Extracellular vesicles (EVs) are commonly studied by flow cytometry. Due to their small size and low refractive index, the scatter intensity of most EVs is below the detection limit of common flow cytometers. Here, we aim to improve forward scatter (FSC) and side scatter (SSC) sensitivity of a common flow cytometer to detect single 100 nm EVs. The effects of the optical and fluidics configuration on scatter sensitivity of a FACSCanto (Becton Dickinson) were evaluated by the separation index (SI) and robust coefficient of variation (rCV) of polystyrene beads (BioCytex). Improvement is defined as increased SI and/or reduced rCV. Changing the obscuration bar improved the rCV 1.9‐fold for FSC. A 10‐fold increase in laser power improved the SI 19‐fold for FSC and 4.4‐fold for SSC, whereas the rCV worsened 0.8‐fold and improved 1.5‐fold, respectively. Confocalization worsened the SI 1.2‐fold for FSC, and improved the SI 5.1‐fold for SSC, while the rCV improved 1.1‐fold and worsened 1.5‐fold, respectively. Replacing the FSC photodiode with a photomultiplier tube improved the SI 66‐fold and rCV 4.2‐fold. A 2‐fold reduction in sample stream width improved both SI and rCV for FSC by 1.8‐fold, and for SSC by 1.3‐ and 2.2‐fold, respectively. Decreasing the sample flow velocity worsened rCVs. Decreasing the flow channel dimensions and the pore size of the sheath filter did not substantially change the SI or rCV. Using the optimal optical configuration and fluidics settings, the SI improved 3.8∙10 4 ‐fold on FSC and 30‐fold on SSC, resulting in estimated detection limits for EVs (assuming a refractive index of 1.40) of 246 and 91 nm on FSC and SSC, respectively. Although a 50‐fold improvement on FSC is still necessary, these adaptions have produced an operator‐friendly, high‐throughput flow cytometer with a high sensitivity on both SSC and FSC. © 2020 The Authors. Cytometry Part A published by Wiley Periodicals, Inc. on behalf of International Society for Advancement of Cytometry.

This study investigates the feasibility of in vivo quantitative optical coherence tomography (OCT) of human brain tissue during glioma resection surgery in six patients. High‐resolution detection of glioma tissue may allow precise and thorough tumor resection while preserving functional brain areas, and improving overall survival. In this study, in vivo 3D OCT datasets were collected during standard surgical procedure, before and after partial resection of the tumor, both from glioma tissue and normal parenchyma. Subsequently, the attenuation coefficient was extracted from the OCT datasets using an automated and validated algorithm. The cortical measurements yield a mean attenuation coefficient of 3.8 ± 1.2 mm−1 for normal brain tissue and 3.6 ± 1.1 mm−1 for glioma tissue. The subcortical measurements yield a mean attenuation coefficient of 5.7 ± 2.1 and 4.5 ± 1.6 mm−1 for, respectively, normal brain tissue and glioma. Although the results are inconclusive with respect to trends in attenuation coefficient between normal and glioma tissue due to the small sample size, the results are in the range of previously reported values. Therefore, we conclude that the proposed method for quantitative in vivo OCT of human brain tissue is feasible during glioma resection surgery.

Background The aim of this systematic review was to identify all methods to quantify intraoperative fluorescence angiography (FA) of the gastrointestinal anastomosis, and to find potential thresholds to predict patient outcomes, including anastomotic leakage and necrosis. Methods This systematic review adhered to the PRISMA guidelines. A PubMed and Embase literature search was performed. Articles were included when FA with indocyanine green was performed to assess gastrointestinal perfusion in human or animals, and the fluorescence signal was analysed using quantitative parameters. A parameter was defined as quantitative when a diagnostic numeral threshold for patient outcomes could potentially be produced. Results Some 1317 articles were identified, of which 23 were included. Fourteen studies were done in patients and nine in animals. Eight studies applied FA during upper and 15 during lower gastrointestinal surgery. The quantitative parameters were divided into four categories: time to fluorescence (20 studies); contrast-to-background ratio (3); pixel intensity (2); and numeric classification score (2). The first category was subdivided into manually assessed time (7 studies) and software-derived fluorescence–time curves (13). Cut-off values were derived for manually assessed time (speed in gastric conduit wall) and derivatives of the fluorescence–time curves (Fmax, T1/2, TR and slope) to predict patient outcomes. Conclusion Time to fluorescence seems the most promising category for quantitation of FA. Future research might focus on fluorescence–time curves, as many different parameters can be derived and the fluorescence intensity can be bypassed. However, consensus on study set-up, calibration of fluorescence imaging systems, and validation of software programs is mandatory to allow future data comparison.