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Four-channel asymmetric Real-Time PCR hybridization probe assay: A rapid pre-screening method for critical BCR-ABL kinase domain mutations
- Source :
- Clinical Biochemistry. 45:345-351
- Publication Year :
- 2012
- Publisher :
- Elsevier BV, 2012.
-
Abstract
- Objectives Within the laboratory protocols, used for the study of BCR-ABL resistance mutations in chronic myeloid leukemia patients treated with Imatinib, direct sequencing remains the reference method. Since the incidence of patients with a mutation-related loss of response is not very high, it is very useful in the routine laboratory to perform a fast pre-screening method. Design and methods With this in mind, we have designed a new technique, based on a single Real-Time FRET-based PCR, followed by a study of melting peaks. This new tool, developed in a LightCycler 2.0, combines four different fluorescence channels for the simultaneous detection, in a single close tube, of critical mutations within the ABL kinase domain. Results Assay evaluation performed on 33 samples, previously genotyped by sequentiation, resulted in full concordance of results. Conclusions This new methodology detects in a few steps the presence of critical mutations associated to Imatinib resistance.
- Subjects :
- Male
Asymmetric
NILOTINIB
DNA Mutational Analysis
Clinical Biochemistry
Fusion Proteins, bcr-abl
Nucleic Acid Denaturation
Piperazines
chemistry.chemical_compound
Bone Marrow
hemic and lymphatic diseases
Fluorescence Resonance Energy Transfer
ponatinib
BCR-ABL
Aged, 80 and over
DASATINIB
ABL
Hybridization probe
Ponatinib
Myeloid leukemia
General Medicine
Middle Aged
Dasatinib
Real-time polymerase chain reaction
Benzamides
Imatinib Mesylate
Female
DNA Probes
medicine.drug
Adult
kinase
Antineoplastic Agents
Biology
Real-Time Polymerase Chain Reaction
IMATINIB
Leukemia, Myelogenous, Chronic, BCR-ABL Positive
medicine
Humans
Aged
Retrospective Studies
Base Sequence
Imatinib
Molecular biology
Pyrimidines
chemistry
Nilotinib
Drug Resistance, Neoplasm
Mutation
FRET
real-time PCR
Multiplex Polymerase Chain Reaction
Subjects
Details
- ISSN :
- 00099120
- Volume :
- 45
- Database :
- OpenAIRE
- Journal :
- Clinical Biochemistry
- Accession number :
- edsair.doi.dedup.....21075cd18bf12e3b30dddababcc1fd07