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We present the first demonstration of shot-noise limited supercontinuum-based spectral domain optical coherence tomography (SD-OCT) with an axial resolution of 5.9 μm at a center wavelength of 1370 nm. Current supercontinuum-based SD-OCT systems cannot be operated in the shot-noise limited detection regime because of severe pulse-to-pulse relative intensity noise of the supercontinuum source. To overcome this disadvantage, we have developed a low-noise supercontinuum source based on an all-normal dispersion (ANDi) fiber, pumped by a femtosecond laser. The noise performance of our 90 MHz ANDi fiber-based supercontinuum source is compared to that of two commercial sources operating at 80 and 320 MHz repetition rate. We show that the low-noise of the ANDi fiber-based supercontinuum source improves the OCT images significantly in terms of both higher contrast, better sensitivity, and improved penetration. From SD-OCT imaging of skin, retina, and multilayer stacks we conclude that supercontinuum-based SD-OCT can enter the domain of shot-noise limited detection.

We present a single-capture multimodal bright-field (BF) and quantitative phase imaging (QPI) approach that enables the analysis of large, connected, or extended samples, such as confluent cell layers or tissue sections. The proposed imaging concept integrates a fiber-optic Mach–Zehnder interferometer-based off-axis digital holographic microscopy (DHM) with an inverted commercial optical BF microscope. Utilizing 8-bit grayscale dynamic range multiplexing, we simultaneously capture both BF images and digital holograms, which are then demultiplexed numerically via Fourier filtering, phase aberration compensation, and weighted image subtraction procedures. Compared to previous BF-DHM systems, our system avoids synchronization challenges caused by multiple image recording devices, improves acquisition speed, and enhances versatility for fast imaging of large, connected, and rapidly moving samples. Initially, we perform a systematic characterization of the system’s multimodal imaging performance by optimizing numerical as well as coherent and incoherent illumination parameters. Subsequently, the application capabilities are evaluated by multimodal imaging of living cells. The results highlight the potential of single-capture BF-DHM for fast biomedical imaging.

Background Engineered nanomaterials display unique properties that may have impact on human health, and thus require a reliable evaluation of their potential toxicity. Here, we performed a standardized in vitro screening of 23 engineered nanomaterials. We thoroughly characterized the physicochemical properties of the nanomaterials and adapted three classical in vitro toxicity assays to eliminate nanomaterial interference. Nanomaterial toxicity was assessed in ten representative cell lines. Results Six nanomaterials induced oxidative cell stress while only a single nanomaterial reduced cellular metabolic activity and none of the particles affected cell viability. Results from heterogeneous and chemically identical particles suggested that surface chemistry, surface coating and chemical composition are likely determinants of nanomaterial toxicity. Individual cell lines differed significantly in their response, dependent on the particle type and the toxicity endpoint measured. Conclusion In vitro toxicity of the analyzed engineered nanomaterials cannot be attributed to a defined physicochemical property. Therefore, the accurate identification of nanomaterial cytotoxicity requires a matrix based on a set of sensitive cell lines and in vitro assays measuring different cytotoxicity endpoints.

Cytotoxicity quantification of nanoparticles is commonly performed by biochemical assays to evaluate their biocompatibility and safety. We explored quantitative phase imaging (QPI) with digital holographic microscopy (DHM) as a time-resolved in vitro assay to quantify effects caused by three different types of organic nanoparticles in development for medical use. Label-free proliferation quantification of native cell populations facilitates cytotoxicity testing in biomedical nanotechnology. Therefore, DHM quantitative phase images from measurements on nanomaterial and control agent incubated cells were acquired over 24 h, from which the temporal course of the cellular dry mass was calculated within the observed field of view. The impact of LipImage™ 815 lipidots® nanoparticles, as well as empty and cabazitaxel-loaded poly(alkyl cyanoacrylate) nanoparticles on the dry mass development of four different cell lines (RAW 264.7, NIH-3T3, NRK-52E, and RLE-6TN), was observed vs. digitonin as cytotoxicity control and cells in culture medium. The acquired QPI data were compared to a colorimetric cell viability assay (WST-8) to explore the use of the DHM assay with standard biochemical analysis methods downstream. Our results show that QPI with DHM is highly suitable to identify harmful or low-toxic nanomaterials. The presented DHM assay can be implemented with commercial microscopes. The capability for imaging of native cells and the compatibility with common 96-well plates allows high-throughput systems and future embedding into existing experimental routines for in vitro cytotoxicity assessment.

