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https://www.readbyqxmd.com/read/28932544/the-detectability-of-the-pretreatment-egfr-t790m-mutations-in-lung-adenocarcinoma-using-cast-pcr-and-digital-pcr
#1
Tsutomu Tatematsu, Katsuhiro Okuda, Ayumi Suzuki, Risa Oda, Tadashi Sakane, Osamu Kawano, Hiroshi Haneda, Satoru Moriyama, Hidefumi Sasaki, Ryoichi Nakanishi
BACKGROUND: A gatekeeper T790M mutation is thought to cause resistance to epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) treatment. The detection of a 2nd mutation is important for planning the next therapy when patients acquire resistance to the first line EGFR-TKI. METHODS: We used a competitive allele-specific polymerase chain reaction (CAST-PCR) to analyze the incidence and clinical significance of T790M mutations in 153 lung adenocarcinomas with EGFR-activating mutations...
August 2017: Journal of Thoracic Disease
https://www.readbyqxmd.com/read/28928368/detection-of-clinically-relevant-copy-number-alterations-in-oral-cancer-progression-using-multiplexed-droplet-digital-pcr
#2
Curtis B Hughesman, X J David Lu, Kelly Y P Liu, Yuqi Zhu, Rebecca M Towle, Charles Haynes, Catherine F Poh
Copy number alterations (CNAs), a common genomic event during carcinogenesis, are known to affect a large fraction of the genome. Common recurrent gains or losses of specific chromosomal regions occur at frequencies that they may be considered distinctive features of tumoral cells. Here we introduce a novel multiplexed droplet digital PCR (ddPCR) assay capable of detecting recurrent CNAs that drive tumorigenesis of oral squamous cell carcinoma. Applied to DNA extracted from oral cell lines and clinical samples of various disease stages, we found good agreement between CNAs detected by our ddPCR assay with those previously reported using comparative genomic hybridization or single nucleotide polymorphism arrays...
September 19, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28923315/development-of-a-digital-droplet-pcr-assay-to-measure-hbv-dna-in-patients-receiving-long-term-tdf-treatment
#3
Yang Liu, Andrea L Cathcart, William E Delaney, Kathryn M Kitrinos
The COBAS TaqMan assay has a lower limit of quantification (LLOQ) of 169 HBV copies/mL and a lower limit of detection (LLOD) of 58 copies/mL. HBV DNA below the TaqMan LLOQ is classified as target not detected (TND) (<58 copies/mL) or target detected (TD) (between 58 to 169 copies/mL). Here we have developed a more sensitive digital droplet PCR (ddPCR) assay to evaluate the impact of long-term tenofovir disoproxil fumarate (TDF) treatment in patients that did or did not achieve HBsAg seroconversion. A ddPCR assay was developed to detect HBV DNA to 8 copies/mL...
September 15, 2017: Journal of Virological Methods
https://www.readbyqxmd.com/read/28921124/droplet-volume-variability-as-a-critical-factor-for-accuracy-of-absolute-quantification-using-droplet-digital-pcr
#4
Alexandra Bogožalec Košir, Carla Divieto, Jernej Pavšič, Stefano Pavarelli, David Dobnik, Tanja Dreo, Roberto Bellotti, Maria Paola Sassi, Jana Žel
Accurate and precise nucleic-acid quantification is crucial for clinical and diagnostic decisions, as overestimation or underestimation can lead to misguided treatment of a disease or incorrect labelling of the products. Digital PCR is one of the best tools for absolute nucleic-acid copy-number determination. However, digital PCR needs to be well characterised in terms of accuracy and sources of uncertainty. With droplet digital PCR, discrepancies between the droplet volume assigned by the manufacturer and measured by independent laboratories have already been shown in previous studies...
