Bayesian modelling of time series data (BayModTS)-a FAIR workflow to process sparse and highly variable data.

MOTIVATION: Systems biology aims to better understand living systems through mathematical modelling of experimental and clinical data. A pervasive challenge in quantitative dynamical modelling is the integration of time series measurements, which often have high variability and low sampling resolution. Approaches are required to utilize such information while consistently handling uncertainties. RESULTS: We present BayModTS (Bayesian modelling of time series data), a new FAIR (findable, accessible, interoperable, and reusable) workflow for processing and analysing sparse and highly variable time series data. BayModTS consistently transfers uncertainties from data to model predictions, including process knowledge via parameterized models. Further, credible differences in the dynamics of different conditions can be identified by filtering noise. To demonstrate the power and versatility of BayModTS, we applied it to three hepatic datasets gathered from three different species and with different measurement techniques: (i) blood perfusion measurements by magnetic resonance imaging in rat livers after portal vein ligation, (ii) pharmacokinetic time series of different drugs in normal and steatotic mice, and (iii) CT-based volumetric assessment of human liver remnants after clinical liver resection. AVAILABILITY AND IMPLEMENTATION: The BayModTS codebase is available on GitHub at https://github.com/Systems-Theory-in-Systems-Biology/BayModTS. The repository contains a Python script for the executable BayModTS workflow and a widely applicable SBML (systems biology markup language) model for retarded transient functions. In addition, all examples from the paper are included in the repository. Data and code of the application examples are stored on DaRUS: https://doi.org/10.18419/darus-3876. The raw MRI ROI voxel data were uploaded to DaRUS: https://doi.org/10.18419/darus-3878. The steatosis metabolite data are published on FairdomHub: 10.15490/fairdomhub.1.study.1070.1.

libRoadRunner 2.0: a high performance SBML simulation and analysis library.

MOTIVATION: This article presents libRoadRunner 2.0, an extensible, high-performance, cross-platform, open-source software library for the simulation and analysis of models expressed using the systems biology markup language (SBML). RESULTS: libRoadRunner is a self-contained library, able to run either as a component inside other tools via its C++, C and Python APIs, or interactively through its Python or Julia interface. libRoadRunner uses a custom just-in-time (JIT) compiler built on the widely used LLVM JIT compiler framework. It compiles SBML-specified models directly into native machine code for a large variety of processors, making it fast enough to simulate extremely large models or repeated runs in reasonable timeframes. libRoadRunner is flexible, supporting the bulk of the SBML specification (except for delay and non-linear algebraic equations) as well as several SBML extensions such as hierarchical composition and probability distributions. It offers multiple deterministic and stochastic integrators, as well as tools for steady-state, sensitivity, stability and structural analyses. AVAILABILITY AND IMPLEMENTATION: libRoadRunner binary distributions for Windows, Mac OS and Linux, Julia and Python bindings, source code and documentation are all available at https://github.com/sys-bio/roadrunner, and Python bindings are also available via pip. The source code can be compiled for the supported systems as well as in principle any system supported by LLVM-13, such as ARM-based computers like the Raspberry Pi. The library is licensed under the Apache License Version 2.0.

BioSimulators: a central registry of simulation engines and services for recommending specific tools.

Computational models have great potential to accelerate bioscience, bioengineering, and medicine. However, it remains challenging to reproduce and reuse simulations, in part, because the numerous formats and methods for simulating various subsystems and scales remain siloed by different software tools. For example, each tool must be executed through a distinct interface. To help investigators find and use simulation tools, we developed BioSimulators (https://biosimulators.org), a central registry of the capabilities of simulation tools and consistent Python, command-line and containerized interfaces to each version of each tool. The foundation of BioSimulators is standards, such as CellML, SBML, SED-ML and the COMBINE archive format, and validation tools for simulation projects and simulation tools that ensure these standards are used consistently. To help modelers find tools for particular projects, we have also used the registry to develop recommendation services. We anticipate that BioSimulators will help modelers exchange, reproduce, and combine simulations.

