Molecular Analysis of Gene Fusions in Bone and Soft Tissue Tumors by Anchored Multiplex PCR–Based Targeted Next-Generation Sequencing
In this study, the applicability of a novel technique termed anchored multiplex PCR (AMP) for next-generation sequencing (NGS), using the Archer FusionPlex Sarcoma kit, aimed at 26 genes, was evaluated and compared with FISH and reverse transcriptase-PCR. In case of discrepant results, further analysis occurred with a third independent technique. Eighty-one samples were subjected to AMP-based targeted NGS, and 86% (n = 70) were successfully conducted and were either fusion positive (n = 48) or fusion negative, but met all criteria for good quality (n = 22). A concordance of 90% was found ...
Source: The Journal of Molecular Diagnostics - August 20, 2018 Category: Pathology Source Type: research

High-Throughput Copy Number Profiling by Digital Multiplex Ligation-Dependent Probe Amplification in Multiple Myeloma
Publication date: Available online 8 August 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Szabolcs Kosztolanyi, Richard Kiss, Lilit Atanesyan, Ambrus Gango, Karel de Groot, Maryvonne Steenkamer, Pal Jakso, Andras Matolcsy, Bela Kajtar, Laszlo Pajor, Karoly Szuhai, Suvi Savola, Csaba Bodor, Donat AlparAbstractMultiple myeloma (MM) is a genetically heterogeneous disease with diverse clinical outcome. Copy number alterations (CNAs) including whole chromosome and subchromosomal gains and losses are common contributors of the pathogenesis and have demonstrated prognostic impact in MM. We tested the performance of d...
Source: The Journal of Molecular Diagnostics - August 8, 2018 Category: Pathology Source Type: research

Curating clinically relevant transcripts for the interpretation of sequence variants
Publication date: Available online 8 August 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Marina T. DiStefano, Sarah E. Hemphill, Brandon J. Cushman, Mark J. Bowser, Elizabeth Hynes, Andrew R. Grant, Rebecca K. Siegert, Andrea M. Oza, Michael A. Gonzalez, Sami S. Amr, Heidi L. Rehm, Ahmad N. Abou TayounAbstractVariant interpretation depends on accurate annotations using biologically relevant transcripts. We have developed a systematic strategy for designating primary transcripts, and applied it to 109 hearing loss–associated genes that were divided into three categories. Category 1 genes (n=38) had a sin...
Source: The Journal of Molecular Diagnostics - August 8, 2018 Category: Pathology Source Type: research

Describing the Reportable Range Is Important for Reliable Treatment Decisions: A Multi-Laboratory Study for Molecular Tumor Profiling Using Next-Generation Sequencing
Publication date: Available online 26 July 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Véronique Tack, Lien Spans, Ed Schuuring, Cleo Keppens, Karen Zwaenepoel, Patrick Pauwels, Jeroen Van Houdt, Elisabeth M.C. DequekerAs interpretation of next-generation sequencing (NGS) data remains challenging, optimization of the NGS process is needed to obtain correct sequencing results. Therefore, extensive validation and continuous monitoring of the quality is essential. NGS performance was compared to traditional detection methods and technical quality of nine NGS technologies was assessed. First, nine formali...
Source: The Journal of Molecular Diagnostics - July 26, 2018 Category: Pathology Source Type: research

Benchmarking of Amplicon-Based Next-Generation Sequencing Panels Combined with Bioinformatics Solutions for Germline BRCA1 and BRCA2 Alteration Detection
In this study, we evaluated the combinations of several BRCA testing assays and bioinformatics solutions for the identification of single nucleotide variants, insertion/deletion variants (indels), and copy number variations (CNVs). Four assays (BRCA Tumor, BRCA HC, Ion AmpliSeq BRCA, and Access Array BRCA) and two commercial bioinformatics solutions (the SeqNext software and the Sophia DDM platform) were tested on a set of 28 previously genotyped samples. All solutions exhibited accurate detection of single nucleotide variants /indels, except for Ion AmpliSeq BRCA, which exhibited drop in coverage. Interestingly, for CNV a...
Source: The Journal of Molecular Diagnostics - July 26, 2018 Category: Pathology Source Type: research

