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Leuk Lymphoma. 2011 May;52(5):896-904. doi: 10.3109/10428194.2011.555569. Epub 2011 Feb 21.

Sensitive quantitation of minimal residual disease in chronic myeloid leukemia using nanofluidic digital polymerase chain reaction assay.

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1
Molecular Genetics Research Institute, The Catholic University of Korea, Seoul, Korea.

Abstract

Undetectable BCR-ABL transcripts in patients with chronic myeloid leukemia (CML) should not be regarded as indicative of a cure, due to the sensitivity limit of current real-time quantitative polymerase chain reaction (RQ-PCR) technology. To demonstrate the feasibility of more sensitive approaches, 62 samples from 43 patients with CML were screened by conventional RQ-PCR, replicate RQ-PCR (rRQ-PCR), and/or nanofluidic digital PCR (dPCR). First, we confirmed the correlation of dPCR to conventional RQ-PCR using 30 patient samples with various minimal residual disease (MRD) levels. When the sensitivity limits were determined using cell line and patient sample dilutions, rRQ-PCR and dPCR with pre-amplification showed 2-3 log improvement compared to conventional RQ-PCR, and 24 of 32 PCR negative samples as assayed by conventional RQ-PCR showed detectable BCR-ABL in rRQ-PCR and/or dPCR. More important, using dPCR in conjunction with a pre-amplification step, a continuous decline in MRD level could be precisely monitored even after it became undetectable by conventional RQ-PCR. In this study, both rRQ-PCR and dPCR demonstrated successful detection of BCR-ABL transcripts not detectable in conventional RQ-PCR, and these data show the potential feasibility of highly sensitive PCR approaches for molecular monitoring and clinical relevance in future CML management by allowing further characterization of patients who achieve PCR negativity in a conventional RQ-PCR assay.

PMID:
21338281
DOI:
10.3109/10428194.2011.555569
[Indexed for MEDLINE]

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