Scientist at Delft University of Technology developed Plasmonic nanoantennas can directly enhance the fluorescence of proteins in living mammalian cells. This is important for instantaneous monitoring of the activities of living organisms, which allows us to study the processes of life itself and its diversity.
Image source: TU Delft
Genetically modified proteins have long been created that simply emit light in living cells in response to the cell’s chemical and electrical activity. For example, such proteins are synthesized in mouse brain cells. In this case, the protein’s glow could be used to monitor the mouse’s response to stimuli without the need for implants and probes in the mouse’s brain, which would be less traumatic for the animal and provide more information about its behavior. But researchers often lack the brightness of this glow and the protein’s sensitivity to electrical impulses in the nervous system, allowing for clearer and more complete visualization of the process.
In the new work, scientists show that it is possible to create unique nanoantennas whose proximity to proteins increases their fluorescence intensity by up to six times, while the cells remain viable and continue to function normally. Previously, similar nanoantennas have been used to enhance luminescence in inanimate environments, but it turns out the idea also works in organic matter.
As nanoantennas, the researchers used gold nanostars—plasmonic nanoparticles with complex geometries capable of focusing electromagnetic fields in nanoscale regions around the surface. When illuminated by laser, local surface plasmons in the particles are excited, generating a strong optical near field. If a fluorescent protein gets close enough to this structure, its interaction with light changes: excitation efficiency and photon emission rate increase.
The key engineering challenge is to select the shape and size of the particles, ensure their chemical stability and achieve the correct positioning relative to membrane proteins. It is the controlled nanocoupling of antennas with fluorophores that makes it possible to transfer the well-known effect of enhancing single-molecule fluorescence into living cells without causing harm to them.
The most unexpected result, however, is not just an increase in brightness. The QuasAr6a protein now responds about ten times faster to changes in membrane voltage. The researchers attribute this to the fact that the plasmonic structure affects not only the intensity of the light emission, but also the photochemical cycle of the protein itself. The researchers believe that the enhanced optical near field changes the rate of transitions between protein states, while increasing the rate of photon emission and accelerating its dynamics.
The latter is important for recording fast signals at synapses, for which transgenic proteins do not have sufficient temporal resolution. All previous efforts to modify the genes of photoproteins have failed to improve their brightness and resolution, something that nanoantennas can easily handle. Engineers beat geneticists. But the benefits are huge – highly sensitive protein-nanantenna complexes will make it possible to monitor the activity of mammalian (mouse) brains and even organs deep within rodent tissue in greater detail. This was previously difficult to achieve, but with the new nanoantennas, this will become a feasible task.
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