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An effective and easy to implement ligand exchange strategy is paramount to the design of stable and multifunctional gold and other inorganic nanocolloids. This is also crucial for their use in bio...

Reacting poly(maleic anhydride)-based polymers with H2N–R nucleophiles is a flexible and highly effective approach for preparing a variety of multifunctional, multicoordinating, and multireactive polymers. The exact transformation of the anhydride ring during this addition reaction is still an open question. In this report, we characterize the transformation of a representative block copolymer, poly(isobutylene-alt-maleic anhydride), with a few H2N–R nucleophiles. In particular, we test the effects of varying a few reaction parameters/conditions (e.g., temperature, solvent, reaction time, and addition of thionyl chloride) on the nature of the anhydride transformation and bond formed between the polymer and the lateral R groups. The resulting polymers are characterized using a combination of analytical techniques including FT-IR, one- and two-dimensional NMR, and gel electrophoresis. We find that the ring opening transformation occurs under mild conditions. Conversely, cyclic imide transformation can take ...

A terminally protected self-assembling pentapeptide Boc-Leu(1)-Val(2)-Phe(3)-Phe(4)-Ala(5)-OMe 1 bearing sequence similarity with Aβ17–21 (the fragment 17–21 of the amyloid β-peptide Aβ42) forms thermoreversible transparent gels in various organic solvents including benzene, toluene, m-xylene and 1,2-dichlorobenzene. A series of its variants have been synthesized in order to study the role of adjacently located phenylalanine residues and the protecting groups for gelation in different organic solvents. Replacement of any of the Phe residues of the Phe-Phe segment with any other hydrophobic α-amino acid residue drastically changes the gel forming properties indicating that both Phe residues have an important role in gel formation. These gels are characterised using field emission scaning electron microscopy (FE-SEM), transmission electron microscopy (TEM), FT-IR and wide angle X-ray scattering (WAXS) studies. WAXS studies of the peptide 1–benzene gel indicate that π–π interaction is responsible for gel formation and it reveals the necessity of the Phe residues in gel formation. Transmission electron microscopy (TEM) of the gels reveals a nanofibrillar morphology, which is obtained from the self-assembled gelators in the gel phase. These gels bind with a physiological dye, Congo red, and show a green birefringence under cross polarizers, which is a characteristic feature of amyloid fibrils.

Hydrodynamic size is a characteristic dimension that reflects the Brownian diffusion of objects, such as proteins, macromolecules, and various colloids when dissolved/dispersed in fluid phases. This property is crucial when investigating the utility of colloidal nanocrystals and polymeric materials in biology. Dynamic light scattering (DLS) has been widely used to measure the diffusion coefficient and hydrodynamic size of such systems. Comparatively, diffusion-ordered NMR spectroscopy (DOSY-NMR) is a relatively new analytical method that has provided researchers with an alternative experimental approach to access such information. Here, we apply DLS and DOSY-NMR simultaneously to characterize the diffusion coefficient and hydrodynamic size of several sets of nanocolloids, including dispersions of gold nanoparticles and luminescent quantum dots that are surface-capped with either hydrophobic or hydrophilic coatings, as well as a monomer and a low-molecular-weight polymer. We compare, side by side, the findings acquired from each measurement, which has allowed us to identify the benefits and constraints of each technique. Our results show that the two approaches provide comparable data when larger size nanocolloids are probed. However, we find that DOSY is substantially more effective in characterizing nanocolloids that are fluorescent and/or have very small dimensions, as well as molecular-scale organic ligands, where DLS reaches its limit. Additionally, we find that, compared to DLS, DOSY tends to require higher solute concentrations and longer collection time to generate data with high signal-to-noise ratios.

We report on the ability of a chemically synthesized anticancer peptide, SVS-1, to promote the rapid uptake of gold nanorods (AuNRs) and gold nanoparticles (AuNPs) by live HeLa cells. For this, AuNPs and AuNRs, surface ligated with a multicoordinating polymer that presents several amine groups per ligand, are simultaneously reacted with SVS-1 and Texas-Red dye; the latter allows fluorescence visualization of the nanocrystals. Using epifluorescence microscopy, we find that incubation of the SVS-1-conjugated AuNPs and AuNRs with a model cancer cell line yields extended staining throughout the cell cytoplasm, even at low conjugate concentrations (∼0.1 nM). Furthermore, uptake is specific to the SVS-1-conjugated nanocrystals. Additional endocytosis inhibition experiments, where cells have been incubated with the conjugates at 4 °C or in the presence of endocytic inhibitors, show that significant levels of conjugate uptake persist. These results combined indicate an uptake mechanism that does not necessarily rely on endocytosis, a promising finding with implications for the use of nanomaterials in the field of biology and nanomedicine.

We detail the design of a new set of multicoordinating polymer ligands based on the phosphonate anchoring motif and apply them for the surface coating of luminescent quantum dots, gold nanoparticles, and iron oxide nanoparticles alike. The ligand is synthesized via a nucleophilic addition reaction between poly(isobutylene-alt-maleic anhydride) and amine-modified phosphonate derivatives and short polyethylene glycol hydrophilic blocks, which allows the flexibility to tune the architecture and stoichiometry of the final compound. We find that these phosphonate-based polymers exhibit a strong coordinating affinity for ZnS-overcoated quantum dots (QDs), Au nanoparticles, and iron oxide nanoparticles, yielding nanocrystal dispersions that exhibit good colloidal stability for all three systems. The affinity of these ligands is also preserved when additional coordinating groups are introduced, such as imidazoles. Furthermore, the resulting polymer-coated nanocrystals are easily functionalized with reactive group...