Next-Generation Sequencing Using S1 Nuclease for Poor-Quality Formalin-Fixed, Paraffin-Embedded Tumor Specimens
In conclusion, the S1 method is optimal for NGS testing of poor-quality FFPE specimens. (Source: The Journal of Molecular Diagnostics)
Source: The Journal of Molecular Diagnostics - July 26, 2018 Category: Pathology Source Type: research

Multicenter Evaluation of the Idylla NRAS-BRAF Mutation Test in Metastatic Colorectal Cancer
Publication date: Available online 26 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Iván Prieto-Potin, Clara Montagut, Beatriz Bellosillo, Matthew Evans, Matthew Smith, Linea Melchior, Werner Reiltin, Michael Bennett, Veronica Pennati, Francesca Castiglione, Karl-Friedrich Bürrig, Ulrike Cooper, Barbara Dockhorn-Dworniczak, Christiana Rossenbach, Claudia M. Luna-Aguirre, Hugo A. Barrera-Saldaña, José C. Machado, José L. Costa, Rinat Yacobi, Hilla Tabibian-KeissarTreatment of colorectal cancer (CRC) with monoclonal antibodies against epidermal growth factor receptor require...
Source: The Journal of Molecular Diagnostics - July 24, 2018 Category: Pathology Source Type: research

Characterization of 108 Genomic DNA Reference Materials for 11 Human Leukocyte Antigen Loci: A GeT-RM Collaborative Project
Publication date: Available online 26 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Maria P. Bettinotti, Deborah Ferriola, Jamie L. Duke, Timothy L. Mosbruger, Nikolaos Tairis, Lawrence Jennings, Lisa V. Kalman, Dimitri MonosThe highly polymorphic human leukocyte antigen (HLA) genes, located in the human major histocompatibility complex, encode the class I and II antigen-presenting molecules. which are centrally involved in the immune response. HLA typing is used for several clinical applications, such as transplantation, pharmacogenetics, and diagnosis of autoimmune disease. HLA typing is highly complex ...
Source: The Journal of Molecular Diagnostics - July 21, 2018 Category: Pathology Source Type: research

Laboratory Information Systems and Instrument Software Lack Basic Functionality for Molecular Laboratories
This study determined functionality gaps of LISs in molecular laboratories and the associated impact to workflow, efficiency, and security by collecting anonymous survey data from clinical laboratory professionals. A 34-question survey (30 required + 4 optional) was compiled using an online survey tool. Participants were recruited through several professional molecular society listservs and given 4 weeks to complete the survey. Data collected included participant demographics, scope of testing, software capabilities for the LIS and molecular instruments, and comments. Eighty respondents completed the entire surve...
Source: The Journal of Molecular Diagnostics - July 21, 2018 Category: Pathology Source Type: research

Multiple Ways to Detect IDH2 Mutations in Angioimmunoblastic T-Cell Lymphoma from Immunohistochemistry to Next-Generation Sequencing
Publication date: Available online 5 July 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Aurélie Dupuy, François Lemonnier, Virginie Fataccioli, Nadine Martin-Garcia, Cyrielle Robe, Romain Pelletier, Elsa Poullot, Anissa Moktefi, Karima Mokhtari, Marie C. Rousselet, Alexandra Traverse-Glehen, Richard Delarue, Olivier Tournilhac, Marie H. Delfau-Larue, Corinne Haioun, Nicolas Ortonne, Christiane Copie-Bergman, Laurence de Leval, Anaïs Pujals, Philippe GaulardAngioimmunoblastic T-cell lymphoma (AITL) is a peripheral T-cell lymphoma associated with chemoresistance and a poor prognosis. Various n...
Source: The Journal of Molecular Diagnostics - July 20, 2018 Category: Pathology Source Type: research

