Fluorescent aptasensor for highly sensitive detection of Staphylococcus aureus based on dual-amplification strategy by integrating DNA walking and hybridization chain reaction.

ABSTRACT

Food-borne diseases caused by bacteria threaten human health. Herein, we presented a new fluorescent aptasensor by coupling DNA walking and hybridization chain reaction (HCR) for convenient and sensitive quantification of bacteria. Staphylococcus aureus (S. aureus) was selected as target. When there was target in the system, the binding of S. aureus with its aptamer caused the disintegration of aptamer/DNA walker on the surface of AuNPs and released DNA walker. With the help of Nt.BsmAI, DNA walker moved along the surface of AuNPs and trigger probe was detached from AuNPs. The trigger probe could initiate hybridization chain reaction (HCR) and opened the stems of H1@AuNPs probe and H2@AuNPs probe. After the addition of nicking endonuclease, the adjacent upconversion nanoparticles (UCNPs, NaYF4Yb3+, Er3+) were further away from the quenchers (AuNPs) of H1 and H2. Therefore, the fluorescence intensity of UCNPs could be restored via fluorescence resonance energy transfer (FRET). Bacteria were thus detected by recording the fluorescence intensity of UCNPs. This method is simple, rapid and sensitive. It can directly detect bacteria in a low background signal. The limit of detection (LOD) was 10 CFU/mL, detection time was less than 3 h. Recovery rates in simulated milk, honey and human serum samples ranged from 93.6 % to 105.8 %. The strategy opens up new paths for early diagnosis of diseases and food monitoring.