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https://www.readbyqxmd.com/read/28088212/ultrasensitive-detection-of-oncogenic-human-papillomavirus-in-oropharyngeal-tissue-swabs
#1
Andre Isaac, Morris Kostiuk, Han Zhang, Cameron Lindsay, Fawaz Makki, Daniel A O'Connell, Jeffrey R Harris, David W J Cote, Hadi Seikaly, Vincent L Biron
BACKGROUND: The incidence of oropharyngeal squamous cell carcinoma (OPSCC) caused by oncogenic human papillomavirus (HPV) is rising worldwide. HPV-OPSCC is commonly diagnosed by RT-qPCR of HPV E6 and E7 oncoproteins or by p16 immunohistochemistry (IHC). Droplet digital PCR (ddPCR) has been recently reported as an ultra-sensitive and highly precise method of nucleic acid quantification for biomarker analysis. To validate the use of a minimally invasive assay for detection of oncogenic HPV based on oropharyngeal swabs using ddPCR...
January 14, 2017: Journal of Otolaryngology—Head & Neck Surgery
https://www.readbyqxmd.com/read/28073896/patient-specific-circulating-tumor-dna-detection-during-neoadjuvant-chemotherapy-in-triple-negative-breast-cancer
#2
Francesca Riva, Francois-Clement Bidard, Alexandre Houy, Adrien Saliou, Jordan Madic, Aurore Rampanou, Caroline Hego, Maud Milder, Paul Cottu, Marie-Paule Sablin, Anne Vincent-Salomon, Olivier Lantz, Marc-Henri Stern, Charlotte Proudhon, Jean-Yves Pierga
BACKGROUND: In nonmetastatic triple-negative breast cancer (TNBC) patients, we investigated whether circulating tumor DNA (ctDNA) detection can reflect the tumor response to neoadjuvant chemotherapy (NCT) and detect minimal residual disease after surgery. METHODS: Ten milliliters of plasma were collected at 4 time points: before NCT; after 1 cycle; before surgery; after surgery. Customized droplet digital PCR (ddPCR) assays were used to track tumor protein p53 (TP53) mutations previously characterized in tumor tissue by massively parallel sequencing...
January 10, 2017: Clinical Chemistry
https://www.readbyqxmd.com/read/28069289/liquid-biopsy-analysis-of-fgfr3-and-pik3ca-hotspot-mutations-for-disease-surveillance-in-bladder-cancer
#3
Emil Christensen, Karin Birkenkamp-Demtröder, Iver Nordentoft, Søren Høyer, Kirstin van der Keur, Kim van Kessel, Ellen Zwarthoff, Mads Agerbæk, Torben Falck Ørntoft, Jørgen Bjerggaard Jensen, Lars Dyrskjøt
BACKGROUND: Disease surveillance in patients with bladder cancer is important for early diagnosis of progression and metastasis and for optimised treatment. OBJECTIVE: To develop urine and plasma assays for disease surveillance for patients with FGFR3 and PIK3CA tumour mutations. DESIGN, SETTING, AND PARTICIPANTS: Droplet digital polymerase chain reaction (ddPCR) assays were developed and tumour DNA from two patient cohorts was screened for FGFR3 and PIK3CA hotspot mutations...
January 6, 2017: European Urology
https://www.readbyqxmd.com/read/28061461/estimation-of-cell-free-circulating-egfr-mutation-concentration-predicts-outcomes-in-nsclc-patients-treated-with-egfr-tkis
#4
Yan-Juan Zhu, Hai-Bo Zhang, Yi-Hong Liu, Fu-Li Zhang, Ya-Zhen Zhu, Yong Li, Jian-Ping Bai, Li-Rong Liu, Yan-Chun Qu, Xin Qu, Xian Chen, Yan Li, Guang-Juan Zheng
Detection of circulating tumor DNA using droplet digital polymerase chain reaction (ddPCR) is a highly-sensitive, minimally invasive alternative to serial biopsies for assessment and management of cancer. We used ddPCR to assess the utility of measuring plasma concentrations of common epidermal growth factor receptor (EGFR) mutations (L858R, exon 19 deletion, and T790M) in 57 non-small cell lung cancer (NSCLC) patients treated with EGFR tyrosine kinase inhibitors (EGFR-TKIs). High baseline plasma EGFR mutation (pEGFRmut) concentrations were associated with shorter progression-free survival (8...
