 听 |
The fluorescence enhancement mechanisms of a series of DNA stains of the oxazole yellow (YO) family have been investigated in detail using steady鈥恠tate and ultrafast time鈥恟esolved fluorescence spectroscopy. The strong increase in the fluorescence quantum yield of these dyes upon DNA binding is shown to originate from the inhibition of two distinct processes: 1)鈥卛somerisation through large鈥恆mplitude motion that non鈥恟adiatively deactivates the excited state within a few picoseconds and 2)鈥協ormation of weakly emitting H鈥恉imers. As the H鈥恉imers are not totally non鈥恌luorescent, their formation is less efficient than isomerisation as a fluorescent contrast mechanism. The propensity of the dyes to form H鈥恉imers and thus to reduce their fluorescence contrast upon DNA binding is shown to depend on several of their structural parameters, such as their monomeric (YO) or homodimeric (YOYO) nature, their substitution and their electric charge. Moreover, these parameters also have a substantial influence on the affinity of the dyes for DNA and on the ensuing sensitivity for DNA detection. The results give new insight into the development and optimisation of fluorescent DNA probes with the highest contrast.
DOI:
Archive ouverte / Open archive: