Zero-mode waveguide nanophotonic structures for single molecule characterization

Abstract

Single-molecule characterization has become a crucial research tool in the chemical and life sciences, but limitations, such as limited concentration range, inability to control molecular distributions in space, and intrinsic phenomena, such as photobleaching, present significant challenges. Recent developments in non-classical optics and nanophotonics offer promising routes to mitigating these restrictions, such that even low affinity ( K D ~ mM) biomolecular interactions can be studied. Here we introduce and review specific nanophotonic devices used to support single molecule studies. Optical nanostructures, such as zero-mode waveguides (ZMWs), are usually fabricated in thin gold or aluminum films and serve to confine the observation volume of optical microspectroscopy to attoliter to zeptoliter volumes. These simple nanostructures allow individual molecules to be isolated for optical and electrochemical analysis, even when the molecules of interest are present at high concentration (μM - mM) in bulk solution. Arrays of ZMWs may be combined with optical probes such as single molecule fluorescence, single molecule fluorescence resonance energy transfer (smFRET), and fluorescence correlation spectroscopy (FCS) for distributed analysis of large numbers of single-molecule reactions or binding events in parallel. Furthermore, ZMWs may be used as multifunctional devices, for example by combining optical and electrochemical functions in a single discrete architecture to achieve electrochemical ZMWs (E-ZMW). In this review, we will describe the optical properties, fabrication, and applications of ZMWs for single-molecule studies, as well as the integration of ZMWs into systems for chemical and biochemical analysis.

Mixed metal zero-mode guides (ZMWs) for tunable fluorescence enhancement

“…Thus, excitation light does not propagate through the wells but creates an evanescent wave at the entrance of the wells that limits the excitation volume to regions just inside the aperture. 33,34 The photophysical properties of uorophores isolated in ZMWs can also be altered through interactions with the metal structure. These interactions can affect both excitation and emission processes.…”

Gold Ion Beam Milled Gold Zero-Mode Waveguides

“…An example of a gold-ion-milled ZMW is shown in Figure 1 . The holes are not perfectly circular as has been observed by others using FIB methods [ 72 ] and lithographic methods [ 71 ]. Improved circularity may be achieved by astigmatic adjustment of the focused ion beam.…”

“…A variety of ZMW fabrication techniques have been reported, including deep UV photolithography [ 69 ], electron beam lithography [ 14 , 18 , 26 , 69 ], colloidal lithography [ 70 ], and focused ion beam (FIB) milling with gallium ions [ 25 , 29 , 71 , 72 ]. Each of these techniques has advantages and disadvantages.…”

Nanopore-Templated Silver Nanoparticle Arrays Photopolymerized in Zero-Mode Waveguides

“…The spatial distribution of the optical field, both within and external to the ZMWs, is important in determining the extent of photopolymerization that will occur within the nanopores. For wavelengths above the cutoff wavelength of the nanoaperture, λ c , where λ c ~ 1.7 d and d is the pore diameter, the evanescent field decays exponentially with distance at a rate that depends on the radius and diameter (Crouch et al, 2018). In addition, the light-blocking efficiency of the Ag optical cladding layer also depends on the thickness of Ag.…”