M.Sc Thesis


M.Sc StudentNasser Maisa
SubjectEnhancing Single-Molecule Fluorescence in Nanopores via
Plasmonic Nanostructure and Time-Resolved Signals
DepartmentDepartment of Biomedical Engineering
Supervisor PROF. Amit Meller


Abstract

Solid state Nanopores (ssNPs) have emerged in recent years as a highly versatile single molecule biosensors for DNA, RNA and proteins. Their advantage lies both in their extremely high sensitivity, as well as their analyte custom fabrication. Typically, nanopore sensing involves electrical sensing of the ion-current flowing through the pore. However, recently it was shown that optical sensing in ssNPs can provide independent and complementary information to the electrical signal, hence highly extending the sensing capabilities of ssNPs. This approach has recently been proposed by our group as a new method for differentiating proteins and towards whole proteome sequencing. To realize high resolution and multi-color single molecule sensing during the fast passage of proteins through ssNPs, a number of challenges must be addressed.

 Primarily, the fluorescence excitation volume must be engineered to allow a high spatial resolution and a sufficient signal to background ratio. We here propose the use of highly optimized plasmonic nanostructure built on top of the nanopore that act to localize the electrical field. I will discuss numerical simulations of a range of constructs that can enhance multiple color fluorophores.

In a complimentary approach we evaluated the possibility to discriminate multiple fluorophore species having identical emission spectra, based on their inherent fluorescence lifetime. Here we present numerical simulations and experiments using Time Correlated Single-Photon Counting (TCSPC) of fluorophore’s mixtures aiming at evaluating the limits of sensing especially if incorporated in single molecule methods like ssNPs. Our results suggest that we can determine the ratio of the two fluorophore species with high confidence using  photons.