10000-Fold Improvement In Protein Detection Using Nanostructured Porous Silicon Interferometric Aptasensors

On-field analysis (e.g. clinical and diagnostics) of nanostructured porous silicon (PSi) for label-free optical biosensing has been so far hindered by insufficient sensitivity of PSi biosensors. Here we report on a label-free PSi interferometric aptasensor able to specifically detect Tumor Necrosis Factor alpha (TNFα, a protein biomarker of inflammation and sepsis) at concentration down to 3.0 nM with signal-to-noise (S/N) ratio of 10.6 and detection limit (DL) of 200 pM. This represents a 10000-fold improvement with respect to direct label-free PSi biosensors and pushes PSi biosensors close to the most sensitive optical and label free transduction techniques, e.g. surface plasmon resonance (SPR) for which a lowest DL of 100 pM in aptasensing has been reported. A factor 1000 in improvement is achieved by introducing a novel signal processing technique for the optical readout of PSi interferometers, namely Interferogram Average over Wavelength (IAW) reflectance spectroscopy. The IAW reflectance spectroscopy is shown to significantly improve both sensitivity and reliability of PSi biosensors with respect to commonly used Fast Fourier Transform (FFT) reflectance spectroscopy. A further factor 10 is achieved by enabling preparation of PSi interferometer with enlarged pore sizes (up to a 3x increase in diameter) at constant current density (i.e. constant porosity and, in turn, constant refractive index). This method is in contrast to standard PSi preparation where pore size is increased by increasing etching current density (i.e. porosity), and allows tackling mass-limited diffusion of biomolecules into the nanopores without worsening PSi interferometer optical features.