Evaluation of a Hepatitis C Virus Core Antigen Assay in Plasma and Dried Blood Spot Samples
This study evaluated the analytical performance of HCV core antigen (HCVcAg) detection in samples of plasma and dried venous blood spots (DBSs). Paired plasma and DBS samples were prepared from remnant diagnostic samples. Plasma HCV RNA was quantified and measured. Sensitivity and specificity for HCVcAg (>3 fmol/L) at two HCV RNA thresholds (≥15 and ≥3000 IU/mL) were calculated. Of 120 paired samples tested, 25 had nonquantifiable HCV RNA and 95 had quantifiable HCV RNA. The median HCV RNA level in plasma was 5.6 log10 IU/mL (interquartile range: 5.2 to 6.2). The median HCVcAg levels in plasma and DBS samples were...
Source: The Journal of Molecular Diagnostics - July 19, 2018 Category: Pathology Source Type: research

Pre-Analytical Handling Conditions and Small RNA Recovery from Urine for miRNA Profiling
We present a comparison of small RNA recovery and stability in urine under alternate pre-analytical handling conditions and extend recommendations on what conditions optimize yield of miRNA from cell-free urine and urine extracellular vesicles (EVs). Using an affinity slurry for isolation of small RNA from urine, we found that urine samples held at room temperature (20°C) for up to 8 hours before processing yield the highest amounts of intact small RNAs from EVs. Some miRNA is lost from urine samples when held 2°C to 4°C and/or frozen before EV isolation, likely because of EV entrapment in uromodulin precipitat...
Source: The Journal of Molecular Diagnostics - July 18, 2018 Category: Pathology Source Type: research

An In-Depth Evaluation of the Validity and Logistics Surrounding the Testing of AR-V7 mRNA Expression in Circulating Tumor Cells
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): Anieta M. Sieuwerts, Bianca Mostert, Michelle van der Vlugt-Daane, Jaco Kraan, Corine M. Beaufort, Mai Van, Wendy J.C. Prager, Bram De Laere, Nick Beije, Paul Hamberg, Hans M. Westgeest, Metin Tascilar, Luc Y. Dirix, Wendy Onstenk, Ronald de Wit, Martijn P. Lolkema, Ron H.J. Mathijssen, John W.M. Martens, Stefan SleijferRecent reports have emphasized the clinical relevance of detecting AR-V7 in circulating tumor cells (CTCs). Our aim was to set up a validated multicenter pipeline to measure AR-V7 by quantitative RT-PCR (RT-...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Chimerism Analysis in the Pediatric Setting: Direct PCR from Bone Marrow, Whole Blood, and Cell Fractions
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): Susanne Kricke, Lana Mhaldien, Rozendo Fernandes, Charizel Villanueva, Alistair Shaw, Paul Veys, Stuart AdamsCertain blood components and anticoagulants interfere with the PCR process and subsequent analysis. Here we demonstrate that reliable test results can be obtained for chimerism analysis despite omitting a DNA-extraction step and performing PCR and fragment analysis directly on bone marrow, whole blood, and individual cell fractions. For chimerism analysis, direct-tissue PCR is possible with the use of a robust, comme...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)/CRISPR-Associated Endonuclease Cas9–Mediated Homology-Independent Integration for Generating Quality Control Materials for Clinical Molecular Genetic Testing
This study demonstrates that CRISPR/Cas9-induced nonhomologous end joining is a valuable and novel method for generating artificial mutants for use in quality control applications in clinical molecular genetics. (Source: The Journal of Molecular Diagnostics)
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Deciphering Elevated Microsatellite Alterations at Selected Tetra/Pentanucleotide Repeats, Microsatellite Instability, and Loss of Heterozygosity in Colorectal Cancers
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): Yang Wang, Cindy L. Vnencak-Jones, Justin M. Cates, Chanjuan ShiElevated microsatellite alterations at selected tetranucleotide repeats (EMAST) are common in colorectal cancers (CRCs). The association between EMAST and classic mono/dinucleotide microsatellite instability (MSI) is unknown. We assessed the stability of 13 tetranucleotide and three pentanucleotide repeat markers in tumor and normal tissue from 22 MSI-high and 107 microsatellite-stable CRC samples. When present, instability was observed at tetra/pentanucleotide...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Validation of a Customized Bioinformatics Pipeline for a Clinical Next-Generation Sequencing Test Targeting Solid Tumor–Associated Variants
We present the validation of an open source tumor amplicon pipeline (OTA-pipeline) for clinical next-generation sequencing targeting solid tumor–associated variants. Raw data generated from 557 TruSight Tumor 26 samples and in silico data were analyzed by the OTA-pipeline and legacy pipeline and compared. Discrepant results were confirmed by orthogonal methods. The OTA-pipeline reported 22 variants that were not detected by the previously validated pipeline, including seven synonymous or intronic single-nucleotide variants, five single-nucleotide variants at frequency
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Targeted Next-Generation Sequencing Is a Sensitive Tool for Differential Diagnosis of Myelodysplastic Syndromes in Bone Marrow Trephines
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): Andreas Bräuninger, Wolfgang Blau, Kristin Kunze, Ann-Kathrin Desch, Alexander Brobeil, Mehmet K. Tur, Benjamin Etschmann, Ulrich Günther, Dieter Körholz, Georg Schliesser, Andreas Käbisch, Michael Kiehl, Mathias Rummel, Stefan GattenlöhnerMyelodysplastic syndromes are hematological neoplasias in which immunohistologic examination of bone marrow trephines is important for a definite diagnosis. Unequivocal distinction from reactive bone marrow changes is, however, sometimes difficult. Because neoplas...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

