Characterization of the binding activity of immobilized DNA aptamers for nucleotide and non-nucleotide targets
Dunaway, Adam Blake
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Deoxyribonucleic acid (DNA) aptamers are oligonucleotides with high specificity and affinity for non-nucleotide targets ranging from molecular species to cellular proteins. Their high affinity, rapid synthesis, and the ease with which they can be chemically modified to include convenient chemical groups (e.g. amine group on 5’ end) make them excellent adaptable ligands for use in colloidal drug delivery vehicles for both uptake and release of therapeutic agents. This work uses pre-identified aptamers for vascular endothelial growth factor (VEGF) to investigate the design of one such vehicle for controlled uptake and release of target therapeutics and analyzes the ability of particle-immobilized aptamers to bind both nucleotide and non-nucleotide targets. Aptamer sequences are immobilized on colloidal microspheres and binding activity of both the primary DNA and protein targets are directly monitored using flow cytometry. Additionally, the dual nature of aptamer-target binding is further investigated by evaluating the effects of simultaneous and serial incubation of the primary targets. Finally, the ability to recover the functionality of the aptamer is evaluated after displacement of the primary DNA target through DNA mediated interactions. It has been shown that the nature of aptamer-target interactions are complex in nature, requiring optimization for each species incorporated into a delivery vehicle; however, partial recovery of aptamer functionality was achieved after hybridization with the primary DNA target.