Colloidal quantum dots, made of semiconductor cores and surface coated with an organic shell, have generated much interest in areas ranging from spectroscopy to charge and energy transfer interactions to device design, and as probes in biology. Despite the remarkable progress in the growth of these materials, rather limited information about the molecular arrangements of the organic coating is available. Here, several nuclear magnetic resonance (NMR) spectroscopic techniques have been combined to characterize the surface ligand structure(s) on biocompatible CdSe-ZnS quantum dots (QDs). These materials have been prepared via a photoinduced ligand exchange method in which the native hydrophobic coating is substituted, in situ, with a series of polyethylene glycol-modified lipoic acid-based ligands. We first combined diffusion ordered spectroscopy with heteronuclear single-quantum coherence measurements to outline the conditions under which the detected proton signals emanate from only surface-bound ligands ...

A series of allylic substituted cyclopentene monomers with increasing steric bulk are investigated by ring opening metathesis polymerization to understand the effects of these substituents on the microstructural outcomes of the resulting polypentenamers. 3-Triethylsiloxycyclopentene is discovered to polymerize to a highly regular microstructure with up to 96% trans olefins and 92% head-to-tail positional isomers. Dynamic resolution after synthetic modifications produced an enantiopure version (∼92% ee) of this monomer which subsequently produced a precision isotactic and regioregular polypentenamer for the first time. Synthetic investigations, mechanistic aspects, and basic thermal properties are discussed.

We have combined optical absorption with the Ellman's test to identify the parameters that affect the transformation of the 5-membered dithiolanes to thiols in lipoic acid (LA) and its derivatives during UV-irradiation. We found that the nature and polarity of the solvent, the structure of the ligands, acidity of the medium and oxygen can drastically affect the amount of photogenerated thiols. These findings are highly relevant to the understanding of the photochemical transformation of this biologically relevant compound, and would benefit the increasing use of LA-based ligands for the surface functionalization of various nanomaterials.

We have combined a few advanced solution phase NMR spectroscopy techniques, namely, 1H, 31P, heteronuclear single quantum coherence (HSQC), and diffusion ordered spectroscopy (DOSY), to probe the composition of the organic capping layer on colloidal CdSe–ZnS core–shell quantum dots grown via the “hot injection” route. Combining solution phase 31P and 1H NMR with DOSY, we are able to distinguish between free ligands and those coordinated on the QD surfaces. Furthermore, when those NMR measurements are complemented with matrix-assisted laser desorption ionization (MALDI) and FTIR data, we find that the organic shell of the as-prepared QDs consists of a mixture of tri-n-octylphosphine oxide (TOPO), tri-n-octylphosphine (TOP), alkyl amine, and alkyl phosphonic acid (L- and X-type ligands); the latter molecules are usually added during growth at a rather small concentration to improve the quality of the prepared nanocrystals. However, NMR data collected from QD dispersions subjected to two or three rounds of p...

Nonspecific interactions in biological media can lead to the formation of a protein corona around nanocolloids, which tends to alter their behavior and limit their effectiveness when used as probes for imaging or sensing applications. Yet, understanding the corona buildup has been challenging. We hereby investigate these interactions using luminescent quantum dots (QDs) as a model nanocolloid system, where we carefully vary the nature of the hydrophilic block in the surface coating, while maintaining the same dihydrolipoic acid (DHLA) bidentate coordinating motif. We first use agarose gel electrophoresis to track changes in the mobility shift upon exposure of the QDs to protein-rich media. We find that QDs capped with DHLA (which presents a hydrophobic alkyl chain terminated with a carboxyl group) promote corona formation, in a concentration-dependent manner. However, when a polyethylene glycol block or a zwitterion group is appended onto DHLA, it yields a coating that prevents corona buildup. Our results clearly confirm that nonspecific interactions with protein-rich media are strongly dependent on the nature of the hydrophilic motif used. Additional gel experiments using SDS-PAGE have allowed further characterization of the corona protein, and showed that mainly a soft corona forms around the DHLA-capped QDs. These findings will be highly informative when designing nanocolloids that can find potential use in biological applications.

Reacting poly(maleic anhydride)-based polymers with H

The steady progress made over the past three decades in growing a variety of inorganic nanomaterials, with discreet control over their size and photophysical properties, has been exploited to develop several imaging and sensing applications. However, full integration of these materials into biology has been hampered by the complexity of delivering them into cells. In this report, we demonstrate the effectiveness of a chemically synthesized anticancer peptide to facilitate the rapid delivery of luminescent quantum dots (QDs) into live cells. We combine fluorescence imaging microscopy, flow cytometry, and specific endocytosis inhibition experiments to probe QD-peptide conjugate uptake by different cell lines. We consistently find that a sizable fraction of the internalized conjugates does not co-localize with endosomes or the nuclei. These findings are extremely promising for the potential integration of various nanomaterials into biological systems.

Using a few solution phase NMR spectroscopy techniques, including 1H NMR and 31P NMR, we have characterized the organic shell on CdSe-ZnS core-shell quantum dots and tracked changes in its composition when the QD dispersions are subjected to varying degrees of purification. Combining solution phase NMR with diffusion ordered spectroscopy (DOSY), we were able to distinguish between freely diffusing ligands in the sample from those bound on the surfaces. Additionally, matrix assisted laser desorption ionization (MALDI) and FTIR measurements were used to provide complementary and supporting information on the organic ligand coating for these nanocrystals. We found that the organic shell is dominated by monomeric or oligomeric n-hexylphosphonic acid (HPA), along with small portion of 1-hexadecyl amine (HDA). The presence of TOP/TOPO (tri-n-octylphosphine / tri-noctylphosphine oxide) molecules is much smaller, even though large excess of TOP/TOPO were used during the QD growth. These results indicate that HPA (alkyl phosphonate) exhibits the strongest coordination affinity to ZnS-rich QD surfaces grown using the high temperature injection route.

