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Leukemia. 2015 Feb;29(2):369-76. doi: 10.1038/leu.2014.217. Epub 2014 Jul 18.

A certified plasmid reference material for the standardisation of BCR-ABL1 mRNA quantification by real-time quantitative PCR.

Author information

1
1] National Genetics Reference Laboratory (Wessex), Salisbury District Hospital, Salisbury, UK [2] Faculty of Medicine, University of Southampton, Southampton, UK.
2
European Commission, Joint Research Centre, Institute for Reference Materials and Measurements, Geel, Belgium.
3
National Genetics Reference Laboratory (Wessex), Salisbury District Hospital, Salisbury, UK.
4
Department of Haematology, Aarhus University Hospital, Aarhus, Denmark.
5
Hungarian National Blood Transfusion Service, Budapest, Hungary.
6
Regional Genetics Laboratory, Birmingham Women's NHS Foundation Trust, Birmingham, UK.
7
Department of Molecular Medicine and Surgery, Clinical Genetics Karolinska Institutet, Stockholm, Sweden.
8
1] Department of Laboratory Medicine, UZ Leuven, Belgium [2] Department of Oncology, KU Leuven, Belgium.
9
Molecular Malignancy Laboratory and Haemato-Oncology Diagnostic Service, Cambridge University Hospitals NHS Foundation Trust, Cambridge, UK.
10
Haematology Department, Belfast City Hospital, Belfast, UK.
11
Haematology Laboratory and EA3518, University Hospital Saint-Louis, AP-HP, University Paris Diderot, Paris, France.
12
Department of Molecular Haematology, Yorkhill NHS Trust, Glasgow, UK.
13
Molecular Haematology, Bristol Royal Infirmary, Bristol, UK.
14
Hematopathology Unit, Hospital Clinic, IDIBAPS, Barcelona, Spain.
15
Department of Clinical and Biological Science, University of Turin, Turin, Italy.
16
Laboratoire Hematologie, CHU Bordeaux, Hematopoiese Leucemique et Cibles Therapeutiques, INSERM U1035, Universite Bordeaux, Bordeaux, France.
17
Combined Laboratories, Derriford Hospital, Plymouth, UK.
18
Department of Genetics and Molecular Pathology, SA Pathology, Adelaide, SA, Australia.
19
Imperial Molecular Pathology, Centre for Haematology, Imperial College London, London, UK.
20
Molecular Pathology, University Hospitals Southampton NHS Foundation Trust, Southampton, UK.
21
Hematology, Oncology and Transfusion Medicine Center, Vilnius University Hospital Santariskiu Clinics, Vilnius, Lithuania.
22
Laboratory of Molecular Biology and UMR5239, Centre Hospitalier Lyon-Sud, Hospices Civils de Lyon, Pierre Bénite, France.
23
Medical Genetics Department, Erasme Hospital, Brussels, Belgium.
24
Department of Clinical Medicine and Surgery, University 'Federico II' of Naples, Naples, Italy.
25
Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden.
26
Department of Clinical Genetics, University and Regional Laboratories, Lund, Sweden.
27
Department of Internal Medicine-Hematology and Oncology, Masaryk University and University Hospital Brno, Brno, Czech Republic.
28
Turku University Hospital, TYKSLAB, Laboratory of Molecular Genetics, Turku, Finland.
29
Laboratory for Molecular Haemato-Oncology, Kings College Hospital, London, UK.
30
Cancer Research Institute, The Catholic University of Korea, Seoul, South Korea.
31
Abteilung für Hämatologie und internistische Onkologie, Universität Leipzig, Leipzig, Germany.
32
Children's Cancer Research Institute/LabDia Labordiagnostik and Medical University, Vienna, Austria.
33
Institute of Hematology and Blood Transfusion, Prague, Czech Republic.
34
Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Bologna, Italy.
35
Cancer Molecular Diagnostics, St James's Hospital, Dublin, Ireland.
36
VU Medical Centre, Department of Haematology, Amsterdam, The Netherlands.
37
Department of Medical Genetics, NHS-Grampian, Aberdeen, UK.
38
Department of Laboratory Medicine, Medical University of Vienna, Vienna, Austria.
39
Laboratory of Hematology, Sheba Medical Center, Tel Hashomer, Israel.
40
Molecular Oncology Diagnostics Unit, Guy's Hospital, London, UK.
41
Lab Hematologia, Hospital de la Santa Creu i Sant Pau, Barcelona, Spain.
42
Division of Pathology, Rikshospital, Oslo University Hospital, Oslo, Norway.
43
Molecular Diagnostics Laboratory, Department of Hematology, University Hospital Bern, Bern, Switzerland.
44
Genetics Department, Institute of Experimental Medicine (DETAE), Istanbul University, Istanbul, Turkey.
45
Specialized Haematology Laboratory, Division of Internal Medicine, Department of Haematology, University Medical Centre, Ljubljana, Slovenia.
46
Department of Internal Medicine, Hematology/Oncology, Goethe University, Frankfurt, Germany.
47
Laboratoire d'hématologie, CHU Lille, Lille, France.
48
United Laboratories of Tartu University Hospitals, Tartu, Estonia.
49
Molecular Haematology Laboratory, PathWest Laboratory Medicine, Royal Perth Hospital, Perth, WA, Australia.
50
Hematology Department, Jagiellonian University, Krakow, Poland.
51
Hematology Department, Fundeni Clinical Institute, University of Medicine and Pharmacy 'Carol Davila', Bucharest, Romania.
52
Hematology Department and HCT Unit, G. Papanicolaou Hospital, Thessaloniki, Greece.
53
Department of Immunology, Erasmus MC, Rotterdam, The Netherlands.
54
Bristol Genetics Laboratory, Southmead Hospital, Bristol, UK.
55
Department of Haematology, Royal Liverpool University Hospital, Liverpool, UK.
56
HMDS, Leeds Institute of Oncology, St James's University Hospital, Leeds, UK.
57
Sheffield Diagnostic Genetics Service, Sheffield Children's NHS Foundation Trust, Sheffield, UK.
58
Molecular Diagnostics, The Royal Marsden NHS Foundation Trust, Sutton, UK.
59
Department of Laboratory Diagnostics, Clinical Hospital Center, Zagreb University School of Medicine, Zagreb, Croatia.
60
Department of Hematology/Oncology, Jena University Hospital, Jena, Germany.
61
Haematology Research Laboratory, Biomedical Research Foundation, Academy of Athens, Athens, Greece.
62
III. Medizinische Klinik, Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany.

