DNA-barcoded labeling probes for highly multiplexed Exchange-PAINT imaging† †Electronic supplementary information (ESI) available. See DOI: 10.1039/c6sc05420j Click here for additional data file.

Recent advances in super-resolution fluorescence imaging allow researchers to overcome the classical diffraction limit of light, and are already starting to make an impact in biology. However, a key challenge for traditional super-resolution methods is their limited multiplexing capability, which prevents a systematic understanding of multi-protein interactions on the nanoscale. Exchange-PAINT, a recently developed DNA-based multiplexing approach, in theory facilitates spectrally-unlimited multiplexing by sequentially imaging target molecules using orthogonal dye-labeled ‘imager’ strands. While this approach holds great promise for the bioimaging community, its widespread application has been hampered by the availability of DNA-conjugated ligands for protein labeling. Herein, we report a universal approach for the creation of DNA-barcoded labeling probes for highly multiplexed Exchange-PAINT imaging, using a variety of affinity reagents such as primary and secondary antibodies, nanobodies, and small molecule binders. Furthermore, we extend the availability of orthogonal imager strands for Exchange-PAINT to over 50 and assay their orthogonality in a novel DNA origami-based crosstalk assay. Using our optimized conjugation and labeling strategies, we demonstrate nine-color super-resolution imaging in situ in fixed cells

Quantitative Super-Resolution Imaging with qPAINT using Transient Binding Analysis

Current super-resolution techniques offer unprecedented spatial resolution, but quantitative counting of spatially unresolvable molecules remains challenging. Here, we use the programmable and specific binding of dye-labeled DNA probes to count integer numbers of targets. This method, called quantitative Points Accumulation In Nanoscale Topography (qPAINT), avoids the challenging task of analyzing the environmentally sensitive hard-to-predict photophysics of dyes, and enables robust counting by analyzing the predictable binding kinetics of dye-labeled DNA probes. We benchmarked qPAINT in vitro and in situ by counting strands on DNA nanostructures, Nup98 protein clusters in the nuclear pore complex, Bruchpilot proteins in Drosophila, and finally the number of fluorescence in situ hybridization probes on single mRNA targets in fixed cells. We achieved high accuracy (~98–99 %), high precision (~80–95 %), and multiplexed detection over a large dynamic range

Tailoring nanoparticle surface monolayers for biomolecular recognition and delivery applications

Engineering the interfaces between biomolecules and nanomaterials is central to the creation of materials for diverse areas of biomedical applications, including therapy, sensing and imaging. The goal of this research has been oriented toward the tailoring of the interfaces through the atomic level control provided by the organic synthesis. By employing a synergistic approach in the research, combining colloids, surface science, organic synthesis and biology, gold nanoparticles with tailored monolayers have been developed for bio-applications. This thesis illustrates the design and synthesis of these surface functionalized gold nanoparticles and their use in protein surface recognition and delivery systems for therapeutic applications. For protein surface recognition, we have fabricated gold nanoparticles bearing a diversity of amino acids termini and studied their interactions with proteins to elucidate the parameters affecting their interactions and catalytic behavior. In therapeutic applications, we have demonstrated the use of organically tailored nanoparticles for the creation of delivery systems featuring tunable stability and regulated drug release. Additionally, gold nanoparticles functionalized with molecular recognition motif have been used to demonstrate host-guest chemistry inside the living cells for the activation of therapeutic gold nanoparticles

Transient dormant monomer states for supramolecular polymers with low dispersity

Monodisperse and well-defined self-assembled materials can be obtained by fuel-driven temporally controlled supramolecular polymerization via the buffered release of monomers. Here the authors show that a redox-responsive transient dormant state of monomer generated by redox reaction can lead to supramolecular polymers with low dispersity

Self-Assembly and Cross-linking of FePt Nanoparticles at Planar and Colloidal Liquid−Liquid Interfaces

Terpyridine thiol functionalized FePt and Au NPs were self-assembled and cross-linked at the liquid−liquid interfaces using Fe(II) metal ion. Complexation of terpyridine with Fe(II) metal ion leads to NP network and affords stable membranes and colloidal shells at the liquid−liquid interfaces

Direct photopatterning of light-activated gold nanoparticles

Photoactivatable gold NPs were patterned via photolithography. In this approach, charge reversal of the ligands on NPs upon UV irradiation induces crosslinking to generate stable NP patterns

Nanoparticles for detection and diagnosis

Nanoparticle-based platforms for identification of chemical and biological agents offer substantial benefits to biomedical and environmental science. These platforms benefit from the availability of a wide variety of core materials as well as the unique physical and chemical properties of these nanoscale materials. This review surveys some of the emerging approaches in the field of nanoparticle based detection systems, highlighting the nanoparticle based screening methods for metal ions, proteins, nucleic acids, and biologically relevant small molecules

Solvent adaptive dynamic metal-organic soft hybrid for imaging and biological delivery

A solvent responsive dynamic nanoscale metal‐organic framework (NMOF) [Zn(1 a)(H$_2$O)$_2$] has been devised based on the self‐assembly of Zn$^{II}$ and asymmetric bola‐amphiphilic oligo‐(p‐phenyleneethynylene) (OPE) dicarboxylate linker 1 a having dodecyl and triethyleneglycolmonomethylether (TEG, polar) side chains. In THF solvent, NMOF showed nanovesicular morphology (NMOF‐1) with surface decorated dodecyl chains. In water and methanol, NMOF exhibited inverse‐nanovesicle (NMOF‐2) and nanoscroll (NMOF‐3) morphology, respectively, with surface projected TEG chains. The pre‐formed NMOFs also unveiled reversible solvent responsive transformation of different morphologies. The flexible NMOF showed cyan emission and no cytotoxicity, allowing live cell imaging. Cisplatin (14.4 wt %) and doxorubicin (4.1 wt %) were encapsulated in NMOF‐1 by non‐covalent interactions and, in vitro and in vivo drug release was studied. The drug loaded NMOFs exhibited micromolar cytotoxicity