10 p.-8 fig.-1 graph. abst., Coating inorganic nanoparticles with polyethylene glycol (PEG)-appended ligands, as means to preserve their physical characteristics and promote steric interactions with biological systems, including enhanced aqueous solubility and reduced immunogenicity, has been explored by several groups. Conversely, macromolecules present in the human serum and on the surface of cells are densely coated with hydrophilic glycans that act to reduce nonspecific interactions, while facilitating specific binding and interactions. In particular, N-linked glycans are abundant on the surface of most serum proteins and are composed of a branched architecture that is typically characterized by a significant level of molecular heterogeneity. Here we provide two distinct methodologies, covalent bioconjugation and self-assembly, to functionalize two types of Quantum Dots with a homogeneous, complex-type N-linked glycan terminated with a sialic acid moiety. A detailed physical and functional characterization of these glycan-coated nanoparticles has been performed. Our findings support the potential use of such fluorescent platforms to sense glycan-involved biological processes, such as lectin recognition and sialidase-mediated hydrolysis., Funding was provided by the National Institutes of Health (R01NS087070), National Science Foundation (NSF-CHE and#1058957 and #1508501), Fundacion Ramón Areces, and Asahi-Kasei Corp.

We introduce a new set of multicoordinating polymers as ligands that combine two distinct metal-chelating groups, lipoic acid and imidazole, for the surface functionalization of QDs. These ligands combine the benefits of thiol and imidazole coordination to reduce issues of thiol oxidation and weak binding affinity of imidazole. The ligand design relies on the introduction of controllable numbers of lipoic acid and histamine anchors, along with hydrophilic moieties and reactive functionalities, onto a poly(isobutylene-alt-maleic anhydride) chain via a one-step nucleophilic addition reaction. We further demonstrate that this design is fully compatible with a novel and mild photoligation strategy to promote the in situ ligand exchange and phase transfer of hydrophobic QDs to aqueous media under borohydride-free conditions. Ligation with these polymers provides highly fluorescent QDs that exhibit great long-term colloidal stability over a wide range of conditions, including a broad pH range (3-13), storage at nanomolar concentration, under ambient conditions, in 100% growth media, and in the presence of competing agents with strong reducing property. We further show that incorporating reactive groups in the ligands permits covalent conjugation of fluorescent dye and redox-active dopamine to the QDs, producing fluorescent platforms where emission is controlled/tuned by Förster Resonance Energy Transfer (FRET) or pH-dependent charge transfer (CT) interactions. Finally, the polymer-coated QDs have been coupled to cell-penetrating peptides to facilitate intracellular uptake, while subsequent cytotoxicity tests show no apparent decrease in cell viability.

We explored the charge transfer interactions between CdSe–ZnS core–shell quantum dots (QDs) and the redox active neurotransmitter dopamine, using covalently assembled QD–dopamine conjugates. We combined steady-state fluorescence, time-resolved fluorescence, and transient absorption bleach measurements to probe the effects of changing the QD size (thus the QD energy levels) and the conjugate valence on the rate of QD photoluminescence quenching when the pH of the medium was adjusted from acidic to alkaline. We measured substantially larger quenching efficiencies, combined with more pronounced shortening of the carrier dynamics of these assemblies for smaller size QDs and in alkaline pH. Moreover, we found that changes in the QD size alter the electron and hole relaxation of photoexcited QDs but with different extents. For instance, a pronounced change in the hole relaxation was measured in alkaline buffers. Moreover, the hole relaxation was faster for conjugates of green-emitting QDs as compared to their r...

Interfacing inorganic nanoparticles and biological systems with the aim of developing novel imaging and sensing platforms has generated great interest and much activity. However, the effectiveness of this approach hinges on the ability of the surface ligands to promote water-dispersion of the nanoparticles with long term colloidal stability in buffer media. These surface ligands protect the nanostructures from the harsh biological environment, while allowing coupling to target molecules, which can be biological in nature (e.g., proteins and peptides) or exhibit specific photo-physical characteristics (e.g., a dye or a redox-active molecule). Amphiphilic block polymers have provided researchers with versatile molecular platforms with tunable size, composition and chemical properties. Hence, several groups have developed a wide range of polymers as ligands or micelle capsules to promote the transfer of a variety of inorganic nanomaterials to buffer media (including magnetic nanoparticles and semiconductor nanocrystals) and render them biocompatible. In this review, we first summarize the established synthetic routes to grow high quality nanocrystals of semiconductors, metals and metal oxides. We then provide a critical evaluation of the recent developments in the design, optimization and use of various amphiphilic copolymers to surface functionalize the above nanocrystals, along with the strategies used to conjugate them to target biomolecules. We finally conclude by providing a summary of the most promising applications of these polymer-coated inorganic platforms in sensor design, and imaging of cells and tissues.

The understanding of nanoparticle (NP) cytotoxicity is challenging because of incomplete information about physicochemical changes particles undergo once they come into contact with biological fluids. It is therefore essential to characterize changes in NP properties to better understand their biological fate and effects in mammalian cells. In this paper, we present a study on the effect of particle surface oxidation and dissolution rates of Cu NPs. Particle dissolution, cell-associated Cu doses, and oxidative stress responses in A549 luciferase reporter cells were examined for Cu NPs modified with mercaptocarboxylic acids with different carbon chain lengths and a thiotic acid appended-PEG ligand (TA). We found that these Cu NPs released ionic species together with small particles upon oxidation and that surface chemistry influenced the morphology and dissolution rate. The dissolution rate was also shown to impact both the cellular Cu dosimetry and associated oxidative stress responses. The convergent results from dissolution and dosimetry measurements demonstrate that both intracellular and extracellular (i.e., NP uptake-independent) release of ionic species from Cu NPs greatly affect the cytotoxicity.

As control over the growth, stabilization and functionalization of inorganic nanoparticles continue to advance, interest in integrating these materials with biological systems has steadily grown in the past decade. Much attention has been directed towards identifying effective approaches to promote cytosolic internalization of the nanoparticles while avoiding endocytosis. We describe the use of NωV virus derived gamma peptide and a chemically synthesized anticancer peptide, SVS-1 peptide, as vehicles to promote the non-endocytic uptake of luminescent quantum dots (QDs) inside live cells. The gamma peptide is expressed in E. coli as a fusion protein with poly-his tagged MBP (His-MBP-γ) to allow self-assembly onto QDs via metal-histidine conjugation. Conversely, the N-terminal cysteine residue of the SVS-1 peptide is attached to the functionalized QDs via covalent coupling chemistry. Epi-fluorescence microscopy images show that the QD-conjugate staining is distributed throughout the cytoplasm of cell cultures. Additionally, the QD staining does not show co-localization with transferrin-dye-labelled endosomes or DAPI stained nuclei. The QD uptake observed in the presence of physical and pharmacological endocytosis inhibitors further suggest that a physical translocation of QDs through the cell membrane is the driving mechanism for the uptake.

