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Leukemia (2003) 17, 22572317
& 2003 Nature Publishing Group All rights reserved 0887-6924/03 $25.00www.nature.com/leuLEADING ARTICLEDesign and standardization of PCR primers and protocols for detection of clonal
immunoglobulin and T-cell receptor gene recombinations in suspect
lymphoproliferations: Report of the BIOMED-2 Concerted Action BMH4-CT98-3936JJM van Dongen1, AW Langerak1, M Bruggemann2, PAS Evans3, M Hummel4, FL Lavender5, E Delabesse6, F Davi7,E Schuuring8,9, R Garca-Sanz10, JHJM van Krieken11, J Droese2, D Gonzalez10, C Bastard12, HE White5, M Spaargaren13,
M Gonzalez10, A Parreira14, JL Smith5, GJ Morgan3, M Kneba2 and EA Macintyre61Department of Immunology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands;2II Medizinische4lnstitute of Pathology, Free University Berlin, Berlin, Germany; 5Wessex Immunology
Service, Molecular Pathology Unit, Southampton University Hospitals NHS Trust, Southampton, UK;6Laboratoire dHematologieKlinik des Universitatsklinikums Schleswig-Holstein, Campus Kiel, Kiel, Germany;3Academic Unit of Haematology andOncology, University of Leeds, Leeds, UK;and INSERM EMI 210, Hopital Necker-Enfants Malades, Paris, France;7Department of Hematology, Hopital Pitie-Salpetriere,8Department of Pathology, Leiden University Medical Center, Leiden, The Netherlands; 9Department of Pathology
and Laboratory Medicine, University Medical Center Groningen, Groningen, The Netherlands;10Servicio de Hematologia,Paris, France;12Laboratoire de Genetique Oncologique, Centre Henri Becquerel, Rouen, France; 13Department of
Pathology, Academic Medical Center, Amsterdam, The Netherlands; and14Department of Hematology, Instituto Portugues deHospital Universitario de Salamanca, Salamanca, Spain;11Department of Pathology, University Medical Center Nijmegen,Nijmegen, The Netherlands;Oncologia, Lisboa, PortugalIn a European BIOMED-2 collaborative study, multiplex PCR
assays have successfully been developed and standardized for
the detection of clonally rearranged immunoglobulin (Ig) and T-
cell receptor (TCR) genes and the chromosome aberrations
t(11;14) and t(14;18). This has resulted in 107 different primers
in only 18 multiplex PCR tubes: three VHJH, two DHJH, two Ig
kappa (IGK), one Ig lambda (IGL), three TCR beta (TCRB), two
TCR gamma (TCRG), one TCR delta (TCRD), three BCL1-Ig
heavy chain (IGH), and one BCL2-IGH. The PCR products of Ig/
TCR genes can be analyzed for clonality assessment by
heteroduplex analysis or GeneScanning. The detection rate of
clonal rearrangements using the BIOMED-2 primer sets is
unprecedentedly high. This is mainly based on the complementarity of the various BIOMED-2 tubes. In particular,
combined application of IGH (VHJH and DHJH) and IGK
tubes can detect virtually all clonal B-cell proliferations, even in
B-cell malignancies with high levels of somatic mutations. The
contribution of IGL gene rearrangements seems limited.
Combined usage of...