January 4, 2017: Oncotarget
https://www.readbyqxmd.com/read/28059126/an-exploratory-study-of-predisposing-genetic-factors-for-digeorge-velocardiofacial-syndrome
#5
Laia Vergés, Francesca Vidal, Esther Geán, Alexandra Alemany-Schmidt, Maria Oliver-Bonet, Joan Blanco
DiGeorge/velocardiofacial syndrome (DGS/VCFS) is a disorder caused by a 22q11.2 deletion mediated by non-allelic homologous recombination (NAHR) between low-copy repeats (LCRs). We have evaluated the role of LCR22 genomic architecture and PRDM9 variants as DGS/VCFS predisposing factors. We applied FISH using fosmid probes on chromatin fibers to analyze the number of tandem repeat blocks in LCR22 in two DGS/VCFS fathers-of-origin with proven 22q11.2 NAHR susceptibility. Results revealed copy number variations (CNVs) of L9 and K3 fosmids in these individuals compared to controls...
January 6, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28052016/total-dna-input-is-a-crucial-determinant-of-the-sensitivity-of-plasma-cell-free-dna-egfr-mutation-detection-using-droplet-digital-pcr
#6
Yu Zhang, Yan Xu, Wei Zhong, Jing Zhao, Minjiang Chen, Li Zhang, Longyun Li, Mengzhao Wang
We evaluated the use of droplet digital PCR (ddPCR) to detect plasma cell-free DNA (cfDNA) epidermal growth factor receptor (EGFR) mutations in advanced non-small cell lung cancer (NSCLC) patients. Compared with tumor-tissue-based detection, the sensitivity of ddPCR for detecting plasma cfDNA tyrosine kinase inhibitor (TKI)-sensitizing EGFR mutations was 61.3%, the specificity was 96.7%, and the consistency rate was 81.4% (κ=0.605, 95% confidence interval: 0.501-0.706, p <0.0001). The sensitivity declined from 82...
December 30, 2016: Oncotarget
https://www.readbyqxmd.com/read/28039535/use-of-droplet-digital-pcr-for-quantitative-and-automatic-analysis-of-the-her2-status-in-breast-cancer-patients
#7
Kazutaka Otsuji, Takeshi Sasaki, Atsushi Tanaka, Akiko Kunita, Masako Ikemura, Keisuke Matsusaka, Keiichiro Tada, Masashi Fukayama, Yasuyuki Seto
PURPOSE: Digital polymerase chain reaction (dPCR) has been used to yield an absolute measure of nucleic acid concentrations. Recently, a new method referred to as droplet digital PCR (ddPCR) has gained attention as a more precise and less subjective assay to quantify DNA amplification. We demonstrated the usefulness of ddPCR to determine HER2 gene amplification of breast cancer. METHODS: In this study, we used ddPCR to measure the HER2 gene copy number in clinical formalin-fixed paraffin-embedded samples of 41 primary breast cancer patients...
December 30, 2016: Breast Cancer Research and Treatment
https://www.readbyqxmd.com/read/28012713/accurate-quantification-of-t-cells-by-measuring-loss-of-germline-t-cell-receptor-loci-with-generic-single-duplex-droplet-digital-pcr-assays
#8
Willem H Zoutman, Rogier J Nell, Mieke Versluis, Debby van Steenderen, Rajshri N Lalai, Jacoba J Out-Luiting, Mark J de Lange, Maarten H Vermeer, Anton W Langerak, Pieter A van der Velden
Quantifying T cells accurately in a variety of tissues of benign, inflammatory, or malignant origin can be of great importance in a variety of clinical applications. Flow cytometry and immunohistochemistry are considered to be gold-standard methods for T-cell quantification. However, these methods require fresh, frozen, or fixated cells and tissue of a certain quality. In addition, conventional and droplet digital PCR (ddPCR), whether followed by deep sequencing techniques, have been used to elucidate T-cell content by focusing on rearranged T-cell receptor (TCR) genes...
