Improved immunoassay sensitivity and specificity using single-molecule colocalization

Enzyme-linked immunosorbent assays (ELISAs) are a cornerstone of modern molecular detection, but the technique still suffers some notable challenges. One of the biggest problems is discriminating true signal generated by target molecules versus non-specific background arising from the interaction of detection antibodies with the assay substrate or interferents in the sample matrix. Single-Molecule Colocalization Assay (SiMCA) overcomes this problem by employing total internal reflection fluorescence (TIRF) microscopy to quantify target proteins based on the colocalization of fluorescent signal from orthogonally labeled capture and detection antibodies. By specifically counting colocalized fluorescent signals, we can essentially eliminate the confounding effects of background produced by non-specific binding of detection antibodies. We further employed a normalization strategy to account for the heterogeneous distribution of the capture antibodies, greatly improving the reproducibility of our measurements. In a series of experiments with TNF-α, we show that SiMCA can achieve a three-fold lower limit of detection compared to conventional single-color assays using the same antibodies and exhibits consistent performance for assays performed in complex specimens such as chicken serum and human blood. Our results help define the pernicious effects of non-specific background in immunoassays and demonstrate the diagnostic gains that can be achieved by eliminating those effects.

Numerical Algorithms for Simulation of a Fluid-Filed Fracture Evolution in a Poroelastic Medium

The paper deals with the computational framework for the numerical simulation of the three dimensional fluid-filled fracture evolution in a poroelastic medium. The model consists of several groups of equations including the Biot poroelastic model to describe a bulk medium behavior, Reynold’s lubrication equations to describe a flow inside fracture and corresponding bulk/fracture interface conditions. The geometric model of the fracture assumes that it is described as an arbitrary sufficiently smooth surface with a boundary. Main attention is paid to describing numerical algorithms for particular problems (poroelasticity, fracture fluid flow, fracture evolution) as well as an algorithm for the coupled problem solution. An implicit fracture mid-surface representation approach based on the closest point projection operator is a particular feature of the proposed algorithms. Such a representation is used to describe the fracture mid-surface in the poroelastic solver, Reynold’s lubrication equation solver and for simulation of fracture evolutions. The poroelastic solver is based on a special variant of X-FEM algorithms, which uses the closest point representation of the fracture. To solve Reynold’s lubrication equations, which model the fluid flow in fracture, a finite element version of the closet point projection method for PDEs surface is used. As a result, the algorithm for the coupled problem is purely Eulerian and uses the same finite element mesh to solve equations defined in the bulk and on the fracture mid-surface. Finally, we present results of the numerical simulations which demonstrate possibilities of the proposed numerical techniques, in particular, a problem in a media with a heterogeneous distribution of transport, elastic and toughness properties.

An Experimental and Numerical Study of Polyelectrolyte Hydrogel Ionic Diodes: Towards Electrical Detection of Charged Biomolecules

Polyelectrolyte hydrogel ionic diodes (PHIDs) have recently emerged as a unique set of iontronic devices. Such diodes are built on microfluidic chips that feature polyelectrolyte hydrogel junctions and rectify ionic currents owing to the heterogeneous distribution and transport of ions across the junctions. In this paper, we provide the first account of a study on the ion transport behavior of PHIDs through an experimental investigation and numerical simulation. The effects of bulk ionic strength and hydrogel pore confinement are experimentally investigated. The ionic current rectification (ICR) exhibits saturation in a micromolar regime and responds to hydrogel pore size, which is subsequently verified in a simulation. Furthermore, we experimentally show that the rectification is sensitive to the dose of immobilized DNA with an exhibited sensitivity of 1 ng/μL. We anticipate our findings would be beneficial to the design of PHID-based biosensors for electrical detection of charged biomolecules.

Integrin αE(CD103)β7 in Epithelial Cancer

Interactions of both the innate and the adaptive immune system with tumors are complex and often influence courses and therapeutic treatments in unanticipated ways. Based on the concept that CD8+T cells can mediate important antitumor effects, several therapies now aim to amplify their specific activity. A subpopulation of CD8+ tissue-resident T lymphocytes that express the αE(CD103)β7 integrin has raised particular interest. This receptor presumably contributes to the recruitment and retention of tumor-infiltrating immune cells through interaction with its ligand, E-cadherin. It appears to have regulatory functions and is thought to be a component of some immunological synapses. In TGF-rich environments, the αE(CD103)β7/E-cadherin-interaction enhances the binding strength between tumor cells and infiltrating T lymphocytes. This activity facilitates the release of lytic granule contents and cytokines as well as further immune responses and the killing of target cells. Expression of αE(CD103)β7 in some tumors is associated with a rather favorable prognosis, perhaps with the notable exception of squamous cell carcinoma of the skin. Although epithelial skin tumors are by far the most common tumors of fair-skinned people, there have been very few studies on the distribution of αE(CD103)β7 expressing cells in these neoplasms. Given this background, we describe here that αE(CD103)β7 is scarcely present in basal cell carcinomas, but much more abundant in squamous cell carcinomas with heterogeneous distribution. Notwithstanding a substantial number of studies, the role of αE(CD103)β7 in the tumor context is still far from clear. Here, we summarize the essential current knowledge on αE(CD103)β7 and outline that it is worthwhile to further explore this intriguing receptor with regard to the pathophysiology, therapy, and prognosis of solid tumors.

