Functional magnetic nanoparticle-based trapping and sensing approaches for label-free fluorescence detection of DNA

In this study, a label-free fluorescence detection method for DNA was designed using functional magnetic nanoparticles (MNPs) as affinity probes. With the advantage of magnetic feature, MNP-based affinity probes can be easily manipulated for trapping and sensing target species. Two types of MNP-based nanoprobes for trapping and detecting target DNAs were fabricated. The basic strategy for this approach is the use of trapping probes to concentrate target DNAs selectively from complex samples. The detection probes are then used as fluorescence reporters to explore the level of the target species. Trapping probes were constructed by covalently immobilizing probe DNA molecules complementary to the target DNA. Detection nanoprobes were made by linking a fluorescent dye, riboflavin-5'-monophosphate (RFMP), onto the surface of the core/shell Fe(3)O(4)@Al(2)O(3) MNPs via Al-phosphate chelation. The fluorescence derived from RFMP molecules became invisible when molecules were attached onto the MNP surface. However, after phosphorylated species (e.g., DNA molecules) replaced RFMP from the surface of the RFMP-Fe(3)O(4)@Al(2)O(3) MNPs under microwave heating for 15s, the RFMP molecules released from the MNPs enhanced the fluorescence intensity in the solution. Based on the measurement of the fluorescence intensity, the level of target DNA in the samples was determined. The remaining DNA molecules on the RFMP-Fe(3)O(4)@Al(2)O(3) MNPs were characterized by using matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). The detection limit for DNA was as low as 40 pM using this approach.