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Søren Feddersen, Lars Bastholt, Susanne Møller Pedersen
BACKGROUND: The clinical utility of serum thyroglobulin (TG) in the follow-up of patients with differentiated thyroid carcinoma (DTC) may be compromised by the presence of endogenous anti-thyroglobulin antibodies (TGAb). To prevent interference by TGAb several groups have developed real-time PCR (RT-PCR) based assays for quantification of blood TG mRNA levels. For accurate quantification of TG mRNA in blood preanalytical factors must be recognized and controlled. In this study we evaluate the effect of different blood RNA stabilizing systems - the Tempus(T) (M) Blood RNA system and the PAXgene(T) (M) Blood RNA system - and storage time on RNA yield and quality, and TG mRNA stability...
September 13, 2016: Annals of Clinical Biochemistry
Sarah D Linnstaedt, JunMei Hu, Andrea Y Liu, April C Soward, Kenneth A Bollen, Henry E Wang, Phyllis L Hendry, Erin Zimny, Christopher Lewandowski, Marc-Anthony Velilla, Kathia Damiron, Claire Pearson, Robert Domeier, Sangeeta Kaushik, James Feldman, Mark Rosenberg, Jeffrey Jones, Robert Swor, Niels Rathlev, Samuel A McLean
INTRODUCTION: A motor vehicle collision (MVC) is one of the most common life-threatening events experienced by individuals living in the USA. While most individuals recover following MVC, a significant proportion of individuals develop adverse post-traumatic sequelae such as post-traumatic stress disorder or persistent musculoskeletal pain. Adverse post-traumatic sequelae are common, morbid and costly public health problems in the USA and other industrialised countries. The pathogenesis of these disorders following MVC remains poorly understood...
2016: BMJ Open
Iacopo Fabiani, Cristian Scatena, Chiara Maria Mazzanti, Lorenzo Conte, Nicola Riccardo Pugliese, Sara Franceschi, Francesca Lessi, Michele Menicagli, Andrea De Martino, Stefano Pratali, Uberto Bortolotti, Antonio Giuseppe Naccarato, Salvatore La Carrubba, Vitantonio Di Bello
AIMS: Myocardial fibrosis (MF) is a deleterious consequence of aortic valve stenosis (AVS). Global longitudinal strain (GLS) is a novel left ventricular (LV) functional parameter potentially useful to non-invasively estimate MF. MicroRNAs (miRNAs) are non-coding small ribonucleic acids (RNA) modulating genes function, mainly through RNA degradation. miRNA-21 is a biomarker associated with MF in pressure overload. The aim of the present study was to find an integrated algorithm for detection of MF using a combined approach with both bio- and functional markers...
2016: Journal of Translational Medicine
Conny Mathay, Gaël Hamot, Estelle Henry, Kathleen Mommaerts, Audur Thorlaksdottir, Johanna Trouet, Fay Betsou
BACKGROUND: This article is the fifth in a series of publications providing formal method validation for biospecimen processing. We report the optimization and validation of methodology to obtain nucleic acids of sufficient quantity and quality from blood. METHODS: DNA was extracted using the Chemagic DNA Blood Kit on an MSM I. Extraction was optimized in terms of blood volume, elution buffer volume, and lysis conditions. The optimal protocol was validated for reproducibility, robustness (delay to buffy coat extraction, blood vs...
August 22, 2016: Biopreservation and Biobanking
Xichun Liu, Elisa Ledet, Dongying Li, Ary Dotiwala, Allie Steinberger, Allison Feibus, Jianzhuo Li, Yanfeng Qi, Jonathan Silberstein, Benjamin Lee, Yan Dong, Oliver Sartor, Haitao Zhang
PURPOSE: The majority of prostate cancer mortality can be attributed to metastatic castration-resistant prostate cancer, an advanced stage which remains incurable despite recent advances. The androgen receptor (AR) signaling axis remains active in CRPC. Recent studies suggest that the expression of an AR splice variant, AR-V7, may underlie resistance to abiraterone and enzalutamide. However, controversy exists over the optimal assay. The objective of this study is to develop a fast and sensitive assay for AR splice variants (AR-Vs) in patients...
