Peptide-labeled quantum dots for imaging GPCRs in whole cells and as single molecules

We report a robust and practical method for the preparation of water-soluble luminescent quantum dots (QDs) selectively coupled through an amine or thiol linkage to peptide ligands targeted to G-protein coupling receptors (GPCRs) and demonstrate their utility in whole-cell and single-molecule imaging. We utilized a low molecular weight ( approximately 1200 Da) diblock copolymer with acrylic acids as hydrophilic segments and amido-octyl side chains as hydrophobic segments for facile encapsulation of QDs (QD 595 and QD 514) in aqueous solutions. As proof of principle, these QDs were targeted to the human melanocortin receptor (hMCR) by chemoselectively coupling the polymer-coated QDs to either a hexapeptide analog of alpha-melanocyte stimulating hormone or to the highly potent MT-II ligand containing a unique amine. To label QDs with ligands lacking orthogonal amines, the diblock copolymers were readily modified with water-soluble trioxa-tridecanediamine to incorporate freely available amine functionalities. The amine-functionalized QDs underwent facile reaction with the bifunctional linker NHS-maleimide, allowing for covalent coupling to GPCR-targeted ligands modified with unique cysteines. We demonstrate the utility of these maleimide-functionalized QDs by covalent conjugation to a highly potent Deltorphin-II analog that allowed for selective cell-surface and single-molecule imaging of the human delta-opioid receptor (hDOR).     

In vivo molecular and cellular imaging with quantum dots

Quantum dots (QDs), tiny light-emitting particles on the nanometer scale, are emerging as a new class of fluorescent probe for in vivo biomolecular and cellular imaging. In comparison with organic dyes and fluorescent proteins, QDs have unique optical and electronic properties: size-tunable light emission, improved signal brightness, resistance against photobleaching, and simultaneous excitation of multiple fluorescence colors. Recent advances have led to the development of multifunctional nanoparticle probes that are very bright and stable under complex in vivo conditions. A new structural design involves encapsulating luminescent QDs with amphiphilic block copolymers and linking the polymer coating to tumor-targeting ligands and drug delivery functionalities. Polymer-encapsulated QDs are essentially nontoxic to cells and animals, but their long-term in vivo toxicity and degradation need more careful study. Bioconjugated QDs have raised new possibilities for ultrasensitive and multiplexed imaging of molecular targets in living cells, animal models and possibly in humans.     

抗菌生物材料的研究进展

生物高分子材料(医用缝合线、导尿管等)及人工器官(心瓣膜、人工肾等)的应用日益广泛,使用过程中引发的细菌性感染导致诸多严重后果,不容忽视。基于感染机理,人们通过对生物材料表面进行不同的改性,如通过阻止细菌黏附达到抗菌效果、通过干扰细菌细胞的组成取得杀菌效果等,研究了不同的抗菌机理。本综述了新型有机高分子抗菌剂的发展,提出对生物相容性良好的聚氨酯、聚砜、聚醚砜等材料进行表面改性,使之具有好的抗菌能力,是未来抗菌生物材料的研究与发展方向。     

Amphiphilic biopolymers (amphibiopols) as new surfactants for membrane protein solubilization

The aim of this study was to develop new surfactants for membrane protein solubilization, from a natural, biodegradable polymer: the polysaccharide pullulan. A set of amphiphilic pullulans (HMCMPs), differing in hydrophobic modification ratio, charge ratio, and the nature of the hydrophobic chains introduced, were synthesized and tested in solubilization experiments with outer membranes of Pseudomonas fluorescens. The membrane proteins were precipitated, and then resolubilized with various HMCMPs. The decyl alkyl chain (C(10)) was the hydrophobic graft that gave the highest level of solubilization. Decyl alkyl chain-bearing HMCMPs were also able to extract integral membrane proteins from their lipid environment. The best results were obtained with an amphiphilic pullulan bearing 18% decyl groups (18C(10)). Circular dichroism spectroscopy and membrane reconstitution experiments were used to test the structural and functional integrity of 18C(10)-solubilized proteins (OmpF from Escherichia coli and bacteriorhodopsin from Halobacterium halobium). Whatever their structure type (alpha or beta), 18C(10) did not alter either the structure or the function of the proteins analyzed. Thus, HMCMPs appear to constitute a promising new class of polymeric surfactants for membrane protein studies.     

