales of resolution (e.g. cerebral vasculature and minicolumns). Resulting models are exported as triangulated surface meshes and annotated volumes for multiple applications in in silico neuroscience, allowing scalable supercomputer simulations that can unravel intricate cellular structure-function relationships. Ultraliser implements a high-performance and unconditionally robust voxelization engine adapted to create optimized watertight surface meshes and annotated voxel grids from arbitrary non-watertight triangular soups, digitized morphological skeletons or binary volumetric masks. The framework represents a major leap forward in simulation-based neuroscience, making it possible to employ high-resolution 3D structural models for quantification of surface areas and volumes, which are of the utmost importance for cellular and system simulations. The power of Ultraliser is demonstrated with several use cases in which hundreds of models are created for potential application in diverse types of simulations. Ultraliser is publicly released under the GNU GPL3 license on GitHub (BlueBrain/Ultraliser). SIGNIFICANCE: There is crystal clear evidence on the impact of cell shape on its signaling mechanisms. Structural models can therefore be insightful to realize the function; the more realistic the structure can be, the further we get insights into the function. Creating realistic structural models from existing ones is challenging, particularly when needed for detailed subcellular simulations. We present Ultraliser, a neuroscience-dedicated framework capable of building these structural models with realistic and detailed cellular geometries that can be used for simulations.

Imaging the Retinal Vascular Mural Cells In Vivo: Elucidating the Timeline of Their Loss in Diabetic Retinopathy.

BACKGROUND: Vascular mural cells (VMCs) are integral components of the retinal vasculature with critical homeostatic functions such as maintaining the inner blood-retinal barrier and vascular tone, as well as supporting the endothelial cells. Histopathologic donor eye studies have shown widespread loss of pericytes and smooth muscle cells, the 2 main VMC types, suggesting these cells are critical to the pathogenesis of diabetic retinopathy (DR). There remain, however, critical gaps in our knowledge regarding the timeline of VMC demise in human DR. METHODS: In this study, we address this gap using adaptive optics scanning laser ophthalmoscopy to quantify retinal VMC density in eyes with no retinal disease (healthy), subjects with diabetes without diabetic retinopathy, and those with clinical DR and diabetic macular edema. We also used optical coherence tomography angiography to quantify capillary density of the superficial and deep capillary plexuses in these eyes. RESULTS: Our results indicate significant VMC loss in retinal arterioles before the appearance of classic clinical signs of DR (diabetes without diabetic retinopathy versus healthy, 5.0±2.0 versus 6.5±2.0 smooth muscle cells per 100 µm; P<0.05), while a significant reduction in capillary VMC density (5.1±2.3 in diabetic macular edema versus 14.9±6.0 pericytes per 100 µm in diabetes without diabetic retinopathy; P=0.01) and capillary density (superficial capillary plexus vessel density, 37.6±3.8 in diabetic macular edema versus 45.5±2.4 in diabetes without diabetic retinopathy; P<0.0001) is associated with more advanced stages of clinical DR, particularly diabetic macular edema. CONCLUSIONS: Our results offer a new framework for understanding the pathophysiologic course of VMC compromise in DR, which may facilitate the development and monitoring of therapeutic strategies aimed at VMC preservation and potentially the prevention of clinical DR and its associated morbidity. Imaging retinal VMCs provides an unparalleled opportunity to visualize these cells in vivo and may have wider implications in a range of diseases where these cells are disrupted.

Super-resolution re-scan second harmonic generation microscopy.

Second harmonic generation microscopy (SHG) is generally acknowledged as a powerful tool for the label-free three-dimensional visualization of tissues and advanced materials, with one of its most popular applications being collagen imaging. Despite the great need, progress in super-resolved SHG imaging lags behind the developments reported over the past years in fluorescence-based optical nanoscopy. In this work, we demonstrate super-resolved re-scan SHG, qualitatively and quantitatively showing on collagenous tissues the available resolution advantage over the diffraction limit. We introduce as well super-resolved re-scan two-photon excited fluorescence microscopy, an imaging modality not explored to date.

