Researchers with the U.S. Department of Energy (DOE)'s Lawrence Berkeley National Laboratory (Berkeley Lab) and the University of California (UC) Berkeley, have developed the first practical application of optical nanoantennas in cell membrane biology. A scientific team led by chemist Jay Groves has developed a technique for lacing artificial lipid membranes with billions of gold "bowtie" nanoantennas. Through the phenomenon known as "plasmonics," these nanoantennas can boost the intensity of a fluorescent or Raman optical signal from a protein passing through a plasmonic "hot-spot" tens of thousands of times without the protein ever being touched.
"Our technique is minimally invasive since enhancement of optical signals is achieved without requiring the molecules to directly interact with the nanoantenna," Groves says. "This is an important improvement over methods that rely on adsorption of molecules directly onto antennas where their structure, orientation, and behavior can all be altered."
Fluorescent emissions, in which biomolecules of interest are tagged with dyes that fluoresce when stimulated by light, and Raman spectroscopy, in which the scattering of light by molecular vibrations is used to identify and locate biomolecules, are work-horse optical imaging techniques whose value has been further enhanced by the emergence of plasmonics. In plasmonics, light waves are squeezed into areas with dimensions smaller than half-the-wavelength of the incident photons, making it possible to apply optical imaging techniques to nanoscale objects such as biomolecules. Nano-sized gold particles in the shape of triangles that are paired in a tip-to-tip formation, like a bow-tie, can serve as optical antennas, capturing and concentrating light waves into well-defined hot spots, where the plasmonic effect is greatly amplified. Although the concept is well-established, applying it to biomolecular studies has been a challenge because gold particle arrays must be fabricated with well-defined nanometer spacing, and molecules of interest must be delivered to plasmonic hot-spots (see video).
"We're able to fabricate billions of gold nanoantennas in an artificial membrane through a combination of colloid lithography and plasma processing," Groves says.
Imaging & Microscopy Issue 4 , 2012 as free epaper or pdf download
"Controlled spacing of the nanoantenna gaps is achieved by taking advantage of the fact that polystyrene particles melt together at their contact point during plasma processing. The result is well-defined spacing between each pair of gold triangles in the final array with a tip-to-tip distance between neighboring gold nanotriangles measuring in the 5-to-100 nanometer range."
Until now, Groves says, it has not been possible to decouple the size of the gold nanotriangles, which determines their surface plasmon resonance frequency, from the tip-to-tip distance between the individual nanoparticle features, which is responsible for enhancing the plasmonic effect. With their colloidal lithography approach, a self-assembling hexagonal monolayer of polymer spheres is used to shadow mask a substrate for subsequent deposition of the gold nanoparticles. When the colloidal mask is removed, what remains are large arrays of gold nanoparticles and triangles over which the artificial membrane can be formed.
"When we embed our artificial membranes with gold nanoantennas we can trace the trajectories of freely diffusing individual proteins as they sequentially pass through and are enhanced by the multiple gaps between the triangles," Groves says. "This allows us to study a realistic system, like a cell, which can involve billions of molecules, without the static entrapment of the molecules."
As molecules in living cells are generally in a state of perpetual motion, it is often their movement and interactions with other molecules rather than static positions that determine their functions within the cell. Groves says that any technique requiring direct adsorption of a molecule of interest onto a nanoantenna intrinsically removes that molecule from the functioning ensemble that is the essence of its natural behavior. The technique he and his co-authors have developed allows them to look at individual biomolecules but within the context of their surrounding community.
Theo Lohmuller, Lars Iversen, Mark Schmidt, Christopher Rhodes, Hsiung-Lin Tu, Wan-Chen Lin and Jay T. Groves: Single Molecule Tracking on Supported Membranes with Arrays of Optical Nanoantennas, NanoLetters, 2012, 12 (3), pp 1717-1721
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Keywords: Cell Membrane Fluorescence Microscopy gold nanoparticles Jay T. Groves Lawrence Berkeley National Laboratory Nanotechnology Optical Nanoantennas Plasmonics Raman spectroscopy Signal Transduction University of California