Fluorogenic target competitors for developing label-free and sensitive folding-unswitching aptamer sensors

Abstract

Background

Aptamers have aroused tremendous applications in sensors, drug deliveries, diagnosis, and therapies. In particular, target-induced global structure switching of aptamers has been widely used to develop selective sensors. However, fluorophore and/or quencher modification, sequence elongation, and nano-interface adsorption are required to design such global structure-switching aptamer sensors (SSAS) in order to signal target binding events. Accordingly, these requirements make SSAS at a high cost and expense of sensors’ sensitivity. In this aspect, efforts should be made to overcome these drawbacks of SSAS.

Results

Herein, we tried to develop label-free folding-unswitching aptamer sensors (FUAS) by searching fluorogenic target competitors. Using adenine nucleoside/nucleotide as the proof-of-concept model targets, we screened out berberine (BER) from natural isoquinoline alkaloids (having rings comparable to targets) as the best fluorogenic target competitor. Binding of BER at the conserved nucleotides of intact aptamer foldings turned on this fluorogenic target competitor’ fluorescence. Targets then competed with this fluorogenic target competitor over the same conserved nucleotides to cause its release in favor of a resultant fluorescence change. We found that the developed FUAS are much more sensitive than the previously reported SSAS. The FUAS were successfully applied to assays of ATP and adenosine deaminase in serums, and to screening of the adenosine deaminase's inhibitor, verifying the reliability and applicability of this FUAS platform in variant fields.

Significance

We demonstrate that by designing fluorogenic target competitors, FUAS can be alternatively developed in a label-free manner and with a higher sensitivity than the previously developed SSAS. This work opens a new way to develop high-performance aptamer-based sensors. Furthermore, our developed FUAS should inspire more interest for wide applications incluidng target-triggered drug deliveries when therapeutic fluorogenic target competitors are used.