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https://www.readbyqxmd.com/read/28441386/graft-derived-cell-free-dna-a-noninvasive-early-rejection-and-graft-damage-marker-in-liver-transplantation-a-prospective-observational-multicenter-cohort-study
#1
Ekkehard Schütz, Anna Fischer, Julia Beck, Markus Harden, Martina Koch, Tilo Wuensch, Martin Stockmann, Björn Nashan, Otto Kollmar, Johannes Matthaei, Philipp Kanzow, Philip D Walson, Jürgen Brockmöller, Michael Oellerich
BACKGROUND: Graft-derived cell-free DNA (GcfDNA), which is released into the blood stream by necrotic and apoptotic cells, is a promising noninvasive organ integrity biomarker. In liver transplantation (LTx), neither conventional liver function tests (LTFs) nor immunosuppressive drug monitoring are very effective for rejection monitoring. We therefore hypothesized that the quantitative measurement of donor-derived cell-free DNA (cfDNA) would have independent value for the assessment of graft integrity, including damage from acute rejection...
April 2017: PLoS Medicine
https://www.readbyqxmd.com/read/28439827/droplet-microfluidic-device-fabrication-and-use-for-isothermal-amplification-and-detection-of-microrna
#2
Maria Chiara Giuffrida, Roberta D'Agata, Giuseppe Spoto
Droplet microfluidics combined with the isothermal circular strand displacement polymerization (ICSDP) represents a powerful new technique to detect both single-stranded DNA and microRNA sequences. The method here described helps in overcoming some drawbacks of the lately introduced droplet polymerase chain reaction (PCR) amplification when implemented in microfluidic devices. The method also allows the detection of nanoliter droplets of nucleic acids sequences solutions, with a particular attention to microRNA sequences that are detected at the picomolar level...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28437435/infective-respiratory-syncytial-virus-is-present-in-human-cord-blood-samples-and-most-prevalent-during-winter-months
#3
Angela Mary Fonceca, Abha Chopra, Avram Levy, Paul Stanton Noakes, Matthew Wee-Peng Poh, Natasha Leanne Bear, Susan Prescott, Mark Lloyd Everard
BACKGROUND: Human respiratory syncytial virus (RSV) remains the most common cause of severe lower respiratory tract disease amongst infants, and continues to cause annual epidemics of respiratory disease every winter worldwide. Demonstrating placental transmission of viable RSV in human samples is a major paradigm shift in respiratory routes considered likely for RSV transmission. METHODS: Droplet digital PCR (ddPCR) was used to identify RSV present in cord blood mononucleocytes (CBM)...
2017: PloS One
https://www.readbyqxmd.com/read/28437026/analyzing-epidermal-growth-factor-receptor-mutation-status-changes-in-advanced-non-small-cell-lung-cancer-at-different-sampling-time-points-of-blood-within-one%C3%A2-day
#4
Jin Wang, Hua Bai, Chaoyu Hong, Jie Wang, Tong-Hua Mei
BACKGROUND: We investigated whether different sampling time-points within one day would influence epidermal growth factor receptor mutation (EGFRm) status in plasma and evaluated the clinical outcomes according to the quantity analysis of EGFRm in circulating tumor DNA (ctDNA) in non-small-cell lung cancer (NSCLC). METHODS: EGFR-tyrosine kinase inhibitor naïve advanced NSCLC patients who carried EGFRm in both tissues and ctDNA were enrolled in this study. Plasma samples were collected at three time-points within one day (at 8 am, 11 am and 2 pm) for EGFRm analysis by droplet digital PCR...
