Thursday, January 19, 2012

peptide arrays | What is peptide arrays|Papers on peptide arrays|Research on peptide arrays| Publications on peptide arrays

    Results: 1 to 20 of 3328

    1.
    Clin Vaccine Immunol. 2012 Jan 11. [Epub ahead of print]

    Evaluation of Biological Sample Preparation for Immunosignature Based Diagnostics.

    Source

    Center for Innovations in Medicine, The Biodesign Institute, Arizona State University, Tempe, AZ 85287-5901.

    Abstract

    To address the need for a universal system to assess health status, we previously described a method termed immunosignaturing which splays the entire humoral antibody repertoire across a peptide microarray. Two important issues relative to the potential broad use of immunosignatures are sample preparation and stability. In the present study we compared the immunosignatures developed from serum, plasma, saliva and antibodies eluted from blood dried onto filter paper. We found serum and plasma provide identical immunosignatures. Dried blood also correlated well with non-dried serum from the same individual. Immunosignatures derived from dried blood were capable of distinguishing naïve mice from those infected with influenza. Saliva was applied to the arrays and the IgA immunosignature correlated strongly with that from dried blood. Finally, we demonstrate that dried blood retains immunosignature information even when exposed to high temperature. This work expands the potential diagnostic uses for immunosignatures. These features suggest that different forms of archival samples can be used for diagnosis development and that in prospective studies samples can be easily procured.

    PMID:
    22237890
    [PubMed - as supplied by publisher]
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    2.
    Exp Neurol. 2012 Jan 2. [Epub ahead of print]

    Protective action of NDP-MSH in experimental subarachnoid hemorrhage.

    Source

    Center for Surgical Research, Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico, Milan, Italy.

    Abstract

    Subarachnoid hemorrhage (SAH) is still a major cause of morbidity and mortality. α-Melanocyte stimulating hormone (α-MSH) and other melanocortin peptides exert potent neuroprotective action and they might modulate key molecules involved in SAH-induced vasospasm. The aim of this research was to determine whether treatment with the α-MSH analog Nle4,DPhe7-α-MSH (NDP-MSH) exerts protective effects in experimental SAH in the rat. Initial experiments examined effects of NDP-MSH on the basilar artery phenotype in the absence of injury. In these tests intrathecal injection of small concentrations (10ng) of the peptide induced a tolerant phenotype similar to that observed after ischemic preconditioning. Then the effect of systemic treatment with NDP-MSH (100μg i.v.) on experimental SAH was evaluated. SAH was induced by a single-blood injection into the cisterna magna. The basilar artery phenotype was examined at 4h and the artery caliber at 5days following SAH. Expression of 96 genes was analyzed by real-time reverse transcription polymerase chain reaction (RT-PCR) using Custom Taqman Low-Density Arrays. Four hours after SAH, the transcriptional profile of the basilar artery was deeply disrupted. Transcript alteration included genes involved in inflammation, stress response, apoptosis, and vascular remodeling. Treatment with NDP-MSH prevented most of these transcription changes and decreased phosphorylation of extracellular-signal-regulated kinases (ERK1/2) and inhibitor protein IκBα. Vasospasm on day 5 was significantly reduced by NDP-MSH administration. These results combine with others on CNS inflammation to suggest that the melanocortins could be safe and effective therapeutic candidates to treat SAH-related complications.

    Copyright © 2011. Published by Elsevier Inc.

    PMID:
    22230666
    [PubMed - as supplied by publisher]
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    3.
    Ann Rheum Dis. 2012 Jan 5. [Epub ahead of print]

    Association of ferritin autoantibodies with giant cell arteritis/polymyalgia rheumatica.

    Source

    1Department of Immunology and Rheumatology, Medical University, Hannover, Germany.

    Abstract

    OBJECTIVES:

    Polymyalgia rheumatica (PMR) and giant cell arteritis (GCA) are relatively common inflammatory disorders. Establishing the diagnosis however may be difficult, since so far no specific biomarkers of the disorders are available.

