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This review discusses the present knowledge and recent advances in the techniques used to analyze nanomaterial surfaces, including traditional techniques that are easily accessible to researchers as well as advanced techniques and recent developments. Each technique will only be discussed in the context of its utility in nanomaterial surface characterization. We highlight both advantages and drawbacks of the techniques, and review examples from literature with the focus on recent publications. The following topics will be discussed: (1) ligand structure and conformation, (2) ligand density, (3) surface charge, (4) hydrophobicity, (5) ligand shell thickness, (6) binding affinity, and (7) surface morphology. This list is by no means complete, but rather serves as a discussion platform to draw attention to this important yet challenging field of nanomaterial surface characterization. Techniques that can provide quantitative information on the nanomaterial surfaces will be discussed whenever possible. As most nanomaterials are in the form of nanoparticles, both terms will be used interchangeably throughout the text.

Water-soluble gold nanoclusters (AuNCs) are popular in biomedical applications such as bioimaging, labelling, drug delivery, and biosensing. Despite their widespread applications, the synthesis of water-soluble phosphine-capped AuNCs is not as straightforward as their organic-soluble equivalents. Organic soluble phosphine-passivated [Au(9)(L)(8)](3+) are 6-electron closed-shell AuNCs that are generally prepared via the reduction of phosphine-Au(I) complex by NaBH(4). A similar approach attempted for the water-soluble ligand triphenylphosphine monosulfonate (TPPMS) using [AuTPPMS]Cl resulted in a mixture of cluster sizes that required gel electrophoresis or fractional precipitation to isolate the Au(9) product. In this work, we report the synthesis of water-soluble [Au(9)(L)(8)](3+) nanoclusters in high yield through the biphasic ligand exchange of [Au(11)(PPh(3))(8)Cl(2)]Cl with water-soluble phosphines such as TPPMS and 4-(diphenylphosphino)benzoic acid (DPPBA). The small molecule byproducts can be completely removed by size-based separation methods, like size exclusion chromatography or dialysis, as confirmed by (31)P and (1)H nuclear magnetic resonance (NMR) as well as diffusion ordered spectroscopy (DOSY). Furthermore, [Au(9)(DPPBA)(8)]Cl(3) underwent a visible pH- and temperature-induced isomerization in ethanol between the ‘crown’ and ‘butterfly’ isomers of [Au(9)(L)(8)](3+) that has not been previously reported. Cytotoxicity evaluation of these water-soluble nanoclusters gave CC(50) of 36 μg/mL and 70 μg/mL against A549 human alveolar epithelial cells, and 30 μg/mL and 40 μg/mL against NIH/3T3 mouse fibroblast cells for [Au(9)(TPPMS)(8)]Cl(3) and [Au(9)(DPPBA)(8)]Cl(3), respectively. For comparison, auranofin, an FDA-approved gold drug, is more than an order of magnitude more toxic with a CC(50) of 7.7 μg/mL against A549 cells.

Carbohydrate-mediated interactions are involved in many cellular activities including immune responses and infections. These interactions are relatively weak, and as such, cells employ multivalency, i.e., the presentation of multiple monovalent carbohydrate ligands within a close proximity, for cooperative binding thus drastically enhanced binding affinity. In the past two decades, the field of glyconanomaterials has emerged where nanomaterials are used as multivalent scaffolds to present multiple copies of carbohydrate ligands on the nanomaterial surface. At the core of glyconanomaterial research is the ability to control and modulate multivalency through ligand display. For the quantitative evaluation of multivalency, the binding affinity must be determined. Quantification of the binding parameters provides insights for not only the fundamental glyconanomaterial – lectin interactions, but also the rational design of effective diagnostics and therapeutics. Several methods have been developed to determine the binding affinity of glyconanomaterials with lectins, including fluorescence competitive assays in solution or on microarrays, Förster resonance energy transfer, fluorescence quenching, isothermal titration calorimetry, surface plasmon resonance spectroscopy, quartz crystal microbalance and dynamic light scattering. This Feature Article discusses each of these techniques, as well as how each technique is applied to determine the binding affinity of glyconanomaterials with lectins, and the data analysis. Although the results differed depending on the specific method used, collectively, they showed that nanomaterials as multivalent scaffolds could amplify the binding affinity of carbohydrate-lectin interactions by several orders of magnitude, the extent of which depending on the structure of the carbohydrate ligand, the ligand density, the linker length and the particle size.

Liposomes, a nanoscale drug delivery system, are well known for their ability to improve pharmacokinetics and reduce drug toxicity. In this work, maltoheptaose (G7)-presenting glycoliposomes were synthesized and evaluated in the delivery of the antibiotic rifampicin. Two types of liposomes were prepared: nonfluid liposomes from l-α-phosphatidylcholine (PC) and cholesterol, and fluid liposomes from 1,2-dipalmitoyl-sn-glycero-3-phosphocholine and 1,2-dimyristoyl-sn-glycero-3-phospho-(1′-rac-glycerol). G7-derivatized glycolipid, G7-DPPE (DPPE: 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine), was incorporated into the liposomes at 21 and 14 μmol/mg to form nanoparticles of 75 ± 12 and 146 ± 14 nm for the nonfluid and fluid G7-glycoliposomes, respectively. The multivalent G7-glycoliposomes were characterized by lectin binding with concanavalin A (Con A). The dissociation constant K(d) between Con A and the nonfluid or fluid G7-glycoliposomes was 0.93 or 0.51 μM, which represented ~900- or 1600-fold stronger affinity than the binding between Con A and G7. The G7-glycoliposomes were loaded with rifampicin at 6.6 and 16 wt % encapsulation for the nonfluid and fluid G7-glycoliposomes, respectively. Introducing a carbohydrate in the liposomes slowed down the release of rifampicin, with the G7-glycoliposomes having the slowest release rate and the lowest permeability coefficient among the liposome formulations. The fluid G7-glycoliposomes lowered the minimal inhibitory concentration (MIC) of rifampicin against E. coli ORN208 by about 3 times, whereas liposomes without G7 or Man (d-mannose)-glycoliposomes showed no improvement in MIC. The rifampicin-loaded fluid G7-glycoliposomes demonstrated the best sustained antibacterial activity against E. coli, with up to 2 log reduction in the colony forming units at 4 × MIC after 24 h. Fluorescence resonance energy transfer and confocal fluorescence microscopy revealed stronger interactions of the bacterium with the fluid G7-glycoliposomes than other liposome formulations.