December 21, 2016: Journal of Molecular Diagnostics: JMD
https://www.readbyqxmd.com/read/28007639/in-vitro-combinatory-effects-of-the-alternaria-mycotoxins-alternariol-and-altertoxin-ii-and-potentially-involved-mirnas
#9
Katharina Vejdovszky, Matej Sack, Katharina Jarolim, Georg Aichinger, Mark M Somoza, Doris Marko
Alternariol (AOH) and altertoxin II (ATX II) are mycotoxins formed by Alternaria spp. Since they are expected to co-occur in Alternaria-infested food and feed, we addressed the question of combinatory effects. In addition, potentially involved regulatory microRNAs were surveyed in an exploratory approach. Cytotoxicity measurements in constant ratio combinations of 1:10 or 1:1 (ATX II: AOH) mainly revealed additive effects in HepG2, HT29 and HCEC-1CT cells. Yet, in specific high doses antagonism was found. Microarray analysis of miRNA expression profiles in HepG2 cells indicated different patterns of miRNA regulation by AOH and ATX II, including several miRNA species for which no distinct functions are currently known...
December 19, 2016: Toxicology Letters
https://www.readbyqxmd.com/read/28004117/a-blood-based-gene-expression-and-signaling-pathway-analysis-to-differentiate-between-high-and-low-grade-gliomas
#10
Stephen N Ponnampalam, Nor Rizan Kamaluddin, Zubaidah Zakaria, Vickneswaran Matheneswaran, Dharmendra Ganesan, Mohammed Saffari Haspani, Mina Ryten, John A Hardy
The aims of the present study were to undertake gene expression profiling of the blood of glioma patients to determine key genetic components of signaling pathways and to develop a panel of genes that could be used as a potential blood-based biomarker to differentiate between high and low grade gliomas, non-gliomas and control samples. In this study, blood samples were obtained from glioma patients, non-glioma and control subjects. Ten samples each were obtained from patients with high and low grade tumours, respectively, ten samples from non-glioma patients and twenty samples from control subjects...
January 2017: Oncology Reports
https://www.readbyqxmd.com/read/28002565/large-deletions-of-tspan12-cause-familial-exudative-vitreoretinopathy-fevr
#11
Soo Hyun Seo, Man Jin Kim, Sung Wook Park, Jeong Hun Kim, Young Suk Yu, Ji Yun Song, Sung Im Cho, Joo Hyun Ahn, Yeon Hee Oh, Jee-Soo Lee, Seungjun Lee, Moon-Woo Seong, Sung Sup Park, Ji Yeon Kim
Purpose: Familial exudative vitreoretinopathy (FEVR) is a rare, hereditary visual disorder. The gene TSPAN12 is associated with autosomal dominant inheritance of FEVR. The prevalence and impact of large deletions/duplications of TSPAN12 on FEVR patients is unknown. To glean better insight of TSPAN12 on FEVR pathology, herein, we describe three FEVR patients with TSPAN12 deletions. Methods: Thirty-three Korean FEVR patients, who previously screened negative for TSPAN12 mutations, mutations in other FEVR-associated genes such as NDP, FZD4, LRP5, and large deletions and duplications of NDP, FZD4, and LRP5, were selected for TSPAN12 large deletion and duplication analyses...
December 1, 2016: Investigative Ophthalmology & Visual Science
https://www.readbyqxmd.com/read/28000387/a-comparison-of-ddpcr-and-arms-for-detecting-egfr-t790m-status-in-ctdna-from-advanced-nsclc-patients-with%C3%A2-acquired-egfr-tki-resistance
#12
Wenxian Wang, Zhengbo Song, Yiping Zhang
A sensitive and convenient method for detecting epidermal growth factor receptor (EGFR) T790M mutations from circulating tumor DNA (ctDNA) in advanced non-small cell lung cancer (NSCLC) patients with acquired EGFR-TKI resistance would be desirable to direct patient sequential treatment strategy. A comparison of two platforms for detecting EGFR mutations in plasma ctDNA was undertaken. Plasma samples and tumor samples were collected from patients with acquired EGFR-TKI resistance in Zhejiang Cancer Hospital from December 2014 to December 2015...
