Rapid and sensitive ethidium bromide fluorescence quenching assay of polyamine conjugate-DNA interactions for the analysis of lipoplex formation in gene therapy

Abstract

A rapid and sensitive fluorescent assay method is reported for assessing polyamine conjugate calf thymus DNA binding affinity using cholesterol polyamine carbamates with ethidium bromide as a probe. A reproducible method has been developed with an optimal excitation wavelength. Salt concentration is shown to be a critical parameter for both the observed fluorescence intensity of ethidium intercalated in DNA, and also for the binding of positively charged polyammonium ions to DNA, effecting charge neutralisation. This charge neutralisation precedes DNA condensation, a key first step in gene therapy.