    METHODS:

    As a screening procedure, the authors used protein arrays for the detection of new autoantigens in GCA and PMR. The results of the protein array were confirmed by different ELISAs detecting IgG antibodies against the human ferritin heavy chain, N-terminal 27 amino acids of the human ferritin heavy chain or the homologous peptide of Staphylococcus epidermidis. Sera of patients with only GCA (n=64), only PMR (n=47) and both PMR and GCA (n=31) were used.

    RESULTS:

    In the ELISA using the human ferritin peptide, the sensitivity of IgG antibodies against ferritin was 92% in 36 GCA and/or PMR patients before initiation of treatment, 22/32 (69%) in patients with disease flares and 64/117 (55%) in the total cohort including treated and inactive patients. In controls, the false positive rate was 11/38 (29%) in systemic lupus erythematosus, 1/36 (3%) in rheumatoid arthritis, 0/31 (0%) in late onset rheumatoid arthritis, 3/46 (6.5%) in B-non-Hodgkin's lymphoma and 1/100 (1%) in blood donors. In the ELISA using the ferritin peptide of S epidermidis, 89% of 27 patients with untreated GCA and PMR were positive.

    CONCLUSION:

    Antibodies against the ferritin peptide were present in up to 92% of untreated, active GCA and PMR patients. They can be useful as a diagnostic marker of PMR and GCA.

    PMID:
    22228484
    [PubMed - as supplied by publisher]
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    4.
    J Pharm Sci. 2011 Dec 28. doi: 10.1002/jps.23036. [Epub ahead of print]

    The effects of media on pharmaceutically relevant transporters in the human HT-29 adenocarcinoma cell line: Does culture media need to be controlled?

    Source

    Department of Industrial and Physical Pharmacy, College of Pharmacy, Purdue University, West Lafayette, Indiana 47907.

    Abstract

    The HT-29 cell line forms a confluent monolayer with tight junctions, but displays different phenotypes when cultured for 21 days in galactose-supplemented media (differentiated) versus glucose-supplemented media (dedifferentiated). This study is aimed at elucidating how media differences might affect selected drug transporter expression and peptide-based substrate transport toward reducing this variability. A vial of HT-29 cells was amplified and cultured over several passages in four different mediums (American Type Culture Collection recommended McCoy's 5A versus Dulbecco's modified Eagle's media containing glucose, galactose, or neither carbohydrate) with normal supplementation. Transporter mRNA expression was characterized at days 5 and 21 postseeding utilizing SABiosciences quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) drug transporter arrays. Transport studies using [H]histidine, [(3) H]glycylsarcosine, [(3) H]valacyclovir, and [(3) H]carnosine were performed to assess the functional effects of oligopeptide transporter expression changes in HT-29 cells grown in each media. qRT-PCR arrays illustrated variable, media-dependent transporter expression between both the initial and differentiated time points. Permeability studies illustrated considerable media-dependent differences in both paracellular and transcellular substrate fluxes. The results demonstrate that these cells exhibit differing monolayer characteristics and genotypic/phenotypic profile properties when cultured under different media. The results suggest a need for standardization of culture methodologies for reducing inter- and intralaboratory variability. © 2011 Wiley Periodicals, Inc.

    Copyright © 2011 Wiley Periodicals, Inc.

    PMID:
    22213613
    [PubMed - as supplied by publisher]
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    5.
    Mol Cell Proteomics. 2011 Dec 28. [Epub ahead of print]

    Systems kinomics demonstrates congo basin monkeypox virus infection selectively modulates host cell signaling responses as compared to West African monkeypox virus.

    Source

    NIAID, United States;

    Abstract

    Monkeypox virus (MPXV) is comprised of two clades: Congo Basin MPXV, with an associated case fatality rate of 10%, and Western African MPXV, which is associated with less severe infection and minimal lethality. We thus postulated that Congo Basin and West African MPXV would differentially modulate host cell responses and, as many host responses are regulated through phosphorylation independent of transcription or translation, we employed systems kinomics with peptide arrays to investigate these functional host responses. Using this approach we have demonstrated that Congo Basin MPXV infection selectively downregulates host responses as compared to West African MPXV, including growth factor- and apoptosis-related responses. These results were confirmed using FACS analysis demonstrating that West African MPXV infection resulted in a significant increase in apoptosis in human monocytes as compared to Congo Basin MPXV. Further, differentially phosphorylated kinases were identified through comparison of our MPXV data sets and validated as potential targets for pharmacological inhibition of Congo Basin MPXV infection, including increased Akt S473 phosphorylation and decreased p53 S15 phosphorylation. Inhibition of Akt S473 phosphorylation resulted in a significant decrease in Congo Basin MPXV virus yield (261-fold) but did not affect West African MPXV. In addition, treatment with staurosporine, an apoptosis activator resulted in a 49-fold greater decrease in Congo Basin MPXV yields as compared to West African MPXV. Thus, using a systems kinomics approach, our investigation demonstrates that West African and Congo Basin MPXV differentially modulate host cell responses and has identified potential host targets of therapeutic interest.

