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Critical to managing the spread of COVID-19 is the ability to diagnose infection and define the acquired immune response across the population. While genomic tests for the novel Several Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) detect the presence of viral RNA for a limited time frame, when the virus is shed in the upper respiratory tract, tests able to define exposure and infection beyond this short window of detectable viral replication are urgently needed. Following infection, antibodies are generated within days, providing a durable read-out and archive of exposure and infection. Several antibody tests have emerged to diagnose SARS-CoV-2. Here we report on a qualified quantitative ELISA assay that displays all the necessary characteristics for high-throughput sample analysis. Collectively, this test offers a quantitative opportunity to define both exposure and levels of immunity to SARS-CoV-2., Competing Interests: Declaration of Competing Interest Galit Alter is a founder of SeromYx., (Copyright © 2020. Published by Elsevier B.V.)

Humoral immunity is key to protection for nearly all licensed vaccines. Yet, the design of vaccines has been more difficult for some of our most deadly killers (e.g. HIV, influenza, Dengue virus, etc.), likely due to our incomplete understanding of the precise immunological mechanisms associated with protection. Humoral immunity is governed both by B-cells and their bi-functional secreted antibodies, all of which have a unique capacity to evolve during an immune response. Current OMIC technologies capture individual features of the humoral immune response, providing a glimpse into humoral components (Fab/Fc/B-cell-omic), but fail to provide a wholistic view of the humoral response as a collective functional arm. Here, we dissect current OMIC strategies reviewing experimental and computational approaches, that if integrated could provide a true systems-level view of the humoral immune response., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Cellular heterogeneity is known to have important effects on signal processing and cellular decision making. To understand these processes, multiple classes of mathematical models have been introduced. The hierarchical population model builds a novel class which allows for the mechanistic description of heterogeneity and explicitly takes into account subpopulation structures. However, this model requires a parametric distribution assumption for the cell population and, so far, only the normal distribution has been employed. Here, we incorporate alternative distribution assumptions into the model, assess their robustness against outliers and evaluate their influence on the performance of model calibration in a simulation study and a real-world application example. We found that alternative distributions provide reliable parameter estimates even in the presence of outliers, and can in fact increase the convergence of model calibration., (Copyright © 2019. Published by Elsevier Ltd.)

Intracerebral injection of brain extracts from Alzheimer's disease (AD) patients into appropriate mouse models was previously found to drastically accelerate the deposition of Aβ amyloid in the recipient animals indicating a prion-like activity. In this study we show that this prion-like activity can be also identified by using a cell culture model of Aβ plaque formation. Analysis of biochemical fractions of AD brain extract indicate that the seeding-activity correlated with the presence of Aβ peptide and Aβ-derived aggregates. In vitro-formed fibrils were also active but their activity was low and depending on the fibril structure and conditions of fibril formation. Our data indicate a conformational basis of the observed seeding effect and suggest the utility of our cell model for further studies on the prion-like activity of AD extracts., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Plaque hemorrhage, inflammation and microvessel density are key determinants of plaque vulnerability in native coronary atherosclerosis (ATS). This study investigates the role of intraplaque hemorrhage (IPH) and its relation with inflammation and microvessels in cardiac allograft vasculopathy (CAV) in posttransplanted patients. Seventy coronary plaques were obtained from 12 patients who died because of CAV. For each patient we collected both native heart and the allograft, at the time of transplantation and autopsy, respectively. Intralesion inflammation, microvessels and IPH were assessed semi-quantitatively. IPH was observed in 21/35 (60%) CAV lesions and in 8/35 (22.9%) native ATS plaques, with a strong association between fibrocellular lesions and IPH (p = 0.0142). Microvessels were detected in 26/35 (74.3%) of CAV lesions with perivascular leakage as sign of endothelial damage in 18/26 (69.2%). IPH was strongly associated with microvessels (p < 0.0001). Inflammation was present in 31/35 (88.6%) of CAV lesions. CAV IPH+ lesions were characterized by presence of both fresh and old hemorrhage in 12/21 (57.1%). IPH, associated with microvessel damage and inflammation, is an important feature of CAV. Fresh and old intralesion hemorrhage suggests ongoing remodeling processes promoting the lesion progression and vulnerability., (© Copyright 2013 The American Society of Transplantation and the American Society of Transplant Surgeons.)