Artificial intelligence-aided optical imaging for cancer theranostics.

The use of artificial intelligence (AI) to assist biomedical imaging have demonstrated its high accuracy and high efficiency in medical decision-making for individualized cancer medicine. In particular, optical imaging methods are able to visualize both the structural and functional information of tumors tissues with high contrast, low cost, and noninvasive property. However, no systematic work has been performed to inspect the recent advances on AI-aided optical imaging for cancer theranostics. In this review, we demonstrated how AI can guide optical imaging methods to improve the accuracy on tumor detection, automated analysis and prediction of its histopathological section, its monitoring during treatment, and its prognosis by using computer vision, deep learning and natural language processing. By contrast, the optical imaging techniques involved mainly consisted of various tomography and microscopy imaging methods such as optical endoscopy imaging, optical coherence tomography, photoacoustic imaging, diffuse optical tomography, optical microscopy imaging, Raman imaging, and fluorescent imaging. Meanwhile, existing problems, possible challenges and future prospects for AI-aided optical imaging protocol for cancer theranostics were also discussed. It is expected that the present work can open a new avenue for precision oncology by using AI and optical imaging tools.

Beyond primary visual cortex: The leading role of lateral occipital complex in early conscious visual processing.

The study of the neural substrates that serve conscious vision is one of the unsolved questions of cognitive neuroscience. So far, consciousness literature has endeavoured to disentangle which brain areas and in what order are involved in giving rise to visual awareness, but the problem of consciousness still remains unsolved. Availing of two different but complementary sources of data (i.e., Fast Optical Imaging and EEG), we sought to unravel the neural dynamics responsible for the emergence of a conscious visual experience. Our results revealed that conscious vision is characterized by a significant increase of activation in extra-striate visual areas, specifically in the Lateral Occipital Complex (LOC), and that, more interestingly, such activity occurred in the temporal window of the ERP component commonly thought to represent the electrophysiological signature of visual awareness, i.e., the Visual Awareness Negativity (VAN). Furthermore, Granger causality analysis, performed to further investigate the flow of activity occurring in the investigated areas, unveiled that neural processes relating to conscious perception mainly originated in LOC and subsequently spread towards visual and motor areas. In general, the results of the present study seem to advocate for an early contribution of LOC in conscious vision, thus suggesting that it could represent a reliable neural correlate of visual awareness. Conversely, striate visual areas, showing awareness-related activity only in later stages of stimulus processing, could be part of the cascade of neural events following awareness emergence.

Animal models to study cognitive impairment of chronic kidney disease.

Mild cognitive impairment (MCI) is common in people with chronic kidney disease (CKD), and its prevalence increases with progressive loss of kidney function. MCI is characterized by a decline in cognitive performance greater than expected for an individual age and education level but with minimal impairment of instrumental activities of daily living. Deterioration can affect one or several cognitive domains (attention, memory, executive functions, language, and perceptual motor or social cognition). Given the increasing prevalence of kidney disease, more and more people with CKD will also develop MCI causing an enormous disease burden for these individuals, their relatives, and society. However, the underlying pathomechanisms are poorly understood, and current therapies mostly aim at supporting patients in their daily lives. This illustrates the urgent need to elucidate the pathogenesis and potential therapeutic targets and test novel therapies in appropriate preclinical models. Here, we will outline the necessary criteria for experimental modeling of cognitive disorders in CKD. We discuss the use of mice, rats, and zebrafish as model systems and present valuable techniques through which kidney function and cognitive impairment can be assessed in this setting. Our objective is to enable researchers to overcome hurdles and accelerate preclinical research aimed at improving the therapy of people with CKD and MCI.

A novel HER2-specific sensor based on DARPin_9-29 and albumin binding domain for real-time fluorescence-guided tumor detection in animal model of cancer.