On the design of composite protein-quantum dot biomaterials via self-assembly

Incorporation of nanoparticles during the hierarchical self-assembly of protein-based materials can impart function to the resulting composite materials. Herein we demonstrate that the structure and nanoparticle distribution of composite fibers are sensitive to the method of nanoparticle addition and the physicochemical properties of both the nanoparticle and the protein. Our model system consists of a recombinant enhanced green fluorescent protein-Ultrabithorax (EGFP-Ubx) fusion protein and luminescent CdSe-ZnS core-shell quantum dots (QDs), allowing us to optically assess the distribution of both the protein and nanoparticle components within the composite material. Although QDs favorably interact with EGFP-Ubx monomers, the relatively rough surface morphology of composite fibers suggests EGFP-Ubx-QD conjugates impact self-assembly. Indeed, QDs templated onto EGFP-Ubx film post-self-assembly can be subsequently drawn into smooth composite fibers. Additionally, the QD surface charge impacts QD distribution within the composite material, indicating that surface charge plays an important role in self-assembly. QDs with either positively or negatively charged coatings significantly enhance fiber extensibility. Conversely, QDs coated with hydrophobic moieties and suspended in toluene produce composite fibers with a heterogeneous distribution of QDs and severely altered fiber morphology, indicating that toluene severely disrupts Ubx self-assembly. Understanding factors that impact the protein-nanoparticle interaction enables manipulation of the structure and mechanical properties of composite materials. Since proteins interact with nanoparticle surface coatings, these results should be applicable to other types of nanoparticles with similar chemical groups on the surface.     

Isonitrile derivatives of polysaccharides as supports for the covalent fixation of proteins and other ligands.

A method for the introduction of side chains containing isonitrile (isocyanide, functional group) on the backbone of polysaccharides and other hydroxylic polymers was developed. The method was based on (a) ionization of some of the hydroxyl groups on the polymer by treatment with a strong base (tert-butoxide) in a polar aprotic solvent (dimethylsulfoxide), and (b) introduction of side chains containing isonitrile groups by nucleophilic attack of the polymeric alkoxide ions on a low molecular weight isonitrile containing a good leaving group in the omega-position, (1-tosyl-3-isocyanopropane). By this method, the side chains containing the-NC functional groups are attached to the polymeric backbone via stable ether bonds. The isonitrile derivatives of cellulose, linear and cross-linked dextran and cross-linked agarose utilized for the covalent fixation of high and low molecular weight ligands by four-component reactions carried out in aqueous medium, at neutral pH.     

Immobilization of Candida bombicola cells on free-standing organic-gold nanoparticle membranes and their use as enzyme sources in biotransformations

Preparation of chemically functionalized biocompatible surfaces is of current interest, with application in the immobilization of various bioactive species such as DNA, enzymes, whole cells, etc. We report herein the one-step synthesis of a self-supporting gold nanoparticle membrane, its surface modification, and application in the immobilization of Candida bombicola (yeast) cells. The gold nanoparticle membrane is prepared by the spontaneous reduction of aqueous chloroaurate ions by a diamine at a liquid-liquid interface. The gold nanoparticles in the polymeric membrane may be capped with octadecylamine (ODA) molecules, thereby rendering the nanoparticle membrane hydrophobic. Exposure of the hydrophobized organic-gold nanoparticle membrane to C. bombicola yeast cells results in their binding to the membrane, possibly through nonspecific interactions such as hydrophobic interactions between the yeast cell walls and the ODA molecules. The enzyme cytochrome P450 present in the yeast cells immobilized on the organic-gold nanoparticle membrane was then used in the transformation of the arachidonic acid (AA) to sophorolipids followed by acid hydrolysis to form 20-hydroxyeicosatetraneoic acid (20-HETE). The organic-gold nanoparticle membrane-C. bombicola bioconjugate could be easily separated from the reaction medium and reused a number of times.     