Bacteria of the genus Burkholderia include pathogenic Burkholderia mallei, Burkholderia pseudomallei and the Burkholderia cepacia complex (Bcc). These Gram-negative pathogens have intrinsic drug resistance, which makes treatment of infections difficult. Bcc affects individuals with cystic fibrosis (CF) and the species B. cenocepacia is associated with one of the worst clinical outcomes. Following the repurposing of auranofin as an antibacterial against Gram-positive bacteria, we previously synthetized auranofin analogs with activity against Gram-negatives. In this work, we show that two auranofin analogs, MS-40S and MS-40, have antibiotic activity against Burkholderia clinical isolates. The compounds are bactericidal against B. cenocepacia and kill stationary-phase cells and persisters without selecting for multistep resistance. Caenorhabditis elegans and Galleria mellonella tolerated high concentrations of MS-40S and MS-40, demonstrating that these compounds have low toxicity in these model organisms. In summary, we show that MS-40 and MS-40S have antimicrobial properties that warrant further investigations to determine their therapeutic potential against Burkholderia infections.

Auranofin, an FDA-approved arthritis drug, has recently been repurposed as a potential antimicrobial agent; it performed well against many Gram-positive bacteria, including multidrug resistant strains. It is, however, inactive toward Gramnegative bacteria, for which we are in dire need of new therapies. In this work, 40 auranofin analogues were synthesized by varying the structures of the thiol and phosphine ligands, and their activities were tested against ESKAPE pathogens. The study identified compounds that exhibited bacterial inhibition (MIC) and killing (MBC) activities up to 65 folds higher than that of auranofin, making them effective against Gram-negative pathogens. Both thiol and the phosphine structures influence the activities of the analogues. The trimethylphosphine and triethylphosphine ligands gave the highest activities against Gramnegative and Gram-positive bacteria, respectively. Our SAR study revealed that the thiol ligand is also very important, the structure of which can modulate the activities of the Au(I) complexes for both Gram-negative and Gram-positive bacteria. Moreover, these analogues had mammalian cell toxicities either similar to or lower than that of auranofin.

Auranofin, a gold(I)-complex with tetraacetylated thioglucose and triethylphosphine ligands, is an FDA-approved drug used as an anti-inflammatory aid in the treatment of rheumatoid arthritis. In repurposing auranofin for other diseases, it was found that the drug showed significant activity against Gram-positive bacteria but was inactive against Gram-negative bacteria. Herein, we report the design and synthesis of gold nanoclusters (AuNCs) based on the structural motif of auranofin. Phosphine-capped AuNCs were synthesized and glycosylated, yielding auranofin AuNC analogues with mixed phosphine/thioglucose ligand shells. These AuNCs were active against both Gram-negative and Gram-positive bacteria, including a panel of resistant ESKAPE pathogens. Notably, an auranofin analogue, a mixed-ligand 1.6 nm AuNC (4b) was ~4 times more active than auranofin against Pseudomonas aeruginosa, while exhibiting 24 times lower toxicity against human A549 cells. The enhanced antibacterial activity of these AuNCs was characterized by a greater uptake of Au by the bacteria compared to AuI-complexes (20% for AuNC 4b). Additional factors include increased oxidative stress, moderate inhibition of thioredoxin reductase (TrxR), and DNA damage. Most intriguingly, the AuNCs were not affected by the bacterial outer membrane (OM) barrier or by extracellular proteins. This contrasts with AuI-complexes like auranofin that are susceptible to protein binding and hindered by the OM barrier.

We report on a potential method to separate sugars by using the specific interaction between fullerenes and saccharides in liquid chromatography (LC). Aromatic rings with high electron density are believed to interact strongly with saccharides due to CH–π and/or OH–π interactions. In this study, the fullerene-bonded columns were used to separate saccharides by LC under aqueous conditions. As a result, 2-aminobenzamide-labeled glucose homopolymer (Glcs) was effectively separated by both C60 and C70 columns in the range of Glc-1 to Glc-20 and high blood glucose level being retained in greater quantity. Furthermore, similar separations were identified by LC–mass spectrometry with non-labeled glucose homopolymers. Theoretical study based on molecular dynamics and DFT calculation demonstrated that a supramolecular complex of saccharide–fullerene was formed through CH–π and/or OH–π interactions, and that the interactions between saccharide and fullerene increase with the increase units of the saccharide. Additionally, the C60 column retained disaccharides containing maltose, trehalose, and sucrose. In this case, it was assumed that the retention rates were determined by the difference of the dipole moment in each saccharide. These results suggest that the dipole-induced dipole interaction was dominant, and that maltose—with the higher dipole moment—was more strongly retained compared to other disaccharides having lower dipole moment., フラーレンを用いた糖鎖の分離手法を開発 --抗体等の糖タンパク質分離への応用に期待--. 京都大学プレスリリース. 2020-08-26.

We report a fast Staudinger reaction between perfluoroaryl azides (PFAAs) and aryl phosphines, which occurs readily under ambient conditions. A rate constant as high as 18 m−1 s−1 was obtained between methyl 4-azido-2,3,5,6-tetrafluorobenzoate and methyl 2-(diphenylphosphanyl)benzoate in CD3CN/D2O. Furthermore, the iminophosphorane product was stable toward hydrolysis and aza-phosphonium ylide reactions. This PFAA Staudinger reaction proved to be an excellent bioothorgonal reaction. PFAA-derivatized mannosamine and galactosamine were successfully transformed into cell-surface glycans and efficiently labeled with phosphine-derivatized fluorophore-conjugated bovine serum albumin.

Helicobacter pylori infection poses a worldwide public health crisis, as chronic infection is rampant and can lead to gastric ulcers, gastritis, and gastric cancer. Unfortunately, frontline therapies cause harmful side effects and are often ineffective due to antibiotic resistance. The FDA-approved drug auranofin is a gold complex with a Au(I) core coordinated with triethylphosphine and peracetylated thioglucose as the ligands. Auranofin is used for the treatment of rheumatoid arthritis and also displays potent activity against H. pylori. One of auranofin’s modes of action involves cell death by disrupting cellular thiol-redox balance maintained by thioredoxin reductase (TrxR), but this disruption leads to unwanted side effects due to mammalian cell toxicity. Here we developed and tested sugar-modified analogs of auranofin as potential antibiotics against H. pylori, with the rationale that modulating the sugar moiety would bias uptake by targeting bacterial cells and mitigating mammalian cell toxicity. Sugar-modified auranofin analogs displayed micromolar minimum inhibitory concentrations against H. pylori, maintained nanomolar inhibitory activity against the target enzyme TrxR, and caused reduced toxicity to mammalian cells. Taken together, our results suggest that structurally modifying the sugar component of auranofin has the potential to yield superior antibiotics for the treatment of H. pylori infection. Broadly, glyco-tailoring is an attractive approach for repurposing approved drugs.

