Characterization of a reference material for BCR-ABL (M-BCR) mRNA quantitation by real-time amplification assays: towards new standards for gene expression measurements.

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.

• Authors: J Saldanha, M Silvy, N Beaufils, R Arlinghaus, G Barbany, S Branford, J-M Cayuela, G Cazzaniga, M Gonzalez, D Grimwade, V Kairisto, K Miyamura, M Lawler, T Lion, E Macintyre, F-X Mahon, M C Muller, M Ostergaard, H Pfeifer, G Saglio, C Sawyers, O Spinelli, V H J van der Velden, J Q Wang, K Zoi, V Patel, P Phillips, P Matejtschuk, J Gabert
• Journal: Leukemia (Leukemia) Vol. 21 Issue 7 Pg. 1481-7 (Jul 2007) ISSN: 0887-6924 [Print] England
• PMID: 17476280 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Indicators and Reagents
• RNA, Messenger
• Protein-Tyrosine Kinases
• Fusion Proteins, bcr-abl

Topics

• Freeze Drying
• Fusion Proteins, bcr-abl
• Gene Expression Profiling (methods, standards)
• Humans
• Indicators and Reagents
• K562 Cells
• Polymerase Chain Reaction (methods, standards)
• Protein-Tyrosine Kinases (analysis, genetics)
• Quality Control
• RNA, Messenger (analysis)
• Reference Standards