Murine iPSC-Loaded Scaffold Grafts Improve Bone Regeneration in Critical-Size Bone Defects.

In certain situations, bones do not heal completely after fracturing. One of these situations is a critical-size bone defect where the bone cannot heal spontaneously. In such a case, complex fracture treatment over a long period of time is required, which carries a relevant risk of complications. The common methods used, such as autologous and allogeneic grafts, do not always lead to successful treatment results. Current approaches to increasing bone formation to bridge the gap include the application of stem cells on the fracture side. While most studies investigated the use of mesenchymal stromal cells, less evidence exists about induced pluripotent stem cells (iPSC). In this study, we investigated the potential of mouse iPSC-loaded scaffolds and decellularized scaffolds containing extracellular matrix from iPSCs for treating critical-size bone defects in a mouse model. In vitro differentiation followed by Alizarin Red staining and quantitative reverse transcription polymerase chain reaction confirmed the osteogenic differentiation potential of the iPSCs lines. Subsequently, an in vivo trial using a mouse model (n = 12) for critical-size bone defect was conducted, in which a PLGA/aCaP osteoconductive scaffold was transplanted into the bone defect for 9 weeks. Three groups (each n = 4) were defined as (1) osteoconductive scaffold only (control), (2) iPSC-derived extracellular matrix seeded on a scaffold and (3) iPSC seeded on a scaffold. Micro-CT and histological analysis show that iPSCs grafted onto an osteoconductive scaffold followed by induction of osteogenic differentiation resulted in significantly higher bone volume 9 weeks after implantation than an osteoconductive scaffold alone. Transplantation of iPSC-seeded PLGA/aCaP scaffolds may improve bone regeneration in critical-size bone defects in mice.

The systems biology simulation core library.

SUMMARY: Studying biological systems generally relies on computational modeling and simulation, e.g., model-driven discovery and hypothesis testing. Progress in standardization efforts led to the development of interrelated file formats to exchange and reuse models in systems biology, such as SBML, the Simulation Experiment Description Markup Language (SED-ML) or the Open Modeling EXchange format. Conducting simulation experiments based on these formats requires efficient and reusable implementations to make them accessible to the broader scientific community and to ensure the reproducibility of the results. The Systems Biology Simulation Core Library (SBSCL) provides interpreters and solvers for these standards as a versatile open-source API in JavaTM. The library simulates even complex bio-models and supports deterministic Ordinary Differential Equations; Stochastic Differential Equations; constraint-based analyses; recent SBML and SED-ML versions; exchange of results, and visualization of in silico experiments; open modeling exchange formats (COMBINE archives); hierarchically structured models; and compatibility with standard testing systems, including the Systems Biology Test Suite and published models from the BioModels and BiGG databases. AVAILABILITY AND IMPLEMENTATION: SBSCL is freely available at https://draeger-lab.github.io/SBSCL/ and via Maven Central. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

PK-DB: pharmacokinetics database for individualized and stratified computational modeling.

A multitude of pharmacokinetics studies have been published. However, due to the lack of an open database, pharmacokinetics data, as well as the corresponding meta-information, have been difficult to access. We present PK-DB (https://pk-db.com), an open database for pharmacokinetics information from clinical trials. PK-DB provides curated information on (i) characteristics of studied patient cohorts and subjects (e.g. age, bodyweight, smoking status, genetic variants); (ii) applied interventions (e.g. dosing, substance, route of application); (iii) pharmacokinetic parameters (e.g. clearance, half-life, area under the curve) and (iv) measured pharmacokinetic time-courses. Key features are the representation of experimental errors, the normalization of measurement units, annotation of information to biological ontologies, calculation of pharmacokinetic parameters from concentration-time profiles, a workflow for collaborative data curation, strong validation rules on the data, computational access via a REST API as well as human access via a web interface. PK-DB enables meta-analysis based on data from multiple studies and data integration with computational models. A special focus lies on meta-data relevant for individualized and stratified computational modeling with methods like physiologically based pharmacokinetic (PBPK), pharmacokinetic/pharmacodynamic (PK/PD), or population pharmacokinetic (pop PK) modeling.