Formulas derived from theoretical physics provide important insights about the nematocyst discharge process of Cnidaria (Hydra, jellyfishes, box-jellyfishes and sea-anemones). Our model description of the fastest process in living nature raises and answers questions related to the material properties of the cell- and tubule-walls of nematocysts including their polysialic acid (polySia) dependent target function. Since a number of tumor-cells, especially brain-tumor cells such as neuroblastoma tissues carry the polysaccharide chain polySia in similar concentration as fish eggs or fish skin, it makes sense to use these findings for new diagnostic and therapeutic approaches in the field of nanomedicine. Therefore, the nematocyst discharge process can be considered as a bionic blue-print for future nanomedical devices in cancer diagnostics and therapies. This approach is promising because the physical background of this process can be described in a sufficient way with formulas presented here. Additionally, we discuss biophysical and biochemical experiments which will allow us to define proper boundary conditions in order to support our theoretical model approach. PolySia glycans occur in a similar density on malignant tumor cells than on the cell surfaces of Cnidarian predators and preys. The knowledge of the polySia-dependent initiation of the nematocyst discharge process in an intact nematocyte is an essential prerequisite regarding the further development of target-directed nanomedical devices for diagnostic and therapeutic purposes. The theoretical description as well as the computationally and experimentally derived results about the biophysical and biochemical parameters can contribute to a proper design of anti-tumor drug ejecting vessels which use a stylet-tubule system. Especially, the role of nematogalectins is of interest because these bridging proteins contribute as well as special collagen fibers to the elastic band properties. The basic concepts of the nematocyst discharge process inside the tubule cell walls of nematocysts were studied in jellyfishes and in Hydra which are ideal model organisms. Hydra has already been chosen by Alan Turing in order to figure out how the chemical basis of morphogenesis can be described in a fundamental way. This encouraged us to discuss the action of nematocysts in relation to morphological aspects and material requirements. Using these insights, it is now possible to discuss natural and artificial nematocyst-like vessels with optimized properties for a diagnostic and therapeutic use, e.g., in neurooncology. We show here that crucial physical parameters such as pressure thresholds and elasticity properties during the nematocyst discharge process can be described in a consistent and satisfactory way with an impact on the construction of new nanomedical devices.

Glycopolymers have gained increasing importance in investigating glycan-lectin interactions, as drug delivery vehicles and in modulating interactions with proteins. The synthesis of these glycopolymers is still a challenging and rigorous exercise. In this regard, the highly efficient click reaction, copper (I)-catalyzed alkyne-azide cycloaddition, has been widely applied not only for its efficiency but also for its tolerance of the appended carbohydrate groups. However, a significant drawback of this method is the use of the heavy metal catalyst which is difficult to remove completely, and ultimately toxic to biological systems. In this work, we present the synthesis of carbohydrate-grafted glycopolymers utilizing a mild and catalyst-free perfluorophenyl azide (PFPA)-mediated Staudinger reaction. Using this strategy, mannose (Man) and maltoheptaose (MH) were grafted onto the biodegradable poly(lactic acid) (PLA) by stirring a PFAA-functionalized PLA with a phosphine-derivatized Man or MH in DMSO at room temperature within an hour. The glycopolymers were characterized by 1H-NMR, 19F-NMR, 31P-NMR and FTIR.

Mesoporous silica nanomaterials of different shapes (film, platelet, sphere, rod) were synthesized simply by tuning the mole ratio of dual cationic surfactant templates, cetyltrimethylammonium bromide (CTAB) and tetrabutylammonium iodine (TBAI). The film showed the most potent antibacterial activities against mycobacteria.

A tri-component reaction, involving an electrophilically-activated perfluoroaryl azide, an enolizable aldehyde and an amine, reacts readily at room temperature without any catalysts in solvents including aqueous conditions to yield a stable amidine conjugate. The versatility of this reaction is demonstrated in the conjugation of an amino acid without prior protection of the carboxyl group, and in the synthesize antibiotic-nanoparticle conjugates.

Multifunctional cellulose nanocrystals have been synthesized and applied as a new type of glyconanomaterial in lectin binding and bacterial imaging. The cellulose nanocrystals were prepared by TEMPO-mediated oxidation and acidic hydrolysis, followed by functionalization with a quinolone fluorophore and carbohydrate ligands. The cellulose nanocrystals were subsequently applied in interaction studies with carbohydrate-binding proteins and in bacterial imaging. The results show that the functional cellulose nanocrystals could selectively recognize the corresponding cognate lectins. In addition, mannosylated nanocrystals were shown to selectively interact with FimH-presenting E. coli, as detected by TEM and confocal fluorescence microscopy. These glyconanomaterials provide a new application of cellulose nanocrystals in biorecognition and imaging.

Silica and iron oxide nanoparticles with sizes ranging from 6 to 40 nm were functionalized with trehalose. The trehalose-conjugated nanoparticles showed strong interactions with Mycobacterium smegmatis (M. smegmatis) and minimal interactions with macrophage (RAW 264.7) or A549 cells. In addition, trehalose-conjugated silica nanoparticles selectively interacted with M. smegmatis on M. smegmatis-treated A549 cells, demonstrating high potential of trehalose in developing targeted therapy for treating mycobacterial infection.

Fluorescein-doped silica nanoparticles (FSNPs) functionalized with D-arabinose (Ara) showed strong interactions with Mycobacterium smegmatis (M. smegmatis) and caused the bacteria to aggregate. This aggregate formation was used as a means to detect M. smegmatis at the concentration of 10(4) CFU per mL.

X-ray Photoelectron Spectroscopy (XPS) was used to characterize the nitrogen species in perfluorophenylazide (PFPA) self-assembled monolayers. PFPA chemistry is a novel immobilization method for tailoring the surface properties of materials. It is a simple route for the efficient immobilization of graphene, proteins, carbohydrates and synthetic polymers onto a variety of surfaces. Upon light irradiation, the azido group in PFPA is converted to a highly reactive singlet nitrene species that readily undergoes CH insertion and C=C addition reactions. Here, the challenge of characterizing the PFPA modified surfaces was addressed by detailed XPS experimental analyses. The three nitrogen peaks detected in the XPS N1s spectra were assigned to amine/amide (400.5 eV) and azide (402.1 and 405.6 eV) species. The observed 2:1 ratio of the areas from the 402.1 eV to 405.6 eV peaks suggests the assignment of the peak at 402.1 eV to the two outer nitrogen atoms in the azido group and assignment of the peak at 405.6 eV to the central nitrogen atom in the azido group. The azide decomposition as the function of x-ray exposure was also determined. Finally, XPS analyses were conducted on patterned graphene to investigate the covalent bond formation between the PFPA and graphene. This study provides strong evidence for the formation of covalent bonds during the PFPA photocoupling process.

