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https://www.readbyqxmd.com/read/28714107/the-braf-and-nras-mutation-prevalence-in-dermoscopic-subtypes-of-acquired-naevi-reveals-constitutive-mapk-pathway-activation
#1
J M Tan, L N Tom, K Jagirdar, D Lambie, H Schaider, R A Sturm, H P Soyer, M S Stark
BACKGROUND: Acquired naevi can have unique dermoscopic patterns that correspond to distinct microanatomical growth patterns. Previous studies on acquired naevi stratified according to dermoscopic pattern, focused on the frequency of somatic BRAF mutations, whereas NRAS mutations remained to be elucidated. OBJECTIVES: To investigate the BRAF and NRAS mutation prevalence and activation of the MAPK pathway in distinct dermoscopic subtypes of acquired naevi. METHODS: Common mutations present in BRAF and NRAS were assessed in 40 globular, reticular, and peripheral rim of globules (PG) subtypes of acquired naevi from 27 participants (19 male, 8 female; mean age 46...
July 17, 2017: British Journal of Dermatology
https://www.readbyqxmd.com/read/28711990/assessment-of-braf-v600e-mutation-in-pulmonary-langerhans-cell-histiocytosis-in-tissue-biopsies-and-bronchoalveolar-lavages-by-droplet-digital-polymerase-chain-reaction
#2
Clémence Pierry, Charline Caumont, Elodie Blanchard, Camille Brochet, Gael Dournes, Audrey Gros, Thomas Bandres, Séverine Verdon, Marion Marty, Hugues Bégueret, Jean-Philippe Merlio
The neoplastic nature of pulmonary Langerhans cell histiocytosis (PLCH) is still debated. As the detection of BRAF (V600E) and MAP2K1 mutations in patients with PCLH is now considered for such assessment, the aim of our study was to evaluate digital droplet polymerase chain reaction (ddPCR) in PCLH diagnosis. We retrospectively analyzed BRAF(V600E) detection in a cohort of 42 PCLH tissues and 18 bronchoalveolar lavages (BALs) by ddPCR, immunohistochemistry, high-resolution melting PCR (HRM), and next-generation sequencing (NGS)...
July 15, 2017: Virchows Archiv: An International Journal of Pathology
https://www.readbyqxmd.com/read/28711173/presymptomatic-diagnosis-of-spinal-muscular-atrophy-through-newborn-screening
#3
Yin-Hsiu Chien, Shu-Chuan Chiang, Wen-Chin Weng, Ni-Chung Lee, Ching-Jie Lin, Wu-Shiun Hsieh, Wang-Tso Lee, Yuh-Jyh Jong, Tsang-Ming Ko, Wuh-Liang Hwu
OBJECTIVE: To demonstrate the feasibility of presymptomatic diagnosis of spinal muscular atrophy (SMA) through newborn screening (NBS). STUDY DESIGN: We performed a screening trial to assess all newborns who underwent routine newborn metabolic screening at the National Taiwan University Hospital newborn screening center between November 2014 and September 2016. A real-time polymerase chain reaction (RT-PCR) genotyping assay for the SMN1/SMN2 intron 7 c.888+100A/G polymorphism was performed to detect homozygous SMN1 deletion using dried blood spot (DBS) samples...
July 12, 2017: Journal of Pediatrics
https://www.readbyqxmd.com/read/28694163/mice-maintain-predominantly-maternal-g%C3%AE-s-expression-throughout-life-in-brown-fat-tissue-bat-but-not-other-tissues
#4
Olta Tafaj, Steven Hann, Ugur Ayturk, Matthew L Warman, Harald Jüppner
The murine Gnas (human GNAS) locus gives rise to Gαs and different splice variants thereof. The Gαs promoter is not methylated thus allowing biallelic expression in most tissues. In contrast, the alternative first Gnas/GNAS exons and their promoters undergo parent specific methylation, which limits transcription to the non-methylated allele. Pseudohypoparathyroidism type Ia (PHP1A) or type Ib (PHP1B) are caused by heterozygous maternal GNAS mutations suggesting that little or no Gαs is derived in some tissues from the non-mutated paternal GNAS thereby causing hormonal resistance...