April 24, 2017: Thoracic Cancer
https://www.readbyqxmd.com/read/28434824/multiple-pathogen-biomarker-detection-using-an-encoded-bead-array-in-droplet-pcr
#5
Prem Kumar Periyannan Rajeswari, Lovisa M Soderberg, Alia Yacoub, Mikael Leijon, Helene Andersson Svahn, Haakan N Joensson
We present a droplet PCR workflow for detection of multiple pathogen DNA biomarkers using fluorescent color-coded Luminex® beads. This strategy enables encoding of multiple singleplex droplet PCRs using a commercially available bead set of several hundred distinguishable fluorescence codes. This workflow provides scalability beyond the limited number offered by fluorescent detection probes such as TaqMan probes, commonly used in current multiplex droplet PCRs. The workflow was validated for three different Luminex bead sets coupled to target specific capture oligos to detect hybridization of three microorganisms infecting poultry: avian influenza, infectious laryngotracheitis virus and Campylobacter jejuni...
April 20, 2017: Journal of Microbiological Methods
https://www.readbyqxmd.com/read/28426159/open-microfluidic-gel-electrophoresis-rapid-and-low-cost-separation-and-analysis-of-dna-at-the-nanoliter-scale
#6
Ludwig Gutzweiler, Tobias Gleichmann, Laurent Tanguy, Peter Koltay, Roland Zengerle, Lutz Riegger
Gel electrophoresis is one of the most applied and standardized tools for separation and analysis of macromolecules and their fragments in academic research and in industry. In this work we present a novel approach for conducting on-demand electrophoretic separations of DNA molecules in open microfluidic (OM) systems on planar polymer substrates. The approach combines advantages of slab gel, capillary- & chip-based methods offering low consumable costs (<0.1 $) circumventing cost intensive microfluidic chip fabrication, short process times (5 minutes per analysis) and high sensitivity (4 ng/μl dsDNA) combined with reasonable resolution (17 bases)...
April 20, 2017: Electrophoresis
https://www.readbyqxmd.com/read/28424530/molecular-diagnostic-assays-based-on-cpn60-ut-sequences-reveal-the-geographic-distribution-of-subgroup-16srxiii-a-i-i-phytoplasma-in-mexico
#7
Edel Pérez-López, Douglas Rodríguez-Martínez, Chrystel Y Olivier, Mauricio Luna-Rodríguez, Tim J Dumonceaux
Geographically diverse samples from strawberry exhibiting symptoms of Strawberry Green Petal (SbGP), periwinkle plants with virescence, and blackberry, blueberry, and raspberry plants displaying yellowing and inedible fruits, were assayed for the presence of phytoplasma DNA. PCR targeting the 16S rRNA-encoding gene and chaperonin-60 (cpn60) showed that the plants were infected with phytoplasma subgroup16SrXIII-(A/I)I (SbGP/MPV). To examine the geographic distribution of this pathogen in Mexico, we designed an array of cpn60-targeted molecular diagnostic assays for SbGP/MPV phytoplasma...
April 19, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28423028/four-human-plasmodium-species-quantification-using-droplet-digital-pcr
#8
Suttipat Srisutham, Naowarat Saralamba, Benoit Malleret, Laurent Rénia, Arjen M Dondorp, Mallika Imwong
Droplet digital polymerase chain reaction (ddPCR) is a partial PCR based on water-oil emulsion droplet technology. It is a highly sensitive method for detecting and delineating minor alleles from complex backgrounds and provides absolute quantification of DNA targets. The ddPCR technology has been applied for detection of many pathogens. Here the sensitive assay utilizing ddPCR for detection and quantification of Plasmodium species was investigated. The assay was developed for two levels of detection, genus specific for all Plasmodium species and for specific Plasmodium species detection...