Coupling of polyhistidine-appended biomolecules to inorganic nanocrystals driven by metal-affinity interactions is a greatly promising strategy to form hybrid bioconjugates. It is simple to implement and can take advantage of the fact that polyhistidine-appended proteins and peptides are routinely prepared using well established molecular engineering techniques. A few groups have shown its effectiveness for coupling proteins onto Zn- or Cd-rich semiconductor quantum dots (QDs). Expanding this conjugation scheme to other metal-rich nanoparticles (NPs) such as AuNPs would be of great interest to researchers actively seeking effective means for interfacing nanostructured materials with biology. In this report, we investigated the metal-affinity driven self-assembly between AuNPs and two engineered proteins, a His7-appended maltose binding protein (MBP-His) and a fluorescent His6-terminated mCherry protein. In particular, we investigated the influence of the capping ligand affinity to the nanoparticle surface, its density, and its lateral extension on the AuNP-protein self-assembly. Affinity gel chromatography was used to test the AuNP-MPB-His7 self-assembly, while NP-to-mCherry-His6 binding was evaluated using fluorescence measurements. We also assessed the kinetics of the self-assembly between AuNPs and proteins in solution, using time-dependent changes in the energy transfer quenching of mCherry fluorescent proteins as they immobilize onto the AuNP surface. This allowed determination of the dissociation rate constant, Kd(-1) ∼ 1-5 nM. Furthermore, a close comparison of the protein self-assembly onto AuNPs or QDs provided additional insights into which parameters control the interactions between imidazoles and metal ions in these systems.

We have prepared and characterized a new set of highly fluorescent gold nanoclusters (AuNCs) using one-step aqueous reduction of a gold precursor in the presence of bidentate ligands made of lipoic acid anchoring groups, appended with either a poly(ethylene glycol) short chain or a zwitterion group. The AuNCs fluoresce in the red to near-infrared region of the optical spectrum with emission centered at ∼750 nm and a quantum yield of ∼10-14%, and they exhibit long fluorescence lifetimes (up to ∼300 ns). Dispersions of these AuNCs exhibit great long-term colloidal stability, over a wide range of pHs (2-13) and in the presence of high electrolyte concentrations, and a strong resistance to reducing agents such as glutathione. The growth strategy further permitted the controlled, in situ functionalization of the NCs with reactive groups (e.g., carboxylic acid or amine), making these nanoclusters compatible with common and simple-to-implement coupling strategies, such as carbodiimide chemistry. These properties combined make these fluorescent NCs greatly promising for use in various imaging and sensing applications where NIR and long-lived excitations are desired.

We describe the design and synthesis of two compact multicoordinating (lipoic acid-appended) zwitterion ligands for the capping of luminescent quantum dots, QDs. This design is combined with a novel and easy to implement photoligation strategy to promote the in situ ligand exchange and transfer of the QDs to buffer media. This method involves the irradiation of the native hydrophobic nanocrystals in the presence of the ligands, which promotes in situ cap exchange and phase transfer of the QDs, eliminating the need for a chemical reduction of the dithiolane groups. Applied to the present LA-zwitterion ligands, this route has provided QDs with high photoluminescence yields and excellent colloidal stability over a broad range of conditions, including acidic and basic pH, in the presence of growth media and excess salt conditions. The small lateral extension of the capping layer allowed easy conjugation of the QDs to globular proteins expressing a terminal polyhistidine tag, where binding is promoted by metal-affinity interactions between the accessible Zn-rich surface and imidazoles in the terminal tag of the proteins. The ability to carry out conjugation in acidic as well as basic conditions opens up the possibility to use such self-assembled QD-protein conjugates in various biological applications.

We report a one-phase aqueous growth of fluorescent gold nanoclusters (AuNCs) with tunable emission in the visible spectrum, using a ligand scaffold that is made of poly(ethylene glycol) segment appended with a metal coordinating lipoic acid at one end and a functional group at the other end. This synthetic scheme exploits the ability of the UV-induced photochemical transformation of LA-based ligands to provide DHLA and other thiol byproducts that exhibit great affinity to metal nanoparticles, obviating the need for chemical reduction of the dithiolane ring using classical reducing agents. The influence of various experimental conditions, including the photoirradiation time, gold precursor-to-ligand molar ratios, time of reaction, temperature, and the medium pH, on the growth of AuNCs has been systematically investigated. The photophysical properties, size, and structural characterization were carried out using UV-vis absorption and fluorescence spectroscopy, TEM, DOSY-NMR, and X-ray photoelectron spectroscopy. The hydrodynamic size (RH) obtained by DOSY-NMR indicates that the size of these clusters follows the trend anticipated from the absorption and PL data, with RH(red)RH(yellow)RH(blue). The tunable emission and size of these gold nanoclusters combined with their high biocompatibility would make them greatly promising for potential use in imaging and sensing applications.

Ligands made of lipoic acid (LA) appended with a polyethylene glycol (PEG) chain have been used in the aqueous phase growth of luminescent gold clusters with distinct emission from yellow to near-IR, using two different routes. In the first route, the gold-ligand complex was chemically reduced using sodium borohydride in alkaline medium, which gave near- IR luminescent gold clusters with maximum emission around 745 nm. In the second method, LA-PEG ligand was photochemically modified to a mixture of thiols, oligomers and oxygenated species under UV-irradiation, which was then used as both reducing agent and stabilizing ligand. By adjusting the pH, temperature, and time of the reaction, we were able to obtain clusters with two distinct emission properties. Refluxing the gold-ligand complex in alkaline medium in the presence of excess ligand gave yellow emission within the first two hours and the emission shifted to red after overnight reaction. Mass spectrometry and chemical assay were used to understand the photo-chemical transformation of Lipoic Acid (LA). Mass spectroscopic studies showed the photo-irradiated product contains thiols, oligomers (dimers, trimers and tetramers) as well as oxygenated species. The amount of thiol formed under different conditions of irradiation was estimated using Ellman’s assay.

