Pre-coating cRGD-modified bovine serum albumin enhanced the anti-tumor angiogenesis of siVEGF-loaded chitosan-based nanoparticles by manipulating the protein corona composition.

Chitosan-based nanoparticles inevitably adsorb numerous proteins in the bloodstream, forming a protein corona that significantly influences their functionality. This study employed a pre-coated protein corona using cyclic Arg-Gly-Asp peptide (cRGD)-modified bovine serum albumin (BcR) to confer tumor-targeting capabilities on siVEGF-loaded chitosan-based nanoparticles (CsR/siVEGF NPs) and actively manipulated the serum protein corona composition to enhance their anti-tumor angiogenesis. Consequently, BcR effectively binds to the nanoparticles' surface, generating nanocarriers of appropriate size and stability that enhance the inhibition of endothelial cell proliferation, migration, invasion, and tube formation, as well as suppress tumor proliferation and angiogenesis in tumor-bearing nude mice. Proteomic analysis indicated a significant enrichment of serotransferrin, albumin, and proteasome subunit alpha type-1 in the protein corona of BcR-precoated NPs formed in the serum of tumor-bearing nude mice. Additionally, there was a decrease in proteins associated with complement activation, immunoglobulins, blood coagulation, and acute-phase responses. This modification resulted in an enhanced impact on anti-tumor angiogenesis, along with a reduction in opsonization and inflammatory responses. Therefore, pre-coating of nanoparticles with a functionalized albumin corona to manipulate the composition of serum protein corona emerges as an innovative approach to improve the delivery effectiveness of chitosan-based carriers for siVEGF, targeting the inhibition of tumor angiogenesis.

Sources of biases in the in vitro testing of nanomaterials: the role of the biomolecular corona.

The biological fate of nanomaterials (NMs) is driven by specific interactions through which biomolecules, naturally adhering onto their surface, engage with cell membrane receptors and intracellular organelles. The molecular composition of this layer, called the biomolecular corona (BMC), depends on both the physical-chemical features of the NMs and the biological media in which the NMs are dispersed and cells grow. In this work, we demonstrate that the widespread use of 10% fetal bovine serum in an in vitro assay cannot recapitulate the complexity of in vivo systemic administration, with NMs being transported by the blood. For this purpose, we undertook a comparative journey involving proteomics, lipidomics, high throughput multiparametric in vitro screening, and single molecular feature analysis to investigate the molecular details behind this in vivo/in vitro bias. Our work indirectly highlights the need to introduce novel, more physiological-like media closer in composition to human plasma to produce realistic in vitro screening data for NMs. We also aim to set the basis to reduce this in vitro-in vivo mismatch, which currently limits the formulation of NMs for clinical settings.

Cotransport of nanoplastics and plastic additive bisphenol AF (BPAF) in unsaturated hyporheic zone: Coupling effects of surface functionalization and protein corona.

The ecological risk of combined pollution from microplastics (MPs) and associated contaminants usually depends on their interactions and environmental behavior, which was also disturbed by varying surface modifications of MPs. In this study, the significance of surface functionalization and protein-corona on the cotransport of nanoplastics (NPs; 100 nm) and the related additive bisphenol AF (BPAF) was examined in simulated unsaturated hyporheic zone (quartz sand; 250-425 µm). The electronegative bovine serum albumin (BSA) and electropositive trypsin were chosen as representative proteins, while pristine (PNPs), amino-modified (ANPs), and carboxyl-modified NPs (CNPs) were representative NPs with different charges. The presence of BPAF inhibited the mobility of PNPs/CNPs, but enhanced the release of ANPs in hyporheic zone, which was mainly related to their hydrophobicity changes and electrostatic interactions. Meanwhile, the NPs with high mobility and strong affinity to BPAF became effective carriers, promoting the cotransport of BPAF by 16.4 %-26.4 %. The formation of protein-coronas altered the mobility of NPs alone and their cotransport with BPAF, exhibiting a coupling effect with functional groups. BSA-corona promoted the transport of PNPs/CNPs, but this promoting effect was weakened by the presence of BPAF via increasing particle aggregation and hydrophobicity. Inversely, trypsin-corona aggravated the deposition of PNPs/CNPs, but competition deposition sites and increased energy barrier caused by coexisting BPAF reversed this effect, facilitating the cotransport of trypsin-PNPs/CNPs in hyporheic zone. However, BPAF and protein-coronas synergistically promoted the mobility of ANPs, owing to competition deposition sites and decreased electrostatic attraction. Although all of the NPs with two protein-coronas reduced dissolved BPAF in the effluents via providing deposition sites, the cotransport of total BPAF was improved by the NPs with high mobility (BSA-PNPs/CNPs) or high affinity to BPAF (BSA/trypsin-ANPs). However, the trypsin-PNPs/CNPs inhibited the transport of BPAF due to their weak mobility and adsorption with BPAF. The results provide new insights into the role of varying surface modifications on NPs in the vertical cotransport of NPs and associated contaminants in unsaturated hyporheic zone.

Phospholipid Type Regulates Protein Corona Composition and In Vivo Performance of Lipid Nanodiscs.