Abstract

Serial quantification of BCR-ABL1 mRNA is an important therapeutic indicator in chronic myeloid leukaemia, but there is a substantial variation in results reported by different laboratories. To improve comparability, an internationally accepted plasmid certified reference material (CRM) was developed according to ISO Guide 34:2009. Fragments of BCR-ABL1 (e14a2 mRNA fusion), BCR and GUSB transcripts were amplified and cloned into pUC18 to yield plasmid pIRMM0099. Six different linearised plasmid solutions were produced with the following copy number concentrations, assigned by digital PCR, and expanded uncertainties: 1.08±0.13 × 10(6), 1.08±0.11 × 10(5), 1.03±0.10 × 10(4), 1.02±0.09 × 10(3), 1.04±0.10 × 10(2) and 10.0±1.5 copies/μl. The certification of the material for the number of specific DNA fragments per plasmid, copy number concentration of the plasmid solutions and the assessment of inter-unit heterogeneity and stability were performed according to ISO Guide 35:2006. Two suitability studies performed by 63 BCR-ABL1 testing laboratories demonstrated that this set of 6 plasmid CRMs can help to standardise a number of measured transcripts of e14a2 BCR-ABL1 and three control genes (ABL1, BCR and GUSB). The set of six plasmid CRMs is distributed worldwide by the Institute for Reference Materials and Measurements (Belgium) and its authorised distributors (https://ec.europa.eu/jrc/en/reference-materials/catalogue/; CRM code ERM-AD623a-f).

PMID:
25036192
PMCID:
PMC4320294
DOI:
10.1038/leu.2014.217
[Indexed for MEDLINE]
Free PMC Article

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