A Highly Sensitive and Robust Method for Hepatitis B Virus Covalently Closed Circular DNA Detection in Single Cells and Serum
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): Jing-Tao Huang, Ying Yang, Yi-Min Hu, Xing-Hui Liu, Mei-Yan Liao, Roy Morgan, Er-Feng Yuan, Xia Li, Song-Mei LiuDespite implications of persistence of hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) in the development of hepatocellular carcinoma (HCC), little is known about serum cccDNA in HBV-infected diseases. We developed a cccDNA-selective droplet digital PCR (ddPCR) to assess cccDNA content and dynamics across different stages of HCC development. One hundred forty-seven serum samples and 35 formalin-fix...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Table of Contents
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): (Source: The Journal of Molecular Diagnostics)
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Rapid, Loop-Mediated Isothermal Amplification Detection of Celiac Disease Risk Alleles
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): Michael Erlichster, Jason A. Tye-Din, Michael D. Varney, Efstratios Skafidas, Patrick KwanHuman leukocyte antigen (HLA) genotyping has become a useful investigation in the diagnostic work-up of celiac disease (CD), with utility in risk stratification and screening. However, broad application of this technology has been hindered by the cost and time burden of conventional laboratory-based assays. We have developed and validated CD–loop-mediated isothermal amplification (CD-LAMP), a LAMP assay, which enables rapid ident...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Evaluation of Two Commercial Real-Time PCR Kits for Aspergillus DNA Detection in Bronchoalveolar Lavage Fluid in Patients with Invasive Pulmonary Aspergillosis
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): Julie Denis, Faezeh Forouzanfar, Raoul Herbrecht, Elise Toussaint, Romain Kessler, Marcela Sabou, Ermanno Candolfi, Valérie Letsher-BruInvasive pulmonary aspergillosis (IPA) is a common complication of immunosuppression. Rapid diagnosis using molecular techniques is essential to improve patient survival. PCR techniques are promising in enhancing Aspergillus detection in blood and respiratory samples. We evaluate for the first time the performances of two commercial real-time PCR kits, the A. fumigatus Bio-Evolution a...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Spinocerebellar Ataxia Tethering PCR: A Rapid Genetic Test for the Diagnosis of Spinocerebellar Ataxia Types 1, 2, 3, 6, and 7 by PCR and Capillary Electrophoresis
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): Claudia Cagnoli, Alessandro Brussino, Cecilia Mancini, Marina Ferrone, Laura Orsi, Paola Salmin, Patrizia Pappi, Elisa Giorgio, Elisa Pozzi, Simona Cavalieri, Eleonora Di Gregorio, Marta Ferrero, Alessandro Filla, Giuseppe De Michele, Cinzia Gellera, Caterina Mariotti, Suran Nethisinghe, Paola Giunti, Giovanni Stevanin, Alfredo BruscoSpinocerebellar ataxia (SCA) types 1, 2, 3, 6, and 7, associated with a (CAG)n repeat expansion in coding sequences, are the most prevalent autosomal dominant ataxias worldwide (approximately 6...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Reference Size Matching, Whole-Genome Amplification, and Fluorescent Labeling as a Method for Chromosomal Microarray Analysis of Clinically Actionable Copy Number Alterations in Formalin-Fixed, Paraffin-Embedded Tumor Tissue
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): Shelly R. Gunn, Shailin Govender, Cynthe L. Sims, Aditi Khurana, Samuel Koo, Jayne Scoggin, Mathew W. Moore, Philip D. CotterCancer genome copy number alterations (CNAs) assist clinicians in selecting targeted therapeutics. Solid tumor CNAs are most commonly evaluated in formalin-fixed, paraffin-embedded (FFPE) tissue by fluorescence in situ hybridization. Although fluorescence in situ hybridization is a sensitive and specific assay for interrogating preselected genomic regions, it provides no information about coexisting c...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Hepatitis B Virus Covalently Closed Circular DNA–Selective Droplet Digital PCR: A Sensitive and Noninvasive Method for Hepatocellular Carcinoma Diagnosis?
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): Fan Shen, Consolato Sergi, Hui-lung Sun (Source: The Journal of Molecular Diagnostics)
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Recommendations for Clinical CYP2C19 Genotyping Allele Selection: A Report of the Association for Molecular Pathology
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): Victoria M. Pratt, Andria L. Del Tredici, Houda Hachad, Yuan Ji, Lisa V. Kalman, Stuart A. Scott, Karen E. WeckThis document was developed by the Pharmacogenomics (PGx) Working Group of the Association for Molecular Pathology Clinical Practice Committee, whose aim is to recommend variants for inclusion in clinical pharmacogenomic testing panels. The goals of the Association for Molecular Pathology PGx Working Group are to define the key attributes of PGx alleles recommended for clinical testing and to define a minimum set o...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Editorial Board
Publication date: May 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 3Author(s): (Source: The Journal of Molecular Diagnostics)
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Optimal Reference Gene Selection for Expression Studies in Human Reticulocytes
This study aimed at selecting optimal reference genes for gene-expression analysis of reticulocytes and at validating them in hereditary spherocytosis (HS) and β-thalassemia intermedia (βTI) patients. Seven reference genes (PGK1, MPP1, HPRT1, ACTB, GAPDH, RN18S1, and SDHA) were selected because of published reports. Real-time quantitative PCR was performed on reticulocytes in 20 healthy volunteers, 15 HS patients, and 10 βTI patients. Threshold cycle values were compared with fold-change method and RefFinder software. The stable reference genes recommended by RefFinder were validated with SLC4A1 and flow cyt...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Clinical Implementation and Validation of Automated Human Genome Variation Society (HGVS) Nomenclature System for Next-Generation Sequencing–Based Assays for Cancer
Publication date: Available online 21 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Keith M. Callenberg, Lucas Santana-Santos, Liang Chen, Wayne L. Ernst, Michelle Barbi De Moura, Yuri E. Nikiforov, Marina N. Nikiforova, Somak RoyAbstractHuman Genome Variation Society (HGVS) nomenclature is a de facto clinical standard for reporting DNA sequence variants. With increasing use of high-throughput sequencing, manual generation of HGVS nomenclatures for all variants is impractical and error-prone. It is therefore beneficial to include one or more HGVS generator tools in next-generation sequencing (NGS) bioinfo...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Validation and Implementation of BRCA1/2 Variant Screening in Ovarian Tumor Tissue
Publication date: Available online 21 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Marthe M. de Jonge, Dina Ruano, Ronald van Eijk, Nienke van der Stoep, Maartje Nielsen, Juul T. Wijnen, Natalja T. ter Haar, Astrid Baalbergen, Monique E.M.M. Bos, Marjolein J. Kagie, Maaike P.G. Vreeswijk, Katja N. Gaarenstroom, Judith R. Kroep, Vincent T.H.B.M. Smit, Tjalling Bosse, Tom van Wezel, Christi J. van AsperenAbstractBRCA1/2 variant analysis in tumor tissue could streamline the referral of patients with epithelial ovarian, fallopian tube, or primary peritoneal cancer to genetic counselors and select patients wh...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

