Fluorescent biosensor based on FRET and catalytic hairpin assembly for sensitive detection of polysialic acid by using a new screened DNA aptamer.

Abstract

A fast and efficient dsDNA library-immobilized magnetic bead-based SELEX technique was employed for selection of the aptamers against polysialic acid (PSA). Overall twelve rounds of screening, the pooled library was subjected to high-throughput sequencing. Five aptamer candidates with low Gibbs binding free energy and high abundance were selected for affinity evaluation. Apt3 was demonstrated to be the optimal aptamer for PSA with Kd of 114.0nM. Furthermore, an ultrasensitive fluorescence resonance energy transfer (FRET)-based biosensor for PSA was constructed by employing the newly selected aptamer and catalytic hairpin assembly (CHA) amplification strategy. The linear detection range for PSA is from 10 pM to 1μM and the limit of detection is 0.63 pM. The fluorescent biosensor is able to detect the target in the complex biological samples, which indicates that Apt3 has good application prospect for the biological detection and clinical diagnostics.