Clinical biochemistry · 2012
Four-channel asymmetric Real-Time PCR hybridization probe assay: a rapid pre-screening method for critical BCR-ABL kinase domain mutations.
СтатьяВалидационное исследованиеНазвание и аннотация: Перевод пока не подготовлен
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Martinez-Serra Jordi, Gutiérrez Antonio, Marcús Toni F, Soverini Simona, Amat Juan Carlos, Navarro-Palou María, Ros Teresa, Bex Teresa, Ballester Carmen, Bauça Josep Miquel, SanFelix Sara, Novo Andrés, Vidal Carmen, Santos Carmen, Besalduch Joan
Аннотация
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.
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.
Предметные рубрики MeSH
AdultAgedAged, 80 and overAntineoplastic AgentsBase SequenceBenzamidesBone MarrowDNA Mutational AnalysisDNA ProbesDrug Resistance, NeoplasmFemaleFluorescence Resonance Energy TransferFusion Proteins, bcr-ablHumansImatinib MesylateLeukemia, Myelogenous, Chronic, BCR-ABL PositiveMaleMiddle AgedMultiplex Polymerase Chain ReactionMutationNucleic Acid DenaturationPiperazinesPyrimidinesReal-Time Polymerase Chain ReactionRetrospective Studies