Pelvic tilt impacts cup orientation on CT: how accurate is the gold standard?

BACKGROUND: Computed tomography (CT) is the gold standard for the accurate measurement of cup inclination and anteversion in total hip arthroplasty (THA). However, functional cup position strongly depends on the tilt of the pelvis and thus pelvic tilt in the supine position on the CT table harbors the risk of misinterpretation. PURPOSE: To evaluate the influence of pelvic tilt on cup measurements in postoperative CT scans. MATERIAL AND METHODS: This is a secondary outcome analysis of a prospective study. In 123 patients undergoing THA, anatomic cup inclination and anteversion as measured on postoperative 3D-CT scans were compared between: (i) the anterior pelvic plane (APP) resembling neutral pelvic tilt; and (ii) the coronal plane representing the standard planes on CT. Furthermore, the effect of the variation on cup positions within Lewinnek's safe zone in relation to the applied reference plane was assessed. RESULTS: Mean pelvic tilt in supine position was -3.5° ± 5.6°. This resulted in a mean difference for anteversion between APP und coronal plane of 2.1° ± 3.7° and of 2.1° ± 1.9° for cup inclination, respectively. The change varied up to 20.2° for cup inclination and up 12.2° for cup anteversion. The overall conversion rate for inclination and/or anteversion regarding Lewinnek's safe zone was high at 23.6% (29/123). CONCLUSION: Pelvic tilt affects cup orientation on CT scans depending on the applied reference plane. Standard CT planes should be corrected regarding pelvic tilt before measurements to avoid errors in measurement.

A novel radiological classification for displaced os coccyx: the Benditz-König classification.

BACKGROUND: Treatment of coccygodynia is still a challenging entity. Clear surgical selection criteria are still lacking. The aim of the investigation was to establish a novel radiological classification for surgical decision-making in coccygodynia cases. MATERIAL AND METHODS: Retrospective analysis of standing and sitting X-rays of coccygodynia patients referred to a single centre from 2018 to 2020. The sacro-coccygeal angle (SCA), the intra-coccygeal angle (ICA) and the difference of the intervertebral disc height (∆IDH) were measured. All coccyges were distributed in subtypes and correlated with the patients' treatment. RESULTS: In total, 138 patients (female/male: 103/35) with a mean age of 45.6 ± 15.4 years were included in the study. In total, 49 patients underwent coccygectomy. Four different subtypes of displaced coccyges were identified: Type I with a non-segmented coccyx, anterior pivot, increased SCA and ICA from standing to sitting, ∆IDH = 1.0 ± 1.5 mm. Type II with a multisegmented coccyx, anterior pivot, increased SCA and ICA standing/sitting, ∆IDH = 1.1 ± 1.6 mm. Type III showed a posterior pivoted coccyx, negative SCA and ICA, ∆IDH = 0.6 ± 1.6 mm. Type IV is characterized by an anterior-posterior dissociation of the tail bone with a positive SCA, and the ICA shifted from a posterior to an anterior orientation. ∆IDH was - 0.6 ± 1.8 mm. CONCLUSION: The presented radiological classification could help to facilitate the surgical decision-making for patients with displaced os coccyx. In addition, lateral and sitting X-rays were easy to perform and did not need unnecessary ionizing radiation like in CT scans and were more cost-effective than MRI investigations. The subtypes III and especially IV were more likely leading to surgery.

Increase in Lower Limb Strength after Multimodal Pain Management in Patients with Low Back Pain.