July 20, 2016: Journal of Urology
Ying Liu, Deepak P Edward
AIM: To compare the effect of the two fixatives on tissue morphology and utility to obtain good quality nucleic acids for molecular analysis from micro-dissected retinal samples. METHODS: Enucleated specimens from New Zealand white rabbits were fixed in formalin or PAXgene fixative according to standard protocols, and then processed and embedded in paraffin for sectioning. Sections were stained with hematoxylin and eosin to assess the structural integrity of retina...
July 13, 2016: Current Eye Research
William Mathieson, Nathalie Marcon, Laurent Antunes, David A Ashford, Fay Betsou, Sonia G Frasquilho, Olga A Kofanova, Siobhan C McKay, Stephan Pericleous, Colleen Smith, Kristian M Unger, Constanze Zeller, Geraldine A Thomas
OBJECTIVES: To evaluate the PAXgene tissue fixation system. METHODS: Clinical biospecimens (n = 46) were divided into PAXgene-fixed paraffin-embedded (PFPE), formalin-fixed paraffin-embedded (FFPE), and fresh-frozen (FF) blocks. PFPE and FFPE sections were compared for histology (H&E staining) and immunohistochemistry (14 antibodies) using tissue microarrays. PFPE, FFPE, and FF samples were compared in terms of RNA quality (RNA integrity number, polymerase chain reaction [PCR] amplicon length, and quantitative reverse transcription PCR), DNA quality (gel electrophoresis and methylation profiling) and protein quality (liquid chromatography-mass spectrometry [LC-MS/MS])...
July 2016: American Journal of Clinical Pathology
Satu Luhtala, Synnöve Staff, Mark Barok, Minna Tanner, Jorma Isola
BACKGROUND: Growth factor receptor HER3 (ErbB3) lacks standardized immunohistochemistry (IHC)-based methods for formalin-fixed paraffin-embedded (FFPE) tissue samples. We compared 4 different anti-HER3 antibodies to explain the differences found in the staining results reported in the literature. MATERIALS AND METHODS: Four commercial HER3 antibodies were tested on FFPE samples including mouse monoclonal antibody clones, DAK-H3-IC and RTJ1, rabbit monoclonal antibody clone SP71, and rabbit polyclonal antibody (SAB4500793)...
July 6, 2016: Applied Immunohistochemistry & Molecular Morphology: AIMM
Vlasta Korenkova, Jana Slyskova, Vendula Novosadova, Sara Pizzamiglio, Lucie Langerova, Jens Bjorkman, Ondrej Vycital, Vaclav Liska, Miroslav Levy, Karel Veskrna, Pavel Vodicka, Ludmila Vodickova, Mikael Kubista, Paolo Verderio
Successful molecular analyses of human solid tissues require intact biological material with well-preserved nucleic acids, proteins, and other cell structures. Pre-analytical handling, comprising of the collection of material at the operating theatre, is among the first critical steps that influence sample quality. The aim of this study was to compare the experimental outcomes obtained from samples collected and stored by the conventional means of snap freezing and by PAXgene Tissue System (Qiagen). These approaches were evaluated by measuring rRNA and mRNA integrity of the samples (RNA Quality Indicator and Differential Amplification Method) and by gene expression profiling...
2016: Scientific Reports
Thomas W McDade, Kharah M Ross, Ruby L Fried, Jesusa M G Arevalo, Jeffrey Ma, Gregory E Miller, Steve W Cole
Genome-wide transcriptional profiling has emerged as a powerful tool for analyzing biological mechanisms underlying social gradients in health, but utilization in population-based studies has been hampered by logistical constraints and costs associated with venipuncture blood sampling. Dried blood spots (DBS) provide a minimally invasive, low-cost alternative to venipuncture, and in this article we evaluate how closely the substantive results from DBS transcriptional profiling correspond to those derived from parallel analyses of gold-standard venous blood samples (PAXgene whole blood and peripheral blood mononuclear cells [PBMC])...
