title: Comprehensive analytical comparison of strategies used for small molecule aptamer evaluation.
abstract: 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.
authors.affiliation: Department of Bioengineering, Stanford University , 443 Via Ortega, Stanford, California 94305, United States.
authors.forename: Maureen
authors.initials: M
authors.lastname: McKeague
authors.affiliation: Institute of Sciences of Food Production, National Research Council of Italy , via G. Amendola 122/O, 70126 Bari, Italy.
authors.forename: Annalisa
authors.initials: A
authors.lastname: De Girolamo
authors.affiliation: Institute of Sciences of Food Production, National Research Council of Italy , via G. Amendola 122/O, 70126 Bari, Italy.
authors.forename: Stefania
authors.initials: S
authors.lastname: Valenzano
authors.affiliation: Institute of Sciences of Food Production, National Research Council of Italy , via G. Amendola 122/O, 70126 Bari, Italy.
authors.forename: Michelangelo
authors.initials: M
authors.lastname: Pascale
authors.affiliation: Chemistry Department, Carleton University , 1125 Colonel By Drive, Ottawa, Ontario, Canada K1S 5B6.
authors.forename: Annamaria
authors.initials: A
authors.lastname: Ruscito
authors.affiliation: Chemistry Department, Carleton University , 1125 Colonel By Drive, Ottawa, Ontario, Canada K1S 5B6.
authors.forename: Ranganathan
authors.initials: R
authors.lastname: Velu
authors.affiliation: Chemistry Department, Carleton University , 1125 Colonel By Drive, Ottawa, Ontario, Canada K1S 5B6.
authors.forename: Nadine R
authors.initials: NR
authors.lastname: Frost
authors.affiliation: Chemistry Department, Carleton University , 1125 Colonel By Drive, Ottawa, Ontario, Canada K1S 5B6.
authors.forename: Kayla
authors.initials: K
authors.lastname: Hill
authors.affiliation: Chemistry Department, Carleton University , 1125 Colonel By Drive, Ottawa, Ontario, Canada K1S 5B6.
authors.forename: McKenzie
authors.initials: M
authors.lastname: Smith
authors.affiliation: Chemistry Department, Carleton University , 1125 Colonel By Drive, Ottawa, Ontario, Canada K1S 5B6.
authors.forename: Erin M
authors.initials: EM
authors.lastname: McConnell
authors.affiliation: Chemistry Department, Carleton University , 1125 Colonel By Drive, Ottawa, Ontario, Canada K1S 5B6.
authors.forename: Maria C
authors.initials: MC
authors.lastname: DeRosa
mesh_terms: D052157:Aptamers, Nucleotide / Q000737:chemistry*; D002352:Carrier Proteins / Q000737:chemistry; D002623:Chemistry Techniques, Analytical / Q000379:methods*; D052156:SELEX Aptamer Technique
chemical_list: D052157:Aptamers, Nucleotide; D002352:Carrier Proteins