Nucleic acid aptamers are versatile molecular recognition agents that bind to their targets with high selectivity and affinity. The past few years have seen a dramatic increase in aptamer development and interest for diagnostic and therapeutic applications. As the applications for aptamers expand, the need for a more standardized, stringent, and informative characterization and validation methodology increases. Here we performed a comprehensive analysis of a panel of conventional affinity binding assays using a suite of aptamers for the small molecule target ochratoxin A (OTA). Our results highlight inconsistency between conventional affinity assays and the need for multiple characterization strategies. To mitigate some of the challenges revealed in our head-to-head comparison of aptamer binding assays, we further developed and evaluated a set of novel strategies that facilitate efficient screening and characterization of aptamers in solution. Finally, we provide a workflow that permits rapid and robust screening, characterization, and functional verification of aptamers thus improving their development and integration into novel applications. (Chemical Equation Presented).

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Journal Analytical Chemistry
McKeague, M. (Maureen), De Girolamo, A. (Annalisa), Valenzano, S. (Stefania), Pascale, M. (Michelangelo), Ruscito, A. (Annamaria), Velu, R. (Ranganathan), … DeRosa, M.C. (2015). Comprehensive Analytical Comparison of Strategies Used for Small Molecule Aptamer Evaluation. Analytical Chemistry, 87(17), 8608–8612. doi:10.1021/acs.analchem.5b02102