2017: PloS One
https://www.readbyqxmd.com/read/28419517/comparative-analysis-between-rq-pcr-and-digital-droplet-pcr-of-bcl2-igh-gene-rearrangement-in-the-peripheral-blood-and-bone-marrow-of-early-stage-follicular-lymphoma
#9
Marzia Cavalli, Lucia Anna De Novi, Irene Della Starza, Luca Vincenzo Cappelli, Vittorio Nunes, Alessandro Pulsoni, Ilaria Del Giudice, Anna Guarini, Robin Foà
BCL2/IGH rearrangements were analysed by polymerase chain reaction (PCR) at diagnosis in paired peripheral blood (PB) and bone marrow (BM) samples from 67 patients with stage I/II follicular lymphoma (FL). Real time quantitative PCR (RQ-PCR) and digital droplet PCR (ddPCR) were performed in cases with a major breakpoint region (MBR+) at diagnosis and after localized radiotherapy and rituximab administration in order to investigate the applicability of ddPCR. The overall ddPCR/RQ-PCR concordance was 81·9% (113/138 samples) and 97·5% in the 40/138 with quantifiable disease (RQ-PCR≥10(-5) )...
April 17, 2017: British Journal of Haematology
https://www.readbyqxmd.com/read/28414780/sensitive-quantification-of-the-hiv-1-reservoir-in-gut-associated-lymphoid-tissue
#10
Sara Morón-López, Maria C Puertas, Cristina Gálvez, Jordi Navarro, Anna Carrasco, Maria Esteve, Josep Manyé, Manel Crespo, Maria Salgado, Javier Martinez-Picado
BACKGROUND: The implementation of successful strategies to achieve an HIV cure has become a priority in HIV research. However, the current location and size of HIV reservoirs is still unknown since there are limited tools to evaluate HIV latency in viral sanctuaries such as gut-associated lymphoid tissue (GALT). As reported in the so called "Boston Patients", despite undetectable levels of proviral HIV-1 DNA in blood and GALT, viral rebound happens in just few months after ART interruption...
2017: PloS One
https://www.readbyqxmd.com/read/28414286/-pcr-analysis-of-the-absolute-number-of-copies-of-human-chromosome-18-transcripts-in-liver-and-hepg2-cells
#11
Y Y Kiseleva, K G Ptitsyn, O V Tikhonova, S P Radko, L K Kurbatov, I V Vakhrushev, V G Zgoda, E A Ponomarenko, A V Lisitsa, A I Archakov
Using reverse transcription in conjunction with the quantitative real-time PCR or digital droplet PCR, the transcriptome profiling of human chromosome 18 has been carried out in liver hepatocytes and hepatoblastoma cells (HepG2 cell line) in terms of the absolute number of each transcript per cell. The transcript abundance varies within the range of 0.006 to 9635 and 0.011 to 4819 copies per cell for HepG2 cell line and hepatocytes, respectively. The expression profiles for genes of chromosome 18 in hepatocytes and HepG2 cells were found to significantly correlate: the Spearman's correlation coefficient was equal to 0...
March 2017: Biomedit︠s︡inskai︠a︡ Khimii︠a︡
https://www.readbyqxmd.com/read/28414163/droplet-digital-pcr-ddpcr-vs-quantitative-real-time-pcr-qpcr-approach-for-detection-and-quantification-of-merkel-cell-polyomavirus-mcpyv-dna-in-formalin-fixed-paraffin-embedded-ffpe-cutaneous-biopsies
#12
Rosaria Arvia, Mauro Sollai, Federica Pierucci, Carmelo Urso, Daniela Massi, Krystyna Zakrzewska
BACKGROUND: Merkel cell polyomavirus (MCPyV) is associated with Merkel cell carcinoma and high viral load in the skin was proposed as a risk factor for the occurrence of this tumour. MCPyV DNA was detected, with lower frequency, in different skin cancers but since the viral load was usually low, the real prevalence of viral DNA could be underestimated. OBJECTIVE: To evaluate the performance of two assays (qPCR and ddPCR) for MCPyV detection and quantification in formalin fixed paraffin embedded (FFPE) tissue samples...