Biomimetic Hydrophobic Surfaces with Low or High Adhesion Based on Poly(vinyl alcohol) and SiO2 Nanoparticles

Superhydrophobic surfaces are often found in nature,such as plant leaves and insect wings.Inspired by superhydrophobic phenomenon of the rose petals and the lotus leaves,biomimetic hydrophobic surfaces with high or low adhesion were prepared with a facile drop-coating approach in this paper.Poly(vinyl alcohol) (PVA) was used as adhesive and SiO2 nanoparticles were used to fabricate surface micro-structure.Stearic acid or dodecafluoroheptyl-propyl-trimethoxysilane (DFTMS) were used as low surface energy materials to modify the prepared PVA/SiO2 coating surfaces.The effects of size of SiO2 nanoparticles,concentration of SiO2 nanoparticle suspensions and the modifications on the wettability of the surface were investigated.The morphology of the PVA/SiO2 coating surfaces was observed by using scanning electron microscope.Water contact angle of the obtained superhydrophilic surface could reach to 3°.Stearic acid modified PVA/SiO2 coating surfaces showed hydrophobicity with high adhesion.By mixing the SiO2 nanoparticles with sizes of 40 nm and 200 nm and modifying with DFTMS,water contact angle of the obtained coating surface could be up to 155° and slide angle was only 5°.This work provides a facile and useful method to control surface wettability through changing the roughness and chemical composition of a surface.     

Capping of CdSe-ZnS quantum dots with DHLA and subsequent conjugation with proteins

We provide a detailed protocol for designing water-soluble CdSe-ZnS quantum dots (QDs) based on cap exchange of the native hydrophobic shell with dihydrolipoic acid (DHLA) ligands, and the preparation of functional QD bioconjugates for use in immunoassays. Our conjugation strategy is based on non-covalent self-assembly between DHLA-capped QDs and protein appended with either an electrostatic attachment domain (namely, the basic leucine zipper) or a polyhistidine tag. These bioconjugates combine the properties of the QD and attached biomolecule to create structures with desirable luminescent and biologically specific properties. This method also allows the preparation of mixed surface conjugates, which results in the conjugates gaining multiple biological activities. Conjugation of DHLA-capped QDs to maltose binding protein (MBP), the immunoglobulin-G-binding beta2 domain of streptococcal protein G (PG) and avidin will be described. MBP and PG were modified by genetic fusion with either a charged leucine zipper or a polyhistidine interaction domain.     

Synthesis of biocompatible polymeric hydrogels with tunable adhesion to both hydrophobic and hydrophilic surfaces

We report the synthesis of biocompatible polymeric hydrogels based on poly(vinyl acetate) (PVAc) and poly(methyl vinyl ether-co-maleic anhydride) (PMVE-MA). These polymeric hydrogels show strong and tunable adhesion to both hydrophobic and hydrophilic surfaces and should be ideal candidates as bioadhesives for applications such as denture adhesion. PVAc was partially hydrolyzed and then mixed with PMVE-MA. Crosslinking between these two polymers through reactions between hydroxyl groups in partially hydrolyzed PVAc and maleic anhydride groups in PMVE-MA increased their compatibility and prevented phase separation so transparent hydrogels were formed. The adhesion of these polymeric hydrogels to hydrophobic and hydrophilic surfaces was tailored by regulating the degree of hydrolysis of PVAc and the molecular weights of the polymers. In the vicinity of critical gel point, where the elastic modulus G' and the viscous modulus G' scale as G' approximately G' approximately omega (0.3), polymeric hydrogels show optimal adhesion. Transparent gels are formed in mixed solvents of water and ethanol. The content of ethanol in the mixed solvent can be partially replaced by propylene glycol, glycerol, or poly(ethenyl glycol)-400, and the composition of appropriate mixed solvents can be determined by the calculation of solubility parameters.     

Quantum dots and peptides: a bright future together

Nanocrystalline semi-conductor materials, called quantum dots (QDs), exhibit unique optical and spectroscopic properties, which include, broad absorption, narrow and tunable emission, resistance to photobleaching, strong luminescence, and long luminescent lifetimes. These remarkable properties of QDs have resulted in their use as an alternative to both small-molecule and protein fluorophores in innumerable biological applications. The overlap of QDs with the rich chemistry and biology that is characteristic of the peptide arena is an emerging research area. Peptides engineered with appropriate cysteines or histidines have served as ligands for producing water soluble QDs as well as for tagging protein ligands and biosensors to QD surfaces. Incorporation of cell-penetrating peptides on QD surfaces has allowed for the translocation of functionalized QDs into cells for intracellular imaging applications. QDs containing fluorescently labeled peptide substrates have shown utility in the development of novel protease assays. Moreover, QDs-labeled peptides that serve as ligands for cellular receptors provide an alternative to antibody mediated imaging at the whole-cell and single molecule level to study receptor distribution and trafficking. This review highlights the overlap between QD and peptide chemistry and speculates on future research directions.     