We report the synthesis of a three-dimensional graphene (3DG)–TiO2 nanocomposite by covalently attaching P25 TiO2 nanoparticles onto pristine 3DG through a perfluorophenyl azide-mediated coupling reaction. The TiO2 nanoparticles were robustly attached on the 3DG surface, with minimal particle agglomeration. In photocatalytic CO2 reduction, the 3DG–TiO2 nanocomposite demonstrated excellent activity, about 11 times higher than that of the P25 TiO2 nanoparticles. The enhanced activity can be partially attributed to the highly dispersed state of the P25 TiO2 nanoparticles on the 3DG substrate. This 3DG-based system offers a new platform for fabricating photocatalytic materials with enhanced activities.

Carbohydrate-conjugated fluorescent silica nanoprobes were prepared and used as a platform for galectin-1 and prostate cancer cell detection. The nanoparticles were efficiently conjugated using native, unprotected carbohydrate structures following a photochemical approach, resulting in lactose- and cellobiose-conjugated probes, respectively. The probes were used to challenge binding to galectin-1, an overexpressed galactose-selective lectin at prostate cancer cell surfaces, and the results show that lactose-conjugated nanoprobes exhibit differential binding behavior with prostate cancer cells and normal prostate cells. In particular, lactose-conjugated fluorescent silica nanoparticles showed specific binding to PC3 cells pre-treated with a reducing agent. The results indicate that galectin-1 expression and galectin-1-selective nanoparticles are potentially useful for sensitive and selective detection of prostate cancer.

A multivalent trehalose-grafted biodegradable poly(lactic acid) was successfully synthesized and encapsulated with iron oxide magnetic nanoparticles to give stable micelles (87.8 ± 7.4 nm). The magnetic micelles, when treated with concanavalin A, caused a rapid precipitation, indicating the multivalent presentation of trehalose ligands on the micelles. When the magnetic micelles were treated with Mycobacterium smegmatis mc2 155, orange clusters were observed resulting from the interactions between the magnetic micelles and the mycobacteria. Very little particle interaction was found on Staphylococcus epidermidis 35984 or E. coli ORN 208. With the aid of a household magnet, the orange precipitates could be easily seen by the naked eyes at 106 CFU/mL, and 104 CFU/mL under an optical microscope. The method developed represents a new approach to the detection of mycobacteria, which does not require the use of molecular biology reagents or sophisticated instruments.

Nanomaterials constitute a class of structures that have unique physiochemical properties and are excellent scaffolds for presenting carbohydrates, important biomolecules that mediate a wide variety of important biological events. The fabrication of carbohydrate-presenting nanomaterials, glyconanomaterials, is of high interest and utility, combining the features of nanoscale objects with biomolecular recognition. The structures can also produce strong multivalent effects, where the nanomaterial scaffold greatly enhances the relatively weak affinities of single carbohydrate ligands to the corresponding receptors, and effectively amplifies the carbohydrate-mediated interactions. Glyconanomaterials are thus an appealing platform for biosensing applications. In this review, we discuss the chemistry for conjugation of carbohydrates to nanomaterials, summarize strategies, and tabulate examples of applying glyconanomaterials in in vitro and in vivo sensing applications of proteins, microbes, and cells. The limitations and future perspectives of these emerging glyconanomaterials sensing systems are furthermore discussed. QC 20150715

The photoinitiated radical reactions between thiols and alkenes/alkynes (thiol-ene and thiol-yne chemistry) have been applied to a functionalization methodology to produce carbohydrate-presenting surfaces for analyses of biomolecular interactions. Polymer-coated quartz surfaces were functionalized with alkenes or alkynes in a straightforward photochemical procedure utilizing perfluorophenylazide (PFPA) chemistry. The alkene/alkyne surfaces were subsequently allowed to react with carbohydrate thiols in water under UV-irradiation. The reaction can be carried out in a drop of water directly on the surface without photoinitiator, and any disulfide side products were easily washed away after the functionalization process. The resulting carbohydrate-presenting surfaces were evaluated in real-time studies of protein–carbohydrate interactions using a quartz crystal microbalance (QCM) flow-through system with recurring injections of selected lectins, with intermediate regeneration steps using low pH buffer. The resulting methodology proved fast, efficient and scalable to high-throughput analysis formats, and the produced surfaces showed significant protein binding with expected selectivities of the lectins used in the study.

We report a new type of microarray, based on glyconanoparticles (GNPs), to study glycan-lectin interactions. GNPs, synthesized by conjugating carbohydrate ligands on silica nanoparticles, were printed on a photoactive surface followed by covalent immobilization by light activation. The GNP microarrays could be probed by lectins labeled with fluorescein as well as fluorescein-doped silica nanoparticles (FSNPs). Results showed that FSNP as the label enhanced the signals for the higher affinity ligands than the lower ones.

Poly(2-ethyl-2-oxazoline) (PEOX) of various molecular weights were covalently immobilized on silicon wafers and gold slides to form protein-resistant surfaces via a fast and general photocoupling chemistry based on the CH insertion reaction of light-activated perfluorophenyl azide (PFPA). The thicknesses of the immobilized PEOX films ranged from 23 to 80 Å for molecular weight of 5,000 to 500,000, and the grafting density reached 3.2 × 10−3 Å−2 for PEOX 5,000. The protein-resistant property of the films was studied using bovine serum albumin (BSA) by fluorescence imaging, ellipsometry, and surface plasmon resonance imaging (SPRi). The fluorescence imaging and ellipsometry studies showed the largest amount of BSA adsorbed on PEOX 5,000 and the smallest on PEOX 500,000. This was consistent with the kinetic analysis of BSA adsorption by SPRi showing that PEOX 5,000 exhibited the fastest association rate and the slowest dissociation rate whereas PEOX 500,000 had the slowest association rate and the fastest dissociation rate. The PEOX film was then applied in the fabrication of carbohydrate microarrays to reduce the non-specific adsorption of lectins and thus the background noises. Results showed that the microarray signals were significantly enhanced when the PEOX film was used.