September 18, 2017: Analytical and Bioanalytical Chemistry
https://www.readbyqxmd.com/read/28915283/clinical-evaluation-of-a-multi-modal-facial-serum-that-addresses-hyaluronic-acid-levels-in-skin
#5
Susana Raab, Margarita Yatskayer, Stephen Lynch, Megan Manco, Christian Oresajo
BACKGROUND: Hyaluronic acid (HA), the major glycosaminoglycan present in the human skin, is a key contributor to water retention and mechanical support in skin. The level, size, and functionality of cutaneous HA are known to diminish with age. Topical treatments designed to increase the HA content of skin have been met with limited success. The purpose of this study was to evaluate the tolerance and efficacy of a multi-modal facial serum containing HA, Proxylane (C-Xyloside), purple rice extract, and dipotassium glycyrrhizate in addressing HA levels in skin...
September 1, 2017: Journal of Drugs in Dermatology: JDD
https://www.readbyqxmd.com/read/28915237/dna-replication-stress-restricts-ribosomal-dna-copy-number
#6
Devika Salim, William D Bradford, Amy Freeland, Gillian Cady, Jianmin Wang, Steven C Pruitt, Jennifer L Gerton
Ribosomal RNAs (rRNAs) in budding yeast are encoded by ~100-200 repeats of a 9.1kb sequence arranged in tandem on chromosome XII, the ribosomal DNA (rDNA) locus. Copy number of rDNA repeat units in eukaryotic cells is maintained far in excess of the requirement for ribosome biogenesis. Despite the importance of the repeats for both ribosomal and non-ribosomal functions, it is currently not known how "normal" copy number is determined or maintained. To identify essential genes involved in the maintenance of rDNA copy number, we developed a droplet digital PCR based assay to measure rDNA copy number in yeast and used it to screen the yeast conditional temperature-sensitive mutant collection of essential genes...
September 15, 2017: PLoS Genetics
https://www.readbyqxmd.com/read/28910375/application-of-droplet-digital-pcr-for-quantitative-detection-of-spiroplasma-citri-in-comparison-with-real-time-pcr
#7
Yogita Maheshwari, Vijayanandraj Selvaraj, Subhas Hajeri, Raymond Yokomi
Droplet digital polymerase chain reaction (ddPCR) is a method for performing digital PCR that is based on water-oil emulsion droplet technology. It is a unique approach to measure the absolute copy number of nucleic acid targets without the need of external standards. This study evaluated the applicability of ddPCR as a quantitative detection tool for the Spiroplasma citri, causal agent of citrus stubborn disease (CSD) in citrus. Two sets of primers, SP1, based on the spiral in housekeeping gene, and a multicopy prophage gene, SpV1 ORF1, were used to evaluate ddPCR in comparison with real time (quantitative) PCR (qPCR) for S...
2017: PloS One
https://www.readbyqxmd.com/read/28906277/effect-of-cytomegalovirus-and-epstein-barr-virus-replication-on-intestinal-mucosal-gene-expression-and-microbiome-composition-of-hiv-infected-and-uninfected-individuals
#8
Sara Gianella, Antoine Chaillon, Ece A Mutlu, Phillip A Engen, Robin M Voigt, Ali Keshavarzian, John Losurdo, Prachi Chakradeo, Steven M Lada, Masato Nakazawa, Alan L Landay
BACKGROUND: HIV-infection is associated with dramatic changes in the intestinal mucosa. The impact of other viral pathogens is unclear. METHODS: One hundred and eight (108) biopsies from left and right colon (n = 79) and terminal ileum (n = 29) were collected from 19 HIV-infected and 22 HIV-uninfected participants. Levels of cytomegalovirus (CMV) and Epstein-Barr virus (EBV) DNA were measured by droplet digital PCR. Mucosal gene expression was measured via multiplex-assay...