We describe the synthesis of two metal-coordinating ligands that present one or two lipoic acid (LA) anchors, a hydrophilic polyethylene glycol (PEG) segment and a terminal reactive group made of an azide or an aldehyde, two functionalities with great utility in bio-orthogonal coupling techniques. These ligands were introduced onto the QD surfaces using a combination of photochemical ligation and mixed cap exchange strategy, where control over the fraction of azide and aldehyde groups per nanocrystal can be easily achieved: LA-PEG-CHO, LA-PEG-N3, and bis(LA)-PEG-CHO. We then demonstrate the application of two novel bio-orthogonal coupling strategies directly on luminescent quantum dot (QD) surfaces that use click chemistry and hydrazone ligation under catalyst-free conditions. We applied the highly efficient hydrazone ligation to couple 2-hydrozinopyridine (2-HP) to aldehyde-functionalized QDs, which produces a stable hydrazone chromophore with a well-defined optical signature. This unique optical feature has enabled us to extract a measure for the ligand density on the QDs for a few distinct sizes and for different ligand architectures, namely mono-LA-PEG and bis(LA)-PEG. We found that the foot-print-area per ligand was unaffected by the nanocrystal size but strongly depended on the ligand coordination number. Additionally, we showed that when the two bio-orthogonal functionalities (aldehyde and azide) are combined on the same QD platform, the nanocrystal can be specifically reacted with two distinct targets and with great specificity. This design yields QD platforms with distinct chemoselectivities that are greatly promising for use as carriers for in vivo imaging and delivery.

We report a new strategy for the photomediated phase transfer of luminescent quantum dots, QDs, and potentially other inorganic nanocrystals, from hydrophobic to polar and hydrophilic media. In particular, we demonstrate that UV-irradiation (λ400 nm) promotes the in situ ligand exchange on hydrophobic CdSe QDs with lipoic acid (LA)-based ligands and their facile QD transfer to polar solvents and to buffer media. This convenient method obviates the need to use highly reactive agents for chemical reduction of the dithiolane groups on the ligands. It maintains the optical and spectroscopic properties of the QDs, while providing high photoluminescence yield and robust colloidal stability in various biologically relevant conditions. Furthermore, development of this technique significantly simplifies the preparation and purification of QDs with sensitive functionalities. Application of these QDs to imaging the brain of live mice provides detailed information about the brain vasculature over the period of a few hours. This straightforward approach offers exciting possibilities for expanded functional compatibilities and reaction orthogonality on the surface of inorganic nanocrystals.

We investigated the charge transfer interactions between luminescent quantum dots (QDs) and redox active dopamine. For this, we used pH-insensitive ZnS-overcoated CdSe QDs rendered water-compatible using poly (ethylene glycol)-appended dihydrolipoic acid (DHLA-PEG), where a fraction of the ligands was amine-terminated to allow for controlled coupling of dopamine-isothiocyanate onto the nanocrystal. Using this sample configuration, we probed the effects of changing the density of dopamine and the buffer pH on the fluorescence properties of these conjugates. Using steady-state and time-resolved fluorescence, we measured a pronounced pH-dependent photoluminescence (PL) quenching for all QD-dopamine assemblies. Several parameters affect the PL loss. First, the quenching efficiency strongly depends on the number of dopamines per QD-conjugate. Second, the quenching efficiency is substantially increased in alkaline buffers. Third, this pH-dependent PL loss can be completely eliminated when oxygen-depleted buffers are used, indicating that oxygen plays a crucial role in the redox activity of dopamine. We attribute these findings to charge transfer interactions between QDs and mainly two forms of dopamine: the reduced catechol and oxidized quinone. As the pH of the dispersions is changed from acidic to basic, oxygen-catalyzed transformation progressively reduces the dopamine potential for oxidation and shifts the equilibrium toward increased concentration of quinones. Thus, in a conjugate, a QD can simultaneously interact with quinones (electron acceptors) and catechols (electron donors), producing pH-dependent PL quenching combined with shortening of the exciton lifetime. This also alters the recombination kinetics of the electron and hole of photoexcited QDs. Transient absorption measurements that probed intraband transitions supported those findings where a simultaneous pronounced change in the electron and hole relaxation rates was measured when the pH was changed from acidic to alkaline.

In this report we review some of the recent progress made for enhancing the biocompatibility of luminescent quantum dots (QDs) and for developing targeted bio-inspired applications centered on live cell imaging and sensing. We start with a detailed analysis of the surface functionalization strategies developed thus far, and discuss their effectiveness for providing long term stability of the quantum dots in biological media, to changes in pH and to added electrolytes. We then discuss the available conjugation techniques to couple QDs to a variety of biological receptors and compare their effectiveness. In particular, we highlight the implementation of new strategies such as the use of copper-free cyclo-addition reaction (CLICK) chemistry and chemo-selective ligation. We then discuss the advances made for intracellular delivery where ideas such as receptor-driven endocytosis and uptake promoted by cell penetrating peptides are used. We then describe a few representative examples where QDs have been used to investigate specific cell biology processes. Such processes include binding of QDs conjugated to the nerve growth factor to membrane specific receptors and intracellular uptake, tracking of membrane protein at the single molecule level, and recognition of ligand bound QDs by T cell receptors. We conclude by discussing issues of toxicity associated with the use of QDs in biology.