    PMID:
    22205724
    [PubMed - as supplied by publisher]
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    6.
    Nucleic Acids Res. 2011 Dec 23. [Epub ahead of print]

    Type III restriction endonuclease EcoP15I is a heterotrimeric complex containing one Res subunit with several DNA-binding regions and ATPase activity.

    Source

    Institute of Medical Virology, Helmut-Ruska-Haus, Charité - Universitätsmedizin Berlin, Charitéplatz 1, D-10117 Berlin, Institute for Biophysical Chemistry, Hannover Medical School, Carl-Neuberg-Straße 1, D-30625 Hannover and JPT Peptide Technologies GmbH, Volmerstraße 5, D-12489 Berlin, Germany.

    Abstract

    For efficient DNA cleavage, the Type III restriction endonuclease EcoP15I communicates with two inversely oriented recognition sites in an ATP-dependent process. EcoP15I consists of methylation (Mod) and restriction (Res) subunits forming a multifunctional enzyme complex able to methylate or to cleave DNA. In this study, we determined by different analytical methods that EcoP15I contains a single Res subunit in a Mod(2)Res stoichiometry. The Res subunit comprises a translocase (Tr) domain carrying functional motifs of superfamily 2 helicases and an endonuclease domain with a PD..D/EXK motif. We show that the isolated Tr domain retains ATP-hydrolyzing activity and binds single- and double-stranded DNA in a sequence-independent manner. To localize the regions of DNA binding, we screened peptidearrays representing the entire Res sequence for their ability to interact with DNA. We discovered four DNA-binding regions in the Tr domain and two DNA-binding regions in the endonuclease domain. Modelling of the Tr domain shows that these multiple DNA-binding regions are located on the surface, free to interact with DNA. Interestingly, the positions of the DNA-binding regions are conserved among other Type III restriction endonucleases.

    PMID:
    22199260
    [PubMed - as supplied by publisher]
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    7.
    Trends Biotechnol. 2011 Dec 22. [Epub ahead of print]

    Short self-assembling peptides as building blocks for modern nanodevices.

    Source

    Institute of Bioengineering and Nanotechnology, 31 Biopolis Way, The Nanos, Singapore 138669.

    Abstract

    Short, self-assembling peptides form a variety of stable nanostructures used for the rational design of functional devices.Peptides serve as organic templates for conjugating biorecognition elements, and assembling ordered nanoparticlearrays and hybrid supramolecular structures. We are witnessing the emergence of a new phase of bionanotechnology, particularly towards electronic, photonic and plasmonic applications. Recent advances include self-assembly of photoluminescent semiconducting nanowires and peptide-conjugated systems for sensing, catalysis and energy storage. Concurrently, methods and tools have been developed to control and manipulate the self-assembled nanostructures. Furthermore, there is growing knowledge on nanostructure properties such as piezoelectricity, dipolar electric field and stability. This review focuses on the emerging role of short, linear self-assembling peptides as simple and versatile building blocks for nanodevices.

    Copyright © 2011 Elsevier Ltd. All rights reserved.

    PMID:
    22197260
    [PubMed - as supplied by publisher]
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    8.
    PLoS One. 2011;6(12):e28263. Epub 2011 Dec 14.

    Hypoxia Due to Cardiac Arrest Induces a Time-Dependent Increase in Serum Amyloid β Levels in Humans.

    Source

    Department of Psychiatry and Neurochemistry, The Sahlgrenska Academy at the University of Gothenburg, Mölndal, Sweden.

