Abstract
Prototropic and solvatochromatic properties of fluorescein (FL) were employed to detect the presence of microenvironments in polyplexes consisting of polycationic polymer (POCP) and a fluorescein-conjugated RNA, the HIV-1 transactivation response element (TAR-FL). Results reveal new aspects of polyplex structure with respect to polyplex-bound RNA existing in the following local microenvironments: (a) RNA associated with the polyplex that experiences local pH changes in a manner dependent on POCP nitrogen to RNA phosphate ratio (N:P), (b) RNA experiencing relatively acidic local pH environment that remains constant in polyplexes formed after a charge-neutral ratio, and (c) RNA packed close enough to mediate fluorophore/fluorophore quenching. The magnitude of these changes observed as a function of POCP to nucleic acid N:P ratio is polymer dependent. Assessment of the different microenvironments can help elucidate the functional hierarchy of polyplex-bound oligonucleotides and additionally characterize POCPs based on the resulting local pH and solvent properties upon polyplex formation.
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