This study aimed to compare different pancreatic enzyme preparations (PEPs) available in Germany regarding particle geometry and size, and to evaluate enzyme activity under physiologically relevant conditions in vitro. Pancreatic endocrine insufficiency is characterized by deficiency of pancreatic enzymes resulting in maldigestion. It is orally treated by pancreatic enzyme replacement therapy. The formulations differ in their physical properties and enzyme release behavior, potentially resulting in inconsistent dosages and poor interchangeability of products. A total of 25 products were analyzed for particle size and number of particles per capsule. Enzyme activities of lipase, amylase, and protease were measured by digestion of olive oil emulsion, starch, and casein, respectively. To analyze enzyme release, gastric environments were simulated by incubating PEPs at pH 1, 4, or 5. Duodenal conditions were simulated by subsequent incubation at pH 6. Regarding physical properties and enzyme release kinetics, considerable differences between different PEPs were found. Furthermore, compared to the label claim, excess lipase activity was observed for most products, reaching up to 148%. These in vitro results suggest poor interchangeability of PEPs, potentially explained by physical and release characteristics. Physicians and patients should be aware of the potential gap between label claims and the real-life performance of different PEPs.

The incorporation of nanomaterials into products can improve performance, efficiency and durability both in industrial applications and in consumer articles. This book presents the state of the art in nanosafety research from a lifecycle perspective. It is divided into four parts: characterization, hazard, release & exposure, and real-life case studies. To improve coherence throughout the book, various chapters review the same suite of well-characterized, judiciously chosen, and identical industrial nanomaterials.

The assessment of nanoparticle cytotoxicity is challenging due to the lack of customized and standardized guidelines for nanoparticle testing. Nanoparticles, with their unique properties, can interfere with biochemical test methods, so multiple tests are required to fully assess their cellular effects. For a more reliable and comprehensive assessment, it is therefore imperative to include methods in nanoparticle testing routines that are not affected by particles and allow for the efficient integration of additional molecular techniques into the workflow. Digital holographic microscopy (DHM), an interferometric variant of quantitative phase imaging (QPI), has been demonstrated as a promising method for the label-free assessment of the cytotoxic potential of nanoparticles. Due to minimal interactions with the sample, DHM allows for further downstream analyses. In this study, we investigated the capabilities of DHM in a multimodal approach to assess cytotoxicity by directly comparing DHM-detected effects on the same cell population with two downstream biochemical assays. Therefore, the dry mass increase in RAW 264.7 macrophages and NIH-3T3 fibroblast populations measured by quantitative DHM phase contrast after incubation with poly(alkyl cyanoacrylate) nanoparticles for 24 h was compared to the cytotoxic control digitonin, and cell culture medium control. Viability was then determined using a metabolic activity assay (WST-8). Moreover, to determine cell death, supernatants were analyzed for the release of the enzyme lactate dehydrogenase (LDH assay). In a comparative analysis, in which the average half-maximal effective concentration (EC50) of the nanocarriers on the cells was determined, DHM was more sensitive to the effect of the nanoparticles on the used cell lines compared to the biochemical assays.