    Abstract

    Amyloid β (Aβ) peptides are proteolytic products from amyloid precursor protein (APP) and are thought to play a role in Alzheimer disease (AD) pathogenesis. While much is known about molecular mechanisms underlying cerebral Aβ accumulation in familial AD, less is known about the cause(s) of brain amyloidosis in sporadic disease. Animal and postmortem studies suggest that Aβ secretion can be up-regulated in response to hypoxia. We employed a new technology (Single Molecule Arrays, SiMoA) capable of ultrasensitive protein measurements and developed a novel assay to look for changes in serum Aβ42 concentration in 25 resuscitated patients with severe hypoxia due to cardiac arrest. After a lag period of 10 or more hours, very clear serum Aβ42 elevations were observed in all patients. Elevations ranged from approximately 80% to over 70-fold, with most elevations in the range of 3-10-fold (average approximately 7-fold). The magnitude of the increase correlated with clinical outcome. These data provide the first direct evidence in living humans that ischemia acutely increases Aβ levels in blood. The results point to the possibility that hypoxia may play a role in the amyloidogenic process of AD.

    PMID:
    22194817
    [PubMed - in process]
    PMCID: PMC3237426
    Free PMC Article
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    9.
    Biotechnol Bioeng. 2011 Dec 14. doi: 10.1002/bit.24405. [Epub ahead of print]

    Fabrication of hierarchical hybrid structures using bio-enabled layer-by-layer self-assembly.

    Source

    Department of Material Science and Engineering, Genetically Engineered Materials Science and Engineering Center (GEMSEC), University of Washington, Seattle, Washington 98195; telephone: 206-616-6980; fax: 206-543-3100.

    Abstract

    Development of versatile and flexible assembly systems for fabrication of functional hybrid nanomaterials with well-defined hierarchical and spatial organization is of a significant importance in practical nanobiotechnology applications. Here we demonstrate a bio-enabled self-assembly technique for fabrication of multi-layered protein and nanometallic assemblies utilizing a modular gold-binding (AuBP1) fusion tag. To accomplish the bottom-up assembly we first genetically fused the AuBP1 peptide sequence to the C'-terminus of maltose-binding protein (MBP) using two different linkers to produce MBP-AuBP1 hetero-functional constructs. Using various spectroscopic techniques, surface plasmon resonance (SPR) and localized surface plasmon resonance (LSPR), we verified the exceptional binding and self-assembly characteristics of AuBP1 peptide. The AuBP1 peptide tag can direct the organization of recombinant MBP protein on various gold surfaces through an efficient control of the organic-inorganic interface at the molecular level. Furthermore using a combination of soft-lithography, self-assembly techniques and advanced AuBP1 peptide tag technology, we produced spatially and hierarchically controlled protein multi-layered assemblies on gold nanoparticlearrays with high molecular packing density and pattering efficiency in simple, reproducible steps. This model system offers layer-by-layer assembly capability based on specific AuBP1 peptide tag and constitutes novel biological routes for biofabrication of various protein arrays, plasmon-active nanometallic assemblies and devices with controlled organization, packing density and architecture. Biotechnol. Bioeng. © 2011 Wiley Periodicals, Inc.

    Copyright © 2011 Wiley Periodicals, Inc.

    PMID:
    22170333
    [PubMed - as supplied by publisher]
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    10.
    Chem Commun (Camb). 2012 Jan 4;48(8):1132-4. Epub 2011 Dec 14.

    Electron transfer through α-peptides attached to vertically aligned carbon nanotube arrays: a mechanistic transition.

    Source

    School of Chemistry and Physics, The University of Adelaide, Adelaide, SA 5005, Australia. andrew.abell@adelaide.edu.au.

    Abstract

    The mechanism of electron transfer in α-aminoisobutyric (Aib) homoligomers is defined by the extent of secondary structure, rather than just chain length. Helical structures (Aib units ≥3) undergo an electron hopping mechanism, while shorter disordered sequences (Aib units <3) undergo an electron superexchange mechanism.

    PMID:
    22166913
    [PubMed - in process]
    11.
    Langmuir. 2011 Dec 22. [Epub ahead of print]

    Phage-Chips for Novel Optically Readable Tissue Engineering Assays.