September 24, 2017: AIDS
https://www.readbyqxmd.com/read/28905136/highly-sensitive-detection-of-esr1-mutations-in-cell-free-dna-from-patients-with-metastatic-breast-cancer-using-molecular-barcode-sequencing
#9
Nanae Masunaga, Naofumi Kagara, Daisuke Motooka, Shota Nakamura, Tomohiro Miyake, Tomonori Tanei, Yasuto Naoi, Masafumi Shimoda, Kenzo Shimazu, Seung Jin Kim, Shinzaburo Noguchi
PURPOSE: We aimed to develop a highly sensitive method to detect ESR1 mutations in cell-free DNA (cfDNA) using next-generation sequencing with molecular barcode (MB-NGS) targeting the hotspot segment (c.1600-1713). METHODS: The sensitivity of MB-NGS was tested using serially diluted ESR1 mutant DNA and then cfDNA samples from 34 patients with metastatic breast cancer were analyzed with MB-NGS. The results of MB-NGS were validated in comparison with conventional NGS and droplet digital PCR (ddPCR)...
September 13, 2017: Breast Cancer Research and Treatment
https://www.readbyqxmd.com/read/28902411/functional-changes-in-the-neural-retina-occur-in-the-absence-of-mitochondrial-dysfunction-in-a-rodent-model-of-diabetic-retinopathy
#10
Dustin R Masser, Laura Otalora, Nicholas W Clark, Michael T Kinter, Michael H Elliott, Willard M Freeman
Diabetic retinopathy is a neurovascular diabetes complication resulting in vision loss. A wealth of literature reports retinal molecular changes indicative of neural deficits, inflammation, and vascular leakage with chronic diabetes, but the mechanistic causes of disease initiation and progression are unknown. Microvascular mitochondrial DNA (mtDNA) damage leading to mitochondrial dysfunction has been proposed to drive vascular dysfunction in retinopathy. However, growing evidence suggests that neural retina dysfunction precedes and may cause vascular damage...
September 13, 2017: Journal of Neurochemistry
https://www.readbyqxmd.com/read/28900844/a-pilot-study-on-the-use-of-cerebrospinal-fluid-cell-free-dna-in-intramedullary-spinal-ependymoma
#11
Ian David Connolly, Yingmei Li, Wenying Pan, Eli Johnson, Linya You, Hannes Vogel, John Ratliff, Melanie Hayden Gephart
Cerebrospinal fluid (CSF) represents a promising source of cell-free DNA (cfDNA) for tumors of the central nervous system. A CSF-based liquid biopsy may obviate the need for riskier tissue biopsies and serve as a means for monitoring tumor recurrence or response to therapy. Spinal ependymomas most commonly occur in adults, and aggressive resection must be delicately balanced with the risk of injury to adjacent normal tissue. In patients with subtotal resection, recurrence commonly occurs. A CSF-based liquid biopsy matched to the patient's spinal ependymoma mutation profile has potential to be more sensitive then surveillance MRI, but the utility has not been well characterized for tumors of the spinal cord...
September 12, 2017: Journal of Neuro-oncology
https://www.readbyqxmd.com/read/28899979/clinical-performance-evaluation-of-the-idylla-nras-braf-mutation-test-on-retrospectively-collected-formalin-fixed-paraffin-embedded-colorectal-cancer-tissue
#12
Louise Johnston, Michael Power, Philip Sloan, Anna Long, Angela Silmon, Ben Chaffey, Andrea Jane Lisgo, Liam Little, Ellen Vercauteren, Torben Steiniche, Tine Meyer, John Simpson
AIMS: Understanding the molecular mechanisms of underlying disease has led to a movement away from the one-drug-fits-all paradigm towards treatment tailored to the genetic profile of the patient. The Biocartis Idylla platform is a novel fully automated, real-time PCR-based in vitro diagnostic system. The Idylla NRAS-BRAF mutation test has been developed for the qualitative detection of mutations in NRAS and BRAF oncogenes, facilitating genetic profiling of patients with cancer. The aim of this study was to carry out a formal clinical performance evaluation...