Background and Objectives: The aim of the present study was to evaluate the efficacy of a multimodal pain therapy (MPM) regarding the objective parameter muscle strength of segment-dependent lower limb muscle groups before and after such a treatment. Materials and Methods: 52 patients with a history of low back pain and/or leg pain received standardized multimodal pain management. Strength of segment indicating lower limb muscles were assessed for each patient before and after ten days of treatment by handheld dynamometry. Results: Overall strength increased significantly from 23.6 kg ± 6.6 prior to treatment to 25.4 ± 7.3 after treatment, p ≤ 0.001. All muscle groups significantly increased in strength with exception of great toe extensors. Conclusions: Despite lower basic strength values at the beginning of treatment, all investigated muscle groups, except for the great toe extensors, showed a significant increase of overall strength after completion of the multimodal pain management concept. Increased overall strength could help with avoiding further need of medical care by supporting patients' autonomy in daily life activities, as well as maintaining working abilities. Thus, our study is the first to show a significant positive influence on lower limb strength in patients with low back pain after a conservative MPM program.

Harmonizing semantic annotations for computational models in biology.

Life science researchers use computational models to articulate and test hypotheses about the behavior of biological systems. Semantic annotation is a critical component for enhancing the interoperability and reusability of such models as well as for the integration of the data needed for model parameterization and validation. Encoded as machine-readable links to knowledge resource terms, semantic annotations describe the computational or biological meaning of what models and data represent. These annotations help researchers find and repurpose models, accelerate model composition and enable knowledge integration across model repositories and experimental data stores. However, realizing the potential benefits of semantic annotation requires the development of model annotation standards that adhere to a community-based annotation protocol. Without such standards, tool developers must account for a variety of annotation formats and approaches, a situation that can become prohibitively cumbersome and which can defeat the purpose of linking model elements to controlled knowledge resource terms. Currently, no consensus protocol for semantic annotation exists among the larger biological modeling community. Here, we report on the landscape of current annotation practices among the COmputational Modeling in BIology NEtwork community and provide a set of recommendations for building a consensus approach to semantic annotation.

SBML Level 3: an extensible format for the exchange and reuse of biological models.

Systems biology has experienced dramatic growth in the number, size, and complexity of computational models. To reproduce simulation results and reuse models, researchers must exchange unambiguous model descriptions. We review the latest edition of the Systems Biology Markup Language (SBML), a format designed for this purpose. A community of modelers and software authors developed SBML Level 3 over the past decade. Its modular form consists of a core suited to representing reaction-based models and packages that extend the core with features suited to other model types including constraint-based models, reaction-diffusion models, logical network models, and rule-based models. The format leverages two decades of SBML and a rich software ecosystem that transformed how systems biologists build and interact with models. More recently, the rise of multiscale models of whole cells and organs, and new data sources such as single-cell measurements and live imaging, has precipitated new ways of integrating data with models. We provide our perspectives on the challenges presented by these developments and how SBML Level 3 provides the foundation needed to support this evolution.

The rupture of an anterior communicating artery aneurysm does not associate with an asymmetry in the A1 or A2 arteries: a retrospective study of radiological features.

BACKGROUND AND OBJECTIVES: Although the formation and rupture risk of an anterior communicating artery (ACoA) aneurysm has been the subject of many studies, no previous study has primarily searched for the relationship of the parent and daughter vessels and the impact of their size/diameter ratio on the potential rupture risk of an AcoA aneurysm. The objective of this study is to explore this link and to further analyse the surrounding vasculature of the anterior communicating artery aneurysm. MATERIALS AND METHODS: We conducted a retrospective analysis of 434 patients: 284 patients with an ACoA aneurysm (121 unruptured and 162 ruptured) and 150 control patients without an ΑCoA aneurysm. Radiological angiography investigations were used to assess the diameter ratios of the parent vessels in addition to ACoA aneurysm morphology parameters. RESULTS: When comparing the ruptured to the unruptured cases, we observed no significant difference in the parent or daughter vessel diameter ratios. Younger patient age (OR 0.96, p = 0.00) and a higher aneurysm size ratio (OR 1.10, p = 0.02) were of prognostic importance concerning the rupture risk of the aneurysm. The A1 diameter ratio and the A2 diameter were not statistically significant (OR 1.00, p = 0.99, and OR 3.38, p = 0.25 respectively). CONCLUSIONS: In our study, we focused on asymmetry in the parent and daughter vessels as well as traditional ACoA aneurysm morphological characteristics. We were able to label younger patient age and a greater size ratio as independent prognostic factors for ACoA aneurysm rupture. We were unable to label parent and daughter vessel asymmetry as prognostic factors. To validate our findings, parent and daughter vessel asymmetry should be subjected to future prospective studies.