April 14, 2017: Journal of Virological Methods
https://www.readbyqxmd.com/read/28412176/human-parvovirus-b19-nosocomial-outbreak-in-healthcare-personnel-in-a-paediatric-ward-at-a-national-tertiary-referral-centre-in-thailand
#13
S Sungkate, W Phongsamart, S Rungmaitree, K Lapphra, O Wittawatmongkol, V Pumsuwan, N Wiruchkul, S Assanasen, Y Rongrungruang, N Onlamoon, N Horthongkham, W Lermankul, N Kongstan, K Chokephaibulkit
BACKGROUND: Nosocomial outbreaks of parvovirus B19 (pB19) have been reported, but they rarely occur among healthcare personnel (HCP). Susceptibility among pregnant HCP was the major concern. METHODS: An outbreak of pB19 among HCP is described in a paediatric ward with a cross-sectional serologic study in all HCP and patients exposed to the outbreak. Acute infection was diagnosed by polymerase chain reaction or positive anti-parvovirus B19 IgM. FINDINGS: Among 48 HCP (three pregnant) and 22 patients included in the outbreak serologic study, 11 (23%) HCP and two (9%) patients had acute infection...
March 18, 2017: Journal of Hospital Infection
https://www.readbyqxmd.com/read/28405680/optimal-method-for-quantitative-detection-of-plasma-egfr-t790m-mutation-using-droplet-digital-pcr-system
#14
Ken Suzawa, Hiromasa Yamamoto, Kadoaki Ohashi, Shinsuke Hashida, Shuta Tomida, Toshio Kubo, Yuho Maki, Junichi Soh, Kazunori Tsukuda, Katsuyuki Kiura, Shinichiro Miyoshi, Shinichi Toyooka
Though patients with EGFR mutations are initially responsive to EGFR-tyrosine kinase inhibitors (TKIs), most tumors ultimately acquire resistance to EGFR-TKIs. The most frequently reported mechanism is EGFR T790M mutation. In this study, using a droplet digital PCR (ddPCR) system, we assessed optimal conditions for a mutation detection assay for EGFR T790M obtained from circulating cell-free DNA (cfDNA) in plasma. The advantages of locked nucleic acids (LNA) probe, short amplicon size, and blocking oligo using peptide nucleic acids (PNA) were assessed using control DNAs from cell lines to improve the sensitivity of mutation detection...
May 2017: Oncology Reports
https://www.readbyqxmd.com/read/28402007/interactions-of-bovine-oocytes-with-follicular-elements-with-respect-to-lipid-metabolism
#15
Ewelina Warzych, Piotr Pawlak, Marcin Pszczola, Adam Cieslak, Zofia E Madeja, Dorota Lechniak
Among many factors, lipid metabolism within the follicular environment emerges as an important indicator of oocyte quality. In the literature a crucial significance is described concerning follicular fluid (FF) composition as well as messenger RNA (mRNA) expression in follicular cells. The aim of this study was to describe the relationship between oocyte, FF and follicular cells with regard to lipid metabolism. The set of data originating from individual follicles comprised: lipid droplets (LD) number in oocytes (BODIPY staining), mRNA expression of seven genes in cumulus and granulosa cells (SCD, FADS2, ELOVL2, ELOVL5, GLUT1, GLUT3, GLUT8; real time polymerase chain reaction) and fatty acid (FA) composition in FF (gas chromatography)...
April 12, 2017: Animal Science Journal, Nihon Chikusan Gakkaihō
https://www.readbyqxmd.com/read/28397990/germline-transformation-of-the-western-corn-rootworm-diabrotica-virgifera-virgifera
#16
F Chu, W Klobasa, P Wu, S Pinzi, N Grubbs, S Gorski, Y Cardoza, M D Lorenzen
The western corn rootworm (WCR), a major pest of maize, is notorious for rapidly adapting biochemically, behaviourally and developmentally to a variety of control methods. Despite much effort, the genetic basis of WCR adaptation remains a mystery. Since transformation-based applications such as transposon tagging and enhancer trapping have facilitated genetic dissection of model species such as Drosophila melanogaster, we developed a germline-transformation system for WCR in an effort to gain a greater understanding of the basic biology of this economically important insect...
