1,381 research outputs found

    Enhancement of toxin- and virus-neutralizing capacity of single-domain antibody fragments by N-glycosylation

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    Single-domain antibody fragments (VHHs) have several beneficial properties as compared to conventional antibody fragments. However, their small size complicates their toxin- and virus-neutralizing capacity. We isolated 27 VHHs binding Escherichia coli heat-labile toxin and expressed these in Saccharomyces cerevisiae. The most potent neutralizing VHH (LT109) was N-glycosylated, resulting in a large increase in molecular mass. This suggests that N-glycosylation of LT109 improves its neutralizing capacity. Indeed, deglycosylation of LT109 decreased its neutralizing capacity three- to fivefold. We also studied the effect of glycosylation of two previously isolated VHHs on their ability to neutralize foot-and-mouth disease virus. For this purpose, these VHHs that lacked potential N-glycosylation sites were genetically fused to another VHH that was known to be glycosylated. The resulting fusion proteins were also N-glycosylated. They neutralized the virus at at least fourfold-lower VHH concentrations as compared to the single, non-glycosylated VHHs and at at least 50-fold-lower VHH concentrations as compared to their deglycosylated counterparts. Thus, we have shown that N-glycosylation of VHHs contributes to toxin- and virus-neutralizing capacity

    Using continuous-time spatial capture–recapture models to make inference about animal activity patterns

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    This work was part‐funded by EPSRC Grant EP/I000917/1, by the research fellowship RF‐2018‐213/9, and the fieldwork was funded by the Summerlee Foundation and Panthera.1. Quantifying the distribution of daily activity is an important component of behavioral ecology. Historically, it has been difficult to obtain data on activity patterns, especially for elusive species. However, the development of affordable camera traps and their widespread usage has led to an explosion of available data from which activity patterns can be estimated. 2. Continuous-time spatial capture?recapture (CT SCR) models drop the occasion structure seen in traditional spatial and nonspatial capture?recapture (CR) models and use the actual times of capture. In addition to estimating density, CT SCR models estimate expected encounters through time. Cyclic splines can be used to allow flexible shapes for modeling cyclic activity patterns, and the fact that SCR models also incorporate distance means that space-time interactions can be explored. This method is applied to a jaguar dataset. 3. Jaguars in Belize are most active and range furthest in the evening and early morning and when they are located closer to the network of trails. There is some evidence that females have a less variable pattern than males. The comparison between sexes demonstrates how CT SCR can be used to explore hypotheses about animal behavior within a formal modeling framework. 4. SCR models were developed primarily to estimate and model density, but the models can be used to explore processes that interact across space and time, especially when using the CT SCR framework that models the temporal dimension at a finer resolution.Publisher PDFPeer reviewe

    Studies on the Induction of Antigen-Specific Antibody in Anti-CD40 Cultured Human B Lymphocytes

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    Costimulatory signals provided by T cells are required for B cells to produce specific antibody to T-dependent antigen. We have investigated the suitability of using the CD40 culture system for the proliferation and differentiation of Ag-specific human B cells using cytomegalovirus (CMV) or tetanus toxoid (TT) as antigen. We modified the CD40 culture system (CD32- transfected L cells, anti-CD40, and IL-4) by applying a sequential cytokine stimulation and compared total B-cell cultures with antigen-specific B cells preselected by panning. The detection of specific antibody became possible when antigen-selected B cells were cultured for 7 days in the CD40 system to induce clonal expansion, followed by the addition of IL-2 and IL10 for an additional 7 days to induce plasma-cell differentiation. We conclude that our intial inability to detect specific antibody in the CD40 system is due to overgrowth of nonspecific B cell clones and that selection of antigen-specific B cells by panning overcomes this problem. Induction of antigen-specific antibody production was found to be optimal when the initial contact with antigen during panning was limited to between 1 to 24 hours

    Hyaluronic acid-recombinant gelatin gels as a scaffold for soft tissue regeneration

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    An array of different types of hyaluronic acid (HA)- and collagen-based products is available for filling soft-tissue defects. A major drawback of the current soft-tissue fillers is their inability to induce cell infiltration and new tissue formation. Our aim is to develop novel biodegradable injectable gels which induce soft tissue regeneration, initially resulting in integration and finally replacement of the gel with new autologous tissue. Two reference gels of pure HA, monophasic HA-1 and micronised HA-2, were used. Furthermore, both gels were mixed with recombinant gelatin (RG) resulting in HA-1+RG and HA-2+RG. All gels were subcutaneously injected on the back of rats and explanted after 4 weeks. Addition of RG to HA-1 resulted in stroma formation (neovascularisation and ECM deposition) which was restricted to the outer rim of the HA-1+RG gel. In contrast, addition of RG to HA-2 induced stroma formation throughout the gel. The RG component of the gel was degraded by macrophages and giant cells and subsequently replaced by new vascularised tissue. Immunohistochemical staining showed that the extracellular matrix components collagen I and III were deposited throughout the gel. In conclusion, this study shows the proof of principle that addition of RG to HA-2 results in a novel injectable gel capable of inducing soft tissue regeneration. In this gel HA has a scaffold function whereas the RG component induces new tissue formation, resulting in proper vascularisation and integration of the HA-2+RG gel with the autologous tissue
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