A homogeneous europium cryptate-based assay for the diagnosis of mutations by time-resolved fluorescence resonance energy transfer

摘要: Oligonucleotide ligation assay (OLA) is considered to be a very useful methodology for the detection and characterization of mutations, particularly clinical purposes. The fluorescence resonance energy transfer between fluorescent donor suitable fluorophore as acceptor has been applied in past several scientific fields. This technique well adapted nucleic acid analysis such DNA sequencing, hybridization polymerase chain reaction. We describe here homogeneous format based on use rare earth cryptate label donor: tris-bipyridine-Eu(3+). long-lived this makes it possible reach high sensitivity by using time-resolved mode. A non-radiative technology, known amplification emission (TRACE((R))) characterized temporal spectral selectivity developed. TRACE((R)) single nucleotide polymorphism OLA allelic discrimination proposed. demonstrate potentialities OLA-TRACE((R)) through K-ras oncogene point mutations.