Water-soluble conjugated polymer as a platform for adenosine deaminase sensing based on fluorescence resonance energy transfer technique.

We report a new biosensor for adenosine deaminase (ADA) sensing based on water-soluble conjugated poly(9,9-bis(6'-N,N,N-trimethylammonium)hexyl)fluorine phenylene (PFP) and fluorescence resonance energy transfer technique. In this biosensor, PFP, DNAc-FI labeled with fluorescein (FAM), and ethidium bromide (EB) were used as the fluorescence energy donor, resonance gate, and the final fluorescence energy acceptor, respectively. In the absence of ADA, the adenosine aptamer forms a hairpin-like conformation with adenosine, which is far from its complementary single-stranded DNA (DNAc-FI). When PFP is excited at 380 nm, fluorescein emits strong green fluorescence via one-step FRET while EB has no fluorescence. After addition of ADA, adenosine is hydrolyzed to inosine and then double-stranded DNA (dsDNA) is formed between the aptamer and DNAc-FI, followed by EB intercalating into dsDNA. Once PFP is excited, EB will emit strong yellow fluorescence after two-step FRET from PFP to fluorescein and from fluorescein to EB. The sensitive ADA detection then is realized with a low detection limit of 0.5 U/L by measuring the FRET ratio of EB to fluorescein. Most importantly, the assay is accomplished homogeneously in 25 min without further treatments, which is much more simple and rapid than that reported in literature. Hence, this method demonstrates the sensitive, cost-effective, and rapid detection of ADA activity. It also opens an opportunity for designing promising sensors for other enzymes.