A novel near-infrared, single nanoparticle surface plasmon resonance imaging (SPRI) microscope has been developed to detect single DNA and RNA bioaffinity adsorption events on chemically modified gold thin films. The adsorption of single nanoparticles creates point diffraction patterns in the sequential real-time SPRI differential reflectivity images. These difference images are used to create digital adsorption binding curves of surface bioaffinity measurements, specifically the sequence-selective hybridization adsorption of DNA and RNA-functionalized gold nanoparticles. In this work, microRNA strands are enzymatically captured onto 40 nm gold nanoparticles via a surface ligation reaction. Subsequently, single microRNA molecules are detected through single nanoparticle SPRI measurements by monitoring the hybridization adsorption of the microRNA-functionalized gold nanoparticles onto a complementary DNA-functionalized gold surface. The presence of a single RNA-DNA heteroduplex on the surface is verified by the enzymatic desorption of the nanoparticle from the surface using RNase H to hydrolyze the RNA in the heteroduplex.

Single nanoparticle SPRI microscopy for the enzymatic detection of single microRNA molecules

251st National Meeting (2016)

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