July 8, 2017: Bone
https://www.readbyqxmd.com/read/28692472/quantification-of-immune-dysregulation-by-next-generation-polymerase-chain-reaction-to-improve-sepsis-diagnosis-in-surgical-patients
#5
Raquel Almansa, Alicia Ortega, Ana Ávila-Alonso, Maria Heredia-Rodríguez, Silvia Martín, Diana Benavides, Marta Martín-Fernandez, Lucia Rico, César Aldecoa, Jesús Rico, Iñigo López de Cenarruzabeitia, Juan Beltrán de Heredia, Esther Gomez-Sanchez, Marta Aragón, Cristina Andrés, Dolores Calvo, David Andaluz-Ojeda, Pilar Liu, Francisco Blanco-Antona, Lydia Blanco, Jose Ignacio Gómez-Herreras, Eduardo Tamayo, Jesus F Bermejo-Martin
OBJECTIVES: To quantify immunological dysfunction in surgical patients with presence/absence of sepsis using a droplet digital polymerase chain reaction (ddPCR) transcriptomic analysis. The study also aims to evaluate this approach for improving identification of sepsis in these patients. BACKGROUND: Immune dysregulation is a central event in sepsis. Quantification of the expression of immunological genes participating in the pathogenesis of sepsis could represent a new avenue to improve its diagnosis...
July 7, 2017: Annals of Surgery
https://www.readbyqxmd.com/read/28691390/an-improved-digital-polymerase-chain-reaction-protocol-to-capture-low-copy-kras-mutations-in-plasma-cell-free-dna-by-resolving-subsampling-issues
#6
REVIEW
Yusuke Ono, Ayumu Sugitani, Hidenori Karasaki, Munehiko Ogata, Reo Nozaki, Junpei Sasajima, Tomoki Yokochi, Shingo Asahara, Kazuya Koizumi, Kiyohiro Ando, Katsunori Hironaka, Tsutomu Daito, Yusuke Mizukami
Genetic alterations responsible for the initiation of cancer may serve as immediate biomarkers for early diagnosis. Plasma levels of cell-free DNA (cfDNA) in cancer patients are higher than those in healthy individuals, however, the major technical challenge for the widespread implementation of cfDNA genotyping as a diagnostic tool is the insufficient sensitivity and specificity of detecting early-stage tumors that shed low amounts of cfDNA. To establish a protocol for ultrasensitive droplet digital polymerase chain reaction (ddPCR) for quantification of low-frequency alleles within a limited cfDNA pool, two-step multiplex ddPCR targeting eight clinically relevant mutant KRAS variants was examined...
July 10, 2017: Molecular Oncology
https://www.readbyqxmd.com/read/28673857/development-and-application-of-a-reverse-transcriptase-droplet-digital-pcr-rt-ddpcr-for-sensitive-and-rapid-detection-of-japanese-encephalitis-virus
#7
Xulong Wu, Hua Lin, Shijie Chen, Lu Xiao, Miao Yang, Wei An, Yin Wang, Xueping Yao, Zexiao Yang
Japanese encephalitis (JE) is one of the most common zoonoses caused by Japanese encephalitis virus (JEV). Droplet digital PCR (ddPCR) is a novel sensitive, accurate method that enables absolute quantitation without the need for calibration curves. The aim of this study was to develop a RT-ddPCR method to detect JEV, and to compare its sensitivity with real-time TaqMan RT-PCR by analysis of clinical samples. The methods of JEV real-time RT-PCR and RT-ddPCR were established and optimal reaction conditions were confirmed...
June 30, 2017: Journal of Virological Methods
https://www.readbyqxmd.com/read/28670746/hpv-circulating-tumor-dna-to-monitor-the-efficacy-of-anti-pd-1-therapy-in-metastatic-squamous-cell-carcinoma-of-the-anal-canal-a-case-report
#8
Luc Cabel, François-Clément Bidard, Vincent Servois, Wulfran Cacheux, Pascale Mariani, Emanuela Romano, Mathieu Minsat, Ivan Bieche, Fereshteh Farkhondeh, Emmanuelle Jeannot, Bruno Buecher
Squamous cell carcinoma of the anal canal (SCCA) is a rare HPV-associated cancer with limited sensitivity to standard chemotherapy. In a phase 2 study, nivolumab, an anti PD-1 immune checkpoint inhibitor, demonstrated significant efficacy as single-agent therapy in metastatic SCCA patients. Nevertheless, imaging assessment by standard RECIST criteria of the efficacy of immune therapy can be difficult in some patients due to tumor immune cell infiltration, and biomarkers of treatment efficacy are needed. We have previously developed a quantitative droplet digital PCR (ddPCR) technique to detect HPV circulating tumor DNA (HPV ctDNA), with excellent sensitivity and specificity...
