Determination of the stoichiometry of DNA–dye interaction and application to the study of a bis-cyanine dye–DNA complex

Some, though not all, cationic dyes exhibit enhanced fluorescence in the presence of DNA. Based on this phenomenon a simple and rapid method has been developed for determining the stoichiometry of interaction of such dyes with DNA. The method was validated using the well-characterised dyes Ethidium Bromide and Thiazole Orange dimer TOTO [1,1′-(4,4,7,7-tetramethyl-4,7-diazaundecamethylene)-bis-4-[3-methyl-2,3-dihydro-(benzo-1,3-thiazole)-2-methyl-idene]-quinolinium tetraiodide] as well as the butyl homologue of Thiazole Orange. In each case the stoichiometries thus determined were consistent with those already reported. Using this protocol the stoichiometry of interaction, with DNA, of a novel type of bis-cyanine dye was studied, and shown to involve binding of an unexpectedly large molar ratio of dye to DNA. Possible mechanisms of binding are suggested.