Objective: Exogenous insulin use in patients with type 2 diabetes (DM2) has been associated with an increased risk of cardiovascular events. Through which mechanisms insulin may increase atherosclerotic plaque vulnerability is currently unclear. Because insulin has been suggested to promote angiogenesis in diabetic retinopathy and tumors, we hypothesized that insulin enhances intra-plaque angiogenesis., Methods: An in vitro model of pathological angiogenesis was used to assess the potential of insulin to enhance capillary-like tube formation of human microvascular endothelial cells (hMVEC) into a three dimensional fibrin matrix. In addition, insulin receptor expression within atherosclerotic plaques was visualized in carotid endarterectomy specimens of 20 patients with carotid artery stenosis, using immunohistochemical techniques. Furthermore, microvessel density within atherosclerotic plaques was compared between 68 DM2 patients who received insulin therapy and 97 DM2 patients who had been treated with oral glucose lowering agents only., Results: Insulin, at a concentration of 10(-8)M, increased capillary-like tube formation of hMVEC 1.7-fold (p<0.01). Within human atherosclerotic plaques, we observed a specific distribution pattern for the insulin receptor: insulin receptor expression was consistently higher on the endothelial lining of small nascent microvessels compared to more mature microvessels. There was a trend towards an increased microvessel density by 20% in atherosclerotic plaques derived from patients using insulin compared to plaques derived from patients using oral glucose lowering agents only (p=0.05)., Conclusion: Exogenous insulin use in DM2 patients may contribute to increased plaque vulnerability by stimulating local angiogenesis within atherosclerotic plaques., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Aims: Coronary thrombotic occlusion in ST-segment elevation myocardial infarction (STEMI) patients is often preceded by episodes of progressive growth of the thrombus mass. Similar to wound healing, the organization of thrombus could depend on ingrowth of microvessels in order to stabilize its structure. We investigated the patterns of neovascularization in different stages of coronary thrombus evolution., Material and Methods: Thrombectomy materials obtained from STEMI patients were histologically classified according to thrombus age in three groups: fresh (< 1 day), lytic (1-5 days) or organized (> 5 days) thrombi. Forty thrombi of each group were randomly collected. Neovascularization in the thrombi was evaluated histomorphologically and with immunodouble stains to visualize various differentiation antigens of endothelial cells (ECs) and primitive cells., Results: Morphologically, ECs in the coronary thrombi manifested as: single cells, cell clusters or microvessels. CD31+/CD34+ ECs were present in 98% of all the thrombi. In addition, endothelial clusters were found in 63% of the fresh thrombi (< 1 day). CD105+, Ki67+, or C-kit+ ECs (active, proliferating cells) were observed in all the stages, but significantly more in organized thrombi (> 5 days) compared with fresh and lytic ones (< 5 days), and mainly as cell clusters (P ≤ 0.05 for all). CD133+ primitive cells were found only sporadically in 11% of all the samples., Conclusion: EC proliferation is initiated very early, and gradually progresses during the organization process of thrombus after coronary plaque disruption, with only a limited contribution of primitive cells in this process., (© 2012 International Society on Thrombosis and Haemostasis.)

Idiopathic pulmonary fibrosis constitutes the most devastating form of fibrotic lung disorders and remains refractory to current therapies. The coagulation cascade is frequently activated during pulmonary fibrosis, but this observation has so far resisted a mechanistic explanation. Recent data suggest that protease-activated receptor (PAR)-2, a receptor activated by (among others) coagulation factor (F)Xa, plays a key role in fibrotic disease; consequently, we assessed the role of PAR-2 in the development of pulmonary fibrosis in this study. We show that PAR-2 is up-regulated in the lungs of patients with idiopathic pulmonary fibrosis and that bronchoalveolar lavage fluid from these patients displays increased procoagulant activity that triggers fibroblast survival. Using a bleomycin model of pulmonary fibrosis, we show that bleomycin induces PAR-2 expression, as well as both myofibroblast differentiation and collagen synthesis. In PAR-2-/- mice, both the extent and severity of fibrotic lesions are reduced, whereas myofibroblast differentiation is diminished and collagen expression is decreased. Moreover, fibrin deposition in the lungs of fibrotic PAR-2-/- mice is reduced compared with wild-type mice due to differential tissue factor expression in response to bleomycin. Taken together, these results suggest an important role for PAR-2 in the development of pulmonary fibrosis, and the inhibition of the PAR-2-coagulation axis may provide a novel therapeutic approach to treat this devastating disease.

Modelling of pollen dispersal and cross-pollination is of great importance for the ongoing discussion on thresholds for the adventitious presence of genetically modified material in food and feed. Two different modelling approaches for pollen dispersal are used to simulate the cross-pollination rate of pollen emerged from an adjacent transgenic crop field. The models are applied to cross-pollination data from field experiments with transgenic maize (Zea mays). The data were generated by an experimental setup specifically designed to suit the demands of mathematical modelling. First a Gaussian plume model is used for the simulation of pollen transport in and from plant canopies. This is a semiempirical approach combining the atmospheric diffusion equation and Lagrangian methodology. The second model is derived from the localised near field (LNF) theory and based on the physical processes in the canopy. Both modelling approaches prove to be appropriate for the simulation of the cross-pollination rates at distances of about 7.5m and more from the transgene source. The simulation of the cross-pollination rate is less precise at the edge of the source plot especially with the LNF theory. However, the simulation results lie within the range of variability of the observations. Concluding can be pointed out that both models might be adapted to other pollen dispersal experiments of different crops and plot sizes.