July 3, 2017: International Journal of Cancer. Journal International du Cancer
https://www.readbyqxmd.com/read/28669806/the-development-of-a-sensitive-droplet-digital-pcr-for-quantitative-detection-of-porcine-reproductive-and-respiratory-syndrome-virus
#9
Qi Yang, Jun Xi, Xinxin Chen, Sihong Hu, Nan Chen, Songlin Qiao, Shaogui Wan, Dengke Bao
Porcine reproductive and respiratory syndrome (PRRS) is an infectious disease, resulting in important economic losses in pig farming. Prompt detection of PRRS virus (PRRSV) in the field samples is important for effective PRRS control. Droplet digital PCR (ddPCR) is a novel PCR technology, which offers good precision and direct quantification without using calibration curves. In this study, we established a ddPCR system for the sensitive and accurate quantification of PRRSV. Specificity of the assay was determined by the failure of amplification of other relevant viruses...
June 30, 2017: International Journal of Biological Macromolecules
https://www.readbyqxmd.com/read/28656489/detection-of-brca1-gross-rearrangements-by-droplet-digital-pcr
#10
Elena V Preobrazhenskaya, Ilya V Bizin, Ekatherina Sh Kuligina, Alla Yu Shleykina, Evgeny N Suspitsin, Olga A Zaytseva, Elena I Anisimova, Sergey A Laptiev, Tatiana V Gorodnova, Alexey M Belyaev, Evgeny N Imyanitov, Anna P Sokolenko
PURPOSE: Large genomic rearrangements (LGRs) constitute a significant share of pathogenic BRCA1 mutations. Multiplex ligation-dependent probe amplification (MLPA) is a leading method for LGR detection; however, it is entirely based on the use of commercial kits, includes relatively time-consuming hybridization step, and is not convenient for large-scale screening of recurrent LGRs. MATERIALS AND METHODS: We developed and validated the droplet digital PCR (ddPCR) assay, which covers the entire coding region of BRCA1 gene and is capable to precisely quantitate the copy number for each exon...
June 27, 2017: Breast Cancer Research and Treatment
https://www.readbyqxmd.com/read/28654700/both-absolute-and-relative-quantification-of-urinary-mrna-are-useful-for-non-invasive-diagnosis-of-acute-kidney-allograft-rejection
#11
Jung-Woo Seo, Haena Moon, Se-Yun Kim, Ju-Young Moon, Kyung Hwan Jeong, Yu-Ho Lee, Yang-Gyun Kim, Tae-Won Lee, Chun-Gyoo Ihm, Chan-Duck Kim, Byung Ha Chung, Yeong Hoon Kim, Sang Ho Lee
Urinary mRNA analysis with three-gene set (18S rRNA, CD3ε, and IP-10) has been suggested as a non-invasive biomarker of acute rejection (AR) in kidney transplant recipients using quantitative real-time PCR (qPCR). Application of droplet digital PCR (ddPCR), which has been suggested to provide higher sensitivity, accuracy, and absolute quantification without standard curves, could be a useful method for the quantifying low concentration of urinary mRNA. We investigated the urinary expression of these three genes in Korean patients with kidney transplantation and also evaluated the usefulness of ddPCR...
2017: PloS One
https://www.readbyqxmd.com/read/28637527/rodents-goats-and-dogs-their-potential-roles-in-the-transmission-of-schistosomiasis-in-china
#12
Clare F VAN Dorssen, Catherine A Gordon, Yuesheng Li, Gail M Williams, Yuanyuan Wang, Zhenhua Luo, Geoffrey N Gobert, Hong You, Donald P McManus, Darren J Gray
Schistosomiasis in China has been substantially reduced due to an effective control programme employing various measures including bovine and human chemotherapy, and the removal of bovines from endemic areas. To fulfil elimination targets, it will be necessary to identify other possible reservoir hosts for Schistosoma japonicum and include them in future control efforts. This study determined the infection prevalence of S. japonicum in rodents (0-9·21%), dogs (0-18·37%) and goats (6·9-46·4%) from the Dongting Lake area of Hunan province, using a combination of traditional coproparasitological techniques (miracidial hatching technique and Kato-Katz thick smear technique) and molecular methods [quantitative real-time PCR (qPCR) and droplet digital PCR (ddPCR)]...
