Abstract
We propose two-focus fluorescence correlation spectroscopy (2fFCS) on basis of plasmonic nanoantennas that provide distinct hot spots that are individually addressable through polarization, yet lie within a single diffraction limited microscope focus. The importance of two-focus FCS is that a calibrated distance between foci provides an intrinsic calibration to derive diffusion constants from measured correlation times. Through electromagnetic modelling we analyze a geometry of perpendicular nanorods, and their inverse, i.e., nanoslits. While we find that nanorods are not suited for nano-antenna enhanced 2fFCS due to substantial background signal, a nanoslit geometry is expected to provide a di tinct cross-correlation between orthogonally polarized detection channels. Furthermore, by utilizing a periodic array of nanoslits instead of a single pair, the amplitude of the cross-correlation can be enhanced. To demonstrate this technique, we present a proof of principle experiment on the basis of a periodic array of nanoslits, applied to lipid diffusion in a supported lipid bilayer.
Highlights
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We propose two-focus fluorescence correlation spectroscopy (2fFCS) on basis of plasmonic nanoantennas that provide distinct hot spots that are individually addressable through polarization, yet lie within a single diffraction limited microscope focus
Fluorescence correlation spectroscopy is a common technique to deduce the concentration and mobility of fluorescent particles. It is based on measurements of fluorescence intensity fluctuations, which occur as particles perform a random walk through a single, tight, microscope focus[1,2,3]
Summary
Before we present numerical results on nano-antenna enhanced 2fFCS, we outline the general calculation approach. For the periodic array of nano-apertures we again perform full wave simulations with FDTD using a Gaussian excitation beam corresponding to a tight focus (NA = 0.7), and we obtain fields at the optical pump wavelength and Stokes-shifted fluorescent wavelength For the array of alternating rectangular nano-apertures, the standard FCS trace (single polarization channel) and the cross-correlation of cross-polarized channels shows a large difference (Fig. 5(c)) This measurement demonstrates the feasibility of nano-antenna 2fFCS. We attribute this discrepancy to a difference in electrostatic properties between the glass reference, and the aperture array, due to surface properties of the gold film and planarization layer, and their modification by focused ion beam milling This discrepancy has no bearing on the validation of the optical mechanisms of nano-antenna enhanced 2fFCS per se, as is evidenced by the values for the geometrical fit parameters that we retrieve. Low-quantum efficiency dyes are well-known to be much more effective at singling out hot-spot properties[36], likely leading to improved polarization contrast by removing the background focus contribution
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