Validation of an in Vitro Mismatch Repair Assay Used in the Functional Characterization of Mismatch Repair Variants.

Autor:	González-Acosta M; Hereditary Cancer Program, the Catalan Institute of Oncology (ICO), Hereditary Cancer Group, Molecular Mechanisms and Experimental Therapy in Oncology Program, Institut d'Investigació Biomèdica de Bellvitge (IDIBELL), Ciber Oncología (CIBERONC) Instituto Salud Carlos III, L'Hospitalet de Llobregat, Barcelona, Spain., Hinrichsen I; Biomedical Research Laboratory, Department of Internal Medicine 1, Universitätsklinikum Frankfurt, Frankfurt, Germany., Fernández A; Hereditary Cancer Program, the Catalan Institute of Oncology (ICO), Hereditary Cancer Group, Molecular Mechanisms and Experimental Therapy in Oncology Program, Institut d'Investigació Biomèdica de Bellvitge (IDIBELL), Ciber Oncología (CIBERONC) Instituto Salud Carlos III, L'Hospitalet de Llobregat, Barcelona, Spain., Lázaro C; Hereditary Cancer Program, the Catalan Institute of Oncology (ICO), Hereditary Cancer Group, Molecular Mechanisms and Experimental Therapy in Oncology Program, Institut d'Investigació Biomèdica de Bellvitge (IDIBELL), Ciber Oncología (CIBERONC) Instituto Salud Carlos III, L'Hospitalet de Llobregat, Barcelona, Spain., Pineda M; Hereditary Cancer Program, the Catalan Institute of Oncology (ICO), Hereditary Cancer Group, Molecular Mechanisms and Experimental Therapy in Oncology Program, Institut d'Investigació Biomèdica de Bellvitge (IDIBELL), Ciber Oncología (CIBERONC) Instituto Salud Carlos III, L'Hospitalet de Llobregat, Barcelona, Spain., Plotz G; Biomedical Research Laboratory, Department of Internal Medicine 1, Universitätsklinikum Frankfurt, Frankfurt, Germany., Capellá G; Hereditary Cancer Program, the Catalan Institute of Oncology (ICO), Hereditary Cancer Group, Molecular Mechanisms and Experimental Therapy in Oncology Program, Institut d'Investigació Biomèdica de Bellvitge (IDIBELL), Ciber Oncología (CIBERONC) Instituto Salud Carlos III, L'Hospitalet de Llobregat, Barcelona, Spain. Electronic address: gcapella@iconcologia.net.
Jazyk:	angličtina
Zdroj:	The Journal of molecular diagnostics : JMD [J Mol Diagn] 2020 Mar; Vol. 22 (3), pp. 376-385. Date of Electronic Publication: 2019 Dec 25.
DOI:	10.1016/j.jmoldx.2019.12.001
Abstrakt:	A significant proportion of DNA-mismatch repair (MMR) variants are classified as of unknown significance, precluding diagnosis. The in vitro MMR assay is used to assess their MMR capability, likely the most important function of an MMR protein. However, the robustness of the assay, crucial for its use in the clinical setting, has been rarely evaluated. The aim of the present work was to validate an in vitro MMR assay approach to the functional characterization of MMR variants, as a first step to meeting quality standards of diagnostic laboratories. The MMR assay was optimized by testing a variety of reagents and experimental conditions. Reference materials and standard operating procedures were established. To determine the intra- and interexperimental variability of the assay and its reproducibility among centers, independent transfections of six previously characterized MLH1 variants were performed in two independent laboratories. Reagents and conditions optimal for performing the in vitro MMR assay were determined. The validated assay demonstrated no significant intra- or interexperimental variability and good reproducibility between centers. We set up a robust in vitro MMR assay that can provide relevant in vitro functional evidence for MMR variant pathogenicity assessment, eventually improving the molecular diagnosis of hereditary cancer syndromes associated with MMR deficiency. (Copyright © 2020 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
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