We have developed a drug delivery nanosystem based on chitosan and capsaicin. Both substances have a wide range of biological activities. We investigated the nanosystem's influence on migration and morphology of Madin Darby canine kidney (MDCK-C7) epithelial cells in comparison to the capsaicin-free nanoformulation, free capsaicin, and control cells. For minimally-invasive quantification of cell migration, we applied label-free digital holographic microscopy (DHM) and single-cell tracking. Moreover, quantitative DHM phase images were used as novel stain-free assay to quantify the temporal course of global cellular morphology changes in confluent cell layers. Cytoskeleton alterations and tight junction protein redistributions were complementary analyzed by fluorescence microscopy. Calcium influx measurements were conducted to characterize the influence of the nanoformulations and capsaicin on ion channel activities. We found that both, capsaicin-loaded and unloaded chitosan nanocapsules, and also free capsaicin, have a significant impact on directed cell migration and cellular motility. Increase of velocity and directionality of cell migration correlates with changes in the cell layer surface roughness, tight junction integrity and cytoskeleton alterations. Calcium influx into cells occurred only after nanoformulation treatment but not upon addition of free capsaicin. Our results pave the way for further studies on the biological significance of these findings and potential biomedical applications, e.g. as drug and gene carriers.

In a prospective observational pilot study on patients undergoing elective cardiac surgery with cardiopulmonary bypass, we evaluated label-free quantitative phase imaging (QPI) with digital holographic microscopy (DHM) to describe perioperative inflammation by changes in biophysical cell properties of lymphocytes and monocytes. Blood samples from 25 patients were investigated prior to cardiac surgery and postoperatively at day 1, 3 and 6. Biophysical and morphological cell parameters accessible with DHM, such as cell volume, refractive index, dry mass, and cell shape related form factor, were acquired and compared to common flow cytometric blood cell markers of inflammation and selected routine laboratory parameters. In all examined patients, cardiac surgery induced an acute inflammatory response as indicated by changes in routine laboratory parameters and flow cytometric cell markers. DHM results were associated with routine laboratory and flow cytometric data and correlated with complications in the postoperative course. In a subgroup analysis, patients were classified according to the inflammation related C-reactive protein (CRP) level, treatment with epinephrine and the occurrence of postoperative complications. Patients with regular courses, without epinephrine treatment and with low CRP values showed a postoperative lymphocyte volume increase. In contrast, the group of patients with increased CRP levels indicated an even further enlarged lymphocyte volume, while for the groups of epinephrine treated patients and patients with complicative courses, no postoperative lymphocyte volume changes were detected. In summary, the study demonstrates the capability of DHM to describe biophysical cell parameters of perioperative lymphocytes and monocytes changes in cardiac surgery patients. The pattern of correlations between biophysical DHM data and laboratory parameters, flow cytometric cell markers, and the postoperative course exemplify DHM as a promising diagnostic tool for a characterization of inflammatory processes and course of disease.

Accurate in vitro assessment of nanoparticle cytotoxicity requires a careful selection of the test systems. Due to high adsorption capacity and optical activity, engineered nanoparticles are highly potential in influencing classical cytotoxicity assays. Here, four common in vitro assays for oxidative stress, cell viability, cell death and inflammatory cytokine production (DCF, MTT, LDH and IL-8 ELISA) were assessed for validity using 24 well-characterized engineered nanoparticles. For all nanoparticles, the possible interference with the optical detection methods, the ability to convert the substrates, the influence on enzymatic activity and the potential to bind proinflammatory cytokines were analyzed in detail. Results varied considerably depending on the assay system used. All nanoparticles tested were found to interfere with the optical measurement at concentrations of 50 μg cm −2 and above when DCF, MTT and LDH assays were performed. Except for Carbon Black, particle interference could be prevented by altering assay protocols and lowering particle concentrations to 10 μg cm −2 . Carbon Black was also found to oxidize H 2 DCF-DA in a cell-free system, whereas only ZnO nanoparticles significantly decreased LDH activity. A dramatic loss of immunoreactive IL-8 was observed for only one of the three TiO 2 particle types tested. Our results demonstrate that engineered nanoparticles interfere with classic cytotoxicity assays in a highly concentration-, particle- and assay-specific manner. These findings strongly suggest that each in vitro test system has to be evaluated for each single nanoparticle type to accurately assess the nanoparticle toxicity.