The development and validation of clinical exome-based panels using ExomeSlicer: Considerations and proof of concept using an Epilepsy panel
Publication date: Available online 22 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Rojeen Niazi, Michael A. Gonzalez, Jorune Balciuniene, Perry Evans, Mahdi Sarmady, Ahmad N. Abou TayounAbstractExome-based panels are becoming the preferred diagnostic strategy in clinical laboratories. This approach enables dynamic gene content update and, if needed, cost-effective reflex to whole exome sequencing. There are currently no guidelines or appropriate resources to support the clinical implementation of exome-based panels. Here, we highlight principles and important considerations for the clinical development a...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Analytical validation of a hybrid capture–based next-generation sequencing clinical assay for genomic profiling of cell-free circulating tumor DNA
Publication date: Available online 22 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Travis A. Clark, Jon H. Chung, Mark Kennedy, Jason D. Hughes, Niru Chennagiri, Daniel S. Lieber, Bernard Fendler, Lauren Young, Mandy Zhao, Michael Coyne, Virginia Breese, Geneva Young, Amy Donahue, Dean Pavlick, Alyssa Tsiros, Timothy Brennan, Shan Zhong, Tariq Mughal, Mark Bailey, Jie HeAbstractGenomic profiling of circulating tumor DNA derived from cell-free DNA (cfDNA) in blood can provide a non-invasive method for the detection of genomic biomarkers to guide clinical decision-making for cancer patients. We developed a...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Pre-Analytical Handling Conditions and Small RNA Recovery from Urine for microRNA Profiling
Publication date: Available online 22 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): David A. Armstrong, John A. Dessaint, Carol S. Ringelberg, Haley F. Hazlett, Louisa Howard, Moemen A.K. Abdalla, Roxanna L. Barnaby, Bruce A. Stanton, Mark A. Cervinski, Alix AshareThere are currently no standardized protocols for pre-analytical handling of urine to best preserve small RNA for microRNA profiling studies. MicroRNA is an attractive candidate as a potential biomarker due to high level of stability in body fluids and its ability to be quantified on multiple high-throughput platforms. Here, we present a compari...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Analysis of mutation and loss of heterozygosity by whole-exome sequencing yields insights into pseudomyxoma peritonei
In conclusion, we have investigated the mutation profile of pseudomyxoma peritonei of appendiceal origin, and provided the first report of RNF43 involvement in its progression. (Source: The Journal of Molecular Diagnostics)
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