We have developed a versatile strategy to prepare a series of multicoordinating and multifunctional ligands optimized for the surface-functionalization of luminescent quantum dots (QDs) and gold nanoparticles (AuNPs) alike. Our chemical design relies on the modification of l-aspartic acid precursor to controllably combine, through simple peptide coupling chemistry, one or two lipoic acid (LA) groups and poly(ethylene glycol) (PEG) moieties in the same ligand. This route has provided two sets of modular ligands: (i) bis(LA)-PEG, which presents two lipoic acids (higher coordination) appended onto a single end-functionalized PEG, and (ii) LA-(PEG)2 made of two PEG moieties (higher branching, with various end reactive groups) appended onto a single lipoic acid. These ligands are combined with a new photoligation strategy to yield hydrophilic and reactive QDs that are colloidally stable over a broad range of conditions, including storage at nanomolar concentration and under ambient conditions. AuNPs capped with these ligands exhibit excellent stability in various biological conditions and improved resistance against NaCN digestion. This route also provides compact nanocrystals with tunable surface reactivity. As such, we have covalently coupled QDs capped with bis(LA)-PEG-COOH to transferrin to facilitate intracellular uptake. We have also characterized and quantified the coupling of dye-labeled peptides to QD surfaces using fluorescence resonance energy transfer interactions in QD-peptide-dye assemblies.

We have designed, prepared, and tested a new set of multidentate catechol- and polyethylene glycol (PEG)-derivatized oligomers, OligoPEG-Dopa, as ligands that exhibit strong affinity to iron oxide nanocrystals. The ligands consist of a short poly(acrylic acid) backbone laterally appended with several catechol anchoring groups and several terminally functionalized PEG moieties to promote affinity to aqueous media and to allow further coupling to target molecules (bio and others). These multicoordinating PEGylated oligomers were prepared using a relatively simple chemical strategy based on N,N'-dicyclohexylcarbodiimide (DCC) and N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide (EDC) condensation. The ability of these catechol-functionalized oligomers to impart long-term colloidal stability to the nanoparticles is compared to other control ligands, namely, oligomers presenting several carboxyl groups and monodentate ligands presenting either one catechol or one carboxyl group. We found that the OligoPEG-Dopa ligands provide rapid ligand exchange, and the resulting nanoparticles exhibit greatly enhanced colloidal stability over a broad pH range and in the presence of excess electrolytes; stability is notably improved compared to non-catechol presenting molecular or oligomer ligands. By inserting controllable fractions of azide-terminated PEG moieties, the nanoparticles (NPs) become reactive to complementary functionalities via azide-alkyne cycloaddition (Click), which opens up the possibility of biological targeting of such stable NPs. In particular, we tested the Click coupling of azide-functionalized nanoparticles to an alkyne-modified dye. We also measured the MRI T(2) contrast of the OligoPEG-capped Fe(3)O(4) nanoparticles and applied MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay to test the potential cytotoxicity of these NPs to live cells; we found no measurable toxicity to live cells.

Three new peptide based dendrimers with different generations were synthesized, purified, and characterized. Each of these dendrimers form efficient organogels under suitable conditions and these gels were characterized by field emission scanning electron microscopy (FE-SEM), high resolution transmission electron microscopy (HR-TEM), atomic force microscopy (AFM), Fourier transformed infrared (FT-IR) spectroscopy, differential scanning calorimetry (DSC) and rheology. It has been observed that gel forming propensity increases from first to second generation dendrimer and it decreases from second to third generation. The hydrogen bonding interaction is the main driving force for the formation of aggregated structure that leads to the formation of a fibrillar network, responsible for gelation. The morphology is network type consisting of taped or twisted fibrils spanning throughout the space. DSC measurements show the thermoreversible first-order phase transition. Rheological studies indicate that flow behavior and segmental motion of these gels are different for different peptidic gels, obtained from various generations of dendritic peptides.

Here, we report two synthetic oligopeptide-based, thermoreversible, pH-sensitive hydrogels. In gel phase, these self-assembling tetrapeptides form a long interconnected nanofibrilar network structure, as is evident from various microscopic techniques, including field emission scanning electron microscopy (FE-SEM), transmission electron microscopy (TEM), and atomic force microscopy (AFM). FTIR, circular dichroism, and wide angle X-ray diffraction (WAXD) favor an antiparallel beta-sheet structure of these gelators in the gel state. Finally, these hydrogels have been utilized for entrapment and slow release of an anticancer drug doxorubicin at physiological pH, promising their future application as a drug delivery vehicle.

Two peptide based dendrimers containing l-aspartic acid as the branching unit and succinic acid/terephthalic acid as the core unit were synthesized, characterized, and studied. These dendritic peptides form gels in various organic solvents including n-hexanol, benzene, toluene, chlorobenzene, 1,2-dichlorobenzene, o-xylene, tetralin, and nitrobenzene. Gels were characterized by freeze fracture transmission electron microscopic (FF-TEM), field emission scanning electron microscopic (SEM), transmission electron microscopic (TEM), wide angle X-ray powder diffraction (WAXPD), and variable temperature FTIR (VT-FTIR) studies. The VT-FTIR study indicates that amides and ester groups are involved in intermolecular hydrogen bonding in the gel state. Two transitions have been observed for both the dendrimer gels upon heating: the first one corresponds to the gel to sol transition and corresponds to the breaking of hydrogen bonds between esters and amides; the second one corresponds to the breaking of hydrogen bonds between amides. In the case of dendrimer 1 structural reorganization occurs in the sol state after the first transition, which is absent in the dendrimer 2 in the sol state. FF-TEM observations showed that both dendritic peptides form a platelet structure in gel state. SEM images of these dried gels indicate different geometry in different solvents in their self-assembled gel state.

Luminescent quantum dots (QDs) can potentially be used for many biological experiments, provided that they are constructed in such a way as to be stable in biological matrices. Furthermore, QDs that are compact in size and easy to couple to biomolecules can be readily used for applications ranging from protein tracking to vasculature imaging. In this protocol, we describe the preparation of ligands comprising either one or two lipoic acid (LA) groups chemically linked to a zwitterion moiety. These ligands are then used to functionalize luminescent QDs via a photochemical transformation of LA. This route produces nanocrystals that are compact in size and stable over a broad range of conditions. In addition, the resulting QDs are readily self-assembled with polyhistidine-appended proteins. This mode of conjugation maintains the protein biological activity and its orientation, yielding highly promising fluorescent conjugates that can be used for imaging and sensing. The protocol in its entirety can be completed in 3 weeks.

