Objective comparison of particle tracking methods

Particle tracking is of key importance for quantitative analysis of intracellular dynamic processes from time-lapse microscopy image data. Because manually detecting and following large numbers of individual particles is not feasible, automated computational methods have been developed for these tasks by many groups. Aiming to perform an objective comparison of methods, we gathered the community and organized an open competition in which participating teams applied their own methods independently to a commonly defined data set including diverse scenarios. Performance was assessed using commonly defined measures. Although no single method performed best across all scenarios, the results revealed clear differences between the various approaches, leading to notable practical conclusions for users and developers

A Solve-RD ClinVar-based reanalysis of 1522 index cases from ERN-ITHACA reveals common pitfalls and misinterpretations in exome sequencing

Purpose Within the Solve-RD project (https://solve-rd.eu/), the European Reference Network for Intellectual disability, TeleHealth, Autism and Congenital Anomalies aimed to investigate whether a reanalysis of exomes from unsolved cases based on ClinVar annotations could establish additional diagnoses. We present the results of the “ClinVar low-hanging fruit” reanalysis, reasons for the failure of previous analyses, and lessons learned. Methods Data from the first 3576 exomes (1522 probands and 2054 relatives) collected from European Reference Network for Intellectual disability, TeleHealth, Autism and Congenital Anomalies was reanalyzed by the Solve-RD consortium by evaluating for the presence of single-nucleotide variant, and small insertions and deletions already reported as (likely) pathogenic in ClinVar. Variants were filtered according to frequency, genotype, and mode of inheritance and reinterpreted. Results We identified causal variants in 59 cases (3.9%), 50 of them also raised by other approaches and 9 leading to new diagnoses, highlighting interpretation challenges: variants in genes not known to be involved in human disease at the time of the first analysis, misleading genotypes, or variants undetected by local pipelines (variants in off-target regions, low quality filters, low allelic balance, or high frequency). Conclusion The “ClinVar low-hanging fruit” analysis represents an effective, fast, and easy approach to recover causal variants from exome sequencing data, herewith contributing to the reduction of the diagnostic deadlock

Environmental analysis of polar and non-polar Polycyclic Aromatic Compounds in airborne particulate matter, settled dust and soot: Part II: Instrumental analysis and occurrence

Interests in PAHs and their derivatives (NPAHs, OPAHs, Azaarenes and PASHs) have been growing because of their toxicity. The second part of this review gathers information on the separation and detection of Polycyclic Aromatic Compounds (PACs) and on their occurrence levels in airborne particulate matter, dust and soot. Chromatography is used to separate PACs before their identification and quantification. For both GC and LC, the choice of the stationary phase is crucial to obtain good resolution of PACs, which can be difficult when a lot of compounds are included in the same analysis. Mass Spectrometry is ideal for PACs detection. It can be hyphenated to both GC and LC, is applicable to all subclasses of PACs and its sensitivity and specificity enables environmental assessment of ultratrace levels. PACs are generally around the ng•m-3 level in atmospheric PM and at several µg•g-1 in dust and soot. Some geographical and seasonal trends of their occurrence can be highlighted

Revealing the potential of capillary electrophoresis/mass spectrometry: the tipping point

International audienc

Analysis of Electromagnetic Waves Spatio-Temporal Variability in the Context of Exposure to Mobile Telephony Base Station

International audienceWith the increasing number of mobile phone users, new services and mobile applications, the proliferation of radio antennas has raised concerns about human exposure to electromagnetic waves. This is now a challenging topic to many stakeholders such as local authorities, mobile phone operators, citizen and consumer groups. The study of the spatial and temporal variability of the actual downlink exposure is a very important requirement to find an accurate exposure assessment. In this paper, a concept of exposure areas linked to specific variations of the electric field is introduced. Then a measurement campaign of the temporal variability of the electric field in urban environment is presented, considering different technologies and mobile operators in the previously defined exposure areas. This study allowed to determine updated daytime and nighttime exposure profiles. A second result yielded the averaging duration needed to reach a stable evaluation of the electric field exposure levels, inside each exposure area and according to each technology

Analysis of Electric field spatial variability in simulations of electromagnetic waves exposure to mobile telephony base stations

International audience—this paper focuses on the study of electric field spatial variability in the context of mobile telephony base stations exposure. Electric field is computed using a Uniform Theory of Diffraction (UTD) based simulation technique suitable for large urban areas. Two complementary approaches of spatial variability are proposed here. One based upon spatial autocorrelation and the other one on statistical laws identification to account for electric field distribution in an urban area. The first approach allows us to quantify the spatial dependency of electric field in three representative areas of exposure. The second one demonstrates that statistical laws depending of the area type and urban typology can represent the electric field behavior