In animal models of cancer, targeted fluorescence bioimaging, performed non-invasively and in real time, is indispensable tool for assessing tumor location, spread of metastasis, and the therapeutic potential of anticancer drugs under development. To overcome the limitation of antibodies in bioimaging applications, small artificial scaffold proteins based on ankyrin repeats (DARPins, designed ankyrin repeat proteins) are used as tumor-associated antigen binders. In this study for the first time, we assessed the potential of DARPin_9-29, the human epidermal growth factor receptor 2 (HER2) subdomain I-specific protein, genetically fused with albumin binding domain (ABD) and conjugated with Cyanine5.5 as a NIR sensor for fluorescence bioimaging of HER2-positive cancer in animal model. In vivo biodistribution studies have revealed sufficient tumor-to-background ratios at 48 h (3.17 ± 0.55) and 72 h (3.49 ± 0.64) postinjection, providing excellent contrast between the primary tumor and tissue background and allowing clear breast tumor detection. Ex vivo biodistribution has shown that ABD module in DARP-ABD sensor prevents renal reabsorption and increases tumor accumulation in more than 10-folds compared to excreting organs. To verify if DARP-ABD-Cy5.5 can demarcate HER2-positive tumor in vivo, HER2-positive syngeneic breast cancer cell line with constitutive gene expression of luciferase eFFLuc, was created. The powerful combination of bioluminescence and fluorescence imaging let to track the fluorescent anti-HER2 DARP-ABD sensor in bioluminescent HER2-positive breast tumors. Our results validate DARP-ABD as a promising sensor for fluorescence-guided imaging of HER2-positive solid cancer, which can be used in the development of improved anticancer treatment strategies.

Viewing early life without labels: optical approaches for imaging the early embryo†.

Embryo quality is an important determinant of successful implantation and a resultant live birth. Current clinical approaches for evaluating embryo quality rely on subjective morphology assessments or an invasive biopsy for genetic testing. However, both approaches can be inherently inaccurate and crucially, fail to improve the live birth rate following the transfer of in vitro produced embryos. Optical imaging offers a potential non-invasive and accurate avenue for assessing embryo viability. Recent advances in various label-free optical imaging approaches have garnered increased interest in the field of reproductive biology due to their ability to rapidly capture images at high resolution, delivering both morphological and molecular information. This burgeoning field holds immense potential for further development, with profound implications for clinical translation. Here, our review aims to: (1) describe the principles of various imaging systems, distinguishing between approaches that capture morphological and molecular information, (2) highlight the recent application of these technologies in the field of reproductive biology, and (3) assess their respective merits and limitations concerning the capacity to evaluate embryo quality. Additionally, the review summarizes challenges in the translation of optical imaging systems into routine clinical practice, providing recommendations for their future development. Finally, we identify suitable imaging approaches for interrogating the mechanisms underpinning successful embryo development.

Scattering-Based Light-Sheet Microscopy Imaging of Human Papillomavirus-Associated Squamous Lesions of the Anal Canal: A Proof-of-Principle Study.

Demand for anal cancer screening is expected to rise following the recent publication of the Anal Cancer-HSIL Outcomes Research trial, which showed that treatment of high-grade squamous intraepithelial lesions significantly reduces the rate of progression to anal cancer. While screening for human papillomavirus-associated squamous lesions in the cervix is well established and effective, this is less true for other sites in the lower anogenital tract. Current anal cancer screening and prevention rely on high-resolution anoscopy with biopsies. This procedure has a steep learning curve for providers and may cause patient discomfort. Scattering-based light-sheet microscopy (sLSM) is a novel imaging modality with the potential to mitigate these challenges through real-time, microscopic visualization of disease-susceptible tissue. Here, we report a proof-of-principle study that establishes feasibility of dysplasia detection using an sLSM device. We imaged 110 anal biopsy specimens collected prospectively at our institution's dysplasia clinic (including 30 nondysplastic, 40 low-grade squamous intraepithelial lesion, and 40 high-grade squamous intraepithelial lesion specimens) and found that these optical images are highly interpretable and accurately recapitulate histopathologic features traditionally used for the diagnosis of human papillomavirus-associated squamous dysplasia. A reader study to assess diagnostic accuracy suggests that sLSM images are noninferior to hematoxylin and eosin images for the detection of anal dysplasia (sLSM accuracy = 0.87; hematoxylin and eosin accuracy = 0.80; P = .066). Given these results, we believe that sLSM technology holds great potential to enhance the efficacy of anal cancer screening by allowing accurate sampling of diagnostic tissue at the time of anoscopy. While the current imaging study was performed on ex vivo biopsy specimens, we are currently developing a handheld device for in vivo imaging that will provide immediate microscopic guidance to high-resolution anoscopy providers.