December 20, 2016: Cancer Medicine
https://www.readbyqxmd.com/read/27993791/multiplex-krasg12-g13-mutation-testing-of-unamplified-cell-free-dna-from-the-plasma-of-patients-with-advanced-cancers-using-droplet-digital-polymerase-chain-reaction
#13
F Janku, H J Huang, T Fujii, D N Shelton, K Madwani, S Fu, A M Tsimberidou, S A Piha-Paul, J J Wheler, R G Zinner, A Naing, D S Hong, D D Karp, G Cabrilo, E S Kopetz, V Subbiah, R Luthra, B K Kee, C Eng, V K Morris, G A Karlin-Neumann, F Meric-Bernstam
BACKGROUND: Cell-free DNA (cfDNA) from plasma offers easily obtainable material for KRAS mutation analysis. Novel, multiplex, and accurate diagnostic systems using small amounts of DNA are needed to further the use of plasma cfDNA testing in personalized therapy. PATIENTS AND METHODS: Samples of 16 ng of unamplified plasma cfDNA from 121 patients with diverse progressing advanced cancers were tested with a KRAS (G12/G13) multiplex assay to detect the 7 most common mutations in the hotspot of exon 2 using droplet digital polymerase chain reaction (ddPCR)...
December 19, 2016: Annals of Oncology: Official Journal of the European Society for Medical Oncology
https://www.readbyqxmd.com/read/27992475/a-systematic-investigation-of-parameters-influencing-droplet-rain-in-the-listeria-monocytogenes-prfa-assay-reduction-of-ambiguous-results-in-ddpcr
#14
Anna Kristina Witte, Patrick Mester, Susanne Fister, Matthias Witte, Dagmar Schoder, Peter Rossmanith
The droplet digital polymerase chain reaction (ddPCR) determines DNA amounts based upon the pattern of positive and negative droplets, according to Poisson distribution, without the use of external standards. However, division into positive and negative droplets is often not clear because a part of the droplets has intermediate fluorescence values, appearing as "rain" in the plot. Despite the droplet rain, absolute quantification with ddPCR is possible, as shown previously for the prfA assay in quantifying Listeria monocytogenes...
2016: PloS One
https://www.readbyqxmd.com/read/27982132/sensitive-and-accurate-quantification-of-human-malaria-parasites-using-droplet-digital-pcr-ddpcr
#15
Cristian Koepfli, Wang Nguitragool, Natalie E Hofmann, Leanne J Robinson, Maria Ome-Kaius, Jetsumon Sattabongkot, Ingrid Felger, Ivo Mueller
Accurate quantification of parasite density in the human host is essential for understanding the biology and pathology of malaria. Semi-quantitative molecular methods are widely applied, but the need for an external standard curve makes it difficult to compare parasite density estimates across studies. Droplet digital PCR (ddPCR) allows direct quantification without the need for a standard curve. ddPCR was used to diagnose and quantify P. falciparum and P. vivax in clinical patients as well as in asymptomatic samples...
December 16, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27974721/liquid-biopsy-a-future-tool-for-posttreatment-surveillance-in-head-and-neck-cancer
#16
Joost H van Ginkel, Manon M H Huibers, Rob Noorlag, Remco de Bree, Robert J J van Es, Stefan M Willems
The prognosis of head and neck squamous cell carcinoma (HNSCC) is largely based on disease stage. Despite improvements in treatment, recurrence rates are still considered high. Currently, disease progression or regression after curative treatment is monitored by clinical evaluation combined with flexible endoscopy and/or imaging. However, specificity of imaging is low due to the posttreatment effects. Detection of circulating tumor DNA (ctDNA) from blood samples of HNSCC patients is a minimally invasive technique that could lead to an earlier detection of recurrence...