2016: Biodemography and Social Biology
Lisa Oberauner-Wappis, Martina Loibner, Christian Viertler, Daniel Groelz, Ralf Wyrich, Kurt Zatloukal
Molecular diagnostics in personalized medicine increasingly relies on the combination of a variety of analytical technologies to characterize individual diseases and to select patients for targeted therapies. The gold standard for tissue-based diagnostics is fixation in formalin and embedding in paraffin, which results in excellent preservation of morphology but negatively impacts on a variety of molecular assays. The formalin-free, non-cross-linking PAXgene tissue system preserves morphology in a similar way to formalin, but also preserves biomolecules essentially in a similar way to cryopreservation, which markedly widens the spectrum, sensitivity and accuracy of molecular analytics...
April 2016: International Journal of Experimental Pathology
V Mauffré, B Grimard, C Eozenou, S Inghels, L Silva, C Giraud-Delville, D Capo, O Sandra, F Constant
We investigated the diagnostic reliability of pregnancy detection using changes in interferon stimulated gene (ISG) messenger RNA (mRNA) levels in circulating immune cells in ewes. Two different groups of ewes (an experimental group, experiment 1 and a farm group, experiment 2) were oestrus-synchronized and blood sampled on day 14 (D0=day of insemination in control animals, experiment 1) and day 15 (experiment 2). Real-time PCR were performed to evaluate the abundance of different ISG mRNAs. In the experimental group, peripheral blood mononuclear cells of 29 ewes born and bred in experimental facilities were isolated using a Percoll gradient method...
May 6, 2016: Animal: An International Journal of Animal Bioscience
Nathanael Raschzok, Anja Reutzel-Selke, Rosa B Schmuck, Laura M Tannus, Mehmet H Morgul, Corinna Dietel, Annekatrin Leder, Benjamin Struecker, Steffen Lippert, Hannes Sallmon, Moritz Schmelzle, Michael Bartels, Sven Jonas, Johann Pratschke, Igor M Sauer
OBJECTIVE: We investigated whether microRNA signatures in whole blood samples are associated with acute cellular rejection (ACR) after liver transplantation. MATERIALS AND METHODS: Blood samples were collected using Paxgene technology and analyzed by microarrays and quantitative real-time polymerase chain reaction (qRT-PCR). RESULTS: microRNA signatures failed to distinguish between 19 patients with ACR and 16 controls. Let-7b-5p and let-7c were upregulated in a subgroup of patients with ACR during the 6th and 7th postoperative days but failed in an independent validation of 20 patients...
April 28, 2016: Biomarkers: Biochemical Indicators of Exposure, Response, and Susceptibility to Chemicals
Sayaka Yuzawa, Hiroshi Nishihara, Shigeru Yamaguchi, Hiromi Mohri, Lei Wang, Taichi Kimura, Masumi Tsuda, Mishie Tanino, Hiroyuki Kobayashi, Shunsuke Terasaka, Kiyohiro Houkin, Norihiro Sato, Shinya Tanaka
Recent genetic analyses using next-generation sequencers have revealed numerous genetic alterations in various tumors including meningioma, which is the most common primary brain tumor. However, their use as routine laboratory examinations in clinical applications for tumor genotyping is not cost effective. To establish a clinical sequencing system for meningioma and investigate the clinical significance of genotype, we retrospectively performed targeted amplicon sequencing on 103 meningiomas and evaluated the association with clinicopathological features...