Improving Trip- and Slip-Resisting Skills in Older People: Perturbation Dose Matters.

Aging negatively affects balance recovery responses after trips and slips. We hypothesize that older people can benefit from brief treadmill-based trip and slip perturbation exposure despite reduced muscular capacities, but with neuropathology, their responsiveness to these perturbations will be decreased. Thus, to facilitate long-term benefits and their generalizability to everyday life, one needs to consider the individual threshold for perturbation dose.

Retention and generalizability of balance recovery response adaptations from trip perturbations across the adult life span.

For human locomotion, varying environments require adjustments of the motor system. We asked whether age affects gait balance recovery adaptation, its retention over months, and the transfer of adaptation to an untrained reactive balance task. Healthy adults (26 young, 27 middle-aged, and 25 older; average ages 24, 52, and 72 yr, respectively) completed two tasks. The primary task involved treadmill walking: either unperturbed (control; n = 39) or subject to unexpected trip perturbations (training; n = 39). A single trip perturbation was repeated after a 14-wk retention period. The secondary transfer task, before and after treadmill walking, involved sudden loss of balance in a lean-and-release protocol. For both tasks, the anteroposterior margin of stability (MoS) was calculated at foot touchdown. For the first (i.e., novel) trip, older adults required one more recovery step (P = 0.03) to regain positive MoS compared with younger, but not middle-aged, adults. However, over several trip perturbations, all age groups increased their MoS for the first recovery step to a similar extent (up to 70%) and retained improvements over 14 wk, although a decay over time was found for older adults (P = 0.002; middle-aged showing a tendency for decay: P = 0.076). Thus, although adaptability in reactive gait stability control remains effective across the adult life span, retention of adaptations over time appears diminished with aging. Despite these robust adaptations, the perturbation training group did not show superior improvements in the transfer task compared with age-matched controls (no differences in MoS changes), suggesting that generalizability of acquired fall-resisting skills from gait-perturbation training may be limited.NEW & NOTEWORTHY The human neuromotor system preserves its adaptability across the adult life span. However, although adaptability in reactive gait stability control remains effective as age increases, retention of recovery response adaptations over time appears to be reduced with aging. Furthermore, acquired fall-resisting skills from single-session perturbation training seem task specific, which may limit the generalizability of such training to the variety of real-life falls.

Tellurium notebooks-An environment for reproducible dynamical modeling in systems biology.

The considerable difficulty encountered in reproducing the results of published dynamical models limits validation, exploration and reuse of this increasingly large biomedical research resource. To address this problem, we have developed Tellurium Notebook, a software system for model authoring, simulation, and teaching that facilitates building reproducible dynamical models and reusing models by 1) providing a notebook environment which allows models, Python code, and narrative to be intermixed, 2) supporting the COMBINE archive format during model development for capturing model information in an exchangeable format and 3) enabling users to easily simulate and edit public COMBINE-compliant models from public repositories to facilitate studying model dynamics, variants and test cases. Tellurium Notebook, a Python-based Jupyter-like environment, is designed to seamlessly inter-operate with these community standards by automating conversion between COMBINE standards formulations and corresponding in-line, human-readable representations. Thus, Tellurium brings to systems biology the strategy used by other literate notebook systems such as Mathematica. These capabilities allow users to edit every aspect of the standards-compliant models and simulations, run the simulations in-line, and re-export to standard formats. We provide several use cases illustrating the advantages of our approach and how it allows development and reuse of models without requiring technical knowledge of standards. Adoption of Tellurium should accelerate model development, reproducibility and reuse.

