To facilitate fine-scale phenotyping of whole specimens, we describe here a set of tissue fixation-embedding, detergent-clearing and staining protocols that can be used to transform excised organs ...and whole organisms into optically transparent samples within 1-2 weeks without compromising their cellular architecture or endogenous fluorescence. PACT (passive CLARITY technique) and PARS (perfusion-assisted agent release in situ) use tissue-hydrogel hybrids to stabilize tissue biomolecules during selective lipid extraction, resulting in enhanced clearing efficiency and sample integrity. Furthermore, the macromolecule permeability of PACT- and PARS-processed tissue hybrids supports the diffusion of immunolabels throughout intact tissue, whereas RIMS (refractive index matching solution) grants high-resolution imaging at depth by further reducing light scattering in cleared and uncleared samples alike. These methods are adaptable to difficult-to-image tissues, such as bone (PACT-deCAL), and to magnified single-cell visualization (ePACT). Together, these protocols and solutions enable phenotyping of subcellular components and tracing cellular connectivity in intact biological networks.
This article reports Australia's first confirmed ancient underwater archaeological sites from the continental shelf, located off the Murujuga coastline in north-western Australia. Details on two ...underwater sites are reported: Cape Bruguieres, comprising > 260 recorded lithic artefacts at depths down to -2.4 m below sea level, and Flying Foam Passage where the find spot is associated with a submerged freshwater spring at -14 m. The sites were discovered through a purposeful research strategy designed to identify underwater targets, using an iterative process incorporating a variety of aerial and underwater remote sensing techniques and diver investigation within a predictive framework to map the submerged landscape within a depth range of 0-20 m. The condition and context of the lithic artefacts are analysed in order to unravel their depositional and taphonomic history and to corroborate their in situ position on a pre-inundation land surface, taking account of known geomorphological and climatic processes including cyclone activity that could have caused displacement and transportation from adjacent coasts. Geomorphological data and radiometric dates establish the chronological limits of the sites and demonstrate that they cannot be later than 7000 cal BP and 8500 cal BP respectively, based on the dates when they were finally submerged by sea-level rise. Comparison of underwater and onshore lithic assemblages shows differences that are consistent with this chronological interpretation. This article sets a foundation for the research strategies and technologies needed to identify archaeological targets at greater depth on the Australian continental shelf and elsewhere, building on the results presented. Emphasis is also placed on the need for legislation to better protect and manage underwater cultural heritage on the 2 million square kilometres of drowned landscapes that were once available for occupation in Australia, and where a major part of its human history must lie waiting to be discovered.
The widespread occurrence of antibiotics as contaminants in the aquatic environment has increased attention in the last years. The concern over the release of antibiotics into the environment is ...related primarily to the potential for the development of antimicrobial resistance among microorganisms. This article presents an overview of analytical methodologies for the determination of quinolone (Qs) and fluoroquinolone (FQs), macrolide (MLs), tetracycline (TCs), sulfonamide (SAs) antibiotics and trimethoprim (TMP) in different environmental waters. The analysis of these antibiotics has usually been carried out by high-performance liquid chromatography (HPLC) coupled to mass spectrometry (MS) or tandem mass spectrometry (MS/MS) and to a lesser extent by ultraviolet (UV) or fluorescence detection (FD). A very important step before LC analysis is sample preparation and extraction leading to elimination of interferences and prevention of matrix effect and preconcentration of target analytes.
The rapid spread of the SARS-CoV-2 virus that caused the COVID-19 disease, has highlighted our urgent need for sensitive, fast and accurate diagnostic technologies. In fact, one of the main ...challenges for flatting COVID-19 spread charts is the ability to accurately and rapidly identify asymptomatic cases that result in spreading the virus to close contacts. SARS-CoV-2 virus mutation is also relatively rapid, which makes the detection of COVID-19 diseases still crucial even after the vaccination. Conventional techniques, which are commercially available have focused on clinical manifestation, along with molecular and serological detection tools that can identify the SARS-CoV-2 virus however, owing to various disadvantages including low specificity and sensitivity, a quick, low cost and easy approach is needed for diagnosis of COVID-19. Scientists are now showing extensive interest in an effective portable and simple detection method to diagnose COVID-19. There are several novel methods and approaches that are considered viable advanced systems that can meet the demands. This study reviews the new approaches and sensing technologies that work on COVID-19 diagnosis for easy and successful detection of SARS-CoV-2 virus.
