Ultrasensitive Fluorescent miRNA Biosensor Based on a 'Sandwich' Oligonucleotide Hybridization and Fluorescence Resonance Energy Transfer Process Using an Ln(III)-MOF and Ag Nanoparticles for Early Cancer Diagnosis: Application of Central Composite Design

摘要

Herein, a novel, rapid, highly sensitive, and selective fluorescent biosensor is presented, which is designed based on "sandwich-type" hybridization of oligonucleotides and the fluorescence resonance energy transfer (FRET) strategy. It senses and determines the MicroRNA-155 (miRNA-155) expression levels as a cancer biomarker. In this study, a modified La(III)-metal-organic framework(MOF) and silver nanoparticles (Ag NPs) were used as the energy donor-acceptor pairs in fluorescence quenching through the FRET process. La(III)-MOF was synthesized and then modified by glutaraldehyde as a cross-linking agent. The Ag NPs were also prepared, and then, the surface of both was conjugated with different 5'-amino-labeled ssDNA strands (aptamers). These prepared nanoprobes were characterized by various physicochemical techniques such as X-ray diffraction, energy-dispersive X-ray spectrometry, Fourier transform infrared, field emission scanning electron microscopy, UV-vis spectroscopy, elemental mapping, and gel electrophoresis. To optimize the detection conditions, several factors affecting biosensor performance were assessed by one variable-at-a-time and central composite design methods. Under optimum conditions, this "turn-off" fluorescent biosensor could detect and determine as low as 0.04 ppb (ng. mL(-1)) or 5.5 fM of the miRNA-155 biomarker. Therefore, this biosensor provides highly promising potential for lung and breast cancer diagnosis.