Quality control material for the detection of somatic mutations in fixed clinical specimens by next-generation sequencing

Background Targeted next generation sequencing (NGS) technology to assess the mutational status of multiple genes on formalin-fixed, paraffin embedded (FFPE) tumors is rapidly being adopted in clinical settings, where quality control (QC) practices are required. Establishing reliable FFPE QC materials for NGS can be challenging and/or expensive. Here, we established a reliable and cost-effective FFPE QC material for routine utilization in the Ion AmpliSeq™ Cancer Hotspot Panel v2 (CHP2) assay. Methods The performance characteristics of the CHP2 assay were determined by sequencing various cell line mixtures and 55 different FFPE tumors on the Ion Torrent PGM platform. A FFPE QC material was prepared from a mixture of cell lines derived from different cancers, comprising single nucleotide variants and small deletions on actionable genes at different allelic frequencies. Results The CHP2 assay performed with high precision and sensitivity when custom variant calling pipeline parameters where established. In addition, all expected somatic variants in the QC material were consistently called at variant frequencies ranging from 9.1 % (CV = 11.1 %) to 37.9 % (CV = 2.8 %). Conclusions The availability of a reliable and cost-effective QC material is instrumental in assessing the performance of this or any targeted NGS assay that detects somatic variants in fixed solid tumor specimens

Portuguese 16th to Early 18th Century tin Glazed Ceramics Found at the Tagus Estuary Saltpans

UID/NAN/50024/2019 M-ERA-MNT/0002/2015 UIDB/04209/2020 UIDP/04209/2020Salt was extracted from saltpans, in the South shore of Tagus River by evaporation of the salted sea water which penetrates into the estuary. The walls and the bottom of those saltpans were made of local clays to retain the sea water, and ceramic sherds have been used to endure those walls and also to allow people to walk on the saltpans’ walls. Those sherds constitute an example of the ceramics production in Lisbon workshops, starting in the mid-16th century and reaching high levels of quality in the 17th and 18th century. A detailed archaeometric study of 14 selected sherds dated from the 16th to the early 18th century has been made and the conjugation of the information provided by the micro-Raman, XRD and XRF experiments for those sherds and clays collected locally, allowed us the characterisation of the Lisbon ceramics in that period. Three main groups could be identified in the ceramic bodies; all made with locally collected clays of Miocene origin. In the first group, the potters used raw materials from the clay sources located at Santa Catarina area in Lisbon, most probably mixed with Lapa clays, which were fired at a high temperatures, ranging from 850ºC to 950ºC. Gehlenite and quartz are the dominant minerals of these ceramic bodies. In the second and third groups of Lisbon ceramic bodies, made with Prazeres clays, diopside was also detected but with variable amounts of gehlenite. Clay formations mineralogy mainly include kaolinite, muscovite/illite, quartz, calcite, and feldspars (albite and microcline).publishersversionpublishe

Quality control material for the detection of somatic mutations in fixed clinical specimens by next-generation sequencing

Background: Targeted next generation sequencing (NGS) technology to assess the mutational status of multiple genes on formalin-fixed, paraffin embedded (FFPE) tumors is rapidly being adopted in clinical settings, where quality control (QC) practices are required. Establishing reliable FFPE QC materials for NGS can be challenging and/or expensive. Here, we established a reliable and cost-effective FFPE QC material for routine utilization in the Ion AmpliSeq™ Cancer Hotspot Panel v2 (CHP2) assay. Methods: The performance characteristics of the CHP2 assay were determined by sequencing various cell line mixtures and 55 different FFPE tumors on the Ion Torrent PGM platform. A FFPE QC material was prepared from a mixture of cell lines derived from different cancers, comprising single nucleotide variants and small deletions on actionable genes at different allelic frequencies. Results: The CHP2 assay performed with high precision and sensitivity when custom variant calling pipeline parameters where established. In addition, all expected somatic variants in the QC material were consistently called at variant frequencies ranging from 9.1 % (CV = 11.1 %) to 37.9 % (CV = 2.8 %). Conclusions: The availability of a reliable and cost-effective QC material is instrumental in assessing the performance of this or any targeted NGS assay that detects somatic variants in fixed solid tumor specimens

Delphi-Consensus Weights for Ischemic and Bleeding Events to Be Included in a Composite Outcome for RCTs in Thrombosis Prevention

To weight ischemic and bleeding events according to their severity to be used in a composite outcome in RCTs in the field of thrombosis prevention.Using a Delphi consensus method, a panel of anaesthesiology and cardiology experts rated the severity of thrombotic and bleeding clinical events. The ratings were expressed on a 10-point scale. The median and quartiles of the ratings of each item were returned to the experts. Then, the panel members evaluated the events a second time with knowledge of the group responses from the first round. Cronbach's a was used as a measure of homogeneity for the ratings. The final rating for each event corresponded to the median rating obtained at the last Delphi round.Of 70 experts invited, 32 (46%) accepted to participate. Consensus was reached at the second round as indicated by Cronbach's a value (0.99 (95% CI 0.98-1.00)) so the Delphi was stopped. Severity ranged from under-popliteal venous thrombosis (median = 3, Q1 = 2; Q3 = 3) to ischemic stroke or intracerebral hemorrhage with severe disability at 7 days and massive pulmonary embolism (median = 9, Q1 = 9; Q3 = 9). Ratings did not differ according to the medical specialty of experts.These ratings could be used to weight ischemic and bleeding events of various severity comprising a composite outcome in the field of thrombosis prevention

Application of a correlation correction factor in a microarray cross-platform reproducibility study