A Monochrome Multiplex Real-Time Quantitative PCR Assay for the Measurement of Mitochondrial DNA Content
Publication date: Available online 22 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Anthony Y.Y. Hsieh, Matthew Budd, David Deng, Izabella Gadawska, Hélène C.F. CôtéMitochondrial DNA copies per cell (mtDNA content) can fluctuate with cellular aging, oxidative stress, and mitochondrial dysfunction, and has been investigated in cancer, diabetes, HIV, and metabolic disease. mtDNA content testing in both clinical and basic settings is expected to increase as research uncovers its biological relevance. Herein, we present a novel mtDNA content assay developed on monochrome multiplex r...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Assessing the accuracy of variant detection in cost-effective gene panel testing by next-generation sequencing
Publication date: Available online 25 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Ryoji Fujiki, Makoto Ikeda, Akiko Yoshida, Maeda Akiko, Yue Yao, Motio Nishimura, Kazuyuki Matsushita, Tomohiko Ichikawa, Tomoaki Tanaka, Hiroko Morisaki, Takayuki Morisaki, Osamu OharaAbstractThere is significant debate within the diagnostics community regarding the accuracy of variant identification by next-generation sequencing and the necessity of confirmatory testing of detected variants. Since the quality threshold to discriminate false-positives depends on the nature of the workflow, no regulatory standard regarding...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Multi-center evaluation of the Idylla™ NRAS-BRAF Mutation Test in metastatic colorectal cancer
Publication date: Available online 26 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Iván Prieto-Potin, Clara Montagut, Beatriz Bellosillo, Matthew Evans, Matthew Smith, Linea Melchior, Werner Reiltin, Michael Bennett, Veronica Pennati, Francesca Castiglione, Karl-Friedrich Bürrig, Ulrike Cooper, Barbara Dockhorn-Dworniczak, Christiana Rossenbach, Claudia Maribel Luna-Aguirre, Hugo Alberto Barrera-Saldaña, José Carlos Machado, José Luis Costa, Rinat Yacobi, Hilla Tabibian-KeissarAbstractTreatment of colorectal cancer (CRC) with monoclonal antibodies against epidermal growth...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Characterization of 108 Genomic DNA Reference Materials for 11 Human Leukocyte Antigen (HLA) Loci: A GeT-RM Collaborative Project
Publication date: Available online 26 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Maria P. Bettinotti, Deborah Ferriola, Jamie L. Duke, Timothy L. Mosbruger, Nikolaos Tairis, Lawrence Jennings, Lisa V. Kalman, Dimitri MonosAbstractThe highly polymorphic human leukocyte antigen (HLA) genes, located in the human major histocompatibility complex, encode the class I and II antigen-presenting molecules which are centrally involved in the immune response. HLA typing is used for several clinical applications such as transplantation, pharmacogenetics, and diagnosis of autoimmune disease. HLA typing is highly co...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Determining Performance Metrics for Targeted Next-Generation Sequencing Panels Using Reference Materials
Publication date: Available online 26 June 2018Source: The Journal of Molecular DiagnosticsAuthor(s): Megan H. Cleveland, Justin M. Zook, Marc Salit, Peter M. ValloneAbstractThe National Institute of Standards and Technology has developed reference materials for five human genomes. DNA aliquots are available for purchase and the data, analyses, and high-confidence small variant and homozygous reference calls are freely available on the web. These reference materials are useful for evaluating whole-genome sequencing methods and can also be used to benchmark targeted sequencing panels, which are commonly used in clinical set...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Evaluation of a Hepatitis C Virus Core Antigen Assay in Plasma and Dried-Blood Spot Samples
This study evaluated the analytical performance of HCV core antigen (HCVcAg) detection in plasma and dried blood spot (DBS) samples. Paired plasma and venous DBS samples were prepared from remnant diagnostic samples. Plasma HCV RNA was quantified by AmpliPrep/COBAS Taqman (Roche) and HCVcAg measured by ARCHITECT HCV Ag (Abbott Diagnostics). Sensitivity and specificity for HCVcAg (>3 fmol/L) at two HCV RNA thresholds (≥15 IU/mL and ≥3,000 IU/mL) were calculated. Of 120 paired samples tested, 25 had non-quantifiable HCV RNA and 95 quantifiable HCV RNA. The median HCV RNA level in plasma was 5.6 log10 IU/mL (IQR: 5.2...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Editorial Board
Publication date: July 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 4Author(s): (Source: The Journal of Molecular Diagnostics)
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

