Fluorescent DNA probes with 1,6-hexanediyl as the linker between two pyrenes, phenylpyrenes or phenylethynyl pyrene fluorophores were synthesized (Py-1, Py-2 and Py-3) and their interactions with DNA were studied by UV-vis absorption spectra, fluorescence spectra and viscosity measurements. The probes show red-shifted emission compared with pyrene (up to 20 nm). We found the interaction of these probes with DNA can be either intercalation or groove binding. Ratiometric fluorometry (ratio of the monomer and excimer emission intensity versus concentration of DNA) was achieved with these probes for DNA quantification (with limit of detection, LOD, up to 0.1 microg/mL). We also found that the undesired oxygen sensitivity of the emission intensity of pyrene fluorophore can be greatly suppressed by extending the pi-conjugation framework of pyrene (the I(Ar)/I(air) value is decreased from 8.10 for pyrene to less than 2.20 for the DNA probes described herein).