Purpose: To evaluate whether fluorescence correlation spectroscopy (FCS) can be used to characterize the complexation between oligonucleotides and cationic polymers.
Methods: The features of the complexes between rhodamine labeled oligonucleotides (Rh-ONs) and poly(2-dimethylamino)ethyl methacrylate (pDMAEMA), poly(ethylene glycol)-poly(ethyleneimine) (pEG-pEI), and diaminobutane-dendrimer-(NH2)64 (DAB64) were characterized by light scattering, electrophoretic mobility, electrophoresis, and FCS.
Results: At low polymer/Rh-ON ratios, a decrease of the fluorescence of the Rh-ONs was observed on binding of the Rh-ONs to all cationic polymers. This was explained by the creation of "multimolecular complexes" in which the Rh-labels quench each other. The multimolecular complexes, which are highly fluorescent as they carry a number of Rh-ONs, resulted in high fluorescence peaks in the fluorescence fluctuation profile as measured by FCS. For pDMAEMA and DAB64, at higher polymer/Rh-ON ratios the fluorescence of the polyplexes increased, caused by the formation of "monomolecular complexes," which consist of only one Rh-ON per polymer. In the case of pEG-pEI, the fluorescence stayed constant when the polymer/Rh-ON ratio increased, so multimolecular polyplexes remained. FCS confirmed these results as the high fluorescence peaks disappeared in case of pDMAEMA/Rh-ON and DAB64/Rh-ON dispersions, but remained present for pEG-pEI/Rh-ON dispersions.
Conclusions: FCS seems applicable for study of the interactions between ONs and different types of cationic polymers.