Dynamic detection of the bio-molecular interaction at the surface of plasmonic nanoarrays

Nanophysics and plasmonics have recently become fields of relevant interest in the world of research and, in particular, in biosensing and biochemistry. Nanoparticles of noble metals interact with incident light giving rise to the Localized Surface Plasmon Resonance (LSPR), a sharp peak of the extinction spectra of the nanoparticles as a result of the collective oscillation at a resonant frequency of the conduction electrons. The shape of the peak and its position strongly depend on both nano system properties, as composition, size, shape, orientation, and on the local dielectric environment. A change in the medium in which the nanoparticle is embedded is indeed detected and transduced as a distortion and shift of the peak. This mechanism is at the basis of the biosensing application of plasmonic structures, revealing binding events of molecules to the surface or extremely small variation in concentration of substances in the proximity. For this reason, LSPR plasmonic biosensors gained great popularity in a broad range of applications, in particular as diagnostic devices able to quantitatively detect biomarker molecules. MicroRNA, among the others, are biomolecules of prominent interest associated to thumoral or other kind of diseases. The aim of this project is to realize and test a sensitive, specific and label-free plasmonic nanobiosensor able to detect microRNA target molecules and to investigate the dynamics of the binding of the biomolecules on the surface of the optical transducers. To accomplish this task, Au nanoprisms arrays (NPA) are chosen as reference structure, with a LSPR wavelength around 800 nm and nanofabricated via NanoSphere Lithography (NSL) and thermal evaporation deposition. All the samples are morphologically characterized with AFM or SEM microscopy. Post-treating procedure and functionalization protocols are employed to allow the binding of the analyte molecule to be detected to the sensor, and all the functionalization signals are detected by linear optical spectroscopy in the visible or near-infrared spectral range. Static measurements are performed to control the peak shift of the sample after each functionalization step, and dynamic measurements in a microfluidic setup allow to monitor the temporal evolution of the optical signal and to reconstruct in real-time the hybridization kinetics at the surface of the plasmonic sensor. A 217nm/RIU bulk sensitivity and 50fMoles limit of detection is reached with the employed structures, indicating that both the nanofabrication and functionalization strategy are successful in the detection of analyte molecules down to low concentration limits. Of course, optimization is desirable, to push even further the sensitivity and solve challenges as for example the aspecific target binding on the sensor surface. Another purpose of the work is to extract interesting information about the dynamics of the hybridization reaction that takes place when the analyte microRNA is bound to the surface of the nanoarray. Hybridization kinetics is studied, determining the time and affinity constants characterizing the reaction. The results obtained will prove the non- ideal behaviour of the association, laying the basis for future and advanced outlook about the building of a non-Langmuir association model able to analytically describe the bi-molecular binding system.