    Source

    Department of Bioengineering, and Berkeley Nanoscience and Nanoengineering Institute, University of California, Berkeley , Berkeley, California 94720, United States.

    Abstract

    We report novel phage-based array chips that are optically readable for cell proliferation and morphology assays. Using M13 phages that were engineered to display RGD on its major coat proteins and/or immobilize FGFb on its minor coat proteins, we prepared arrays of phage spot matrices composed of self-assembled nanofibrous network structures. We cultured fibroblasts on the arrays and, using surface plasmon resonance (SPR) spectroscopy, monitored the effects of the biochemical cues displayed by the phage on cell proliferation and morphology. This study demonstrates the utility of engineered phages as promising coating materials for lab-on-a-chip (LOC) platforms, allowing sensitive monitoring of the effects of functional peptides on cell growth. Phage-chips have great potential for use as high-throughput screening systems for biochemical assays and biosensors and the discovery of novel drugs.

    PMID:
    22149649
    [PubMed - as supplied by publisher]
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    12.
    J Biol Chem. 2011 Dec 6. [Epub ahead of print]

    Insights into high affinity SUMO recognition by SUMO-interacting motifs (SIM) revealed by a combination of NMR and peptide array analysis.

    Source

    Beckman Research Institute of the City of Hope, United States;

    Abstract

    The small ubiquitin-like modifiers (SUMO) regulate many essential cellular functions. Only one type of SUMO-interacting motif (SIM) has been identified that can extend the β-sheet of SUMO as either a parallel or an anti-parallel strand. The molecular determinants of the bound orientation and paralogue-specificity of a SIM is unclear. To address this question, we have conducted structural studies of SUMO1 in complex with a SUMO1-specific SIM that binds to SUMO1 with high affinity without post-translational modifications using nuclear magnetic resonance methods. In addition, the SIM sequence requirements have been investigated by peptide arrays in comparison to another high affinity SIM that binds in the opposing orientation. We found that anti-parallel binding SIMs tolerate more diverse sequences, whereas the parallel binding SIMs prefer the more strict sequences consisting of I/V-D-L-T that have a preference in high affinity SUMO2 and 3 binding. Comparison of two high affinity SUMO1-binding SIMs that binds in opposing orientations has revealed common SUMO1-specific interactions needed for high affinity binding. This study has significantly advanced our understanding of the molecular determinants underlining SUMO-SIM recognition.

    PMID:
    22147707
    [PubMed - as supplied by publisher]
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    13.
    Macromol Biosci. 2011 Dec 6. doi: 10.1002/mabi.201100295. [Epub ahead of print]

    Natively Unfolded State for Engineering Nanoscale Fibrillar Arrays.

    Source

    National Physical Laboratory, Teddington, Middlesex, TW11 0LW, UK; School of Physics and Astronomy, University of Edinburgh, Mayfield Road, Edinburgh, EH9 3JZ, UK. max.ryadnov@npl.co.uk.

    Abstract

    A generic rationale for the fabrication of high aspect ratio fibrillar nanoscale arrays is described. The design emulates an intermittence effect observed for β-structured α-synunclein fibrils, reported herein, in a structurally unrelated α-helical fiber. The generated nanoarrays are composed of periodic nanosized segments separated at uniform distances of unfolded regions. These regions can be targeted for conformational binding and refolding with metal nanoparticle-peptideconjugates for the conversion of fibrillar arrays into nanoparticle arrays. The introduced concept opens new strategies for engineering novel nanoscale materials and devices.

    Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

    PMID:
    22147495
    [PubMed - as supplied by publisher]
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    14.
    Electrophoresis. 2012 Jan;33(1):2-13. doi: 10.1002/elps.201100344. Epub 2011 Dec 5.

    Growing trend of CE at the omics level: The frontier of systems biology - An update.

    Source

    Biomolecules Function Research Center, Korea Institute of Science and Technology, Cheongryang, Seoul, Korea.

