One of the hallmarks of B lymphoid malignancies is a B cell clone characterized by a unique footprint of clonal immunoglobulin (IG) gene rearrangements that serves as a diagnostic marker for clonality assessment. The EuroClonality/BIOMED-2 assay is currently the gold standard for analyzing IG heavy chain (IGH) and κ light chain (IGK) gene rearrangements of suspected B cell lymphomas. Here, the EuroClonality-NGS Working Group presents a multicentre technical feasibility study of a novel approach involving next-generation sequencing (NGS) of IGH and IGK loci rearrangements that is highly suitable for detecting IG gene rearrangements in frozen and formalin-fixed paraffin-embedded tissue specimens. By employing gene-specific primers for IGH and IGK amplifying smaller amplicon sizes in combination with deep sequencing technology, this NGS-based IG clonality analysis showed robust performance, even in DNA samples of suboptimal DNA integrity, and a high clinical sensitivity for the detection of clonal rearrangements. Bioinformatics analyses of the high-throughput sequencing data with ARResT/Interrogate, a platform developed within the EuroClonality-NGS Working Group, allowed accurate identification of clonotypes in both polyclonal cell populations and monoclonal lymphoproliferative disorders. This multicentre feasibility study is an important step towards implementation of NGS-based clonality assessment in clinical practice, which will eventually improve lymphoma diagnostics.

Charité Universitätsmedizin Berlin Institute of Pathology D 10117 Berlin Germany

Department of Haematology Belfast City Hospital Belfast BT9 7AB UK

Department of Hematology University Hospital Schleswig Holstein 24105 Kiel Germany

Department of Immunology Laboratory Medical Immunology Erasmus MC University Medical Center Rotterdam 3015 CN Rotterdam The Netherlands

Department of Pathology Radboud University Medical Center 6525 GA Nijmegen The Netherlands

Hematology Department Hospital Pitié Salpêtrière and Sorbonne University 75013 Paris France

Institute of Applied Biosciences GR 57001 Thermi Thessaloniki Greece

Institute of Pathology and Neuropathology Comprehensive Cancer Center University Hospital Tübingen 72076 Tübingen Germany

Molecular Medicine Program Central European Institute of Technology Masaryk University 62500 Brno Czech Republic

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