This work presents analyses and developments in nanofabrication using stencil lithography and its application for electronics and biosensing. Metallic nanostructures are fabricated using stencil lithography studying the pattern transfer from the stencil to the substrate. The physical properties of the deposited nanostructures are analyzed, namely those relevant for electronics and biosensing. Applications of stenciled nanostructures for biosensing are studied based on localized surface plasmon resonance in metallic nanoparticles, and on impedance measurements on discontinuous films. The clogging of the nanoapertures in the stencil due to the accumulation of material on the membrane and its effect on the deposited structures are analyzed. A method to unclog the nanoapertures based on metal wet etching is presented, allowing the reutilization of the stencils. The blurring of the deposited structures related to the stencil-substrate gap, the divergence of the material flux and the surface mobility of the deposited adatoms is analyzed. The obtained results show that the blurring can be reduced optimizing different deposition parameters such as the thickness, temperature and deposition rate. The systematic fabrication of <100 nm metallic nanostructures using stencil lithography is demonstrated. Al and Au nanowires are deposited on SiO2/ Si substrates on a full wafer scale. The nanowires show an ohmic behavior with an electrical resistivity of ∼5 μΩcm for Au and ∼10 μΩcm for Al, higher than bulk values due to size effects on the electron mean free path. Arrays of Au nanodots are also fabricated and their localized surface plasmon spectra measured. Bulk sensitivities of the plasmons of ∼180 nmRIU-1 are obtained. The resonance wavelength also shows a shift when biotin and streptavidin bind to the Au nanodots, demonstrating the application of these structures for biosensing. Au discontinuous films are also deposited exploiting the blurring phenomena in stencil lithography. The morphology and electrical conduction of these films depends on the geometry of the stencil and other deposition parameters. Electrical measurements on these films show a reduction in impedance when biotin and streptavidin are added. The fabrication of nanostructures on polymer substrates, namely polyimide, SU-8, parylene and PDMS, using stencil lithography is also demonstrated. Au nanodots and nanowires down to 50 nm in size are fabricated. The nanodots deposited on polymers show a better resolution than those deposited on silicon oxide. The nanowires show ohmic behavior and their electrical resistivity varies for different substrates, presumably by the effect of the substrate on the grain structure of the nanowires. The Au nanodots also show localized surface plasmon resonance. An array of Au nanodots deposited on PDMS shows a resonance shift upon the addition of biomolecules, showing the potential for fabricating biosensors on flexible substrates. The results presented in this work are an important contribution for the fabrication of nanostructures with stencil lithography, uncovering the associated physical phenomena and showing the systematic fabrication of sub-100 nm structures. The characterization of the relevant physical properties of the nanostructures and the sensing measurements demonstrate the application of stencil lithography for biosensing and electronics. The work for this thesis was financially by the Swiss Federal Office for Science and Education (OFES) in the EC-funded FP6 project NaPa (NMP4-CT-2003-500120), the Swiss National Science Foundation (IC-Nano-200021-112291/1) and the EPFL-STI Seed Funding.