December 15, 2016: Pathobiology: Journal of Immunopathology, Molecular and Cellular Biology
https://www.readbyqxmd.com/read/27974546/copy-number-heterogeneity-of-jc-virus-standards
#17
Alexander L Greninger, Allen C Bateman, Ederlyn E Atienza, Sharon Wendt, Negar Makhsous, Keith R Jerome, Linda Cook
Quantitative PCR is a diagnostic mainstay of clinical virology, and accurate quantitation of viral load among labs requires the use of international standards. However, the use of multiple passages of viral isolates to obtain sufficient material for international standards may result in genomic changes that complicate their use as quantitative standards. We performed next-generation sequencing to gain single-nucleotide resolution and relative copy number of JC virus (JCV) clinical standards. Strikingly, the WHO international standard and Exact™ v1/v2 prototype standards for JCV showed 8-fold and 4-fold variation in genomic coverage between different loci in the viral genome, respectively, due to large deletions in the large T-antigen region...
December 14, 2016: Journal of Clinical Microbiology
https://www.readbyqxmd.com/read/27913434/a-study-of-tp53-rna-splicing-illustrates-pitfalls-of-rna-seq-methodology
#18
Sunali Mehta, Peter Tsai, Annette Lasham, Hamish Campbell, Roger Reddel, Antony Braithwaite, Cristin Print
TP53 undergoes multiple RNA-splicing events, resulting in at least nine mRNA transcripts encoding at least 12 functionally different protein isoforms. Antibodies specific to p53 protein isoforms have proven difficult to develop, thus researchers must rely on the transcript information to infer isoform abundance. In this study, we used deep RNA-seq, droplet digital PCR (ddPCR), and real-time quantitative reverse transcriptase PCR (RT-qPCR) from nine human cell lines and RNA-seq data available for tumors in The Cancer Genome Atlas to analyze TP53 splice variant expression...
December 15, 2016: Cancer Research
https://www.readbyqxmd.com/read/27894866/transcriptional-regulation-of-the-sodium-channel-gene-scn5a-by-gata4-in-human-heart
#19
Anna Tarradas, Mel Lina Pinsach-Abuin, Carlos Mackintosh, Oriol Llorà-Batlle, Alexandra Pérez-Serra, Montserrat Batlle, Félix Pérez-Villa, Thomas Zimmer, Ivan Garcia-Bassets, Ramon Brugada, Pedro Beltran-Alvarez, Sara Pagans
Aberrant expression of the sodium channel gene (SCN5A) has been proposed to disrupt cardiac action potential and cause human cardiac arrhythmias, but the mechanisms of SCN5A gene regulation and dysregulation still remain largely unexplored. To gain insight into the transcriptional regulatory networks of SCN5A, we surveyed the promoter and first intronic regions of the SCN5A gene, predicting the presence of several binding sites for GATA transcription factors (TFs). Consistent with this prediction, chromatin immunoprecipitation (ChIP) and sequential ChIP (Re-ChIP) assays show co-occupancy of cardiac GATA TFs GATA4 and GATA5 on promoter and intron 1 SCN5A regions in fresh-frozen human left ventricle samples...
November 26, 2016: Journal of Molecular and Cellular Cardiology
https://www.readbyqxmd.com/read/27878529/association-between-copy-number-variation-on-metabolic-phenotypes-and-hdl-c-levels-in-patients-with-polycystic-ovary-syndrome
#20
Birgit Knebel, Stefan Lehr, Onno E Janssen, Susanne Hahn, Sylvia Jacob, Ulrike Nitzgen, Dirk Müller-Wieland, Jorg Kotzka
Polygenic diseases with a broad phenotypic spectrum, such as polycystic ovary syndrome (PCOS), present a particular challenge in terms of identifying the underlying genetic mechanisms, nevertheless genetic variants have impact on the individual phenotype. We aimed to determine if next to genetic variations like SNPs further mechanisms might play a role in the pathogenesis of PCOS. We examined the effect of copy-number variations (CNVs) on metabolic phenotypes in PCOS. The intragenic rs1244979, rs2815752 in NEGR1 gene, and rs780094 in GCKR gene were genotyped and CNVs were determined by droplet digital polymerase chain reaction (ddPCR) in PCOS patients (n = 153) and controls without metabolic syndrome (n = 142)...
November 22, 2016: Molecular Biology Reports
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