A universal photochemical method has been established for the immobilization of intact carbohydrates and their analogues, and for the fabrication of carbohydrate microarrays. The method features the use of perfluorophenyl azide (PFPA)-modified substrates and the photochemical reaction of surface azido groups with printed carbohydrates. Various aldoses, ketoses, nonreducing sugars such as alditols, and their derivatives can be directly arrayed on the PFPA-modified chips. The lectin-recognition ability of arrayed mannose, glucose, and their oligo- and polysaccharides were confirmed using surface plasmon resonance imaging and laser-induced fluorescence imaging.

Glyconanomaterials, nanomaterials carrying multiple carbohydrate ligands, provide an excellent platform for sensitive protein recognition. Using nanomaterials as the scaffold, multivalent interactions between glycan ligands and proteins have been demonstrated. However, the quantitative analysis of the binding affinity of these glyconanomaterials has been lacking. In this Article, we report a new method to measure the binding affinity of glyconanoparticle (GNP)-protein interactions based on a fluorescent competition binding assay, which yielded the apparent dissociation constant (K(d)) of GNPs with the interacting protein. Au nanoparticles conjugated with underivatized mono-, oligo-, and polysaccharides were synthesized using our recently developed photocoupling chemistry. The affinities of these GNPs with lectins were measured and were several orders of magnitude higher than the corresponding free ligands with lectins. The effect of ligand display on the binding affinity of GNPs was, furthermore, studied where GNPs of varying linker type, spacer length, ligand density, and nanoparticle size were prepared and K(d) values determined. The long spacer linker containing hydrocarbon and ethylene oxide units gave the highest binding affinity as well as assay sensitivity. The binding affinity increased with ligand density in general, showing a drastic increase in affinity at low ligand density. In addition, the affinity enhancement was more pronounced on smaller NPs than the larger ones. These results not only demonstrate that the binding affinity of GNPs is highly influenced by how the ligands are presented on the nanoparticles but also pave the way for tailor-made glyconanomaterials with tunable affinity by way of ligand display.

Glyconanomaterials, nanomaterials carrying surface-tethered carbohydrate ligands, have emerged and demonstrated increasing potential in biomedical imaging, therapeutics, and diagnostics. These materials combine the unique properties of nanometer-scale objects with the ability to present multiple copies of carbohydrate ligands, greatly enhancing the weak affinity of individual ligands to their binding partners. Critical to the performance of glyconanomaterials is the proper display of carbohydrate ligands, taking into consideration of the coupling chemistry, the type and length of the spacer linkage, and the ligand density. This article provides an overview of the coupling chemistry for attaching carbohydrate ligands to nanomaterials, and discusses the need for thorough characterization of glyconanomaterials, especially quantitative analyses of the ligand density and binding affinities. Using glyconanoparticles synthesized by a versatile photocoupling chemistry, methods for determining the ligand density by colorimetry and the binding affinity with lectins by a fluorescence competition assay are determined. The results show that the multivalent presentation of carbohydrate ligands significantly enhances the binding affinity by several orders of magnitude in comparison to the free ligands in solution. The effect is sizeable even at low surface ligand density. The type and length of the spacer linkage also affect the binding affinity, with the longer linkage promoting the association of bound ligands with the corresponding lectins.

Methods to rapidly functionalize specific polymeric surfaces with alkynes, which can subsequently be linked to azide-containing carbohydrates, are presented. The methods are comprised of two main concepts: azide photoligation and Cu-catalyzed azide-alkyne cycloaddition. 2-Azidoethyl functionalized α-d-mannopyranoside was synthesized, and covalently attached to alkyne-functionalized polymeric surfaces using the techniques. The protein recognition properties of the carbohydrate-presenting surfaces were evaluated using quartz crystal microbalance biosensor instrumentation.

Nanomaterials, possessing unique physical and chemical properties, have attracted much interest and generated wide varieties of applications. Recent investigations of functionalized nanomaterials have expanded into the biological area, providing a versatile platform in biomedical applications such as biomolecular sensing, biological imaging, drug delivery and disease therapy. Bio-functions and bio-compatibility of nanomaterials are realized by introducing synthetic ligands or natural biomolecules onto nanomaterials, and combining ligand-receptor biological interactions with intrinsic nanomaterial properties. Common strategies of engineering nanomaterial surfaces involve physisorption or chemisorption of desired ligands. We developed a photochemically initiated surface coupling chemistry, bringing versatility and simplicity to nanomaterial functionalization. The method was applied to attach underivatized carbohydrates efficiently on gold and iron oxide nanoparticles, and the resulting glyconanoparticles were successfully used as a sensitive biosensing system probing specific interactions between carbohydrates and proteins as well as bacteria.

We present a simple and efficient method to covalently immobilize graphene on silicon wafers using perfluorophenylazide (PFPA) as the coupling agent. Graphene sheets were covalently attached to PFPA-functionalized wafer surface by a simple heat treatment under ambient conditions. The formation of single and multiple layers of graphene were confirmed by Raman spectroscopy, and optical and atomic force microscopy. Evidence of covalent bond formation between graphene and PFPA decorated silicon wafer was given by X-ray photoelectron spectroscopy and sonication treatment.

We report a simple and versatile method for grafting polymers on nanoparticles. A procedure was developed for the synthesis and subsequent functionalization of silica nanoparticles with a perfluorophenylazide. Polymers were then grafted by the photochemically induced insertion reactions of the perfluorophenylnitrene. Polystyrene, poly(4-vinylpyridine), and poly(2-ethyl-2-oxazoline) were successfully grafted on silica nanoparticles. Grafting density was studied with regard to polymer concentration and molecular weight.

Bacterial infections constitute an increasing problem to human health in response to build-up of resistance to present antibiotics and sluggish development of new pharmaceuticals. However, a means to address this problem is to pinpoint the drug delivery to - and into - the bacteria. This results in a high local concentration of the drug, circumventing the increasingly high doses otherwise necessary. Combined with other effectors, such as covalent attachment to carriers, rendering the drugs less degradable, and the combination with efflux inhibitors, old drugs can be revived. In this context, glyconanomaterials offer exceptional potential, since these materials can be tailored to accommodate different effectors. In this feature article, we describe the different advantages of glyconanomaterials, and point to their importance in antibiotic ‘revitalization’.