April 11, 2017: Insect Molecular Biology
https://www.readbyqxmd.com/read/28393575/using-circulating-cell-free-dna-to-monitor-personalized-cancer-therapy
#17
Michael Oellerich, Ekkehard Schütz, Julia Beck, Philipp Kanzow, Piers N Plowman, Glen J Weiss, Philip D Walson
High-quality genomic analysis is critical for personalized pharmacotherapy in patients with cancer. Tumor-specific genomic alterations can be identified in cell-free DNA (cfDNA) from patient blood samples and can complement biopsies for real-time molecular monitoring of treatment, detection of recurrence, and tracking resistance. cfDNA can be especially useful when tumor tissue is unavailable or insufficient for testing. For blood-based genomic profiling, next-generation sequencing (NGS) and droplet digital PCR (ddPCR) have been successfully applied...
April 10, 2017: Critical Reviews in Clinical Laboratory Sciences
https://www.readbyqxmd.com/read/28385781/monitoring-of-multiple-myeloma-by-quantification-of-recurrent-mutations-in-serum
#18
Even Holth Rustad, Eivind Coward, Emilie R Skytøen, Kristine Misund, Toril Holien, Therese Standal, Magne Børset, Vidar Beisvag, Ola Myklebost, Leonardo A Meza-Zepeda, Hong Yan Dai, Anders Sundan, Anders Waage
Circulating tumor DNA (ctDNA) is a promising biomarker to monitor tumor load and genome alterations. We have explored the presence of ctDNA in multiple myeloma patients and its relation to disease activity during long-term follow-up. We used digital droplet PCR to monitor recurrent mutations, mainly in mitogen activated protein kinase pathway genes NRAS, KRAS and BRAF. Mutations were identified by next generation sequencing or PCR of bone marrow plasma cells, and their presence analyzed in 251 archived serum samples obtained from 20 patients during up to 7 years...
April 6, 2017: Haematologica
https://www.readbyqxmd.com/read/28378527/the-prognostic-value-of-simultaneous-tumor-and-serum-ras-raf-mutations-in-localized-colon-cancer
#19
Caroline Emilie B Thomsen, Ane Lindegaard Appelt, Rikke Fredslund Andersen, Jan Lindebjerg, Lars Henrik Jensen, Anders Jakobsen
The impact of RAS/RAF mutations in localized colon cancer needs clarification. Based on analysis of tumor-specific DNA, this study aimed at elucidating the prognostic influence of mutational status in tumor and serum using an extended panel of mutations. The study retrospectively included 294 patients with curatively resected stage I-III adenocarcinoma of the colon. Mutations in tumor and serum were determined at time of surgery. Analyses were performed with droplet digital PCR technology. Hazard ratio (HR) for the association between mutational status and survival was estimated in multivariate analysis taking known prognostic factors into account...
April 4, 2017: Cancer Medicine
https://www.readbyqxmd.com/read/28377872/dna-methylation-alters-transcriptional-rates-of-differentially-expressed-genes-and-contributes-to-pathophysiology-in-mice-fed-a-high-fat-diet
#20
Pili Zhang, Tianjiao Chu, N Dedousis, Benjamin S Mantell, Ian Sipula, Lucy Li, Kimberly D Bunce, Patricia A Shaw, Liora S Katz, Jun Zhu, Carmen Argmann, Robert M O'Doherty, David G Peters, Donald K Scott
OBJECTIVE: Overnutrition can alter gene expression patterns through epigenetic mechanisms that may persist through generations. However, it is less clear if overnutrition, for example a high fat diet, modifies epigenetic control of gene expression in adults, or by what molecular mechanisms, or if such mechanisms contribute to the pathology of the metabolic syndrome. Here we test the hypothesis that a high fat diet alters hepatic DNA methylation, transcription and gene expression patterns, and explore the contribution of such changes to the pathophysiology of obesity...
April 2017: Molecular Metabolism
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