Atherosclerotic plaques contain inflammation, composed largely of macrophages and lymphocytes. A proportion of lymphocytes shows signs of activation, but the question arises whether they are activated in an antigen specific way. The expression of costimulatory molecules-receptors that provide accessory signals during antigen-specific activation is a prerequisite for such a condition. This aspect of inflammation in atherosclerotic lesions has not been investigated. Human arterial segments with diffuse intimal thickening, fatty streaks and atherosclerotic plaques were studied with immuno-single and double staining methods. Macrophages and T lymphocytes were stained with CD68 and CD3, respectively, and pan-B cell markers CD19 and CD22 were also used. Costimulatory molecules B7-1 and B7-2, together with their common ligand CD28, and CD27 with its ligand CD70, were stained with specific monoclonal antibodies. The results show that most T lymphocytes were CD27 positive and that only a subpopulation of these (5-15%) was positive also for B7-1, CD28 and CD70. Macrophages expressed B7-1, B7-2, CD28 and CD70, while macrophages positive for CD28 and CD70 have not been reported yet. The expression of costimulatory molecules was most pronounced in the superficial layers at the fibrous cap, but decreased towards the lipid core. This study shows, therefore, that atherosclerotic plaques provide costimulatory signals generally accepted as a prerequisite for adequate T cell stimulation. In addition, this study reveals that only approximately 5-15% of the lymphocytes appears actively involved in the inflammatory reaction.

The thoughts and feelings expressed by the prenatal hospitalized client are important considerations for care, yet few investigations of this subject have been reported. A qualitative study using descriptive exploratory methods (a questionnaire and tape recorded interview) addressed these questions using a phenomenological approach. The sample comprised 11 women at 26-38 weeks' gestation. Results revealed experiences of loneliness, boredom, and powerlessness. The findings imply that the emotional needs of this particular group may not be met by current nursing interventions.

The complexity of immune response-associated cells present in normal human skin was recently redefined as the skin immune system (SIS). In the present study, the exact immunophenotypes of lymphocyte subpopulations with their localizations in normal human skin were determined quantitatively. B cells were not found to be present in normal human skin. Lymphocytes were always of T-cell type, and 90% of these T cells were clustered in 1-3 rows around postcapillary venules of the papillary vascular plexus or adjacent to cutaneous appendages. In such perivascular localizations, they were found to differ from their circulating counterparts in three ways. First, skin perivascular cells were found to be approximately evenly distributed over CD4+ inducer and CD8+ suppressor-cytotoxic T-cell subsets (mean CD4/CD8 ratio: papillary layer 0.96, reticular layer 0.99). Second, within the category of CD4+ inducer T cells, most were phenotyped as CD4+, 4B4+ helper inducer T lymphocytes, whereas CD4+, 2H4+ suppressor inducer T lymphocytes were found to be relatively rare (less than 5%). Third, the majority of skin perivascular T cells were activated as they expressed HLA-DR and interleukin 2 receptors. Intraepidermal, directly subepidermal, and other ("free") lymphocytes were mostly of the CD8+ suppressor-cytotoxic T-cell subset but accounted for less than 10% of the total number of lymphocytes. Intraepidermally localized T cells accounted for less than 2% of the total number of lymphocytes present in normal skin. Our results indicate that preferential perivascular localization of activated T lymphocytes is the characteristic of normal human skin. This might be a reflection of continuous antigen recognition upon endothelial cell presentation and/or continuous T cell-mediated endothelial cell activation thereby inducing enhanced antigen clearing by the skin's endothelium.

Simultaneous detection of multiple tissue epitopes with an overlapping distribution pattern by monoclonal antibodies is sometimes needed for routine immunohistological evaluations. Therefore, multistep double and triple immunoenzymatic methods using antibodies from the same species or Ig (sub)class have been developed. Since only commercially available monoclonal antibodies (either unlabelled, biotinylated or as fluorescein conjugate) have been used, the techniques may be regarded as generally applicable. The staining protocol for double staining consists of six incubation steps: (1) unlabelled monoclonal antibody 1; (2) enzyme I-conjugated anti-mouse Ig; (3) normal mouse serum--for blocking; (4) fluoresceinated monoclonal antibody 2; (5) rabbit anti-fluorescein isothiocyanate--employing the fluorochrome as hapten; (6) enzyme II-conjugated anti-rabbit Ig. For enzymes I and II, peroxidase, alkaline phosphatase and beta-galactosidase can be applied; excellent results were obtained with the following colour combinations: peroxidase activity in red/alkaline phosphatase in blue and beta-galactosidase in green/alkaline phosphatase in violet. Moreover, this double staining method can be extended to provide an immunoenzyme triple staining technique by mixing biotinylated monoclonal antibody 3 and avidin-biotin enzyme III complex with the steps 4 and 5 reagents, respectively. In this way three tissue epitopes can simultaneously be detected clearly and selectively in green (beta-galactosidase), blue (alkaline phosphatase) and red (peroxidase).