High-level production and long-term storage of engineered antibodies in transgenic tobacco seeds

We have used transgenic tobacco seeds to produce large amounts of a functionally active engineered antibody. A gene infusion encoding an antigen-binding single chain Fv protein (scFv) that recognizes the hapten oxazolone was constructed and used as a model. After characterization in a bacterial expression system ,the scFv gene was cloned into a plant expression cassette conferring seed specific expression, and transferred using Agrobacterium-mediated transformation, into Nicotiana tabacum. The expressed scFv could be detected in the developing as well as ripe seeds of regenerated transgenic plants, and the functionally active scFv is stabaly deposited and accumulates up to 0.67% of the total soluble seed protein. After storage of ripe transgenic tobacco seeds for one year at room temperature there was no loss of scFv protein or its antigen-binding activity.     

Synthesis of corneal keratan sulfate proteoglycans by bovine keratocytes in vitro

Keratan sulfate proteoglycans (KSPGs) are the major proteoglycans of the cornea and are secreted by keratocytes in the corneal stroma. Previous studies have been able to show only transient secretion of KSPG in cell culture. In this study, cultures of bovine keratocytes were found to secrete the three previously characterized KSPG proteins into culture medium. Reactivity with monoclonal antibody I22 demonstrated substitution of these proteins with keratan sulfate chains. KSPG constituted 15% of the proteoglycan metabolically labeled with [35S]sulfate in keratocyte culture medium. This labeled KSPG contained keratan sulfate chains of 4700 Da compared to 21,000 Da for bovine corneal keratan sulfate. Labeled keratan sulfate from cultures contained nonsulfated, monosulfated, and disulfated disaccharides that were released by digestion with endo-beta-galactosidase or keratanase II. Nonsulfated disaccharides were relatively more abundant in keratan sulfate from culture than in corneal keratan sulfate. These results show that cultured bovine keratocytes maintain the ability to express all three of the known KSPG proteins, modified with keratan sulfate chains and sulfated on both N-acetylglucosamine and galactose moieties. KSPG made in vitro differs from that found in vivo in the length and sulfation of its keratan sulfate chains. The availability of cell cultures secreting corneal keratan sulfate proteoglycans provides an opportunity to examine biosynthesis and control of this important class of molecules.     

A URI-based approach for addressing fragments of media resources on the Web

To make media resources a prime citizen on the Web, we have to go beyond simply replicating digital media files. The Web is based on hyperlinks between Web resources, and that includes hyperlinking out of resources (e.g., from a word or an image within a Web page) as well as hyperlinking into resources (e.g., fragment URIs into Web pages). To turn video and audio into hypervideo and hyperaudio, we need to enable hyperlinking into and out of them. The W3C Media Fragments Working Group is taking on the challenge to further embrace W3C??s mission to lead the World Wide Web to its full potential by developing a Media Fragment protocol and guidelines that ensure the long-term growth of the Web. The major contribution of this paper is the introduction of Media Fragments as a media-format independent, standard means of addressing media resources using URIs. Moreover, we explain how the HTTP protocol can be used and extended to serve Media Fragments and what the impact is for current Web-enabled media formats.     

Complementary Effects of Carbamylated and Citrullinated LL37 in Autoimmunity and Inflammation in Systemic Lupus Erythematosus