Spreading of disturbances in realistic models of transmission grids in dependence on topology, inertia and heterogeneity

The energy transition towards more renewable energy resources (RER) profoundly affects the frequency dynamics and stability of electrical power networks. Here, we investigate systematically the effect of reduced grid inertia, due to an increase in the magnitude of RER, its heterogeneous distribution and the grid topology on the propagation of disturbances in realistic power grid models. These studies are conducted with the DigSILENT PowerFactory software. By changing the power generation at one central bus in each grid at a specific time, we record the resulting frequency transients at all buses. Plotting the time of arrival (ToA) of the disturbance at each bus versus the distance from the disturbance, we analyse its propagation throughout the grid. While the ToAs are found to be distributed, we confirm a tendency that the ToA increases with geodesic distance linearly. Thereby, we can measure an average velocity of propagation by fitting the data with a ballistic equation. This velocity is found to decay with increasing inertia. Characterising each grid by its meshedness coefficient, we find that the distribution of the ToAs depends in more meshed grids less strongly on the grid inertia. In order to take into account the inhomogeneous distribution of inertia, we introduce an effective distance $$r_{\mathrm{eff}}$$ r eff , which is weighted with a factor which strongly depends on local inertia. We find that this effective distance is more strongly correlated with the ToAs, for all grids. This is confirmed quantitatively by obtaining a larger Pearson correlation coefficient between ToA and $$r_{\mathrm{eff}}$$ r eff than with r. Remarkably, a ballistic equation for the ToA with a velocity, as derived from the swing equation, provides a strict lower bound for all effective distances $$r_{\mathrm{eff}}$$ r eff in all power grids. thereby yielding a reliable estimate for the smallest time a disturbance needs to propagate that distance as function of system parameters, in particular inertia. We thereby conclude that in the analysis of contingencies of power grids it may be advisable that system designers and operators use the effective distance $$r_{\mathrm{eff}}$$ r eff , taking into account inhomogeneous distribution of inertia as introduced in Eq. (12), to locate a disturbance. Moreover, our results provide evidence for the importance of the network topology as quantified by the meshedness coefficient $$\beta$$ β .

An Experimental Study Investigating the Impact of Miscible and Immiscible Nitrogen Injection on Asphaltene Instability in Nano Shale Pore Structure

Miscible gas injection has become the most used enhanced oil recovery (EOR) method in the oil and gas industry. The deposition and precipitation of aspahltene during the gas injection process is one of the problems during the oil production process. The asphaltene can deposit and plug the pores, which reduces the permeability in a reservoir; thus, decreasing the oil recovery and increasing the production costs. This research investigates the nitrogen (N2) miscible and immiscible pressure injections on asphaltene instability in shale pore structures . First, a slim-tube was used to determine the minimum miscibility pressure (MMP) of N2to ensure that the effect of both miscible and immiscible gas injection was achievable. Second, filtration experiments were conducted using a specially designed filtration apparatus to investigate the effect of nano pore sizes on asphaltene deposition. Heterogeneous distribution of the filter paper membranes was used in all experiments. The factors studied include miscible/immiscible N2injection and pore size distribution. Visualization tests were conducted to highlight the asphaltene precipitation process over time. The results showed that increasing the pressure increased the asphaltene weight percentage. The miscible N2injection pressure had a significant effect on asphaltene instability. However, the immiscible N2injection pressure had a lower effect on the asphaltene deposition, which resulted in less asphaltene weight percentage. For both miscible/immiscible N2injection pressures, the asphaltene weight percentage increased as the pore size of the filter membranes decreased. Visualization tests showed that after one hour the asphaltene clusters were clearly noticed and suspended in the solvent of heptane, and the asphaltene was fully deposited after 12 hours. Microscopy imaging of filter membranes indicated significant pore plugging from asphaltene, especially for smaller pore sizes.

Species richness of the vascular plants of the Bijagual high Andean forest, Colombia

Background and Aims: The high Andean forest in Colombia corresponds to a plant community located between 2400-3200(-3500) m a.s.l., characterized by the presence of arboreal species with restricted distributions that determine their physiognomy and composition. Nevertheless, these species have been affected by agricultural activity that has generated a loss of connectivity and richness. This research records the species richness, composition, and distributional data for the vascular flora in the high Andean forest of the Bijagual Massif, Bocayá, Colombia. Methods: Sampling was carried out in 27 transects. The specimens were organized under the APG IV classification system, curated and deposited in the UPTC herbarium (Universidad Pedagógica y Tecnológica de Colombia). The number of species, list of taxa, distributions for Colombia, and conservation categories according to the IUCN were noted. Results: We registered 327 species distributed in 151 genera and 69 families; 187 species (56%) were native to the Andean region, 51 (15%) were endemic to the Andes and 24 (7%) were endemic to the Eastern cordillera of the Andes, Colombia. Schefflera bejucosa is a new distributional record for Boyacá. Greigia stenolepis, Hieronyma rufa, Puya goudotiana and Tillandsia pallescens are Near Threatened (NT). Plutarchia guascensis, Quercus humboldtii and Symplocos venulosa are classified as Vulnerable (VU), and Diplostephium oblongifolium and Dunalia trianaei as Endangered (EN). Conclusions: Bijagual is a biological corridor with great potential for biodiversity compared to other high Andean areas in Colombia. Species richness is influenced by other localities near the Massif (Tota and Mampacha), in addition to a possible stochastic and heterogeneous distribution between sampling units. Miconia, Pentacalia, Epidendrum, Elaphoglossum, Tillandsia, and some other genera are species-rich. Clusia alata, Weinmannia balbisiana, Weinmannia rollottii, Ternstroemia cf. camelliifolia, and Brunellia comocladifolia define the arboreal and shrub physiognomy of Bijagual.