Fabrication, characterization, and a comparative study have been performed for chitosan-based polymer electrolytes using two different dispersion media. Chitosan gel film (solid) electrolytes are fabricated using acetic acid or adipic acid as the dispersant for chitosan in combination with ionic liquid and lithium salt. This quaternary system of chitosan, acetic acid or adipic acid, 1-butyl-3-methylimadazolium tetrafluoroborate (ionic liquid), and lithium chloride is formed as an electrolyte for potential secondary energy storage applications. The ionic conductivities, thermal, structural, and morphological properties for these electrolytes are compared. The ionic conductivities for chitosan/adipic acid (CHAD) and for chitosan/acetic acid (CHAC) systems are in the range of 3.71 × 10−4−4.6 × 10−3 and 1.3 × 10−4 −3.2 × 10−3 S cm−1, respectively. The thermal stability of CHAD-based electrolytes is determined to be higher than that of CHAC-based electrolytes. Preliminary studies are performed to determine the electrochemical stability of these materials as solid film electrolytes for electrochemical supercapacitors. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015, 132, 42143.

Interactions between luminescent fluorophores and redox active molecules often involve complex charge transfer processes, and have great ramifications in biology. Dopamine is a redox active neurotransmitter involved in a range of brain activities. We used steady-state and time-resolved fluorescence along with transient absorption bleach measurements, to probe the effects of changing the QD size and valence on the rate of photoluminescence quenching in QD-dopamine conjugates, when the pH of the medium was varied. In particular, we measured substantially larger quenching efficiencies, combined with more pronounced shortening in the PL lifetime decay when smaller size QDs and/or alkaline pH were used. Moreover, we found that changes in the nanocrystal size alter both the electron and hole relaxation of photoexcited QDs but with very different extents. For instance, a more pronounced change in the hole relaxation was recorded in alkaline buffers and for green-emitting QDs compared to their red-emitting counterparts. We attributed these results to the more favorable electron transfer pathway from the reduced form of the complex to the valence band of the QD. This process benefits from the combination of lower oxidation potential and larger energy mismatch in alkaline buffers and for green-emitting QDs. In comparison, the effects on the rate of electron transfer from excited QDs to dopamine are less affected by QD size. These findings provide new insights into the mechanisms that drive charge transfer interactions and the ensuing quenching of QD emission in such assemblies.

We have designed a new set of coordinating ligands made of a lipoic acid (LA) anchor and poly(ethylene glycol) (PEG) hydrophilic moiety appended with a terminal aldehyde for the surface functionalization of QDs. This ligand design was combined with a recently developed photoligation strategy to prepare hydrophilic CdSe-ZnS QDs with good control over the fraction of intact aldehyde (-CHO) groups per nanocrystal. We further applied the efficient hydrazone ligation to react aldehyde-QDs with 2-hydrazinopyridine (2-HP). This covalent modification produces QD-conjugates with a well-defined absorption feature at 350 nm ascribed to the hydrazone chromophore. We exploited this unique optical signature to accurately measure the number of aldehyde groups per QD when the fraction of LA-PEG-CHO per nanocrystal was varied. This allowed us to extract an estimate for the number of LA-PEG ligands per QD. These results suggest that hydrazone ligation has the potential to provide a simple and general analytical method to estimate the number of surface ligands for a variety of nanocrystals such as metal, metal oxide and semiconductor nanocrystals.

We have recently reported that photoinduced ligation of ZnS-overcoated quantum dots (QDs) offers a promising strategy to promote the phase transfer of these materials to polar and aqueous media using multidentate lipoic acid (LA)-modified ligands. In this study we investigate the importance of the underlying parameters that control this process, in particular, whether or not photoexcited QDs play a direct role in the photoinduced ligation. We find that irradiation of the ligand alone prior to mixing with hydrophobic QDs is sufficient to promote ligand exchange. Furthermore, photoligation onto QDs can also be carried out simply by using sunlight. Combining the use of Ellman's test with matrix-assisted laser desorption/ionization and electrospray ionization mass spectrometry, we probe the nature of the photochemical transformation of the ligands. We find that irradiation (using either a UV photoreactor or sunlight) alters the nature of the disulfide groups in the lipoic acid, yielding a different product mixture than what is observed for chemically reduced ligands. Irradiation of the ligand in solution generates a mixture of monomeric and oligomeric compounds. Ligation onto the QDs selectively favors oligomers, presumably due to their higher coordination onto the metal-rich QD surfaces. We also show that photoligation using mixed ligands allows the preparation of reactive nanocrystals. The resulting QDs are coupled to proteins and peptides and tested for cellular staining. This optically controlled ligation of QDs combined with the availability of a variety of multidentate and multifunctional LA-modified ligands open new opportunities for developing fluorescent platforms with great promises for use in imaging and sensor design.

We describe the design and synthesis of a series of compact ligands made of lipoic acid (LA)-based coordinating anchors and hydrophilic zwitterion groups. This ligand design is combined with a novel photoligation strategy to promote the transfer of QDs to polar and buffer media. This approach has provided hydrophilic QDs that exhibit great colloidal stability over a broad range of pHs and in the presence of cell culture media. Our photoligation strategy drastically improves previous phase transfer methods by eliminating the need for chemical reduction of the dithiolane ring using NaBH4 prior to the cap exchange, and it is adapted to several LA-based ligands. We also found that QDs stabilized with these compact zwitterionic ligands are fully compatible with metal-histidine-driven self-assembly where the protein activity is maintained after forming conjugation with the QDs.