Selective N-alkylation of β-alanine facilitates the synthesis of a poly(amino acid)-based theranostic nanoagent

The development of functional amino acid-based polymeric materials is emerging as a platform to create biodegradable and nontoxic nanomaterials for medical and biotechnology applications. In particular, facile synthetic routes for these polymers and their corresponding polymeric nanomaterials would have a positive impact in the development of novel biomaterials and nanoparticles. However, progress has been hampered by the need to use complex protection-deprotection methods and toxic phase transfer catalysts. In this study, we report a facile, single-step approach for the synthesis of an N-alkylated amino acid as an AB-type functional monomer to generate a novel pseudo-poly(amino acid), without using the laborious multistep, protection-deprotection methods. This synthetic strategy is reproducible, easy to scale up, and does not produce toxic byproducts. In addition, the synthesized amino acid-based polymer is different from conventional linear polymers as the butyl pendants enhance its solubility in common organic solvents and facilitate the creation of hydrophobic nanocavities for the effective encapsulation of hydrophobic cargos upon nanoparticle formation. Within the nanoparticles, we have encapsulated a hydrophobic DiI dye and a therapeutic drug, Taxol. In addition, we have conjugated folic acid as a folate receptor-targeting ligand for the targeted delivery of the nanoparticles to cancer cells expressing the folate receptor. Cell cytotoxicity studies confirm the low toxicity of the polymeric nanoparticles, and drug-release experiments with the Taxol-encapsulated nanoparticles only exhibit cytotoxicity upon internalization into cancer cells expressing the folate receptor. Taken together, these results suggested that our synthetic strategy can be useful for the one-step synthesis of amino acid-based small molecules, biopolymers, and theranostic polymeric nanoagents for the targeted detection and treatment of cancer.     

Molecular profiling of single cancer cells and clinical tissue specimens with semiconductor quantum dots

Semiconductor quantum dots (QDs) are a new class of fluorescent labels with broad applications in biomedical imaging, disease diagnostics, and molecular and cell biology. In comparison with organic dyes and fluorescent proteins, quantum dots have unique optical and electronic properties such as size-tunable light emission, improved signal brightness, resistance against photobleaching, and simultaneous excitation of multiple fluorescence colors. Recent advances have led to multifunctional nanoparticle probes that are highly bright and stable under complex in vitro and in vivo conditions. New designs involve encapsulating luminescent QDs with amphiphilic block copolymers, and linking the polymer coating to tumor-targeting ligands and drug-delivery functionalities. These improved QDs have opened new possibilities for real-time imaging and tracking of molecular targets in living cells, for multiplexed analysis of biomolecular markers in clinical tissue specimens, and for ultrasensitive imaging of malignant tumors in living animal models. In this article, we briefly discuss recent developments in bioaffinity QD probes and their applications in molecular profiling of individual cancer cells and clinical tissue specimens.     

A new polymeric coating for protein microarrays

Despite the increasing interest in arraying proteins in a high-density format, several technical issues still impede the development of protein microarray technology. One of the major problems is the availability of substrates that are able to bind native proteins with high density. In this study, we investigated the suitability of a novel surface as a support for protein microarrays. A polymeric glass coating is obtained by physical adsorption of a N,N-dimethylacrylamide (DMA), N,N-acryloyloxysuccinimide (NAS), and [3-(methacryloyl-oxy)propyl]trimethoxysilyl (MAPS) copolymer. The coating procedure provides a fast and inexpensive method of producing hydrophilic functional surfaces. The slide performance was investigated in a protein-protein interaction experiment and in the assessment of rheumatoid factor (RF) in human serum samples. The results demonstrate that the ligands immobilized on the polymeric surface maintain an active conformation and are easily accessible, providing a detection limit of 54amol/spot. Moreover, in the RF assay, after hybridization with the sera, the slides have a low background, leading to a detection limit of 900amol/spot.     