Revisiting hemodynamics and blood oxygenation in a microfluidic microvasculature replica.

The complexity of microvascular circulation has led to the development of advanced imaging techniques and biomimetic models. This study developed a multifaceted microfluidic-based microdevice as an in vitro model of microvasculature to replicate important geometric and functional features of in vivo perfusion in mice. The microfluidic device consisted of a microchannel for blood perfusion, mirroring the natural hierarchical branching vascular structures found in mice. Additionally, the device incorporated a steady gradient of oxygen (O2) which diffused through the polydimethylsiloxane (PDMS) layer, allowing for dynamic blood oxygenation. The assembled multi-layered microdevice was accompanied by a dual-modal imaging system that combined laser speckle contrast imaging (LSCI) and intrinsic signal optical imaging (ISOI) to visualize full-field blood flow distributions and blood O2 profiles. By closely reproducing in vivo blood perfusion and oxygenation conditions, this microvasculature model, in conjunction with numerical simulation results, can provide quantitative information on physiologically relevant hemodynamics and key O2 transport parameters that are not directly measurable in traditional animal studies.

Mapping cutaneous field carcinogenesis of nonmelanoma skin cancer using mesoscopic imaging of pro-inflammation cues.

Nonmelanoma skin cancers remain the most widely diagnosed types of cancers globally. Thus, for optimal patient management, it has become imperative that we focus our efforts on the detection and monitoring of cutaneous field carcinogenesis. The concept of field cancerization (or field carcinogenesis), introduced by Slaughter in 1953 in the context of oral cancer, suggests that invasive cancer may emerge from a molecularly and genetically altered field affecting a substantial area of underlying tissue including the skin. A carcinogenic field alteration, present in precancerous tissue over a relatively large area, is not easily detected by routine visualization. Conventional dermoscopy and microscopy imaging are often limited in assessing the entire carcinogenic landscape. Recent efforts have suggested the use of noninvasive mesoscopic (between microscopic and macroscopic) optical imaging methods that can detect chronic inflammatory features to identify pre-cancerous and cancerous angiogenic changes in tissue microenvironments. This concise review covers major types of mesoscopic optical imaging modalities capable of assessing pro-inflammatory cues by quantifying blood haemoglobin parameters and hemodynamics. Importantly, these imaging modalities demonstrate the ability to detect angiogenesis and inflammation associated with actinically damaged skin. Representative experimental preclinical and human clinical studies using these imaging methods provide biological and clinical relevance to cutaneous field carcinogenesis in altered tissue microenvironments in the apparently normal epidermis and dermis. Overall, mesoscopic optical imaging modalities assessing chronic inflammatory hyperemia can enhance the understanding of cutaneous field carcinogenesis, offer a window of intervention and monitoring for actinic keratoses and nonmelanoma skin cancers and maximise currently available treatment options.

Line-field confocal optical coherence tomography in dermato-oncology: A literature review towards harmonized histopathology-integrated terminology.