    Abstract

    Omics is the study of proteins, peptides, genes, and metabolites in living organisms. Systems biology aims to understand the system through the study of the relationship between elements such as genes and proteins in biological system. Recently, systems biology emerged as the result of the advanced development of high-throughput analysis technologies such as DNA sequencers, DNA arrays, and mass spectrometry for omics studies. Among a number of analytical tools and technologies, CE and CE coupled to MS are promising and relatively rapidly developing tools with the potential to provide qualitative and quantitative analyses of biological molecules. With an emphasis on CE for systems biology, this review summarizes the method developments and applications of CE for the genomic, transcriptomic, proteomic, and metabolomic studies focusing on the drug discovery and disease diagnosis and therapies since 2009.

    Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

    PMID:
    22139583
    [PubMed - in process]
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    15.
    Exp Neurol. 2011 Nov 23. [Epub ahead of print]

    NF-кB-regulated micro RNAs (miRNAs) in primary human brain cells.

    Abstract

    Micro RNAs (miRNAs), small and labile ~22 nucleotide-sized fragments of single stranded RNA, are important regulators of messenger (mRNA) complexity and in shaping the transcriptome of a cell. In this communication, we utilized amyloid beta 42 (Aβ42) peptides and interleukin-1beta (IL-1β) as a combinatorial, physiologically-relevant stress to induce miRNAs in human primary neural (HNG) cells (a co-culture of neurons and astroglia). Specific miRNA up-regulation was monitored using miRNA arrays, Northern micro-dot blots and RT-PCR. Selective NF-кB translocation and DNA binding inhibitors, including the chelator and anti-oxidant pyrollidine dithiocarbamate (PDTC) and the polyphenolic resveratrol analog CAY10512 (trans-3,5,4'-trihydroxystilbene), indicated the NF-кB sensitivity of several brain miRNAs, including miRNA-9, miRNA-125b and miRNA-146a. The inducible miRNA-125b and miRNA-146a, and their verified mRNA targets, including 15-lipoxygenase (15-LOX), synapsin-2 (SYN-2), complement factor H (CFH) and tetraspanin-12 (TSPAN12), suggests complex and highly interactive roles for NF-кB, miRNA-125b and miRNA-146a. These data further indicate that just two NF-кB-mediated miRNAs have tremendous potential to contribute to the regulation of neurotrophic support, synaptogenesis, neuroinflammation, innate immune signaling and amyloidogenesis in stressed primary neural cells of the human brain.

    Copyright © 2011 Elsevier Inc. All rights reserved.

    PMID:
    22138609
    [PubMed - as supplied by publisher]
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    16.
    Biosens Bioelectron. 2012 Jan 15;31(1):233-9. Epub 2011 Oct 20.

    High sensitivity carbon nanotube based electrochemiluminescence sensor array.

    Source

    Biomedical Diagnostic Institute, National Center for Sensor Research, School of Chemical Sciences, Dublin City University, Dublin 9, Ireland.

    Abstract

    Ink jet printed carbon nanotube forest arrays capable of detecting picomolar concentrations of immunoglobulin G (IgG) using electrochemiluminescence (ECL) are described. Patterned arrays of vertically aligned single walled carbon nanotube (SWCNT) forests were printed on indium tin oxide (ITO) electrodes. Capture anti-IgG antibodies were then coupled through peptide bond formation to acidic functional groups on the vertical nanotubes. IgG immunoassays were performed using silica nano particles (Si NP) functionalized with the ECL luminophore [Ru(bpy)(2)PICH(2)](2+)], and IgG labelled G1.5 acid terminated PAMAM dendrimers. PAMAM is poly(amido amine), bpy is 2,2'-bipyridyl and PICH(2) is (2-(4-carboxyphenyl)imidazo[4,5-f][1,10]phenanthroline). The carboxyl terminal of [Ru(bpy)(2)PICH(2)](2+) (fluorescence lifetime≈682±5ns) dye was covalently coupled to amine groups on the 800nm diameter silica spheres in order to produce significant ECL enhancement in the presence of sodium oxalate as co-reactant in PBS at pH 7.2). Significantly, this SWCNT-based sensor array shows a wide linear dynamic range for IgG coated spheres (10(6) to 10(12) spheres) corresponding to IgG concentrations between 20 pM and 300nM. A detection limit of 1.1±0.1pM IgG is obtained under optimal conditions.

    Copyright © 2011 Elsevier B.V. All rights reserved.

