Terminal protection of a small molecule-linked loop DNA probe for turn-on label-free fluorescence detection of proteins

• We proposed a simple and fast fluorescence method to detect proteins by loop DNA.
• The DNA can form a self-complementary structure with two stem-loop structures.
• The target protein can bind to biotin protecting DNA from cleavage by exonuclease.
• The DNA sequence could be randomly coded with part self-hybridization.
• This method is a signal off-on fluorescent strategy.

A novel label-free turn-on fluorescence biosensor for the determination of streptavidin (SA) was proposed. Using terminal protection of small molecule-linked DNA chimeras, which can protect DNA from degradation by various exonucleases when the small molecule moieties are bound to their protein target, we designed a loop probe, where the 3′-end was modified with biotin to resist digestion by exonucleases in the presence of target SA. Coupled with an intercalating dye, SYBR Green I, strong enhancement of the fluorescence signals was obtained compared with that in the absence of SA. A linear correlation equation was obtained for SA from 0 to 200 nM with a limit detection of 0.4 nM. This strategy holds great promise for practical applications with good specificity and sensitivity.