Non-invasive diagnostics like line-field confocal optical coherence tomography (LC-OCT) are being implemented in dermato-oncology. However, unification of terminology in LC-OCT is lacking. By reviewing the LC-OCT literature in the field of dermato-oncology, this study aimed to develop a unified terminological glossary integrated with traditional histopathology. A PRISMA-guided literature-search was conducted for English-language publications on LC-OCT of actinic keratosis (AK), keratinocyte carcinoma (KC), and malignant melanoma (MM). Study characteristics and terminology were compiled. To harmonize LC-OCT terminology and integrate with histopathology, synonymous terms for image features of AK, KC, and MM were merged by two authors, organized by skin layer and lesion-type. A subset of key LC-OCT image-markers with histopathological correlates that in combination were typical of AK, squamous cell carcinoma in situ (SCCis), invasive squamous cell carcinoma (SCC), basal cell carcinoma (BCC), and MM in traditional histopathology, were selected from the glossary by an experienced dermatopathologist. Seventeen observational studies of AK (7 studies), KC (13 studies), MM (7 studies) utilizing LC-OCT were included, with 117 terms describing either AK, KC, or MM. These were merged to produce 45 merged-terms (61.5% reduction); 5 assigned to the stratum corneum (SC), 23 to the viable epidermis, 2 to dermo-epidermal junction (DEJ) and 15 to the dermis. For each lesion, mandatory key image-markers were a well-defined DEJ and presence of mild/moderate but not severe epidermal dysplasia for AK, severe epidermal dysplasia and well-defined DEJ for SCCis, interrupted DEJ and/or dermal broad infiltrative strands for invasive SCC, dermal lobules connected and/or unconnected to the epidermis for BCC, as well as single atypical melanocytes and/or nest of atypical melanocytes in the epidermis or dermis for MM. This review compiles evidence on LC-OCT in dermato-oncology, providing a harmonized histopathology-integrated terminology and key image-markers for each lesion. Further evaluation is required to determine the clinical value of these findings.

Oligonucleotide probes for imaging and diagnosis of bacterial infections.

The rise of infectious diseases as a public health concern has necessitated the development of rapid and precise diagnostic methods. Imaging techniques like nuclear and optical imaging provide the ability to diagnose infectious diseases within the body, eliminating delays caused by sampling and pre-enrichments of clinical samples and offering spatial information that can aid in a more informed diagnosis. Traditional molecular probes are typically created to image infected tissue without accurately identifying the pathogen. In contrast, oligonucleotides can be tailored to target specific RNA sequences, allowing for the identification of pathogens, and even generating antibiotic susceptibility profiles by focusing on drug resistance genes. Despite the benefits that nucleic acid mimics (NAMs) have provided in terms of stabilizing oligonucleotides, the inadequate delivery of these relatively large molecules into the cytoplasm of bacteria remains a challenge for widespread use of this technology. This review summarizes the key advancements in the field of oligonucleotide probes for in vivo imaging, highlighting the most promising delivery systems described in the literature for developing optical imaging through in vivo hybridization.

First-in-man study of the PSMA Minibody IR800-IAB2M for molecularly targeted intraoperative fluorescence guidance during radical prostatectomy.

PURPOSE: Prostate-specific membrane antigen (PSMA) is increasingly used to image prostate cancer in clinical practice. We sought to develop and test a humanised PSMA minibody IAB2M conjugated to the fluorophore IRDye 800CW-NHS ester in men undergoing robot-assisted laparoscopic radical prostatectomy (RARP) to image prostate cancer cells during surgery. METHODS: The minibody was evaluated pre-clinically using PSMA positive/negative xenograft models, following which 23 men undergoing RARP between 2018 and 2020 received between 2.5 mg and 20 mg of IR800-IAB2M intravenously, at intervals between 24 h and 17 days prior to surgery. At every step of the procedure, the prostate, pelvic lymph node chains and extra-prostatic surrounding tissue were imaged with a dual Near-infrared (NIR) and white light optical platform for fluorescence in vivo and ex vivo. Histopathological evaluation of intraoperative and postoperative microscopic fluorescence imaging was undertaken for verification. RESULTS: Twenty-three patients were evaluated to optimise both the dose of the reagent and the interval between injection and surgery and secure the best possible specificity of fluorescence images. Six cases are presented in detail as exemplars. Overall sensitivity and specificity in detecting non-lymph-node extra-prostatic cancer tissue were 100% and 65%, and 64% and 64% respectively for lymph node positivity. There were no side-effects associated with administration of the reagent. CONCLUSION: Intraoperative imaging of prostate cancer tissue is feasible and safe using IR800-IAB2M. Further evaluation is underway to assess the benefit of using the technique in improving completion of surgical excision during RARP. REGISTRATION: ISCRCTN10046036: https://www.isrctn.com/ISRCTN10046036 .