We have designed a set of multifunctional and multicoordinating polymer ligands that are optimally suited for surface functionalizing iron oxide and potentially other magnetic nanoparticles (NPs) and promoting their integration into biological systems. The amphiphilic polymers are prepared by coupling (via nucleophilic addition) several amine-terminated dopamine anchoring groups, poly(ethylene glycol) moieties, and reactive groups onto a poly(isobutylene-alt-maleic anhydride) (PIMA) chain. This design greatly benefits from the highly efficient and reagent-free one-step reaction of maleic anhydride groups with amine-containing molecules. The availability of several dopamine groups in the same ligand greatly enhances the ligand affinity, via multiple coordination, to the magnetic NPs, while the hydrophilic and reactive groups promote colloidal stability in buffer media and allow subsequent conjugation with target biomolecules. Iron oxide nanoparticles ligand exchanged with these polymer ligands have a compact hydrodynamic size and exhibit enhanced long-term colloidal stability over the pH range of 4-12 and in the presence of excess electrolytes. Nanoparticles ligated with terminally reactive polymers have been easily coupled to target dyes and tested in live cell imaging with no measurable cytotoxicity. Finally, the resulting hydrophilic nanoparticles exhibit large and size-dependent r2 relaxivity values.

We describe the design and synthesis of two metal-coordinating zwitterion ligands to promote the transfer of hydrophobic QDs to buffer media over broad range of conditions. The ligands are prepared by appending either one or two lipoic acid anchoring groups onto a zwitterion, LA-TEG200-ZW and bis(LA)- ZW. Combining these ligands with a photochemical reduction of the lipoic acid group in the presence of UV irradiation, provides an easy to implement method to transfer luminescent QDs to buffer media, while preserving their optical and spectroscopic properties intact. The resulting zwitterion-QDs have very thin capping shell, which allows their self-assembly with full size proteins via metal-to-histidine coordination. These conjugates have great potential for use in various bio-motivated applications.

We prepared a set of multi-coordinating and reactive amphiphilic polymer ligands and used them for surface-functionalizing magnetic iron oxide nanoparticles. The amphiphilic oligomers were prepared by coupling (via one step nucleophilic addition) several dopamine anchoring groups, polyethylene glycol moieties and reactive groups onto a poly(isobutylene-alt-maleic anhydride) chain. The availability of several anchoring groups in the same ligand greatly enhances the ligand affinity to the nanoparticle surfaces, via multiplecoordination, while the hydrophilic and reactive groups promote colloidal stability in buffer media and allow subsequent conjugation to target biomolecules. The hydrophilic nanoparticles capped with these polymers maintain compact size and exhibit great long term colloidal stability.

We describe the design and synthesis of a series of compact ligands made of lipoic acid (LA)-based coordinating anchors and hydrophilic zwitterion groups. This ligand design is combined with a novel photoligation strategy to promote the transfer of QDs to polar and buffer media. This approach has provided hydrophilic QDs that exhibit great colloidal stability over a broad range of pHs and in the presence of cell culture media. Our photoligation strategy drastically improves previous phase transfer methods by eliminating the need for chemical reduction of the dithiolane ring using NaBH4 prior to the cap exchange, and it is adapted to several LA-based ligands. We also found that QDs stabilized with these compact zwitterionic ligands are fully compatible with metal-histidine-driven self-assembly where the protein activity is maintained after forming conjugation with the QDs.

Hybrid nanomaterials combining inorganic nanostructures with organic molecules such as polymers and biomolecules offer flexible platforms with size- and composition-tunable properties. These materials have shown promising use in an array of applications ranging from optical devices to biologically active platforms, facilitating important functions such as sensing, imaging, and as diagnostic tools. One of the key requirements to the integration of these inorganic nanostructures into biology is the ability to effectively control and tune their interfaces with the surrounding media. Block copolymers provide versatile molecules with dimensions comparable to those of the inorganic nanostructures and the ability to chemically tune their properties and behavior (solubility and such) over multiple length scales. Amphiphilic polymers have been used by several groups as ligands or/and capsules to promote the transfer of a variety of inorganic nanomaterials (Au nanoparticles, magnetic nanocrystals, and semiconducting nanocrystals) to buffer media and to render them biocompatible. In this article, we provide an overview of the recent developments using various amphiphilic polymers to interface inorganic nanocrystals with biological media. We start with a brief description of the most common routes for growing a few representative high quality nanocrystals of semiconductors (ie, quantum dots), metal, and metal oxide materials. We then discuss the strategies to solubilize those nanocrystals in buffer media using amphiphilic polymers and provide a few representative examples where these hydrophilic platforms have been used as sensors and imaging probes. Keywords: Amphiphilic polymers; quantum dots; magnetic nanoparticles; gold nanoparticles; bioconjugation; imaging; sensing

Hydrophilic functional semiconductor nanocrystals that are also compact provide greatly promising platforms for use in bioinspired applications and are thus highly needed. To address this, we designed a set of metal coordinating ligands where we combined two lipoic acid groups, bis(LA)-ZW, (as a multicoordinating anchor) with a zwitterion group for water compatibility. We further combined this ligand design with a new photoligation strategy, which relies on optical means instead of chemical reduction of the lipoic acid, to promote the transfer of CdSe-ZnS QDs to buffer media. In particular, we found that the QDs photoligated with this zwitterion-terminated bis(lipoic) acid exhibit great colloidal stability over a wide range of pHs, to an excess of electrolytes, and in the presence of growth media and reducing agents, in addition to preserving their optical and spectroscopic properties. These QDs are also stable at nanomolar concentrations and under ambient conditions (room temperature and white light exposure), a very promising property for fluorescent labeling in biology. In addition, the compact ligands permitted metal-histidine self-assembly between QDs photoligated with bis(LA)-ZW and two different His-tagged proteins, maltose binding protein and fluorescent mCherry protein. The remarkable stability of QDs capped with these multicoordinating and compact ligands over a broad range of conditions and at very small concentrations, combined with the compatibility with metal-histidine conjugation, could be very useful for a variety of applications, ranging from protein tracking and ligand-receptor binding to intracellular sensing using energy transfer interactions.