Name: DNA Coating of Gold Nanoparticle (AuNP) for DNA Origami Nanoantenna (DONA)
Uploaded: 2023-07-21T12:40:00
Duration: 2 min 13 s
Description: Watch this Scientific Journal Video about DNA Coating of Gold Nanoparticle AuNP for DNA Origami Nanoantenna DONA at JoVE.com

The video demonstrates a method for coating gold nanoparticles with DNA. It involves centrifuging a gold nanoparticle solution, adding thiol-modified DNA and TCEP, incubating, and freezing the mixture. Color changes during the process indicate successful coating, and absorbance spectra show differences between DNA-coated and bare gold nanoparticles.

dna coating of gold nanoparticle (aunp) for dna origami nanoantenna (dona)

Single-Molecule SERS Measurements Enabled by Plasmonic DNA Origami Nanoantennas

DNA origami is a nanotechnology technique that involves folding DNA strands into specific shapes and patterns. The ability to create structures with precise control at the nanoscale is one of the key advantages of DNA origami1. The ability to manipulate matter at such a small scale has the potential to revolutionize a wide range of fields, including medicine, electronics, and materials science2. For example, DNA origami structures have been used to deliver drugs directly to cancer cells3,4, create nanoscale sensors for detecting diseases5,6, and create intricate patterns on the surfaces of materials6,7. Furthermore, the ability to use DNA origami to create complex nanoscale structures has created new opportunities for studying fundamental biological processes at the nanoscale8.
Surface enhanced Raman scattering (SERS) is a robust analytical technique that detects and identifies molecules at extremely low concentrations9. It is based on the Raman effect, which is a change in the wavelength of scattered light10. SERS requires a plasmonic metal substrate to enhance the Raman signal of molecules adsorbed onto it. This enhancement can be up to 1011 times greater than the signal obtained from the same molecule measured by conventional Raman spectroscopy, making SERS a highly sensitive method for analyzing trace amounts of substances11.
The Raman signal enhancement of nanoparticles is mostly due to electromagnetic enhancement based on the excitation of localized surface plasmon resonance (LSPR)12. In this phenomenon, electrons within the metal nanoparticle oscillate collectively around the surface of the metal nanoparticle during light incidence. This results in the creation of a standing wave of electrons known as a surface plasmon, which can resonate with the incident light. The LSPR greatly enhances the electric field near the particle&#39;s surface, and the optical absorption of the particle is maximum at the plasmon resonant frequency. The surface plasmon&#39;s energy depends on the shape and size of the metal nanoparticle, as well as the properties of the surrounding medium13. A higher enhancement could be further achieved by plasmonic couplings, such as when two nanoparticles are close to each other at a distance of approximately 2.5 times the diameter length or less14. The proximity causes the LSPR of both nanoparticles to interact with each other, enhancing the electric field in the gap between the particles by several orders of magnitude, far exceeding the enhancement of a single nanoparticle15,16,17. The magnitude of the enhancement is inversely proportional to the distance between the nanoparticles; as the distance decreases, a critical point appears where the enhancement is sufficient to detect single molecules (SMs)18.
DNA origami represents a key technology that can efficiently arrange plasmonic nanoparticles to exploit and optimize the plasmonic coupling19. At the same time, molecules of interest can be placed precisely at the place where optical detection is most efficient. This has been demonstrated with DNA origami-based plasmonic nanoantennas for fluorescence detection20. In contrast to the detection of fluorescent labels, SERS provides the possibility to detect a direct chemical fingerprint of a molecule, making the single-molecule SERS very attractive for sensing as well as monitoring chemical reactions and mechanistic studies. DNA origami can also be used as a mask to fabricate plasmonic nanostructures with well-defined shapes21, although the possibility to position target molecules precisely into the hot spot is then lost.
Using colocalized atomic force microscopy (AFM) and Raman measurements, we can obtain the Raman spectra from a single DNA origami nanoantenna (DONA) and potentially a single molecule if placed at the position of highest signal enhancement. The DONA comprises a DNA origami fork and two precisely positioned nanoparticles, fully coated with DNA complementary to extended staple strands attached to the DNA origami. Upon DNA hybridization, the nanoparticles are bound to the DNA origami fork with a 1.2-2.0 nm gap between the nanoparticle surfaces22. Such assembly creates a hot spot in between the nanoparticles with a signal enhancement of up to 1011-fold, as calculated from finite difference time domain (FDTD) simulations22, thus permitting SM SERS measurements. However, the volume of the highest enhancement is small (in the 1-10 nm3 range), and consequently the target molecules need to be positioned precisely into this hot spot. The DNA origami fork allows the positioning of a single molecule between the two nanoparticles using a DNA fork bridge and suitable coupling chemistry. Nevertheless, observing such SMs in Raman spectra is very challenging22. Alternatively, the nanoparticles can be fully coated with the target molecule, such as a TAMRA dye, to allow for single DONA measurements with a higher SERS intensity21. In this case, the TAMRA is covalently attached to the DNA coating strand (Figure 1).

