Sensing local biomolecular environments with coherent optical nanoscopy

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The development of novel imaging tools and technologies, including super-resolution optical microscopy, has revolutionised our understanding of biological processes in living cells. Combining high resolution imaging capability with chemical sensing in living cells has the potential to unravel processes virtually impossible to follow with available techniques, such as for example defining the local environment at the specific entry site of endocytic routes of key importance for drug delivery.The main aim of this project is to demonstrate and quantify the applicability of novel optical microscopy techniques beyond state-of-the-art for local biosensing of single biomolecules directly within living cells. Specifically, the method will feature a unique combination of coherent anti-Stokes Raman scattering (CARS) [1] microscopy of endogenous biomolecules and Four-Wave Mixing (FWM) imaging of metallic nanoparticles [2], with the aim to exploit the local field enhancement effect of CARS in the vicinity of a plasmonic nanoparticle for bio-sensing at the nanoscale [3,4]. A unique home-built CARS/FWM microscope is available in Borri's laboratory [4,5], therefore the main emphasis of this project will be to push its applicability for sensing directly in living cells.[1] A Zumbusch, W Langbein, P Borri "Nonlinear vibrational microscopy applied to lipid biology" Progress in lipid research 52 (4), 615-632 (2013).[2] F. Masia, et al., Opt. Lett. 34, 1816 (2009).; ibid Phys Rev B 85, 235403 (2012).[3] H Xu et al "Electromagnetic contribution to single-molecule sensitivity in surface enhanced Raman scattering" Phys. Rev. E 62, 4318 (2000).[4] Lukas Payne, et al. "Optical micro-spectroscopy of single metallic nanoparticles: quantitative extinction and transient resonant four-wave mixing", Faraday Discussions 184, 305 (2015).[5] I Pope, et al. "Simultaneous hyperspectral differential-CARS, TPF and SHG microscopy with a single 5 fs Ti: Sa laser" Optics express 21 (6), 7096-7106 (2013).