July 2016: Modern Pathology: An Official Journal of the United States and Canadian Academy of Pathology, Inc
Martina Loibner, Walter Buzina, Christian Viertler, Daniel Groelz, Anja Hausleitner, Gintare Siaulyte, Iris Kufferath, Bettina Kölli, Kurt Zatloukal
BACKGROUND: Requirements on tissue fixatives are getting more demanding as molecular analysis becomes increasingly relevant for routine diagnostics. Buffered formaldehyde in pathology laboratories for tissue fixation is known to cause chemical modifications of biomolecules which affect molecular testing. A novel non-crosslinking tissue preservation technology, PAXgene Tissue (PAXgene), was developed to preserve the integrity of nucleic acids in a comparable way to cryopreservation and also to preserve morphological features comparable to those of formalin fixed samples...
2016: PloS One
Pekka Ruusuvuori, Mira Valkonen, Matti Nykter, Tapio Visakorpi, Leena Latonen
This paper describes work presented at the Nordic Symposium on Digital Pathology 2015, in Linköping, Sweden. Prostatic intraepithelial neoplasia (PIN) represents premalignant tissue involving epithelial growth confined in the lumen of prostatic acini. In the attempts to understand oncogenesis in the human prostate, early neoplastic changes can be modeled in the mouse with genetic manipulation of certain tumor suppressor genes or oncogenes. As with many early pathological changes, the PIN lesions in the mouse prostate are macroscopically small, but microscopically spanning areas often larger than single high magnification focus fields in microscopy...
2016: Journal of Pathology Informatics
Francesca Malentacchi, Sara Pizzamiglio, Ralf Wyrich, Paolo Verderio, Chiara Ciniselli, Mario Pazzagli, Stefania Gelmini
BACKGROUND: Inappropriate handling of blood samples might induce or repress gene expression and/or lead to RNA degradation affecting downstream analysis. In particular, sample transport is a critical step for biobanking or multicenter studies because of uncontrolled variables (i.e., unstable temperature). We report the results of a pilot study implemented within the EC funded SPIDIA project, aimed to investigate the role of transport and storage of blood samples containing and not containing an RNA stabilizer...
April 2016: Biopreservation and Biobanking
M Abend, C Badie, R Quintens, R Kriehuber, G Manning, E Macaeva, M Njima, D Oskamp, S Strunz, S Moertl, S Doucha-Senf, S Dahlke, J Menzel, M Port
The risk of a large-scale event leading to acute radiation exposure necessitates the development of high-throughput methods for providing rapid individual dose estimates. Our work addresses three goals, which align with the directive of the European Union's Realizing the European Network of Biodosimetry project (EU-RENB): 1. To examine the suitability of different gene expression platforms for biodosimetry purposes; 2. To perform this examination using blood samples collected from prostate cancer patients (in vivo) and from healthy donors (in vitro); and 3...
February 2016: Radiation Research
B Børresen, L N Nielsen, L R Jessen, A T Kristensen, M Fredholm, S Cirera
Cancer is a prevalent cause of mortality in Bernese mountain dogs (BMDs). Circulating microRNAs (miRNAs) are found in blood and have been identified as promising biomarkers in various neoplastic diseases in humans. In the current study, the expression profile of different types of miRNAs was investigated in healthy BMDs and BMDs with cancer. Seven healthy and six non-treated BMDs with cancer [four with disseminated histiocytic sarcomas (DHS)] were enrolled in this study. Clinical evaluations including physical examination, blood analysis, urinalysis and diagnostic imaging were performed on all dogs...
January 21, 2016: Veterinary and Comparative Oncology
Perenlei Enkhbaatar, Christina Nelson, John R Salsbury, Joseph R Carmical, Karen E O Torres, David Herndon, Donald S Prough, Liming Luan, Edward R Sherwood
BACKGROUND: Animal models that mimic human biology are important for successful translation of basic science discoveries into the clinical practice. Recent studies in rodents have demonstrated the efficacy of TLR4 agonists as immunomodulators in models of infection. However, rodent models have been criticized for not mimicking important characteristics of the human immune response to microbial products. The goal of this study was to compare genomic responses of human and sheep blood to the TLR4 agonists lipopolysaccharide (LPS) and monophosphoryl lipid A (MPLA)...
2015: PloS One
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