DNA origami nanorobot fiber optic genosensor to TMV

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Computational Analysis of Mouse piRNA Sequence and Biogenesis

The recent discovery of a new class of 30-nucleotide long RNAs in mammalian testes, called PIWI-interacting RNA (piRNA), with similarities to microRNAs and repeat-associated small interfering RNAs (rasiRNAs), has raised puzzling questions regarding their biogenesis and function. We report a comparative analysis of currently available piRNA sequence data from the pachytene stage of mouse spermatogenesis that sheds light on their sequence diversity and mechanism of biogenesis. We conclude that (i) there are at least four times as many piRNAs in mouse testes than currently known; (ii) piRNAs, which originate from long precursor transcripts, are generated by quasi-random enzymatic processing that is guided by a weak sequence signature at the piRNA 5′ends resulting in a large number of distinct sequences; and (iii) many of the piRNA clusters contain inverted repeats segments capable of forming double-strand RNA fold-back segments that may initiate piRNA processing analogous to transposon silencing

Highly sensitive and specific detection of E. coli by a SERS nanobiosensor chip utilizing metallic nanosculptured thin films

A nanobiosensor chip, utilizing surface enhanced Raman spectroscopy (SERS) on nanosculptured thin films (nSTFs) of silver, was shown to detect Escherichia coli (E. coli) bacteria down to the concentration level of a single bacterium. The sensor utilizes highly enhanced plasmonic nSTFs of silver on a silicon platform for the enhancement of Raman bands as checked with adsorbed 4-aminothiophenol molecules. T-4 bacteriophages were immobilized on the aforementioned surface of the chip for the specific capture of target E. coli bacteria. To demonstrate that no significant non-specific immobilization of other bacteria occurs, three different, additional bacterial strains, Chromobacterium violaceum, Paracoccus denitrificans and Pseudomonas aeruginosa were used. Furthermore, experiments performed on an additional strain of E. coli to address the specificity and reusability of the sensor showed that the sensor operates for different strains of E. coli and is reusable. Time resolved phase contrast microscopy of the E. coli-T4 bacteriophage chip was performed to study its interaction with bacteria over time. Results showed that the present sensor performs a fast, accurate and stable detection of E. coli with ultra-small concentrations of bacteria down to the level of a single bacterium in 10 μl volume of the sample

Fiber-optic immunosensor for detection of Crimean-Congo Hemorrhagic fever IgG antibodies in patients

Crimean-Congo hemorrhagic fever (CCHF) is a severe viral disease with high fatality rate. CCHF virus is endemic in parts of Africa, Asia, Middle East and southeastern Europe. Rapid diagnostics of CCHF is vital for appropriate clinical management and prevention of secondary spread from human-to-human. Currently, diagnostics relies on Real-Time RT-PCR and antibody or antigen detection using ELISA. These methods require trained personnel and expensive equipment and are not appropriate for point-of-care (POC) diagnostics. Furthermore there are no POC assays available for CCHF. We developed a fiber-optic biosensor for the detection of CCHF IgG antibodies. In order to improve sensitivity, we optimized both the bioreceptor immobilization protocol and the chemiluminescence substrate formulation. The resulting protocol showed a 100-fold greater sensitivity for detection of CCHF antibodies. Finally, we evaluated the fiber-optic biosensor with two CCHF patient sera. We showed that the fiber-optic biosensor is 10-times more sensitive that colorimetric ELISA and is able to detect both patients with high and low levels of IgG antibodies. We believe that the fiber-optic biosensor is a suitable alternative to ELISA as it is much more sensitive and makes it possible to detect small amount of antibodies at an early stage of infection, and can be integrated as a point-of-care diagnostic system of CCHF

Alginate-Based Applications in Biotechnology with a Special Mention to Biosensors

The exploitation of alginate and its composites as immobilisation support matrices in multiple applications remains a promising field that has the potential to create advanced functional materials from sustainable natural sources. They are non-toxic, allow sol-gel transformation, are biocompatible, have remarkable ion exchange properties, are biodegradable, and are amenable to chemical functionalisation. Alginate and its derived composites have numerous biotechnological and biomedical applications, including biomolecule or cell immobilisation, tissue engineering, drug delivery, wound dressing, and biosensors. Alginate can rapidly crosslink into a stable 3D water-insoluble network called hydrogel with polyvalent cations. Blending alginate with other materials to produce composite materials with improved or novel physicochemical properties remains an ongoing research endeavour. For instance, natural and synthetic polymers or nanoparticles have been incorporated into alginate-yielding composite material with enhanced physical strength, controlled porosity, improved interaction between the alginate support and the biomolecules, and the impartation of other features such as electrical and magnetic responsiveness, among others. Immobilisation strategies are discussed herein, including their innovations and future research perspectives

Development and Validation of an On-Line Water Toxicity Sensor with Immobilized Luminescent Bacteria for On-Line Surface Water Monitoring.