LL37 acts as T-cell/B-cell autoantigen in Systemic lupus erythematosus (SLE) and psoriatic disease. Moreover, when bound to “self” nucleic acids, LL37 acts as “danger signal,” leading to type I interferon (IFN-I)/pro-inflammatory factors production. T-cell epitopes derived from citrullinated-LL37 act as better antigens than unmodified LL37 epitopes in SLE, at least in selected HLA-backgrounds, included the SLE-associated HLA-DRB1*1501/HLA-DRB5*0101 backgrounds. Remarkably, while “fully-citrullinated” LL37 acts as better T-cell-stimulator, it loses DNA-binding ability and the associated “adjuvant-like” properties. Since LL37 undergoes a further irreversible post-translational modification, carbamylation and antibodies to carbamylated self-proteins other than LL37 are present in SLE, here we addressed the involvement of carbamylated-LL37 in autoimmunity and inflammation in SLE. We detected carbamylated-LL37 in SLE-affected tissues. Most importantly, carbamylated-LL37-specific antibodies and CD4 T-cells circulate in SLE and both correlate with disease activity. In contrast to “fully citrullinated-LL37,” “fully carbamylated-LL37” maintains both innate and adaptive immune-cells’ stimulatory abilities: in complex with DNA, carbamylated-LL37 stimulates plasmacytoid dendritic cell IFN-α production and B-cell maturation into plasma cells. Thus, we report a further example of how different post-translational modifications of a self-antigen exert complementary effects that sustain autoimmunity and inflammation, respectively. These data also show that T/B-cell responses to carbamylated-LL37 represent novel SLE disease biomarkers.     

Monomer and amino resin modified tall oil fatty acid alkyds

Summary Monomer modified tall oil fatty acid trimethylolethane medium type of phthalic alkyds made by the High Polymer Alkyd Technique show improved properties in dry time, mar resistance, and flexibility as compared with monomer modified conventional prepared systems. Greater hardness, less color degradation, greater resistance to extended periods at high temperature, and alkali and detergent resistances result from the urea and melamine resin modifications of tall oil fatty acid High Polymer alkyds than is obtained with similarly modified conventional alkyds.     

Mössbauer studies of illites and heat-treated illite as related to coal-conversion processes

Mossbauer spectra of iron species in the following illites were studied: Grundite, Fithian, Minerva, and New Albany. Spectra of samples of Fithian illite heated at temperatures of 225, 700, and 1000 °C were also obtained. Analyses of these spectra provide Mössbauer parameter values of iron species in the illites and heat-transformed illite for comparison with similar species found in coals containing illites and in coal process residues derived from them. The illites contain both ferric and ferrous species. Mössbauer parameters for one of the ferric species, designated M(2), are virtually the same as those of pyrite. The two species are therefore difficult to distinguish from one another. Values of the concentration of pyrite in coals and shales may be inflated if the pyrite concentration is measured by Mössbauer spectroscopy. Mössbauer spectra of the heat-treated illite samples reveal changes in iron distribution, principally at the 700 and 1000 °C levels, where there are found three and six different iron species respectively. These changes are accompanied by reduction of ferric to ferrous iron. This process should be integrated into any assessment of the iron chemistry which accompanies coal-conversion processes.     

Compressibility of fine-grained soils subjected to closed-system freezing and thaw consolidation

In order to investigate the effects of freezing and thawing on the compressibility of fine-grained soils, freezing and thawing tests and subsequent compression tests with fine-grained soils in an oedometer were carried out. During the freezing, a part of the soils is loosened and another part is over-consolidated under the freezing pressure ?E. The compression curves after the freezing and thaw consolidation are neither different from the normal consolidation curve nor from the rebound curve of an unfrozen soil, until the consolidation pressure ?z = ?E is arrived. Based on the experimental results, a theoretical model has been developed to predict the frost heaves, the thaw-settlements and the compressive deformations of fine-grained soils after the thaw consolidation. The theoretical results are very close to the experimental results.     

Performance models of storage contention in cloud environments

We propose simple models to predict the performance degradation of disk requests due to storage device contention in consolidated virtualized environments. Model parameters can be deduced from measurements obtained inside Virtual Machines (VMs) from a system where a single VM accesses a remote storage server. The parameterized model can then be used to predict the effect of storage contention when multiple VMs are consolidated on the same server. We first propose a trace-driven approach that evaluates a queueing network with fair share scheduling using simulation. The model parameters consider Virtual Machine Monitor level disk access optimizations and rely on a calibration technique. We further present a measurement-based approach that allows a distinct characterization of read/write performance attributes. In particular, we define simple linear prediction models for I/O request mean response times, throughputs and read/write mixes, as well as a simulation model for predicting response time distributions. We found our models to be effective in predicting such quantities across a range of synthetic and emulated application workloads.