The use of thioglycosides and other glycan derivatives with anomeric sulfur linkages is gaining increasing interest, both in synthesis and in various biological contexts. Herein, we demonstrate the occurrence and circumvention of anomerization during 1-S-glycosylation reactions, and present highly efficient and stereocontrolled syntheses of a series of photoprobe-thiosaccharide conjugates. Mutarotation of glycosyl thiols proved to be the origin of the anomeric mixtures formed, and kinetic effects could be used to circumvent anomerization. The synthesized carbohydrate conjugates were then evaluated by both solution- and solid-phase-based techniques. Both binding results showed that the S-linked glyco-sides interact with their cognate lectins comparably to the corresponding O-analogs in the present cases, thus demonstrating the reliability of the solid-support platform built upon our photo-initiated carbohydrate immobilization method for probing protein bindings, and showing the potential of combining these two means for studying carbohydrate–protein interactions.

Engineering nanomaterials with enhanced antibacterial activities remains a critical and practical challenge. Hollow oblate mesoporous silica nanoparticles (HOMSNs) are synthesized by a simple protocol of ammonia hydrothermal treatment of oblate mesoporous silica nanoparticles prepared using dibenzyl ether as a co-solvent. When conjugate with trehalose as the targeting ligand, the antibiotic-encapsulated HOMSNs exhibit high binding affinity and antibacterial efficacy towards mycobacteria.

A stimuli-responsive drug delivery system based on fluorescent, lectin-gated, mesoporous glyconanoparticles has been developed and evaluated in normal- and cancer lung epithelial cells. The gating process proved efficient, exhibiting good sealing properties in the absence of the glutathione redox trigger, avoiding premature release in normal cells. In the presence of higher levels of glutathione in cancer cells, the lectin gate was rapidly opened and the anticancer drug released.

The reactivities of enamines and predistorted (strained) dipolarophiles toward perfluoroaryl azides (PFAAs) were explored experimentally and computationally. Kinetic analyses indicate that PFAAs undergo (3 + 2) cycloadditions with enamines up to 4 orders of magnitude faster than phenyl azide reacts with these dipolarophiles. DFT calculations were used to identify the origin of this rate acceleration. Orbital interactions between the cycloaddends are larger due to the relatively low-lying LUMO of PFAAs. The triazolines resulting from PFAA–enamine cycloadditions rearrange to amidines at room temperature, while (3 + 2) cycloadditions of enamines and phenyl azide yield stable, isolable triazolines. The 1,3-dipolar cycloadditions of norbornene and DIBAC also show increased reactivity toward PFAAs over phenyl azide but are slower than enamine–azide cycloadditions.

Nano- and molecular-scale materials and devices have demonstrated great potential for unique properties and functions that are unattainable at macroscopic scales.[1] Studies of single molecules and their interactions and reactions at the single-molecule level provide new perspectives on important fundamental issues in physics, chemistry, and biology.[2] Often it is desirable to anchor the molecules on suitable substrates to facilitate the study and application of single molecules under various conditions for physical measurement and chemical processing. Two common approaches to single-polymer immobilization are electrostatic adsorption and chemical grafting. The first method involves the deposition of a very dilute polymer solution on a substrate followed by evaporation of the solvent. A key requirement is that the polymer and the substrate possess opposite charges under the experimental conditions for the electrostatic interactions to occur.[3–5] The second method involves chemically grafting the polymer to the substrate. In this case, the polymer is derivatized with a functional group that is subsequently conjugated to the substrates. Examples include Cl-terminated poly(dimethylsiloxane) on silicon,[6] disulfide-modified polystyrene on gold,[7] and polysilanyllithium on a brominated quartz surface.[8] Relative to noncovalent and electrostatic interactions, covalent immobilization offers more stable and robust attachments that can withstand harsh experimental conditions. Herein, we report a general approach to the covalent immobilization of single polymer molecules. This method does not require chemical derivatization of the polymer and is applicable to a variety of materials, including those that do not possess reactive functional groups. The immobilization chemistry is based on the photochemically or thermally initiated C—H/N—H insertion reactions of perfluorophenylazides (PFPAs).[9–11] The presence of fluorine atoms markedly increases the insertion reaction yield relative to their non-fluorinated counterparts.[12] In the example shown in Scheme 1, a glass or silicon wafer is first treated with PFPA–silane, thus attaching azido groups to the surface. The polymer is then coated and the azido groups activated to form covalent linkages to the polymer. We found that this immobilization method is highly reproducible and tolerant of defects. Uniform polymer films were obtained from wafers treated with PFPA–silane at concentrations of a few micromoles, or when more than 100 times of a non-photoactive silane was added,[13] which is because in principle, only one attachment point is necessary to tether the entire polymer chain to the surface. We therefore hypothesize that as the density of the surface azido groups continues to decrease, the immobilized polymer will no longer be uniform. The thickness of the polymer film will decrease and eventually the surface will be populated by isolated single polymer molecules. The density of the surface azido groups can be controlled by the concentration of PFPA–silane or by treating the surface with a mixture of PFPA–silane and a non-photoactive silane.[13] Scheme 1 Covalent immobilization of polymers on silicon oxide using PFPA–silane as the photo-cross-linker. Herein, we report the covalent immobilization of single polymer molecules using dilute PFPA–silane solutions. Comprehensive studies were conducted on polystyrene, a polymer that does not possess any functional groups. In the experiment, cleaned silicon wafers were treated with solutions of PFPA–silane in toluene, the concentrations of which ranged from 5 × 10−1 to 5 × 10−5 mg mL−1. The wafers were then spin-coated with polystyrene. Irradiation followed by removal of the unbound polymer with toluene yielded covalently attached polymers. The thickness and water-contact angles of the resulting films are shown in Figure 1. Figure 1 a) Thickness, measured by ellipsometry, and b) water contact angle of the immobilized polymer as a function of PFPA–silane concentration. The contact angle of the unfunctionalized silicon wafer was measured to be approximately 17°. As the concentration of PFPA–silane decreased, the immobilized polymer became thinner (Figure 1 a). At the PFPA–silane concentration of 5 × 10−3 mg mL−1, the contact angle decreased to approximately 57°, thus indicating the collapse of the film (Figure 1 b). Further dilution resulted in lower contact angles and no film was detected by ellipsom-etry. When the immobilization was carried out on wafers treated with PFPA–silane at 5 × 10−5 mg mL−1, isolated single polystyrene molecules were observed (Figure 2 a). Figure 2 Single polystyrene molecules covalently immobilized on silicon wafers. Monodisperse polystyrene used: a) Mw = 223 200, b) 570000, and c) 1 877000 g mol−1. The wafers were treated with PFPA–silane at concentrations of a) 5×10−5 ... To confirm that the particles were indeed polymer single molecules, polystyrene of various molecular weights was used. The size of a polymer is directly related to its molecular weight: the higher the molecular weight, the larger the radius of gyration (Rg) and thus the size of the molecule. Indeed, we observed that the size of the particles, that is, the immobilized single polymer molecules, increased with the molecular weight of the polymer (Figure 2). We found that the higher the molecular weight, the lower the concentration of PFPA–silane was needed to obtain single-molecule immobilization (see Figure 2). This finding is consistent with the immobilization chemistry shown in Scheme 1. As the molecular weight and thus size of the polymer increases, less surface azido groups are needed to attach the polymer. The concentration of PFPA– silane was therefore lowered to observe isolated single molecules. Atomic force microscopy (AFM) revealed that the immobilized polystyrene adopted a cone shape (Figure 3). The volume of a single polystyrene molecule V can thus be calculated according to Equation (1), where H is the height and D is the diameter of the molecule measured by AFM. Results showed that the measured values (V) are in good agreement with those calculated from the molecular weights of the polymer (Vcalcd; Table 1).[4] Figure 3 AFM topographic image enlarged from Figure 2 a. Table 1 Measured and calculated sizes of single molecules. V=1/3Hπ(D2)2 (1) The versatility of the method was tested with poly(2-ethyl-oxazoline) (PEOX), a hydrophilic polymer that also lacks reactive functional groups. Following a similar procedure, single molecules of PEOX were successfully immobilized (Figure 4). A much higher concentration of PFPA–silane was needed for PEOX (5 × 10−1 mg mL−1) than for polystyrene (10−5 mg mL−1, Mw = 570 000). This difference can be attributed to the difference in the immobilization efficiency of the two polymers, which is in part governed by the local concentration of the polymer in proximity to the azide-functionalized surface.[14] Evaluation of 62 particles from the AFM image yielded the average particle volume of 1764 nm3 (D = (53 ± 8) nm, H = (2.4 ± 1.1) nm), whereas the calculated molecular volume is 830 nm3 based on the average molecular weight of the sample (Mw = 500 000). Unlike polystyrene samples, which are monodisperse, the PEOX obtained from Aldrich is polydisperse (molecular weight distribution is unknown); a polydisperse polymer is more heterogeneous than its monodisperse counterpart with respect to molecular weight. When a polymer containing various sized molecules is coated on a surface with a limited amount of azido groups, larger molecules, that is, polymers of higher molecule weight, are more likely than smaller molecules to be attached. The molecular volume of the immobilized polymer is thus larger than the theoretical value that is calculated from the average molecular weight of the original sample. This preferential immobilization of larger sized polymers was also observed for polystyrene (Table 1), although the deviations were smaller than PEOX as a result of the monodisperse nature of the polystyrene samples. Figure 4 AFM topographic image of immobilized PEOX (Mw = 500000 g mol−1) on a silicon wafer treated with 5×10−1 mg mL−1 of PFPA–silane in toluene. The scan area is 3×3 μm2. Extended single polymer chains were occasionally observed as shown in Figure 5. The curvilinear length was measured to be 430 nm. This value falls in the range of 95– 953 nm, as calculated from the molecular-weight distribution of the polystyrene sample.[15–16] Figure 5 An extended polystyrene molecule (Mw = 223 200 g mol−1). In summary, a simple and general method has been developed for the covalent immobilization of single polymer molecules on silicon wafers. Because the immobilization is based on C—H/N—H insertion reactions, it is versatile and especially suited for molecules that do not possess functional groups and are difficult to be immobilized by other means. Single-polymer immobilization was also achieved by treating the wafer surface with a mixture of PFPA–silane and a non-photoactive silane. Studies are underway to investigate the nature of the attachment, including the polymer attachment point and orientation.

