Fluorescence Detection Methods For Protein And Small Molecules Based On Silver Nanoparticles

Aptamers are single-stranded oligonucleotides generated from an in vitro process known as SELEX. Based on their high binding affinity and specificity towards target molecules, aptamers have been utilized in applications ranging from biosening to diagnostics. Meanwhile, nanomaterials possessing untique optical, electronic and catalytic properties can interact with biomolecules to yield signal amplication and target recognition. By combining both technologies, aptamer conjugate nanoparticles offer great promise in bioanalysis.In this assay, we demonstrated a simple and highly sensitive fluorescence platform for protein detection. Silver nanoparticles (AgNPs) work both as carriers and quenchers for FAM labeled aptamers (FAM-apt). Biotin labeled aptamers (Bio-apt), FAM-apt functionalized AgNPs (Ag-FAM-apt), and a target protein-human platelet-derived growth factor-BB (PDGF-BB) could form a sandwich-type complex. Once the etching solvents were added, AgNPs were dissolved and the fluorescence resonance energy transfer (FRET) between AgNPs and FAM was broken, FAM-apt were no longer quenched and released into the solution in the 96-well microplates, so the fluorescent signal would turn from "off’ state to "on" state. This method has possessed several advantages:increased specificity contributed by the sandwich binding of aptamers; quenching ability of AgNPs was utilized to make signal turn-on; high throughout. This simple and sensitive method would have a promising future for development and application.The exploitation of the metal-enhanced fluorescence (MEF) effect based on the localized surface plasmon resonance (LSPR) of metallic nanostructures has broadened the scope of fluorescence analysis. Because of the unique localized surface plasmon resonance property of AgNPs, AgNPs could enhance fluorescence signal of many fluorophores, so lots of assays have been developed. In this paper, we developed a small moleculars detection biosensor based on the MEF and FRET of AgNPs. Firstly, we functionlized the AgNPs with fluorophore, then covered the fluorophore with a silver shell to make enhanced fluorescent signals. Secondly, we functionlized the aptamers which had a quenching group BHQ to the surface of silver shell through hybridization. Because of FRET effect, the fluorescent signals were quenched. But, upon the target adenosine added, the aptamer with BHQ would recognize adenosine specifically and separate from the silver shell, so the fluorescence signals would recover. The results indicated that the intensity of the recovering fluorescence signals had a linear relationship with the concentrations of adenosine. The assay carried out full use of the properties of AgNPs and had high sensitivity and selectivity, it had great potential.