Design of surfactants suitable for protein extraction by reversed micelles

New surfactants have been synthesized for potential use in reversed micellar protein extraction operations. Preferential solubility of the surfactant in an aliphatic solvent such as hexane, heptane, or isooctane and the formation of reversed micelles accompanied with solubilization of significant quantities of water can be achieved by using strongly hydrophobic, twin alkyl chains as the hydrophobic moiety. Different surfactants having identical water-solubilizing capacities can have significantly different behavior in protein extractions, where extraction efficiency appears to be governed by the nature of the interfacial complex that forms between surfactants and proteins. Bulky surfactant chains provide a steric hindrance to the adsorption of the surfactant to the protein surface, thus inhibiting solvation of the protein/surfactant complex, and hence protein extraction. Under these conditions, a precipitate forms either in the bulk aqueous phase or at the interface. Surfactants that can form a close-packed complex with the protein are excellent protein-solubilizing agents. Dioleyl phosphoric acid (DOLPA) appears to be the best surfactant currently available for protein extraction. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 54: 26-32, 1997.     

Aptamer-functionalized quantum dots as theranostic nanotools against cancer and bacterial infections: A comprehensive overview of recent trends

Aptamers (Apts) are synthetic nucleic acid ligands that can be engineered to target various molecules, including amino acids, proteins, and pharmaceuticals. Through a series of adsorption, recovery, and amplification steps, Apts are extracted from combinatorial libraries of synthesized nucleic acids. Using aptasensors in bioanalysis and biomedicine can be improved by combining them with nanomaterials. Moreover, Apt-associated nanomaterials, including liposomes, polymeric, dendrimers, carbon nanomaterials, silica, nanorods, magnetic NPs, and quantum dots (QDs), have been widely used as promising nanotools in biomedicine. Following surface modifications and conjugation with appropriate functional groups, these nanomaterials can be successfully used in aptasensing. Advanced biological assays can use Apts immobilized on QD surfaces through physical interaction and chemical bonding. Accordingly, modern QD aptasensing platforms rely on interactions between QDs, Apts, and targets to detect them. QD-Apt conjugates can be used to directly detect prostate, ovarian, colorectal, and lung cancers or simultaneously detect biomarkers associated with these malignancies. Tenascin-C, mucin 1, prostate-specific antigen, prostate-specific membrane antigen, nucleolin, growth factors, and exosomes are among the cancer biomarkers that can be sensitively detected using such bioconjugates. Furthermore, Apt-conjugated QDs have shown great potential for controlling bacterial infections such as Bacillus thuringiensis, Pseudomonas aeruginosa, Escherichia coli, Acinetobacter baumannii, Campylobacter jejuni, Staphylococcus aureus, and Salmonella typhimurium. This comprehensive review discusses recent advancements in the design of QD-Apt bioconjugates and their applications in cancer and bacterial theranostics.     

Solubilization and bio-conjugation of quantum dots and bacterial toxicity assays by growth curve and plate count

Quantum dots (QDs) are fluorescent semiconductor nanoparticles with size-dependent emission spectra that can be excited by a broad choice of wavelengths. QDs have attracted a lot of interest for imaging, diagnostics, and therapy due to their bright, stable fluorescence. QDs can be conjugated to a variety of bio-active molecules for binding to bacteria and mammalian cells. QDs are also being widely investigated as cytotoxic agents for targeted killing of bacteria. The emergence of multiply-resistant bacterial strains is rapidly becoming a public health crisis, particularly in the case of Gram negative pathogens. Because of the well-known antimicrobial effect of certain nanomaterials, especially Ag, there are hundreds of studies examining the toxicity of nanoparticles to bacteria. Bacterial studies have been performed with other types of semiconductor nanoparticles as well, especially TiO(2), but also ZnO and others including CuO. Some comparisons of bacterial strains have been performed in these studies, usually comparing a Gram negative strain with a Gram positive. With all of these particles, mechanisms of toxicity are attributed to oxidation: either the photogeneration of reactive oxygen species (ROS) by the particles or the direct release of metal ions that can cause oxidative toxicity. Even with these materials, results of different studies vary greatly. In some studies the Gram positive test strain is reportedly more sensitive than the Gram negative; in others it is the opposite. These studies have been well reviewed. In all nanoparticle studies, particle composition, size, surface chemistry, sample aging/breakdown, and wavelength, power, and duration of light exposure can all dramatically affect the results. In addition, synthesis byproducts and solvents must be considered. High-throughput screening techniques are needed to be able to develop effective new nanomedicine agents. CdTe QDs have anti-microbial effects alone or in combination with antibiotics. In a previous study, we showed that coupling of antibiotics to CdTe can increase toxicity to bacteria but decrease toxicity to mammalian cells, due to decreased production of reactive oxygen species from the conjugates. Although it is unlikely that cadmium-containing compounds will be approved for use in humans, such preparations could be used for disinfection of surfaces or sterilization of water. In this protocol, we give a straightforward approach to solubilizing CdTe QDs with mercaptopropionic acid (MPA). The QDs are ready to use within an hour. We then demonstrate coupling to an antimicrobial agent. The second part of the protocol demonstrates a 96-well bacterial inhibition assay using the conjugated and unconjugated QDs. The optical density is read over many hours, permitting the effects of QD addition and light exposure to be evaluated immediately as well as after a recovery period. We also illustrate a colony count for quantifying bacterial survival.     

