1. Characterization of a reference material for BCR-ABL (M-BCR) mRNA quantitation by real-time amplification assays: towards new standards for gene expression measurements
- Author
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Heike Pfeifer, Ralph B. Arlinghaus, Gisela Barbany, Charles L. Sawyers, Monique Silvy, Katerina Zoi, John Saldanha, Orietta Spinelli, K Miyamura, Martin C. Müller, Jean Gabert, Nathalie Beaufils, Mette Østergaard, Giuseppe Saglio, E Macintyre, V H J van der Velden, Mark Lawler, Jianmin Wang, Paul Matejtschuk, V Patel, Susan Branford, Veli Kairisto, Mahon Fx, Thomas Lion, Giovanni Cazzaniga, P J. Phillips, Jean Michel Cayuela, David Grimwade, Marcos González, Immunology, Saldanha, J, Silvy, M, Beaufils, N, Arlinghaus, R, Barbany, G, Branford, S, Cayuela, J, Cazzaniga, G, Gonzalez, M, Grimwade, D, Kairisto, V, Miyamura, K, Lawler, M, Lion, T, Macintyre, E, Mahon, F, Muller, M, Ostergaard, M, Pfeifer, H, Saglio, G, Sawyers, C, Spinelli, O, van der Velden, V, Wang, J, Zoi, K, Patel, V, Phillips, P, Matejtschuk, P, and Gabert, J
- Subjects
Quality Control ,Cancer Research ,Protein-Tyrosine Kinases/analysis ,Polymerase Chain Reaction/methods ,Fusion Proteins, bcr-abl ,Biology ,Philadelphia chromosome ,Polymerase Chain Reaction ,Indicators and Reagent ,Protein-Tyrosine Kinase ,hemic and lymphatic diseases ,K562 Cell ,medicine ,Humans ,RNA, Messenger ,ABL ,RNA, Messenger/analysis ,Gene Expression Profiling ,Myeloid leukemia ,Hematology ,Nucleic acid amplification technique ,Protein-Tyrosine Kinases ,Reference Standards ,medicine.disease ,Molecular biology ,Minimal residual disease ,Gene expression profiling ,Freeze Drying ,Real-time polymerase chain reaction ,Oncology ,Reference Standard ,Indicators and Reagents ,Gene Expression Profiling/methods ,K562 Cells ,Human ,K562 cells - Abstract
Monitoring of BCR-ABL transcripts has become established practice in the management of chronic myeloid leukemia. However, nucleic acid amplification techniques are prone to variations which limit the reliability of real-time quantitative PCR (RQ-PCR) for clinical decision making, highlighting the need for standardization of assays and reporting of minimal residual disease (MRD) data. We evaluated a lyophilized preparation of a leukemic cell line (K562) as a potential quality control reagent. This was found to be relatively stable, yielding comparable respective levels of ABL, GUS and BCR-ABL transcripts as determined by RQ-PCR before and after accelerated degradation experiments as well as following 5 years storage at -20 degrees C. Vials of freeze-dried cells were sent at ambient temperature to 22 laboratories on four continents, with RQ-PCR analyses detecting BCR-ABL transcripts at levels comparable to those observed in primary patient samples. Our results suggest that freeze-dried cells can be used as quality control reagents with a range of analytical instrumentations and could enable the development of urgently needed international standards simulating clinically relevant levels of MRD.
- Published
- 2007