September 12, 2017: Journal of Clinical Pathology
https://www.readbyqxmd.com/read/28899967/circulating-cell-free-dna-for-metastatic-cervical-cancer-detection-genotyping-and-monitoring
#13
Zhigang Kang, Sanja Stevanović, Christian S Hinrichs, Liang Cao
PURPOSE: Circulating cell-free (ccf) human papillomavirus (HPV) DNA may serve as a unique tumor marker for HPV-associated malignancies, including cervical cancer. We developed a method to genotype and quantify circulating HPV DNA in patients with HPV16- or HPV18-positive metastatic cervical cancer for potential disease monitoring and treatment-related decision making. PATIENTS AND METHODS: In this retrospective study, HPV ccfDNA was measured in serum samples from 19 metastatic cervical cancer patients by duplex digital droplet (dd) PCR...
September 12, 2017: Clinical Cancer Research: An Official Journal of the American Association for Cancer Research
https://www.readbyqxmd.com/read/28898240/prevalence-of-signs-of-trachoma-ocular-chlamydia-trachomatis-infection-and-antibodies-to-pgp3-in-residents-of-kiritimati-island-kiribati
#14
Anaseini Cama, Andreas Müller, Raebwebwe Taoaba, Robert M R Butcher, Iakoba Itibita, Stephanie J Migchelsen, Tokoriri Kiauea, Harry Pickering, Rebecca Willis, Chrissy H Roberts, Ana Bakhtiari, Richard T Le Mesurier, Neal D E Alexander, Diana L Martin, Rabebe Tekeraoi, Anthony W Solomon
OBJECTIVE: In some Pacific Island countries, such as Solomon Islands and Fiji, active trachoma is common, but ocular Chlamydia trachomatis (Ct) infection and trachomatous trichiasis (TT) are rare. On Tarawa, the most populous Kiribati island, both the active trachoma sign "trachomatous inflammation-follicular" (TF) and TT are present at prevalences warranting intervention. We sought to estimate prevalences of TF, TT, ocular Ct infection, and anti-Ct antibodies on Kiritimati Island, Kiribati, to assess local relationships between these parameters, and to help determine the need for interventions against trachoma on Kiribati islands other than Tarawa...
September 12, 2017: PLoS Neglected Tropical Diseases
https://www.readbyqxmd.com/read/28891364/an-update-on-pcr-use-for-minimal-residual-disease-monitoring-in-acute-lymphoblastic-leukemia
#15
Vittorio Nunes, Gianni Cazzaniga, A Biondi
Acute lymphoblastic leukemia (ALL) is the first neoplasm where the assessment of early response to therapy by minimal residual disease (MRD) monitoring has proven to be a fundamental tool for guiding therapeutic choices. In recent years, thanks to real-time quantitative PCR (qPCR), MRD monitoring has further achieved higher levels of sensitivity and standardization. However, some outstanding issues still remain to be addressed and emerging technologies hold the promise of improving MRD detection in ALL patients...
September 11, 2017: Expert Review of Molecular Diagnostics
https://www.readbyqxmd.com/read/28881720/analysis-of-esr1-and-pik3ca-mutations-in-plasma-cell-free-dna-from-er-positive-breast-cancer-patients
#16
Takashi Takeshita, Yutaka Yamamoto, Mutsuko Yamamoto-Ibusuki, Mai Tomiguchi, Aiko Sueta, Keiichi Murakami, Yoko Omoto, Hirotaka Iwase
BACKGROUND: The measurement of ESR1 and PIK3CA mutations in plasma cell-free DNA (cfDNA) has been studied as a non-invasive method to quickly assess and monitor endocrine therapy (ET) resistant metastatic breast cancer (MBC) patients. METHODS: The subjects of this retrospective study were a total of 185 plasma samples from 86 estrogen receptor-positive BC patients, of which 151 plasma samples were from 69 MBC patients and 34 plasma samples were from 17 primary BC (PBC) patients...