Rapid, Quantitative, and Ultrasensitive Point-of-Care Testing: A Portable SERS Reader for Lateral Flow Assays in Clinical Chemistry.

The design of a portable Raman/SERS-LFA reader with line illumination using a custom-made fiber optic probe for rapid, quantitative, and ultrasensitive point-of-care testing (POCT) is presented. The pregnancy hormone human chorionic gonadotropin (hCG) is detectable in clinical samples within only 2-5 s down to approximately 1.6 mIU mL-1 . This acquisition time is several orders of magnitude shorter than those of existing approaches requiring expensive Raman instrumentation, and the method is 15-times more sensitive than a commercially available lateral flow assay (LFA) as the gold standard. The SERS-LFA technology paves the way for affordable, quantitative, and ultrasensitive POCT with multiplexing potential in real-world applications, ranging from clinical chemistry to food and environmental analysis as well as drug and biowarfare agent testing.

Hollow modular anchorage (HMA) screws for anterior transvertebral fixation in high-grade spondylolisthesis cases requiring 360 degrees in-situ fusion.

OBJECTIVE: 360 degrees in-situ fusion for high-grade spondylolisthesis showed satisfying clinical long-term results. Combining anterior with posterior surgery increases fusion rates. Anteriorly inserted transvertebral HMA screws could be an alternative to strut graft constructs or cages, avoiding donor site complications. In addition, complete posterior muscle detachment is avoided and the injury risk of neural structures is minimized. This study investigates the use of HMA screws in this context. MATERIAL AND METHODS: Five consecutive patients requiring L4-S1 in-situ fusion for isthmic spondylolisthesis (four Grade 3 and one Grade 4) were included. The L5/S1 level was fused with an HMA screw filled with local bone and bone morphogenic protein (BMP2), inserted via the L4/5 disc space level. An L4/5 stand-alone interbody fusion with additional minimal invasive posterior screw fixation was added. RESULTS: Transvertebral insertion of the HMA device was accomplished via a retroperitoneal approach to L4/L5 in all cases without exposure of L5/S1. Blood loss ranged from 150 ml-350 ml. No intraoperative complication occurred. One patient developed posterior wound infection requiring debridement. Solid fusion was confirmed with a CT scan after 6 months in all patients. All patients improved to unrestricted activities of daily living with two being limited by occasional back pain. CONCLUSIONS: HMA screws allow for effective lumbosacral fusion via a limited anterior exposure. This is technically easier than posterior exposure of the lumbosacral junction in high-grade spondylolisthesis requiring 360 degrees fusion.

The safe use of long screws in L5/S1 stand-alone anterior interbody fusion for olisthesis cases.