Carbohydrate-functionalized single-walled carbon nanotubes (SWNTs) were synthesized using microwave-assisted reaction of perfluorophenyl azide with the nanotubes. The results showed that microwave radiation provides a rapid and effective means to covalently attach carbohydrates to SWNTs, producing carbohydrate-SWNT conjugates for biorecognition. The carbohydrate-functionalized SWNTs were furthermore shown to interact specifically with cognate carbohydrate-specific proteins (lectins), resulting in predicted recognition patterns. The carbohydrate-presenting SWNTs constitute a new platform for sensitive protein- or cell recognition, which pave the way for glycoconjugated carbon nanomaterials in biorecognition applications.

Carbohydrates constitute the most abundant organic matter in nature, serving as structural components and energy sources, and mediating a wide range of cellular activities. The emergence of nanomaterials with distinct optical, magnetic, and electronic properties has witnessed a rapid adoption of these materials for biomedical research and applications. Nanomaterials of various shapes and sizes having large specific surface areas can be used as multivalent scaffolds to present carbohydrate ligands. The resulting glyconanomaterials effectively amplify the glycan-mediated interactions, making it possible to use these materials for sensing, imaging, diagnosis, and therapy. In this review, we summarize the synthetic strategies for the preparation of various glyconanomaterials. Examples are given where these glyconanomaterials have been used in sensing and differentiation of proteins and cells, as well as in imaging glycan-medicated cellular responses.

Covalent functionalization of pristine graphene poses considerable challenges due to the lack of reactive functional groups. Herein, we report a simple and general method to covalently functionalize pristine graphene with well-defined chemical functionalities. It is a solution-based process where solvent-exfoliated graphene was treated with perfluorophenylazide (PFPA) by photochemical or thermal activation. Graphene with well-defined chemical functionalities was synthesized and the resulting materials were soluble in organic solvents or water depending on the nature of the functional group on PFPA.

Interactions between carbohydrates and proteins are increasingly being recognized as crucial in many biological processes, such as cellular adhesion and communication. In order to investigate the interactions of carbohydrates and proteins, the development of efficient analytic technologies, as well as novel strategies for the synthesis of carbohydrates, have to be explored. To date, several methods have been exploited to analyze interactions of carbohydrates and proteins, for example, biosensors, nuclear magnetic resonance (NMR); enzyme-linked immunosorbent assays (ELISA), X-ray crystallography and array technologies. This thesis describes the development of novel strategies for the synthesis of carbohydrates, as well as new efficient strategies to Quartz Crystal Microbalance- (QCM-) biosensors and carbohydrate microarrays technologies. These methodologies have been used to probe carbohydrate-lectin-interactions for a range of plant and animal lectins.