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•Reviewing all the published manuscripts focusing on COVID-19 detection and monitoring.•Studying and classifying the applied methods for COVID-19 detection and monitoring.•Studying and classifying the biomolecules and analytes that are targeted for COVID-19 detection and monitoring.•Investigation the pros and cons of applied methods and offering new detection and monitoring methods for future studies.
Fiber-Optic Seismology Lindsey, Nathaniel J; Martin, Eileen R
Annual review of earth and planetary sciences,
05/2021, Letnik:
49, Številka:
1
Journal Article
Recenzirano
Odprti dostop
Distributed acoustic sensing (DAS) is an emerging technology that repurposes a fiber-optic cable as a dense array of strain sensors. This technology repeatedly pings a fiber with laser pulses, ...measuring optical phase changes in Rayleigh backscattered light. DAS is beneficial for studies of fine-scale processes over multi-kilometer distances, long-term time-lapse monitoring, and deployment in logistically challenging areas (e.g., high temperatures, power limitations, land access barriers). These benefits have motivated a decade of applications in subsurface imaging and microseismicity monitoring for energy production and carbon sequestration. DAS arrays have recorded microearthquakes, regional earthquakes, teleseisms, and infrastructure signals. Analysis of these wavefields is enabling earthquake seismology where traditional sensors were sparse, as well as structural and near-surface seismology. These studies improved understanding of DAS instrument response through comparison with traditional seismometers. More recently, DAS has been used to study cryosphere systems, marine geophysics, geodesy, and volcanology. Further advancement of geoscience using DAS requires several community efforts related to instrument access, training, outreach, and cyberinfrastructure.
DAS is a seismic acquisition technology repurposing fiber optics as arrays of dynamic strain sensors at 1- to 10-m spacing over kilometers.
Easy DAS installations have availed time-lapse geophysical sensing in formerly inaccessible sites: urban, icy, and offshore areas.
High-frequency wavefields recorded by DAS are analyzed with array-based methods to characterize seismic sources and image the subsurface.
DAS has shown low-frequency sensitivity in the laboratory and field, for slow hydrodynamic and geodynamic processes.
Genetic tools are needed to examine gene expression in the pathogen
Leptospira interrogans
. We developed a reporter plasmid that replicates in
L. interrogans
with green fluorescent protein (GFP) as ...the readout of promoter activity. We demonstrated an application of the new reporter plasmid by identifying an upstream element responsible for the poor basal expression of the
sph2
sphingomyelinase gene in an
L. interrogans
serovar Lai strain. This new tool is useful for the discovery of the molecular determinants of
L. interrogans
gene expression.
Many strains of the spirochete
Leptospira interrogans
serovar Pomona express the osmotically inducible sphingomyelinase gene
sph2
at much higher levels than strains from other serovars. We developed a new green fluorescent protein (GFP) reporter plasmid to examine
sph2
gene expression determinants. The vector enables the fusion of the test promoter to the ribosome-binding site and coding region of
gfp
. We fused the
sph2
promoters from the
L. interrogans
serovar Lai strain 56601 and from the
L. interrogans
serovar Pomona strain LC82-25 to
gfp
to examine the molecular determinants of differential
sph2
expression between the two strains. Similar to what was observed with the native
sph2
genes, the introduction of the plasmids into the Lai 56601 strain resulted in near background levels of
gfp
expression from the Lai
sph2
promoter, while the expression from the Pomona
sph2
promoter was high. The expression of both fusions increased at physiologic levels of osmolarity achieved by adding sodium chloride to the culture medium. We examined the role of a 17-bp upstream element found in all
L. interrogans
strains expressing low basal levels of
sph2
and missing from Pomona strains that express
sph2
at high levels. When the 17-bp sequence present upstream of the Lai
sph2
promoter was deleted or scrambled, the fusion expression increased substantially. Conversely, the insertion of the 17-bp sequence upstream of the Pomona
sph2
promoter diminished fusion expression. In contrast, the removal of an insertion sequence-like element that is found only in the Pomona
sph2
upstream sequence had no effect on the expression from the Pomona
sph2
fusion in the Lai strain. These findings demonstrate the utility of the
gfp
reporter plasmid in analyzing gene expression in
L. interrogans
.
IMPORTANCE
Genetic tools are needed to examine gene expression in the pathogen
Leptospira interrogans
. We developed a reporter plasmid that replicates in
L. interrogans
with green fluorescent protein (GFP) as the readout of promoter activity. We demonstrated an application of the new reporter plasmid by identifying an upstream element responsible for the poor basal expression of the
sph2
sphingomyelinase gene in an
L. interrogans
serovar Lai strain. This new tool is useful for the discovery of the molecular determinants of
L. interrogans
gene expression.
