Lifting the Concentration Limit of Mass Photometry by PEG Nanopatterning.

Mass photometry (MP) is a rapidly growing optical technique for label-free mass measurement of single biomolecules in solution. The underlying measurement principle provides numerous advantages over ensemble-based methods but has been limited to low analyte concentrations due to the need to uniquely and accurately quantify the binding of individual molecules to the measurement surface, which results in diffraction-limited spots. Here, we combine nanoparticle lithography with surface PEGylation to substantially lower surface binding, resulting in a 2 orders of magnitude improvement in the upper concentration limit associated with mass photometry. We demonstrate the facile tunability of degree of passivation, enabling measurements at increased analyte concentrations. These advances provide access to protein-protein interactions in the high nanomolar to low micromolar range, substantially expanding the application space of mass photometry.

A Quantitative Description for Optical Mass Measurement of Single Biomolecules.

Label-free detection of single biomolecules in solution has been achieved using a variety of experimental approaches over the past decade. Yet, our understanding of the magnitude of the optical contrast and its relationship with the underlying atomic structure as well as the achievable measurement sensitivity and precision remain poorly defined. Here, we use a Fourier optics approach combined with an atomic structure-based molecular polarizability model to simulate mass photometry experiments from first principles. We find excellent agreement between several key experimentally determined parameters such as optical contrast-to-mass conversion, achievable mass accuracy, and molecular shape and orientation dependence. This allows us to determine detection sensitivity and measurement precision mostly independent of the optical detection approach chosen, resulting in a general framework for light-based single-molecule detection and quantification.

Template-directed conjugation of heterogeneous oligonucleotides to a homobifunctional molecule for programmable supramolecular assembly.

Nanoscience aspires to mimic nature's control over functional molecular assemblies. Here we present a templating technique for the efficient attachment of two different oligonucleotides to a homobifunctional molecule, enabling its controlled and programmable placement within a DNA nanostructure. We demonstrate its application to a range of organic molecules with different conjugation chemistries and water solubilities. We show that the two oligonucleotide adapters can be used to integrate a bifunctional cyanine dye into a self-assembled three-dimensional DNA origami nanostructure, giving control of both position and orientation. We also demonstrate the use of both adapters to exert dynamic control over the environment of the target molecule by means of a series of strand-displacement reactions.

Chemical ligation of an entire DNA origami nanostructure.

Within the field of DNA nanotechnology, numerous methods were developed to produce complex two- and three-dimensional DNA nanostructures for many different emerging applications. These structures typically suffer from a low tolerance against non-optimal environmental conditions including elevated temperatures. Here, we apply a chemical ligation method to covalently seal the nicks between adjacent 5' phosphorylated and 3' amine-modified strands within the DNA nanostructures. Using a cost-effective enzymatic strand modification procedure, we are able to batch-modify all DNA strands even of large DNA objects, such as origami nanostructures. The covalent strand linkage increases the temperature stability of the structures by ∼10 K. Generally, our method also allows a 'surgical' introduction of covalent strand linkages at preselected positions. It can also be used to map the strand ligation into chains throughout the whole nanostructure and identify assembly defects. We expect that our method can be applied to a large variety of DNA nanostructures, in particular when full control over the introduced covalent linkages and the absence of side adducts and DNA damages are required.

Fabrication of Metal Nanostructures with Programmable Length and Patterns Using a Modular DNA Platform.

Recently introduced DNA nanomolds allow the shape-controlled growth of metallic nanoparticles. Here we demonstrate that this approach can be used to fabricate longer linear metal nanostructures of controlled lengths and patterns. To this end, we establish a set of different interfaces that enable mold interactions with high affinity and specificity. These interfaces enable and control the modular assembly of mold monomers into larger mold superstructure with programmable dimension in which each mold monomer remains uniquely addressable. Preloading the molds with nanoparticle seeds subsequently allows the growth of linear gold nanostructures whose lengths are controlled by the DNA structure. Exploiting the addressability of individual mold monomers furthermore allows achievement of site-specific metallization, that is, to create defined metal patterns. We think that the introduced approach provides a useful basis to fabricate nanomaterials with complex shapes and material composition in a fully programmable and modular fashion.

Chiral DNA Origami Nanotubes with Well-Defined and Addressable Inside and Outside Surfaces.

We report the design and assembly of chiral DNA nanotubes with well-defined and addressable inside and outside surfaces. We demonstrate that the outside surface can be functionalised with a chiral arrangement of gold nanoparticles to create a plasmonic device and that the inside surface can be functionalised with a track for a molecular motor allowing transport of a cargo within the central cavity.

DNA-Mold Templated Assembly of Conductive Gold Nanowires.

We introduce a new concept for the solution-based fabrication of conductive gold nanowires using DNA templates. To this end, we employ DNA nanomolds, inside which electroless gold deposition is initiated by site-specific attached seeds. Using programmable interfaces, individual molds self-assemble into micrometer-long mold superstructures. During subsequent internal gold deposition, the mold walls constrain the metal growth, such that highly homogeneous nanowires with 20-30 nm diameters are obtained. Wire contacting using electron-beam lithography and electrical conductance characterization at temperatures between 4.2 K and room temperature demonstrate that metallic conducting wires were produced, although for part of the wires, the conductance is limited by boundaries between gold grains. Using different mold designs, our synthesis scheme will, in the future, allow the fabrication of complex metal structures with programmable shapes.

Shape-controlled synthesis of gold nanostructures using DNA origami molds.

We introduce a new concept that allows the synthesis of inorganic nanoparticles with programmable shape. Three-dimensional DNA origami nanostructures harboring an internal cavity are used as molds. A small gold nanoparticle within the cavity nucleates solution-based gold deposition leading to mold filling. We demonstrate the fabrication of 40 nm long rodlike gold particles with quadratic cross section and the formation of higher order assemblies of the obtained particles, which is mediated by their DNA shell.