Switchable aptamers for biosensing and bioseparation of viruses (SwAps-V).

Abstract

There is a widespread interest in the development of aptamer-based affinity chromatographic methods for purification of biomolecules. Regardless of the many advantages exhibited by aptamers when compared to other recognition elements, the lack of an efficient regeneration technique that can be generalized to all targets has encumbered further integration of aptamers into affinity-based purification methods. Here we offer switchable aptamers (SwAps) that have been developed to solve this problem and move aptamer-based chromatography forward. SwAps are controlled-affinity aptamers, which have been employed here to purify vesicular stomatitis virus (VSV) as a model case, however this technique can be extended to all biologically significant molecules. VSV is one oncolytic virus out of an arsenal of potential candidates shown to provide selective destruction of cancer cells both in vitro and in vivo. These SwAps were developed in the presence of Ca(2+) and Mg(2+) ions where they cannot bind to their target VSV in absence of these cations. Upon addition of EDTA and EGTA, the divalent cations were sequestered from the stabilized aptameric structure causing a conformational change and subsequently release of the virus. Both flow cytometry and electrochemical impedance spectroscopy were employed to estimate the binding affinities between the selected SwAps and VSV and to determine the coefficient of switching (CoS) upon elution. Among fifteen sequenced SwAps, four have exhibited high affinity to VSV and ability to switch upon elution and thus were further integrated into streptavidin-coated magnetic beads for purification of VSV.