A Reliable Targeted Next-Generation Sequencing Strategy for Diagnosis of Myopathies and Muscular Dystrophies, Especially for the Giant Titin and Nebulin Genes
Publication date: July 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 4Author(s): Reda Zenagui, Delphine Lacourt, Henri Pegeot, Kevin Yauy, Raul Juntas Morales, Corine Theze, François Rivier, Claude Cances, Guilhem Sole, Dimitri Renard, Ulrike Walther-Louvier, Xavier Ferrer-Monasterio, Caroline Espil, Marie-Christine Arné-Bes, Pascal Cintas, Emmanuelle Uro-Coste, Marie-Laure Martin Negrier, Valérie Rigau, Eric Bieth, Cyril GoizetMyopathies and muscular dystrophies (M-MDs) are genetically heterogeneous diseases, with>100 identified genes, including the giant and complex titin (TTN...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Efficient Detection of Copy Number Mutations in PMS2 Exons with a Close Homolog
Publication date: July 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 4Author(s): Daniel S. Herman, Christina Smith, Chang Liu, Cecily P. Vaughn, Selvi Palaniappan, Colin C. Pritchard, Brian H. ShirtsDetection of 3′ PMS2 copy-number mutations that cause Lynch syndrome is difficult because of highly homologous pseudogenes. To improve the accuracy and efficiency of clinical screening for these mutations, we developed a new method to analyze standard capture-based, next-generation sequencing data to identify deletions and duplications in PMS2 exons 9 to 15. The approach captures sequences using PMS2 ...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Validity of Targeted Next-Generation Sequencing in Routine Care for Identifying Clinically Relevant Molecular Profiles in Non–Small-Cell Lung Cancer: Results of a 2-Year Experience on 1343 Samples
Publication date: July 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 4Author(s): Antoine Legras, Marc Barritault, Anne Tallet, Elizabeth Fabre, Alice Guyard, Bastien Rance, William Digan, Nicolas Pecuchet, Etienne Giroux-Leprieur, Catherine Julie, Stéphane Jouveshomme, Véronique Duchatelle, Véronique Giraudet, Laure Gibault, Alain Cazier, Jean Pastre, Françoise Le Pimpec-Barthes, Pierre Laurent-Puig, Hélène BlonsTheranostic assays are based on single-gene testing, but the ability of next-generation sequencing (NGS) to interrogate numerous genetic alterations wi...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

