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- White, H, et al.
(författare)
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A certified plasmid reference material for the standardisation of BCR-ABL1 mRNA quantification by real time quantitative PCR.
- 2015
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Ingår i: Leukemia. - : Springer Science and Business Media LLC. - 1476-5551 .- 0887-6924. ; 29:2, s. 369-376
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Tidskriftsartikel (refereegranskat)abstract
- Serial quantification of BCR-ABL1 mRNA is an important therapeutic indicator in chronic myeloid leukemia, but there is 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 the numbers of measured transcripts of e14a2 BCR-ABL1 and three control genes; ABL1, BCR and GUSB. The set of 6 plasmid CRMs is distributed worldwide by the Institute for Reference Materials and Measurements (Belgium) and its authorised distributors (http://irmm.jrc.ec.europa.eu; CRM code ERM-AD623a-f).Leukemia accepted article preview online, 18 July 2014; doi:10.1038/leu.2014.217.
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- Pfeifer, H., et al.
(författare)
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Standardisation and consensus guidelines for minimal residual disease assessment in Philadelphia-positive acute lymphoblastic leukemia (Ph plus ALL) by real-time quantitative reverse transcriptase PCR of e1a2 BCR-ABL1
- 2019
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Ingår i: Leukemia. - : NATURE PUBLISHING GROUP. - 0887-6924 .- 1476-5551. ; 33:8, s. 1910-1922
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Tidskriftsartikel (refereegranskat)abstract
- Minimal residual disease (MRD) is a powerful prognostic factor in acute lymphoblastic leukemia (ALL) and is used for patient stratification and treatment decisions, but its precise role in Philadelphia chromosome positive ALL is less clear. This uncertainty results largely from methodological differences relating to the use of real-time quantitative PCR (qRT-PCR) to measure BCR-ABL1 transcript levels for MRD analysis. We here describe the first results by the EURO-MRD consortium on standardization of qRT-PCR for the e1a2 BCR-ABL1 transcript in Ph + ALL, designed to overcome the lack of standardisation of laboratory procedures and data interpretation. Standardised use of EAC primer/probe sets and of centrally prepared plasmid standards had the greatest impact on reducing interlaboratory variability. In QC1 the proportion of analyses with BCR-ABL1/ABL1 ratios within half a log difference were 40/67 (60%) and 52/67 (78%) at 10(-3) and 36/67 (53%) and 53/67 (79%) at 10(-4)BCR-ABL1/ABL1. Standardized RNA extraction, cDNA synthesis and cycler platforms did not improve results further, whereas stringent application of technical criteria for assay quality and uniform criteria for data interpretation and reporting were essential. We provide detailed laboratory recommendations for the standardized MRD analysis in routine diagnostic settings and in multicenter clinical trials for Ph + ALL.
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- Juul-Dam, K. L., et al.
(författare)
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Measurable residual disease assessment by qPCR in peripheral blood is an informative tool for disease surveillance in childhood acute myeloid leukaemia
- 2020
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Ingår i: British Journal of Haematology. - : Wiley. - 0007-1048 .- 1365-2141. ; 190:2, s. 198-208
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Tidskriftsartikel (refereegranskat)abstract
- Serial assessments of measurable (or minimal) residual disease (MRD) by qPCR may identify nascent relapse in children with acute myeloid leukaemia (AML) and enable pre-emptive therapy. We investigated the kinetics and prognostic impact of recurrent fusion transcripts (RUNX1-RUNX1T1, CBFB-MYH11, KMT2A-MLLT3 or KMT2A-ELL) in 774 post-induction samples from bone marrow (BM, 347) and peripheral blood (PB, 427) from 75 children with AML. BM MRD persistence during consolidation did not increase the risk of relapse, and MRD at therapy completion did not correlate to outcome (HR=0·64/MRD log reduction (CI: 0·32–1·26), P=0·19). In contrast, 8/8 patients with detectable MRD in PB after first consolidation relapsed. Persistence (n=4) and shifting from negative to positive (n=10) in PB during follow-up predicted relapse in 14/14 patients. All 253PB samples collected during follow-up from 36 patients in continuous complete remission were MRD negative. In core-binding factor AML, persistent low-level MRD positivity in BM during follow-up was frequent but an increment to above 5×10−4 heralded subsequent haematological relapse in 12/12 patients. We demonstrate that MRD monitoring in PB after induction therapy is highly informative and propose an MRD increment above 5×10−4 in PB and BM as a definition of molecular relapse since it always leads to haematological relapse. © 2020 British Society for Haematology and John Wiley & Sons Ltd
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