Author Spotlight: Single-Molecule Surface-Enhanced Raman Scattering Measurements Enabled by Plasmonic DNA Origami Nanoantennas  

Single-Molecule Surface-Enhanced Raman Scattering Measurements Enabled by Plasmonic DNA Origami Nanoantennas

Our research aims to develop new tools to detect single molecules by surface-enhanced Raman scattering or source. This is the only technique that provides a chemical fingerprint of a molecule and is sensitive enough to detect single molecules. In this way, detailed mechanistic information about chemical reactions can be obtained.DNA origami nanostructures have been used to precisely position both plasmonic nanoparticles and target molecules. And this is required because the enhanced Raman scattering originates from a small nanometric volume between the nanoparticles which we call hotspots. And we have now created new plasmonic DNA origami nanoantenna, exactly for this purpose.The main challenge is to place target molecules in such hotspots between two nanoparticles and collect Raman data from exactly one nanoantenna structure. To collect large amounts of data and efficient correlation between atomic force microscopy an Raman spectroscopy needs to be done. The plasmonic DNA origami nanoantennas allow for a reproducible production of a large number of plasmonic dimers in which the target molecule is precisely positioned in between the nanoparticles in the hotspot.And through a correlation of the AFM and Raman data, we can now make sure that there's only a single molecule that is detected. Now we can track single molecules such as dye molecules or proteins in real time, and their behavior in the hotspots and their reactions to chemical changes in the environment. For example, the change of the spin state of single human molecules was recently monitored.In the future, we aim to monitor chemical reactions at the single molecular level and to study their reaction mechanisms. In addition, we can use this technology to detect medically-relevant biomolecules with very high sensitivity.

Watch this Scientific Journal Video about DNA Coating of Gold Nanoparticle AuNP for DNA Origami Nanoantenna DONA at JoVE.com

DNA Coating of Gold Nanoparticle (AuNP) for DNA Origami Nanoantenna (DONA)  

To coat the gold nanoparticles with DNA, take 400 microliters of commercially obtained gold nanoparticle solution and centrifuge it at 3, 500G at room temperature for five minutes. Using a pipette, remove the supernatant and resolubilize the pellet in 25 microliters of UltraPure water. Next, add four microliters of commercially obtained 100 micromolar thiol modified DNA.Add one microliter of 100 millimolar TCEP solution. Incubate the mixture for 10 minutes at room temperature. Following incubation, add five microliters of the mixture to the previously concentrated gold nanoparticle solution.Vortex the resulting mixture for five seconds and freeze for at least two hours at minus 20 degrees Celsius. After thawing the frozen mixture at room temperature, centrifuge the same at 3, 500G for five minutes at room temperature to remove excess added DNA coating. Using a pipette, remove the supernatant and resolubilize the pellet in 10 microliters of water.The changing color of the solution at each step indicated proper coating of the gold nanoparticle. The deep red color of the gold nanoparticles changed to dark purplish red as soon as the DNA was added. Freezing changed the color to purple, and thawing returned it to deep red.The absorbance spectra of the 60 nanometer DNA coated gold nanoparticles showed a shift in absorbance peak compared to that of the bare gold nanoparticles.

Research

JoVE Journal

Chemistry

Surface-enhanced Raman scattering (SERS) has the capability to detect single molecules for which high field enhancement is required. Single-molecule (SM) ...