Surface water used for drinking water production is frequently monitored in The Netherlands using whole organism biomonitors, with for exampleDaphnia magnaorDreissenamussels, which respond to changes in the water quality. However, not all human-relevant toxic compounds can be detected by these biomonitors. Therefore, a new on-line biosensor has been developed, containing immobilized genetically modified bacteria, which respond to genotoxicity in the water by emitting luminescence. The performance of this sensor was tested under laboratory conditions, as well as under field conditions at a monitoring station along the river Meuse in The Netherlands. The sensor was robust and easy to clean, with inert materials, temperature control and nutrient feed for the reporter organisms. The bacteria were immobilized in sol-gel on either an optical fiber or a glass slide and then continuously exposed to water. Since the glass slide was more sensitive and robust, only this setup was used in the field. The sensor responded to spikes of genotoxic compounds in the water with a minimal detectable concentration of 0.01 mg/L mitomycin C in the laboratory and 0.1 mg/L mitomycin C in the field. With further optimization, which should include a reduction in daily maintenance, the sensor has the potential to become a useful addition to the currently available biomonitors

Primary myoblasts from intrauterine growth-restricted fetal sheep exhibit intrinsic dysfunction of proliferation and differentiation that coincides with enrichment of inflammatory cytokine signaling pathways

Intrauterine growth restriction (IUGR) is linked to lifelong reductions in muscle mass due to intrinsic functional deficits in myoblasts, but the mechanisms underlying these deficits are not known. Our objective was to determine if the deficits were associated with changes in inflammatory and adrenergic regulation of IUGR myoblasts, as was previously observed in IUGR muscle. Primary myoblasts were isolated from IUGR fetal sheep produced by hyperthermia-induced placental insufficiency (PI-IUGR; n = 9) and their controls (n = 9) and from IUGR fetal sheep produced by maternofetal inflammation (MI-IUGR; n = 6) and their controls (n = 7). Proliferation rates were less (P \u3c 0.05) for PI-IUGR myoblasts than their controls and were not affected by incubation with IL-6, TNF-α, norepinephrine, or insulin. IκB kinase inhibition reduced (P \u3c 0.05) proliferation of control myoblasts modestly in basal media but substantially in TNF-α-added media and reduced (P \u3c 0.05) PI-IUGR myoblast proliferation substantially in basal and TNF-α-added media. Proliferation was greater (P \u3c 0.05) for MI-IUGR myoblasts than their controls and was not affected by incubation with TNF-α. Insulin increased (P \u3c 0.05) proliferation in both MI-IUGR and control myoblasts. After 72-h differentiation, fewer (P \u3c 0.05) PI-IUGR myoblasts were myogenin+ than controls in basal and IL-6 added media but not TNF-α-added media. Fewer (P \u3c 0.05) PI-IUGR myoblasts were desmin+ than controls in basal media only. Incubation with norepinephrine did not affect myogenin+ or desmin+ percentages, but insulin increased (P \u3c 0.05) both markers in control and PI-IUGR myoblasts. After 96-h differentiation, fewer (P \u3c 0.05) MI-IUGR myoblasts were myogenin+ and desmin+ than controls regardless of media, although TNF-α reduced (P \u3c 0.05) desmin+ myoblasts for both groups. Differentiated PI-IUGR myoblasts had greater (P \u3c 0.05) TNFR1, ULK2, and TNF-α-stimulated TLR4 gene expression, and PI-IUGR semitendinosus muscle had greater (P \u3c 0.05) TNFR1 and IL6 gene expression, greater (P \u3c 0.05) c-Fos protein, and less (P \u3c 0.05) IκBα protein. Differentiated MI-IUGR myoblasts had greater (P \u3c 0.05) TNFR1 and IL6R gene expression, tended to have greater (P = 0.07) ULK2 gene expression, and had greater (P \u3c 0.05) β-catenin protein and TNF-α-stimulated phosphorylation of NFκB. We conclude that these enriched components of TNF-α/TNFR1/NFκB and other inflammatory pathways in IUGR myoblasts contribute to their dysfunction and help explain impaired muscle growth in the IUGR fetus. Lay Summary-- Myoblasts are stems cells whose functional capacity can limit muscle growth. However, stressful intrauterine conditions cause these cells to be intrinsically dysfunctional. This restricts muscle growth capacity, leading to intrauterine growth restriction (IUGR) of the fetus, low birth weight, and less muscle mass after birth. Consequently, meat yield is reduced in IUGR-born food animals and glucose homeostasis is impaired in IUGR-born humans, which contributes to metabolic dysfunction. Intrinsic dysfunction of IUGR myoblasts has been previously observed, but the fetal programming changes (i.e., permanent changes in the development of cellular mechanisms that explains different functional outcomes) have not been identified. This study shows that one mechanism is the enhancement of signaling pathways for TNF-α and other inflammatory cytokines. These cytokines have roles in stress responses and regulation of muscle growth. Programmed enhancement of these pathways means that IUGR myoblasts are more responsive to even normal amounts of circulating cytokines. Unfortunately, the primary response of myoblasts to cytokines is slower differentiation (i.e., cellular transformation necessary for muscle growth). Programmed enhancement of this response directly impedes myoblast-dependent muscle growth, and the deficit is lifelong. However, identifying this mechanism is a fundamental step for developing strategies to improve muscle growth in low birth weight offspring