System for Informatics in the Molecular Pathology Laboratory: An Open-Source End-to-End Solution for Next-Generation Sequencing Clinical Data Management
We describe the features of SIMPL, its clinical validation at University of Chicago Medicine, and its installation and testing within a different academic center laboratory (University of Colorado), and we propose a platform for future community co-development and interlaboratory data sharing. (Source: The Journal of Molecular Diagnostics)
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Table of Contents
Publication date: July 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 4Author(s): (Source: The Journal of Molecular Diagnostics)
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Molecular Minimal Residual Disease Monitoring in Acute Myeloid Leukemia: Challenges and Future Directions
Publication date: July 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 4Author(s): Adrian G. Selim, Andrew S. MooreThe ability to sensitively monitor minimal residual disease (MRD) has played a key role in improving the management and outcomes for a number of leukemias, particularly acute promyelocytic leukemia and childhood acute lymphoblastic leukemia. By contrast, MRD monitoring in acute myeloid leukemia (AML) has been limited by variable assay methodologies and a relative paucity of patient-specific MRD markers. Inter- and intratumor genetic heterogeneity poses significant challenges for the identifi...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Transferring a Quantitative Molecular Diagnostic Test to Multiple Real-Time Quantitative PCR Platforms
Publication date: July 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 4Author(s): Claudia Gürtler, Mark Laible, Wolfgang Schwabe, Heike Steinhäuser, Xingmin Li, Shujin Liu, Kornelia Schlombs, Ugur SahinQuantitative gene expression assays are increasingly used for diagnosis and research, but are often restricted to specific instrumentation. We propose a robust technical and statistical framework that enables transferring of established real-time quantitative PCR assays across real-time quantitative PCR platforms without compromising analytical and clinical validity. The feasibility of our appro...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Suppression of Wild-Type Amplification by Selectivity Enhancing Agents in PCR Assays that Utilize SuperSelective Primers for the Detection of Rare Somatic Mutations
We report here that the inclusion of the selectivity enhancing agents tetramethylammonium chloride or bis-tetramethylammonium oxalate in SuperSelective PCR assays substantially suppresses the amplification of related wild-type fragments. As a result of this selective suppression, assay sensitivity is increased to such an extent that multiplex PCR assays can be performed in which it is highly unlikely that there will be a false-positive or false-negative result. This advance provides a foundation for the development of rapid, low-cost, multiplex PCR assays for noninvasively assessing the presence of relevant mutations in ca...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research

Accurate Typing of Human Leukocyte Antigen Class I Genes by Oxford Nanopore Sequencing
Publication date: July 2018Source: The Journal of Molecular Diagnostics, Volume 20, Issue 4Author(s): Chang Liu, Fangzhou Xiao, Jessica Hoisington-Lopez, Kathrin Lang, Philipp Quenzel, Brian Duffy, Robi D. MitraOxford Nanopore Technologies' MinION has expanded the current DNA sequencing toolkit by delivering long read lengths and extreme portability. The MinION has the potential to enable expedited point-of-care human leukocyte antigen (HLA) typing, an assay routinely used to assess the immunologic compatibility between organ donors and recipients, but the platform's high error rate makes it challenging to type alleles wit...
Source: The Journal of Molecular Diagnostics - July 10, 2018 Category: Pathology Source Type: research