    PMID:
    22137061
    [PubMed - in process]
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    17.
    Mol Genet Metab. 2011 Nov 10. [Epub ahead of print]

    Differentially expressed angiogenic genes in diabetic erectile tissue - Results from a microarray screening.

    Source

    Institute for Molecular and Cell Biology of the University of Porto (IBMC-UP), Rua do Campo Alegre, 823, 4150-180 Porto, Portugal; Department of Biochemistry (U38-FCT), Faculty of Medicine of the University of Porto, Alameda Prof. Hernani Monteiro, 4200-319 Porto, Portugal.

    Abstract

    Diabetes-induced metabolic derangements promote endothelial malfunction, contributing to erectile dysfunction (ED). However, it remains unclear which angiogenic molecular mechanisms are deregulated in diabetic corpus cavernosum (CC). We investigated early and late alterations in cavernosal angiogenic gene expression associated to diabetes. Angiogenic changes were assessed in penile tissue of streptozotocin-induced Wistar rats, in an early (2-week) and established stage (8-week) of diabetes. Differentially expressed genes were identified by microarrays and expression data validated by quantitative real-time PCR (qrt-PCR). At protein level, quantitative immunohistochemistry confirmed thearrays data and dual immunofluorescence for selected alterations and α-smooth muscle actin (α-SMA) identified the cellular location of target proteins. The selected differentially expressed genes were also evaluated in human non-diabetic and diabetic CC by quantitative immunolabeling. At 2-week diabetes there was no differential gene expression between non-diabetic and diabetic CC. At 8-week, 10 genes were found down-regulated in diabetics. The results were validated by qrt-PCR for the insulin-like growth factor-1 (Igf1) and the natriuretic peptide receptor-1 (Npr1) genes. Dual immunofluorescence for IGF-1/ α-SMA showed predominant localization of IGF-1 in SM. NPR-1 expression was diffuse and mostly present in trabecular fibroblasts and SM. Quantitative immunostaining confirmed the decreased expression of both proteins in diabetic tissues. Concordantly, we detected a significant reduction in IGF-1 and NPR-1 protein expressions in human diabetic samples. Microarray analysis identified 10 angiogenic-related molecules deregulated in CC of established diabetes. Among them, IGF-1 and NPR-1 were significantly down-regulated and might result in preventive/therapeutic targets for ED management.

    Copyright © 2011 Elsevier Inc. All rights reserved.

    PMID:
    22133301
    [PubMed - as supplied by publisher]
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    18.
    BMC Bioinformatics. 2011 Nov 30;12(1):462. [Epub ahead of print]

    Segmentation and intensity estimation for microarray images with saturated pixels.

    Abstract

    ABSTRACT:

    BACKGROUND:

    Microarray image analysis processes scanned digital images of hybridized arrays to produce the input spot-level data for downstream analysis, so it can have a potentially large impact on those and subsequent analysis. Signal saturation is an optical effect that occurs when some pixel values for highly expressed genes or peptides exceed the upper detection threshold of the scanner software (2^16 - 1 = 65,535 for 16-bit images). In practice, spots with a sizable number of saturated pixels are often flagged and discarded. Alternatively, the saturated values are used without adjustments for estimating spot intensities. The resulting expression data tend to be biased downwards and can distort high-level analysis that relies on these data. Hence, it is crucial to effectively correct for signal saturation.

    RESULTS:

    We developed a flexible mixture model-based segmentation and spot intensity estimation procedure that accounts for saturated pixels by incorporating a censored component in the mixture model. As demonstrated with biological data and simulation, our method extends the dynamic range of expression data beyond the saturation threshold and is effective in correcting saturation-induced bias when the lost information is not tremendous. We further illustrate the impact of image processing on downstream classification, showing that the proposed method can increase diagnostic accuracy using data from a lymphoma cancer diagnosis study.

    CONCLUSIONS:

    The presented method adjusts for signal saturation at the segmentation stage that identifies a pixel as part of the foreground, background or other. The cluster membership of a pixel can be altered versus treating saturated values as truly observed. Thus, the resulting spot intensity estimates may be more accurate than those obtained from existing methods that correct for saturation based on already segmented data. As a model-based segmentation method, our procedure is able to identify inner holes, fuzzy edges and blank spots that are common in microarray images. The approach is independent of microarray platform and applicable to both single- and dual-channel microarrays.