BACKGROUND: Stand-alone anterior interbody fusion (STALIF) with poly-ether-ether-ketone (PEEK) cages could offer a treatment option in olisthesis cases. The fixation of the PEEK-cage at the L5/S1 inferior endplate with long divergent screws however might endanger neural sacral structures, especially the S1 nerve root. If shorter screws are used, the achieved bony purchase might not be sufficient to resist the pull out and shear forces at the lumbosacral junction. The aim of the present investigation was to evaluate the use of long screws in PEEK-cages for olisthesis cases at the L5/S1 segment and its neurological complications. MATERIAL AND METHODS: 11 Patients (6 males) with a mean age of 47(± 15.1) years between 2013-2015 designated for an STALIF at the L5/S1 level were consecutively included in the present study. All patients had a Grade 1 or 2 slippage according to Meyerding. PEEK cages (SynFix-LR®, Depuy Synthes) were implanted with 30mm screws at the baseplate of L5/S1 in all patients. Perioperative and postoperative long-term complications were evaluated. Furthermore, radiological follow-up was performed (CT-scan at 6 months, X-ray at 6, 12 and 24 months). RESULTS: 6 patients suffered from an isthmic, 1 from a degenerative olisthesis. 4 patients with iatrogenic spondylolisthesis were included. Pre-operative radiculopathy was noted in 10 patients. 9 patients with pre-operative radiculopathy showed relief of symptoms until the last follow-up after 24 months. Fusion was achieved in all patients after 6 months. No screw-displacement, breakage or violation of the neural foramen was noted. Furthermore, no implant failure or pull-out fractures were seen. CONCLUSION: In this investigation, no complication due to the use of long divergent locking screws was noted. In addition, the majority of patients showed permanent relief of radiculopathy symptoms at the 2 years follow-up. This study demonstrates the safe usage of long divergent locking screws in the baseplate of L5/S1 in anterior interbody fusion at the L5/S1 level.

Efficiency and predictive parameters of outcome of a multimodal pain management concept with spinal injections in patients with low back pain: a retrospective study of 445 patients.

Low back pain is one of the most common diseases of modern civilization. Multimodal pain management (MPM) represents a central approach to avoiding surgery. Short-term results are published rarely and often incomparable because of different treatment concepts. This study compared the subjective and objective parameters as well as the anamnestic and clinical parameters of 445 patients with low back pain before and after inpatient MPM to investigate the influence of this type of therapy on short-term outcome. The majority of patients were very satisfied (39%) or satisfied (58%) with the treatment outcome. The median pain reduction for back pain was 3.0 (IQR 2.88) (numeric rating scale, NRS), thus 66% and 2.75 (IQR 3.38, 62%) for leg pain. The main pain reduction occurred within the first 10 days of treatment and was clinically significant from day 5 onwards. The outcome for patients with hospitalization of more than 10 days was significantly worse. The parameters female sex, BMI of > 30, local pain, and pain duration of 3-24 months had a significantly better outcome. In contrast, age, treatment cause, depression, anxiety, and other diseases had no statistically significant influence on outcome. MPM therapy for more than 5 days seems to be an efficient short-term approach to treating low back pain. Knowledge of some of the outcome predictors helps to early identify patients who require more intensive individual care. In the case of no clear indication for surgery, MPM can be an appropriate treatment option.

Fast and reproducible iSERS microscopy of single HER2-positive breast cancer cells using gold nanostars as SERS nanotags.

Speed is often a bottleneck in conventional Raman microscopy on biological specimens. In immuno-Raman microspectroscopy, or for short iSERS microscopy, the acquisition times per pixel have been reduced by more than one order of magnitude over the past decade since its proof of concept. Typically rather high laser power densities are employed with the intention of compensating for the shorter acquisition times, without checking the reproducibility of the results in repeated experiments on the same sample. Here, we systematically analyze this aspect at the single-cell level since it forms the basis of quantification and is very important for reinspection of the same specimen. Specifically, we investigate experimentally the role of the laser power density in conjunction with the acquisition times per pixel in a series of repeated iSERS experiments on the same single cell overexpressing the breast cancer tumor marker human epidermal growth factor receptor 2 (HER2). Confocal iSERS mapping experiments were guided by wide-field fluorescence microscopy for selecting the regions of interest. We demonstrate that the combination of ca. a 1-2 mW laser power (40× objective, NA 0.6), 50 ms acquisition time per pixel and a high EM-CCD signal gain yields highly reproducible iSERS images in a series of four repeated experiments on the same single cell. In contrast, longer acquisition times (0.8 s, no EM gain) and in particular higher laser power (4 mW up to 18 mW) densities lead to non-reproducible iSERS results due to signal degradation.