June 22, 2017: Parasitology
https://www.readbyqxmd.com/read/28629339/droplet-digital-pcr-for-detection-and-quantification-of-circulating-tumor-dna-in-plasma-of-head-and-neck-cancer-patients
#13
Joost H van Ginkel, Manon M H Huibers, Robert J J van Es, Remco de Bree, Stefan M Willems
BACKGROUND: During posttreatment surveillance of head and neck cancer patients, imaging is insufficiently accurate for the early detection of relapsing disease. Free circulating tumor DNA (ctDNA) may serve as a novel biomarker for monitoring tumor burden during posttreatment surveillance of these patients. In this exploratory study, we investigated whether low level ctDNA in plasma of head and neck cancer patients can be detected using Droplet Digital PCR (ddPCR). METHODS: TP53 mutations were determined in surgically resected primary tumor samples from six patients with high stage (II-IV), moderate to poorly differentiated head and neck squamous cell carcinoma (HNSCC)...
June 19, 2017: BMC Cancer
https://www.readbyqxmd.com/read/28626778/single-cell-based-vector-tracing-in-patients-with-ada-scid-treated-with-stem-cell-gene-therapy
#14
Yuka Igarashi, Toru Uchiyama, Tomoko Minegishi, Sirirat Takahashi, Nobuyuki Watanabe, Toshinao Kawai, Masafumi Yamada, Tadashi Ariga, Masafumi Onodera
Clinical improvement in stem cell gene therapy (SCGT) for primary immunodeficiencies depends on the engraftment levels of genetically corrected cells, and tracing the transgene in each hematopoietic lineage is therefore extremely important in evaluating the efficacy of SCGT. We established a single cell-based droplet digital PCR (sc-ddPCR) method consisting of the encapsulation of a single cell into each droplet, followed by emulsion PCR with primers and probes specific for the transgene. A fluorescent signal in a droplet indicates the presence of a single cell carrying the target gene in its genome, and this system can clearly determine the ratio of transgene-positive cells in the entire population at the genomic level...
September 15, 2017: Molecular Therapy. Methods & Clinical Development
https://www.readbyqxmd.com/read/28625519/multiplex-ultrasensitive-genotyping-of-patients-with-non-small-cell-lung-cancer-for-epidermal-growth-factor-receptor-egfr-mutations-by-means-of-picodroplet-digital-pcr
#15
Masaru Watanabe, Tomoya Kawaguchi, Shun-Ichi Isa, Masahiko Ando, Akihiro Tamiya, Akihito Kubo, Hideo Saka, Sadanori Takeo, Hirofumi Adachi, Tsutomu Tagawa, Osamu Kawashima, Motohiro Yamashita, Kazuhiko Kataoka, Yukito Ichinose, Yukiyasu Takeuchi, Katsuya Watanabe, Akihide Matsumura, Yasuhiro Koh
Epidermal growth factor receptor (EGFR) mutations have been used as the strongest predictor of effectiveness of treatment with EGFR tyrosine kinase inhibitors (TKIs). Three most common EGFR mutations (L858R, exon 19 deletion, and T790M) are known to be major selection markers for EGFR-TKIs therapy. Here, we developed a multiplex picodroplet digital PCR (ddPCR) assay to detect 3 common EGFR mutations in 1 reaction. Serial-dilution experiments with genomic DNA harboring EGFR mutations revealed linear performance, with analytical sensitivity ~0...