Reconstruction of surfaces from mixed hydrocarbon and PEG components in water: responsive surfaces aid fouling release

Coatings derived from surface active block copolymers (SABCs) having a combination of hydrophobic aliphatic (linear hydrocarbon or propylene oxide-derived groups) and hydrophilic poly(ethlyene glycol) (PEG) side chains have been developed. The coatings demonstrate superior performance against protein adsorption as well as resistance to biofouling, providing an alternative to coatings containing fluorinated side chains as the hydrophobe, thus reducing the potential environmental impact. The surfaces were examined using dynamic water contact angle, captive air-bubble contact angle, atomic force microscopy, X-ray photoelectron spectroscopy, and near-edge X-ray absorption fine structure analysis. The PS(8K)-b-P(E/B)(25K)-b-PI(10K) triblock copolymer precursor (K3) initially dominated the dry surface. In contrast to previous studies with mixed fluorinated/PEG surfaces, these new materials displayed significant surface changes after exposure to water that allowed fouling resistant behavior. PEG groups buried several nanometers below the surface in the dry state were able to occupy the coating surface after placement in water. The resulting surface exhibits a very low contact angle and good antifouling properties that are very different from those of K3. The surfaces are strongly resistant to protein adsorption using bovine serum albumin as a standard protein challenge. Biofouling assays with sporelings of the green alga Ulva and cells of the diatom Navicula showed the level of adhesion was significantly reduced relative to that of a PDMS standard and that of the triblock copolymer precursor of the SABCs.     

Adsorption of bacterial capsular exopolymers to hydrophilic and hydrophobic surfaces: A 2D‐FTIR analysis

Analysis of the adsorption of capsular exopolymers (EPS) from Pseudomonas sp. NCIMB 2021 to hydrophilic and hydrophobic gold surfaces was examined, in situ, using Fourier transform infrared spectroscopy. The molecular sequence of events occurring upon EPS adsorption to hydrophilic and hydrophobic surfaces has been elucidated using dynamic 2D‐FTIR correlation spectroscopy. This method of analysis enables the enhancement of the resolution of overlapping spectral features and the elucidation of time‐dependent changes. The data reveal the existence of surface dependent adsorption mechanisms. At both surfaces, the aromatic tyrosyl side chains of the protein moiety displace water. This is followed by an adsorption step dominated by carboxylate groups. However, at the hydrophobic surface, the two steps are interrupted by the ingress of water back to the surface. Furthermore, the amount of neutral exopolymer present was greater at the hydrophilic surface than the hydrophobic surface.     

De novo synthesis and cellular uptake of organic nanocapsules with tunable surface chemistry

Water-soluble organic nanocapsules were prepared from bottlebrush copolymers with triblock terpolymer side chains composed of a degradable inner block (polylactide), a cross-linkable middle block (poly(4-butenylstyrene)), and a functional outer block (poly(styrene-co-maleic anhydride)). Bottlebrush copolymers are macromolecules with a long linear backbone and shorter polymeric side chains densely grafted onto the backbone. Hollow cylindrical nanoparticles were prepared by peripheral cross-linking of the bottlebrush copolymers and subsequent selective removal of the core. Reactive anhydride groups of the outer functional layer allowed for the preparation of nanocapsules with tunable surface characteristics. Cellular uptake of negatively charged organic nanocapsules showed strong surface chemistry dependence. The presence of hydrophobic groups on the nanocapsule surface was necessary for their nonspecific association with the cell membrane and subsequent internalization by endocytosis. The length of surface grafted oligoethylene glycol chains also had a dramatic influence on the intracellular accumulation of nanocapsules. Macropinocytosis was shown to be the predominant pathway for the cellular uptake of organic nanocapsules.