Site-Specific Integration of Hexagonal Boron Nitride Quantum Emitters on 2D DNA Origami Nanopores

Abstract

Optical emitters in hexagonal boron nitride (hBN) are promising probes for single-molecule sensing platforms. When engineered in nanoparticle form, they can be integrated as detectors in nanodevices, yet positional control at the nanoscale is lacking. Here we demonstrate the functionalization of DNA origami nanopores with optically active hBN nanoparticles (NPs) with nanometer precision. The NPs are active under three wavelengths of visible illumination and display both stable and blinking emission, enabling their accurate localization by using wide-field optical nanoscopy. Correlative opto-structural characterization reveals deterministic binding of bright, multicolor hBN NPs at the pore rim due to π-π stacking interactions at site-specific locations on the DNA origami. Our work provides a scalable, bottom-up approach toward deterministic assembly of solid-state emitters on arbitrary structural elements based on DNA origami. Such a nanoscale arrangement of optically active components can advance the development of single-molecule platforms, including optical nanopores and nanochannel sensors.

Keywords: DNA origami; hexagonal boron nitride; nanoparticles; quantum emitters.

Figure 1

hBN NP production and structural characterization. (a) Schematic of the cryogenic pretreated liquid phase exfoliation of hBN NPs. The hBN powder was immersed in liquid nitrogen for 1 h and then dispersed into room-temperature IPA/H₂O to generate cracks in the bulk material with thermal shock. The solution was ultrasonicated for 4 h to break the bulk material. To separate the NPs from the dispersions, the resultant solution was centrifuged at 6000 rpm for 30 min and filtered through a filter with a pore size of 100 nm. (b) TEM image of a representative hBN NP (scale bar = 100 nm). Inset: Electron diffraction pattern of the corresponding NP (scale bar = 1/0.03 nm^–1). (c) AFM image of the diluted hBN NPs on a mica substrate (scale bar = 500 nm). (d) Distribution of the thickness (left) and diameter (right) of the NPs acquired from 600 NPs.

Figure 2

Optical characterization of QEs in hBN NPs. (a) Widefield image of the QEs excited by three different channels (excitation wavelengths: 473, 532, and 640 nm, power = 100 mW; scale bar = 10 μm), two representative quantum emitters with different photon dynamics are circled in each channel: (b) intensity–time trace of the stable quantum emitters and (c) intensity–time trace of the blinking quantum emitters. (d) Temporal evolution of the number of localizations per frames. (e–g) Photon dynamics of the hBN QEs excited by three different channels (excitation wavelengths: 473, 532, and 640 nm). Left panel: Visualization of the QEs ON (bright color) and OFF (black) along the time series. Right panel: histogram of the “ON” frame amount, if the amount of the “ON” frame is over 80% of the whole time series (48 s), we define it as a stable QE (see Figure S2 for detailed classification of the QEs).

Figure 3

Spatial and spectral analysis for the quantum emitters in hBN NPs. (a) Widefield merged 3-channel fluorescence microscopy image of the sample (scale bar = 10 μm). (b) Top: zoomed-in image of the highlighted blue square area in (a); the smeared patterns due to the diffraction limit hinder the precise localization for the quantum emitters. Bottom: SMLM reconstructed an image of the same area in the top area with 570 frames in three different channels using the ImageJ plugin ThunderSTORM. (c) Five representative emission spectra of the quantum emitters in the same sample under 514.5 nm excitation.

Figure 4

Characterization of hybrid hBN–NPs/DNA origami nanopores. (a) Schematic illustration of the DNA origami nanopore: the orange strands indicate the position of the 19 “sticky” ssDNA composed of 30 A bases each. (b) MD simulation of the interaction between DNA bases and the hBN flakes in H₂O/IPA. Here we only show the result of adenine bases, and the IPA and H₂O molecules are removed for clarity. After 3 ns, the DNA bases are attached to the hBN surface. The color of the top view is changed for better visualization. (c) Agarose gel electrophoresis of DNA origami and DNA origami–hBN NPs complexes: 1: m13bp18 scaffold, 2: DNA origami, 3: DNA origami + hBN NPs dispersed in H₂O/IPA (IPA volume ratio = 25%), 4: DNA origami + hBN NPs dispersed in H₂O/IPA (IPA volume ratio 16.7%), 5: DNA origami + hBN NPs dispersed in H₂O/IPA (IPA volume ratio 8.3%). (d) Representative AFM measurements and height profiles along the DNA origami surface for (top) the bare DNA origami nanopore and (bottom) hBN NPs-functionalized DNA nanopore. (Scale bar: 40 nm) (e) Histogram of the thickness of the samples: the mean heights of hBN NPs, DNA origami, and the complex are 2.87, 2.48, and 6.05 nm, respectively. The inset map indicates the distribution of the hBN NPs on DNA origami.

Figure 5

Correlative opto-structural characterization of hybrid hBN QE/DNA origami nanopores. (a) Reconstructed SMLM image of hybrid hBN QE/DNA origami nanopores on the polylysine coated coverslip under 532 nm excitation, using the ImageJ plugin ThunderSTORM (scale bar = 500 nm). (b) AFM measurement of the same area (scale bar = 500 nm). (c) Correlative AFM and SMLM measurements after angle and translocation alignment (scale bar = 500 nm). (d) Representative correlative measurements of hBN QE on DNA origami nanopores and the corresponding SMLM/AFM profiles of the NP along the x and y directions.