Background Recent research examining cross-platform correlation of gene expression intensities has yielded mixed results. In this study, we demonstrate use of a correction factor for estimating cross-platform correlations. Results In this paper, three technical replicate microarrays were hybridized to each of three platforms. The three platforms were then analyzed to assess both intra- and cross-platform reproducibility. We present various methods for examining intra-platform reproducibility. We also examine cross-platform reproducibility using Pearson\u27s correlation. Additionally, we previously developed a correction factor for Pearson\u27s correlation which is applicable when X and Y are measured with error. Herein we demonstrate that correcting for measurement error by estimating the disattenuated correlation substantially improves cross-platform correlations. Conclusion When estimating cross-platform correlation, it is essential to thoroughly evaluate intra-platform reproducibility as a first step. In addition, since measurement error is present in microarray gene expression data, methods to correct for attenuation are useful in decreasing the bias in cross-platform correlation estimates

Faraday rotation in graphene

We study magneto--optical properties of monolayer graphene by means of quantum field theory methods in the framework of the Dirac model. We reveal a good agreement between the Dirac model and a recent experiment on giant Faraday rotation in cyclotron resonance. We also predict other regimes when the effects are well pronounced. The general dependence of the Faraday rotation and absorption on various parameters of samples is revealed both for suspended and epitaxial graphene.Comment: 10 pp; v2: typos corrected and references added, v3, v4: small changes and more reference

Natural and human-induced Holocene paleoenvironmental changes on the Guadiana shelf (northern Gulf of Cadiz)

Three contrasting sedimentary environments on the continental shelf off the Guadiana River (northern Gulf of Cadiz) were integrated in a chronological framework and analysed in terms of sedimentology and benthic foraminiferal assemblages to understand the Holocene paleoenvironmental evolution. The analysed environments differ in terms of their depositional regimes and benthic foraminiferal assemblages. However, a dominant fluvial origin of the sand fraction was observed in all three environments. Holocene sedimentary processes were mainly controlled by natural (sea level changes and climate variations) and human-induced processes (e.g. deforestation, agriculture) along four evolutionary stages. The three older stages were mainly influenced by natural processes, such as sea level variations and fluvial inputs, whereas the most recent stage reflects a combination of climatic- and human-induced processes. A deepening of sedimentary environments related to a period of rapid sea level rise, strongly influenced by river discharges occurred from c. 11,500 to c. 10,000 cal. yr BP. A reduction in sediment export to the shelf, as a result of the continuous and rapid sea level rise and enhanced estuary infilling reflects the second stage, from c. 10,000 to c. 5000 cal. yr BP. The beginning of the third stage, from c. 5000 to c. 1500–1000 cal. yr BP, is marked by a sea-level slowdown and the relatively stable climate and environmental conditions. The fourth stage, from c. 1500–1000 cal. yr BP to Recent times, reflects the intensification of human-induced processes and climatic variability in the Guadiana River basin. This stage also reflects modern depositional conditions, with the formation of a proximal prodeltaic wedge and a distal muddy body

Quaternary structure of a G-protein coupled receptor heterotetramer in complex with Gi and Gs

Background: G-protein-coupled receptors (GPCRs), in the form of monomers or homodimers that bind heterotrimeric G proteins, are fundamental in the transfer of extracellular stimuli to intracellular signaling pathways. Different GPCRs may also interact to form heteromers that are novel signaling units. Despite the exponential growth in the number of solved GPCR crystal structures, the structural properties of heteromers remain unknown. Results: We used single-particle tracking experiments in cells expressing functional adenosine A1-A2A receptors fused to fluorescent proteins to show the loss of Brownian movement of the A1 receptor in the presence of the A2A receptor, and a preponderance of cell surface 2:2 receptor heteromers (dimer of dimers). Using computer modeling, aided by bioluminescence resonance energy transfer assays to monitor receptor homomerization and heteromerization and G-protein coupling, we predict the interacting interfaces and propose a quaternary structure of the GPCR tetramer in complex with two G proteins. Conclusions: The combination of results points to a molecular architecture formed by a rhombus-shaped heterotetramer, which is bound to two different interacting heterotrimeric G proteins (Gi and Gs). These novel results constitute an important advance in understanding the molecular intricacies involved in GPCR function

A Cautionary Tale: MARVELS Brown Dwarf Candidate Reveals Itself To Be A Very Long Period, Highly Eccentric Spectroscopic Stellar Binary

We report the discovery of a highly eccentric, double-lined spectroscopic binary star system (TYC 3010-1494-1), comprising two solar-type stars that we had initially identified as a single star with a brown dwarf companion. At the moderate resolving power of the MARVELS spectrograph and the spectrographs used for subsequent radial-velocity (RV) measurements (R ~ <30,000), this particular stellar binary mimics a single-lined binary with an RV signal that would be induced by a brown dwarf companion (Msin(i)~50 M_Jup) to a solar-type primary. At least three properties of this system allow it to masquerade as a single star with a very low-mass companion: its large eccentricity (e~0.8), its relatively long period (P~238 days), and the approximately perpendicular orientation of the semi-major axis with respect to the line of sight (omega~189 degrees). As a result of these properties, for ~95% of the orbit the two sets of stellar spectral lines are completely blended, and the RV measurements based on centroiding on the apparently single-lined spectrum is very well fit by an orbit solution indicative of a brown dwarf companion on a more circular orbit (e~0.3). Only during the ~5% of the orbit near periastron passage does the true, double-lined nature and large RV amplitude of ~15 km/s reveal itself. The discovery of this binary system is an important lesson for RV surveys searching for substellar companions; at a given resolution and observing cadence, a survey will be susceptible to these kinds of astrophysical false positives for a range of orbital parameters. Finally, for surveys like MARVELS that lack the resolution for a useful line bisector analysis, it is imperative to monitor the peak of the cross-correlation function for suspicious changes in width or shape, so that such false positives can be flagged during the candidate vetting process.Comment: 16 pages, 11 figures, 6 table