Abstract
A bifunctional europium complex of Eu(TTA)(3)(5-NH(2)-phen) using 2-thenoyltrifluoroacetonate (TTA) and 5-amino-1,10-phenanthroline (5-NH(2)-phen) as ligand reagents was applied in DNA detection assays for the first time. The complex has a long fluorescence lifetime, high fluorescence quantum yield, and is easy to label oligonucleotides for time-resolved fluorescence bioanalysis. A two-probe tandem DNA hybridization assay including capture DNA(1), probe DNA(2), and target DNA(3) was employed to detect microbial pathogens. The DNA sequences in the assay were designed using software Primer Premier 5.0 based on published specific nucleotide sequences of Staphylococcus aureus and Escherichia coli. 3'-Amino-modified capture DNA(1) was covalently immobilized on the common glass slide surface and the 5'-amino-modified probe DNA(2) was combined with the functionalized Eu(TTA)(3)(5-NH(2)-phen) via glutaraldehyde. The detection was done by monitoring the fluorescence intensity from the glass surface after the hybridization reaction with complementary target DNA(3). The optimal concentration of capture DNA(1) of 1.0 x 10(-6) mol l(-1) dropped onto the glass slides and optimal hybridization temperatures of 48 degrees C and 39 degrees C respectively for Staphylococcus aureus and Escherichia coli were obtained. The proposed DNA detection system showed higher sensitivity than such a complex doped nanoparticle-based detection system in our previous study for the better uniformity and dispersity of monomolecular labels. The sensing system presented a short hybridization time of 2 h, satisfactory stability, and high selectivity. The results demonstrate that this complex might be a potentially excellent dye in area of biochemical analysis.