June 7, 2017: EBioMedicine
https://www.readbyqxmd.com/read/28615231/single-nucleotide-polymorphism-leading-to-false-allelic-fraction-by-droplet-digital-pcr
#16
Eric S Christenson, William B Dalton, David Chu, Ian Waters, Karen Cravero, Daniel J Zabransky, Amy DeZern, Ben Ho Park
BACKGROUND: Molecular-based diagnostics have great utility for cancer detection. We have used droplet digital PCR (ddPCR) as a platform for identifying mutations in circulating plasma tumor DNA (ptDNA). We present the unexpected finding of a spurious mutant allele fraction that was discovered to be artifactual because of the presence of a single-nucleotide polymorphism (SNP) in a patient sample. DESIGN AND METHODS: Probe and primer combinations for the K700 and V701 loci of the SF3B1 spliceosome gene were designed for ddPCR to identify the percentage of mutant and wild-type alleles...
June 14, 2017: Clinical Chemistry
https://www.readbyqxmd.com/read/28602964/droplet-digital-pcr-improved-the-egfr-mutation-diagnosis-with-pleural-fluid-samples-in-non-small-cell-lung-cancer-patients
#17
Xiaoyan Li, Yi Liu, Weiwei Shi, Huayan Xu, Haixu Hu, Zhengwei Dong, Guanshan Zhu, Yun Sun, Bing Liu, Hongjun Gao, Chuanhao Tang, Xiaoqing Liu
BACKGROUND: Droplet digital polymerase chain reaction (ddPCR) is a promising method for analyzing minor amounts of nucleic acid. However, its application has not been reported in pleural fluid, which is an ideal sample source for epidermal growth factor receptor (EGFR) mutation analysis in non-small-cell lung cancer (NSCLC) patients. METHODS: The extracted DNA from supernatants of pleural fluid was selected from our sample bank and re-analyzed by our previously established ddPCR assay...
June 8, 2017: Clinica Chimica Acta; International Journal of Clinical Chemistry
https://www.readbyqxmd.com/read/28590119/parallel-evaluation-of-melting-temperatures-of-dnas-in-the-arrayed-droplets-through-the-fluorescence-from-dna-intercalators
#18
Yuzuru Shimazaki, Junko Tanaka, Yoshinobu Kohara, Masao Kamahori, Takeshi Sakamoto
Parallel evaluation of melting temperatures (Tm's) of DNA molecules in multiple floating droplets (20 μm in diameter) was demonstrated. The Tm values were evaluated from the melting curves which were observed through the fluorescence from the DNA intercalators. The Tm values measured in the droplets corresponded well to those measured in the bulk, indicating the validity of the measurement. The parallel evaluation of Tm's was realized by observing melting curves of DNAs in the different droplets at the same time using the "droplet guide", which guided and fixed the floating droplets to the designated points in the observing plane...
June 7, 2017: Analytical Chemistry
https://www.readbyqxmd.com/read/28577941/quantitative-cell-free-circulating-egfr-mutation-concentration-is-correlated-with-tumor-burden-in-advanced-nsclc-patients
#19
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
OBJECTIVES: Droplet digital polymerase chain reaction (ddPCR) has shown sufficient concordance in detecting plasma epidermal growth factor receptor (EGFR) status in non-small cell lung cancer (NSCLC), compared to tumor tissues. However, the clinical significance of the quantitative plasma mutated EGFR concentration remains unknown. The purpose of this study was to explore the relationship of plasma mutated EGFR concentration with tumor burden in advanced NSCLC patients. MATERIALS AND METHODS: Using ddPCR, plasma DNA samples prior to administration of therapies from 113 consecutive NSCLC patients were analyzed for EGFR L858R substitution and deletion of exon19 (ex19del)...
July 2017: Lung Cancer: Journal of the International Association for the Study of Lung Cancer
https://www.readbyqxmd.com/read/28562660/a-droplet-digital-pcr-ddpcr-assay-to-detect-helicoverpa-armigera-lepidoptera-noctuidae-in-bulk-trap-samples
#20
Frida A Zink, Luke R Tembrock, Alicia E Timm, Roxanne E Farris, Omaththage P Perera, Todd M Gilligan
Moths in the genus Helicoverpa are some of the most important agricultural pests in the world. Two species, H. armigera (Hübner) and H. zea (Boddie), cause the majority of damage to crops and millions of dollars are spent annually on control of these pests. The recent introduction of H. armigera into the New World has prompted extensive survey efforts for this species in the United States. Surveys are conducted using bucket traps baited with H. armigera pheromone, and, because the same pheromone compounds attract both species, these traps often capture large numbers of the native H...
2017: PloS One
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