August 8, 2017: Oncotarget
https://www.readbyqxmd.com/read/28881596/a-prediction-model-for-distinguishing-lung-squamous-cell-carcinoma-from-adenocarcinoma
#17
Hui Li, Zhengran Jiang, Qixin Leng, Fan Bai, Juan Wang, Xiaosong Ding, Yuehong Li, Xianghong Zhang, HongBin Fang, Harris G Yfantis, Lingxiao Xing, Feng Jiang
Accurate classification of squamous cell carcinoma (SCC) from adenocarcinoma (AC) of non-small cell lung cancer (NSCLC) can lead to personalized treatments of lung cancer. We aimed to develop a miRNA-based prediction model for differentiating SCC from AC in surgical resected tissues and bronchoalveolar lavage (BAL) samples. Expression levels of seven histological subtype-associated miRNAs were determined in 128 snap-frozen surgical lung tumor specimens by using reverse transcription-polymerase chain reaction (RT-PCR) to develop an optimal panel of miRNAs for acutely distinguishing SCC from AC...
August 1, 2017: Oncotarget
https://www.readbyqxmd.com/read/28868565/comparison-of-the-amplification-refractory-mutation-system-super-amplification-refractory-mutation-system-and-droplet-digital-pcr-for-t790%C3%A2-m-mutation-detection-in-non-small-cell-lung-cancer-after-failure-of-tyrosine-kinase-inhibitor-treatment
#18
Lucheng Zhu, Shirong Zhang, Yanping Xun, Yanping Jiang, Bing Xia, Xueqin Chen, Limin Wang, Hong Jiang, Shenglin Ma
Plasma mutation detection has the advantages of non-invasiveness and accessibility. Here, we evaluated three methods, the amplification refractory mutation system (ARMS), second-generation ARMS (SuperARMS), and droplet digital PCR (ddPCR), to assess their concordance and feasibility for the detection of mutations in plasma samples. Non-small lung cancer patients with stage IIIB/IV that were resistant to epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) treatment were enrolled. Blood samples were collected within 14 days after TKI resistance...
September 3, 2017: Pathology Oncology Research: POR
https://www.readbyqxmd.com/read/28866273/exploring-gene-expression-changes-in-the-amphioxus-gill-after-poly-i-c-challenge-using-digital-expression-profiling
#19
Qi-Lin Zhang, Han-Yue Qiu, Ming-Zhong Liang, Bang Luo, Xiu-Qiang Wang, Jun-Yuan Chen
Amphioxus, a cephalochordate, is a key model animal for studying the evolution of vertebrate immunity. Recently, studies have revealed that microRNA (miRNA) expression profiles change significantly in the amphioxus gill after immune stimulation, but it remains largely unknown how gene expression responds to immune stress. Elucidating gene expression changes in the amphioxus gill will provide a deeper understanding of the evolution of gill immunity in vertebrates. Here, we used high-throughput RNA sequencing technology (RNA-seq) to conduct tag-based digital gene expression profiling (DGE) analyses of the gills of control Branchiostoma belcheri and of those exposed to the viral mimic, poly(I:C) (pIC)...
September 1, 2017: Fish & Shellfish Immunology
https://www.readbyqxmd.com/read/28860183/single-step-qpcr-and-dpcr-detection-of-diverse-crispr-cas9-gene-editing-events-in-vivo
#20
Micol Falabella, Linqing Sun, Justin Barr, Andressa Z Pena, Erin E Kershaw, Sebastien Gingras, Elena A Goncharova, Brett A Kaufman
CRISPR-Cas9 based technology is currently the most flexible means to create targeted mutations by recombination or indel mutations by non-homologous end joining. During mouse transgenesis, recombinant and indel alleles are often pursued simultaneously. Multiple alleles can be formed in each animal to create significant genetic complexity that complicates the CRISPR-Cas9 approach and analysis. Currently, there are no rapid methods to measure the extent of on-site editing with broad mutation sensitivity. In this study, we demonstrate the allelic diversity arising from targeted CRISPR-editing in founder mice...
August 31, 2017: G3: Genes—Genomes—Genetics
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