    PMID:
    22129216
    [PubMed - as supplied by publisher]
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    19.
    PLoS Comput Biol. 2011 Nov;7(11):e1002288. Epub 2011 Nov 17.

    Deciphering the Arginine-binding preferences at the substrate-binding groove of Ser/Thr kinases by computational surface mapping.

    Source

    Institute of Biochemistry, Food Science and Nutrition, The Robert H. Smith Faculty of Agriculture, Food and Environment and The Fritz Haber Center for Molecular Dynamics, The Hebrew University, Israel.

    Abstract

    Protein kinases are key signaling enzymes that catalyze the transfer of γ-phosphate from an ATP molecule to a phospho-accepting residue in the substrate. Unraveling the molecular features that govern the preference of kinases for particular residues flanking the phosphoacceptor is important for understanding kinase specificities toward their substrates and for designing substrate-like peptidic inhibitors. We applied ANCHORSmap, a new fragment-based computational approach for mapping amino acid side chains on protein surfaces, to predict and characterize the preference of kinases toward Arginine binding. We focus on positions P-2 and P-5, commonly occupied by Arginine (Arg) in substrates of basophilic Ser/Thr kinases. The method accurately identified all the P-2/P-5 Arg binding sites previously determined by X-ray crystallography and produced Arg preferences that corresponded to those experimentally found by peptide arrays. The predicted Arg-binding positions and their associated pockets were analyzed in terms of shape, physicochemical properties, amino acid composition, and in-silico mutagenesis, providing structural rationalization for previously unexplained trends in kinase preferences toward Arg moieties. This methodology sheds light on several kinases that were described in the literature as having non-trivial preferences for Arg, and provides some surprising departures from the prevailing views regarding residues that determine kinase specificity toward Arg. In particular, we found that the preference for a P-5 Arg is not necessarily governed by the 170/230 acidic pair, as was previously assumed, but by several different pairs of acidic residues, selected from positions 133, 169, and 230 (PKA numbering). The acidic residue at position 230 serves as a pivotal element in recognizing Arg from both the P-2 and P-5 positions.

    PMID:
    22125489
    [PubMed - in process]
    PMCID: PMC3219626
    Free PMC Article
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    20.
    Am J Hum Biol. 2012 Jan;24(1):81-6. doi: 10.1002/ajhb.21229. Epub 2011 Nov 28.

    Validation of a new multiplex assay against individual immunoassays for the quantification of reproductive, stress, and energetic metabolism biomarkers in urine specimens.

    Source

    Faculty of Health Sciences, Simon Fraser University, Burnaby, British Columbia, Canada V5A 1S6.

    Abstract

    Measuring multiple hormones simultaneously in a single assay saves sample volume, labor, time, reagents, money, and consumables. Thus, multiplex arrays represent a faster, more economically and ecologically sound alternative to singleton assays.

    OBJECTIVES:

     To validate a new, commercially available multiplex female array produced by Quansys Biosciences against individual immunoassays for the quantification of six hormones in urine samples from women in different reproductive stages.

    METHODS:

     Urine samples were analyzed using the new Quansys multiplex female hormone array and compared with well-established individual immunoassays for adiponectin, free cortisol, c-peptide, estrone-3-glucuronide (E(1) G), follicle stimulating hormone beta-subunit (FSH-beta), and human chorionic gonadotropin beta-subunit (hCG-beta). Correlations between assays were assessed using Pearson correlation, linear regression and Bland-Altman analysis. The temporal profiles of free cortisol, E1G, FSH-beta, and hCG-beta were also compared.

    RESULTS:

     The multiplex array was highly correlated with the individual immunoassays for five of the tested hormones (Pearson's correlation coefficient ≥0.75), and yielded temporal patterns of hormone profiles consistent with the individual immunoassays for free cortisol, E1G, FSH-beta, and hCG-beta.

    CONCLUSIONS:

      The Quansys multiplex female hormone array is a valid alternative method to individual immunoassays for the quantification of stress, reproductive and energetic hormones and metabolites in human urine samples and can be used to examine the dynamic interactions between these hormones. Am. J. Hum. Biol., 2012. © 2011 Wiley Periodicals, Inc.

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