Carbohydrate-functionalized gold nanoparticles were employed to differentiate plant-legume lectins using a statistical analysis method of linear discriminant analysis (LDA). Various carbohydrates were conjugated on gold nanoparticles, and the resulting glyconanoparticles were treated with lectins. Changes in the localized surface plasmon resonance of the glyconanoparticles upon lectin binding were recorded, and the data were subjected to LDA. Results showed that the glyconanoparticles successfully differentiated all lectins.

Because of their synthetic accessibility, molecularly imprinted polymer (MIP) nanoparticles are ideal building blocks for preparing multifunctional composites. In this work, we developed a general photocoupling chemistry to enable simple conjugation of MIP nanoparticles with inorganic magnetic nanoparticles. We first synthesized MIP nanoparticles using propranolol as a model template and perfluorophenyl azide-modified silica-coated magnetic nanoparticles. Using a simple photoactivation followed by facile purification with a magnet, we obtained magnetic composite particles that showed selective uptake of propranolol. We characterized the nanoparticles and composite materials using FT-IR, TEM, fluorescence spectroscopy, and radioligand binding analysis. Through the high molecular selectivity of the magnetic composite, we demonstrated the nondestructive feature and the high efficiency of the photocoupling chemistry. The versatile photoconjugation method developed in this work should also be very useful for combining organic MIPs with other inorganic nanoparticles to enable new chemical sensors and high efficiency photocatalysts.

A new microarray platform, based on lectin super-microarrays and glycans labeled with dye-doped nanoparticles, has been developed to study glycan-lectin interactions. Glycan ligands were conjugated onto fluorescein-doped silica nanoparticles (FSNPs) using a general photocoupling chemistry to afford FSNP-labeled glycan probes. Lectins were printed on epoxy slides in duplicate sets to generate lectin super-microarrays where multiple assays could be carried out simultaneously in each lectin microarray. Thus, the lectin super-microarray was treated with FSNP-labeled glycans to screen for specific binding pairs. Furthermore, a series of ligand competition assays were carried out on a single lectin super-microarray to generate the dose-response curve for each glycan-lectin pair, from which the apparent affinity constants were obtained. Results showed 4-7 orders of magnitude increase in affinity over the free glycans with the corresponding lectins. Thus, the glycan epitope structures having weaker affinity than the parent glycans could be readily identified and analyzed from the lectin super-microarrays.

Nanoparticles conjugated with D-maltoheptaose (G7) showed a striking increase in the surface adherence and internalization by E. coli. This applies to silica nanoparticles (SNP), magnetic nanoparticles (MNP), silica-coated magnetic nanoparticles (SMNP) and silica-coated quantum dots (SQDs) ranging from a few to over a hundred nanometers in size, as well as wild type E. coli ATCC 33456, ORN 178, ORN 208 with the maltodextrin transport channel and the LamB mutant JW 3392-1 (Fig. 1). TEM images including the thin section samples revealed the uptake of nanoparticles in cell walls and inside the cytoplasm (Fig. 2). Unfunctionalized nanoparticles and nanoparticles functionalized with β-cyclodextrin (CD) showed little or no binding to the E. coli cell surface, and no obvious internalization of the nanoparticles was observed. D-Mannose-functionalized nanoparticles bound to the pili of E. coli ORN 178 through the well-known Man-binding lectin (FimH) rather than cell internalization. Surface ligands that can improve the uptake of nanomaterials to bacterial cells should provide a powerful means of targeting a payload delivery to a potential disease causing strain. Work is underway to develop nanomaterial delivery systems for multidrug resistance bacteria.

We report that proteins labeled with fluorescein-doped silica nanoparticles (FSNPs) showed drastically different fouling behavior than those labeled with the fluorescein dye. Arrays of polymer films were covalently immobilized on silicon wafers and were treated with protein conjugated on FSNPs. Fluorescence imaging showed that the protein-FSNP conjugate adsorbed strongly on hydrophilic polymers such as poly(ethylene oxide) (PEO) and weakly on hydrophobic polymers such as polystyrene (PS), and the extent of adsorption decreased with increasing hydrophobicity of the polymer film. Thus, carbohydrate microarrays probed with FSNP-labeled lectin showed significantly enhanced signals when PS was used as the antifouling coating than when PEO was used, or when using bovine serum albumin as the blocking agent.

Glyconanomaterials have shown high potential in applications including bioanalysis and nanomedicine. Here, a quantitative analytical technique, based on isothermal titration calorimetry, was developed to characterize the interactions between glyconanoparticles and lectins. By titrating lectins into the glyconanoparticle solution, the apparent dissociation constant, thermodynamic parameters, and the number of binding sites were derived simultaneously. For the glyconanoparticles-lectin binding pairs investigated, a 3–5 orders of magnitude affinity enhancement over the free ligand-lectin interactions was observed which can be attributed to the multivalent ligand presentation on the nanoparticles. The impact of ligand density was also studied, and results showed that the affinity increased with the number of glycans on the nanoparticle.

The interaction between synthetic polymer nanoparticles (NPs) and biomacromolecules (e.g., proteins, lipids, and polysaccharides) can profoundly influence the NPs fate and function. Polysaccharides (e.g., heparin/heparin sulfate) are a key component of cell surfaces and the extracelluar matrix and play critical roles in many biological processes. We report a systematic investigation of the interaction between synthetic polymer nanoparticles and polysaccharides by ITC, SPR, and an anticoagulant assay to provide guidelines to engineer nanoparticles for biomedical applications. The interaction between acrylamide nanoparticles (~30 nm) and heparin is mainly enthalpy driven with submicromolar affinity. Hydrogen bonding, ionic interactions, and dehydration of polar groups are identified to be key contributions to the affinity. It has been found that high charge density and cross-linking of the NP can contribute to high affinity. The affinity and binding capacity of heparin can be significantly diminished by an increase in salt concentration while only slightly decreased with an increase of temperature. A striking difference in binding thermodynamics has been observed when the main component of a polymer nanoparticle is changed from acrylamide (enthalpy driven) to N-isopropylacryalmide (entropy driven). This change in thermodynamics leads to different responses of these two types of polymer NPs to salt concentration and temperature. Select synthetic polymer nanoparticles have also been shown to inhibit protein-heparin interactions and thus offer the potential for therapeutic applications.

Low-mannose (LM) structures were coupled to gold nanoparticles (Au NPs) to amplify the affinity of LMs with Cyanovirin-N (CV-N) lectins and to study the structures of CV-N variants CVN(Q50C) and CVN(MutDB).