The use of pseudo‐MRM for a sensitive and selective detection and quantification of polycyclic aromatic compounds by tandem mass spectrometry

Rationale: Multiple Reaction Monitoring (MRM) is a sensitive and selective detection mode for target trace-level analysis. However, it requires the fragmentation of labile bonds which are not present in molecules such as Polycyclic Aromatic Hydrocarbons (PAHs) and their heterocyclic derivatives (PANHs, PASHs).Methods: We present the application of an alternative tandem mass spectrometry (MS/MS) mode called "pseudo-MRM" for the GCMS/MS analysis of Polycyclic Aromatic Compounds (PACs). This mode is based on the monitoring of transitions with no mass loss between the precursor and the product ion. Pseudo-MRM peak areas were compared with those of classic MRM on three different mass spectrometers: two triple quadrupoles and an ion trap.Results: For all non-polar PACs studied here (PAHs, PANHs and PASHs), the pseudo-MRM transition was always the most intense. The classic MRM transitions exhibited peak areas 2 to 5 times lower. On the contrary, for the functionalized PACs (oxygenated and nitrated PAHs), classic MRM was favored over pseudo-MRM. These observations were confirmed on two triple quadrupoles (QqQs), and the real-world applicability of pseudo-MRM on QqQs was validated by the successful analysis of Diesel PM. However, a comparison with an ion trap showed that pseudo-MRM was never favored on that instrument, which caused fragmentation of non-polar PACs in MS/MS.Conclusions: The results of this study show an important gain in sensitivity when using pseudo-MRM instead of MRM for non-polar PACs on QqQ instruments. The selectivity of MRM is preserved in pseudo-MRM by applying non-zero collision energies to which only these non-polar PACs are resistant, not the isobaric interferences. No interference issue was observed when analyzing Diesel PM, a complex matrix, with our pseudo-MRM method. Therefore, we advise for a broader use of this MS/MS mode for trace-level determination of non-polar PAHs

Simultaneous Determination of 79 Polar and Non-Polar Polycyclic Aromatic Compounds in Airborne Particulate Matter by Gas Chromatography – Tandem Mass Spectrometry

Research on Polycyclic Aromatic Compounds (PACs) in the atmospheric environment has been mainly focused on the 16 EPA’s PAHs. Due to toxicological relevance and particular emission sources, there is a research need in broadening the scope of PACs analysis to include polar and semi-polar PACs in new analytical methods. Here, we describe a single GC-MS/MS method to simultaneously determine 79 polar, semi-polar and non-polar PACs. Temperature gradient and injection conditions were optimized. Optimal Multiple Reaction Monitoring (MRM) and pseudo-MRM mass transitions were searched to enhance the detection. PACs were extracted from particulate samples by Pressurized Liquid Extraction (PLE). The choice of the optimal extraction solvent was carefully evaluated. Instrumental LOQ were from 1 to 2 ng. The method was validated against linearity of the calibration, intra-day and inter-day variability, LOQ and recoveries. The analysis of a Diesel Particulate Matter Certified Reference Material enabled a firm method validation. The method was applied to PM10 samples from a residential parking lot. PACs total amount varied from 2.3 to 14 ng/m3. The most abundant PACs were phthalic anhydride, 1,8-naphthalic anhydride and benzo(b)fluoranthene. PAHs diagnostic ratios enabled the identification of fuel combustion and vehicular traffic as the main source of PACs.</p

Monoclonal antibody N-glycosylation profiling using capillary electrophoresis – Mass spectrometry: Assessment and method validation

Characterization of therapeutic proteins represents a major challenge for analytical sciences due to their heterogeneity caused by post-translational modifications (PTM). Among these PTM, glycosylation which is possibly the most prominent, require comprehensive identification because of their major influence on protein structure and effector functions of monoclonal antibodies (mAbs). As a consequence, glycosylation profiling must be deeply characterized. For this application, several analytical methods such as separation-based or MS-based methods, were evaluated. However, no CE-ESI-MS approach has been assessed and validated. Here, we illustrate how the use of CE-ESI-MS method permits the comprehensive characterization of mAbs N-glycosylation at the glycopeptide level to perform relative quantitation of N-glycan species. Validation of the CE-ESI-MS method in terms of robustness and reproducibility was demonstrated through the relative quantitation of glycosylation profiles for ten different mAbs produced in different cell lines. Glycosylation patterns obtained for each mAbs were compared to Hydrophilic Interaction Chromatography of 2-aminobenzamide labelled glycans with fluorescence detector (HILIC-FD) analysis considered as a reference method. Very similar glycoprofiling were obtained with the CE-ESI-MS and HILIC-FD demonstrating the attractiveness of CE-ESI-MS method to characterize and quantify the glycosylation heterogeneity of a wide range of therapeutic mAbs with high accuracy and precisio