Ionic liquids in separation techniques Berthod, A.; Ruiz-Ángel, M.J.; Carda-Broch, S.
Journal of Chromatography A,
03/2008, Letnik:
1184, Številka:
1
Journal Article
Recenzirano
The growing interest in ionic liquids (ILs) has resulted in an exponentially increasing production of analytical applications. The potential of ILs in chemistry is related to their unique properties ...as non-molecular solvents: a negligible vapor pressure associated to a high thermal stability. ILs found uses in different sub-disciplines of analytical chemistry. After drawing a rapid picture of the physicochemical properties of selected ILs, this review focuses on their use in separation techniques: gas chromatography (GC), liquid chromatography (LC) and electrophoretic methods (CE). In LC and CE, ILs are not used as pure solvents, but rather diluted in aqueous solutions. In this situation ILs are just salts. They are dual in nature. Too often the properties of the cations are taken as the properties of the IL itself. The lyotropic theory is recalled and the effects of a chaotropic anion are pointed out. Many results can be explained considering all ions present in the solution. Ion-pairing and ion-exchange mechanisms are always present, associated with hydrophobic interactions, when dealing with IL in diluted solutions. Chromatographic and electrophoretic methods are also mainly employed for the control and monitoring of ILs. These methods are also considered. ILs will soon be produced on an industrial scale and it will be necessary to develop reliable analytical procedures for their analysis and control.
Adult stem cell-based organoid technology is a versatile tool for the generation and long-term maintenance of near-native 3D epithelial tissues in vitro. The generation of cancer organoids from ...primary patient material enables a range of therapeutic agents to be tested in the resulting organoid cultures. Patient-derived cancer organoids therefore hold great promise for personalized medicine. Here, we provide an overview of the protocols used by different groups to establish organoids from various epithelial tissues and cancers, plus the different protocols subsequently used to test the in vitro therapy sensitivity of these patient-derived organoids. We also provide an in-depth protocol for the generation of head and neck squamous cell carcinoma organoids and their subsequent use in semi-automated therapy screens. Establishment of organoids and subsequent screening can be performed within 3 months, although this timeline is highly dependent on a.o. starting material and the number of therapies tested. The protocol provided may serve as a reference to successfully establish organoids from other cancer types and perform drug screenings thereof.
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•An overview on recent trends in the sample preparation and determination of sulfonamides is given.•A comparison of different methods of real samples preparation is made.•The general ...chromatographic and other methods of SAs determination are discussed.•Examples of SAs determination in different matrices are given.
Sulfonamides (SAs) have been the most widely used antimicrobial drugs for more than 70 years, and their residues in foodstuffs and environmental samples pose serious health hazards. For this reason, sensitive and specific methods for the quantification of these compounds in numerous matrices have been developed. This review intends to provide an updated overview of the recent trends over the past five years in sample preparation techniques and methods for detecting SAs. Examples of the sample preparation techniques, including liquid–liquid and solid-phase extraction, dispersive liquid–liquid microextraction and QuEChERS, are given. Different methods of detecting the SAs present in food and feed and in environmental, pharmaceutical and biological samples are discussed.
We introduce a new dataset, Human3.6M, of 3.6 Million accurate 3D Human poses, acquired by recording the performance of 5 female and 6 male subjects, under 4 different viewpoints, for training ...realistic human sensing systems and for evaluating the next generation of human pose estimation models and algorithms. Besides increasing the size of the datasets in the current state-of-the-art by several orders of magnitude, we also aim to complement such datasets with a diverse set of motions and poses encountered as part of typical human activities (taking photos, talking on the phone, posing, greeting, eating, etc.), with additional synchronized image, human motion capture, and time of flight (depth) data, and with accurate 3D body scans of all the subject actors involved. We also provide controlled mixed reality evaluation scenarios where 3D human models are animated using motion capture and inserted using correct 3D geometry, in complex real environments, viewed with moving cameras, and under occlusion. Finally, we provide a set of large-scale statistical models and detailed evaluation baselines for the dataset illustrating its diversity and the scope for improvement by future work in the research community. Our experiments show that our best large-scale model can leverage our full training set to obtain a 20% improvement in performance compared to a training set of the scale of the largest existing public dataset for this problem. Yet the potential for improvement by leveraging higher capacity, more complex models with our large dataset, is substantially vaster and should stimulate future research. The dataset together with code for the associated large-scale learning models, features, visualization tools, as well as the evaluation server, is available online at http://vision.imar.ro/human3.6m.
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