Over the years, there has been significant interest in PEGylated lipid-based nanocarriers within the drug delivery field. The inevitable interplay between the nanocarriers and plasma protein plays a pivotal role in their in vivo biological fate. Understanding the factors influencing lipid-based nanocarrier and protein corona interactions is of paramount importance in the design and clinical translation of these nanocarriers. Herein, discoid-shaped lipid nanodiscs (sNDs) composed of different phospholipids with varied lipid tails and head groups were fabricated. We investigated the impact of phospholipid components on the interaction between sNDs and serum proteins, particle stability, and biodistribution. The results showed that all of these lipid nanodiscs remained stable over a 15 day storage period, while their stability in the blood serum demonstrated significant differences. The sND composed of POPG exhibited the least stability due to its potent complement activation capability, resulting in rapid blood clearance. Furthermore, a negative correlation between the complement activation capability and serum stability was identified. Pharmacokinetic and biodistribution experiments indicated that phospholipid composition did not influence the capability of sNDs to evade the accelerated blood clearance phenomenon. Complement deposition on the sND was inversely associated with the area under the curve. Additionally, all lipid nanodiscs exhibited dominant adsorption of apolipoprotein. Remarkably, the POPC-based lipid nanodisc displayed a significantly higher deposition of apolipoprotein E, contributing to an obvious brain distribution, which provides a promising tool for brain-targeted drug delivery.

The in vitro gastrointestinal digestion-associated protein corona of polystyrene nano- and microplastics increases their uptake by human THP-1-derived macrophages.

BACKGROUND: Micro- and nanoplastics (MNPs) represent one of the most widespread environmental pollutants of the twenty-first century to which all humans are orally exposed. Upon ingestion, MNPs pass harsh biochemical conditions within the gastrointestinal tract, causing a unique protein corona on the MNP surface. Little is known about the digestion-associated protein corona and its impact on the cellular uptake of MNPs. Here, we systematically studied the influence of gastrointestinal digestion on the cellular uptake of neutral and charged polystyrene MNPs using THP-1-derived macrophages. RESULTS: The protein corona composition was quantified using LC‒MS-MS-based proteomics, and the cellular uptake of MNPs was determined using flow cytometry and confocal microscopy. Gastrointestinal digestion resulted in a distinct protein corona on MNPs that was retained in serum-containing cell culture medium. Digestion increased the uptake of uncharged MNPs below 500 nm by 4.0-6.1-fold but did not affect the uptake of larger sized or charged MNPs. Forty proteins showed a good correlation between protein abundance and MNP uptake, including coagulation factors, apolipoproteins and vitronectin. CONCLUSION: This study provides quantitative data on the presence of gastrointestinal proteins on MNPs and relates this to cellular uptake, underpinning the need to include the protein corona in hazard assessment of MNPs.

Effect of Lipid Corona on Phenylalanine-Functionalized Gold Nanoparticles to Develop Stable and Corona-Free Systems.

Conventional gold nanoparticles (Au NPs) have many limitations, such as aggregation and subsequent precipitation in the medium of high ionic strength and protein molecules. Furthermore, when exposed to biological fluids, nanoparticles form a protein corona, which controls different biological processes such as the circulation lifetime, drug release profile, biodistribution, and in vivo cellular distribution. These limitations reduce the functionality of Au NPs in targeted delivery, bioimaging, gene delivery, drug delivery, and other biomedical applications. To circumvent these problems, there are numerous attempts to design corona-free and stable nanoparticles. Here, we report for the first time that lipid corona (coating of lipid) formation on phenylalanine-functionalized Au NPs (AuPhe NPs) imparts excellent stability against the high ionic strength of bivalent metal ions, amino acids, and proteins of different charges as compared to bare nanoparticles. Moreover, this work is focused on the ability of lipid corona formation on AuPhe NPs to prevent protein adsorption in the presence of cell culture medium (CCM), oppositely charged protein (e.g., histone 3), and human serum albumin (HSA). The results demonstrate that the lipid corona successfully protects the AuPhe NPs from protein adsorption, leading to the development of corona-free character. This unique achievement has profound implications for enhancing the biomedical utility and safety of these nanoparticles.

Separation of protein corona from nanoparticles under intracellular acidic conditions: effect of protonation on nanoparticle-protein and protein-protein interactions.

Protein coronas separate from nanoparticles under intracellular acidic conditions however, competitive adsorption of multiple proteins and their protein network formation under different pH conditions have not yet been systematically studied at the atomic scale. Herein, we report all-atom molecular dynamics simulations of plasma proteins (human serum albumin and immunoglobulin gamma-1 chain C) adsorbed to 10 nm-sized carboxyl-terminated polystyrene (PS) nanoparticles at different protonation states that mimic extracellular and intracellular pH conditions of 7, 6-5, and 4.5. Binding free energies are calculated from umbrella sampling simulations, showing the significantly weakened binding between PS particles and proteins at the protonation state at pH 4.5, in agreement with experiments showing the separation of protein corona from nanoparticles at pH 4.5. Mixtures of multiple proteins and PS particles are also simulated, showing much less protein adsorption and protein cluster formation at the protonation state at pH 4.5 than that at higher pH values, which are further confirmed by calculating the diffusivities and hydrodynamic radii of individual proteins. In particular, electrostatic particle-protein and protein-protein interactions are significantly weakened by a combination of particle and protein protonation rather than by particle protonation alone, to an extent dependent on different proteins. These findings help explain the experimental observations regarding separation of protein corona from nanoparticles under intracellular acidic conditions at pH 4.5 but not at higher pH, supporting that acidification cannot be the only reason for this separation during the process of endosome maturation.

Development of a fast and simple method for the isolation of superparamagnetic iron oxide nanoparticles protein corona from protein-rich matrices.

The adsorption of proteins onto the surface of nanoparticle (NP) leads to the formation of the so-called "protein corona" as consisting both loosely and tightly bound proteins. It is well established that the biological identity of NPs that may be acquired after exposure to a biological matrix is mostly provided by the components of the hard corona as the pristine surface is generally less accessible for binding. For that reason, the isolation and the characterisation of the NP-corona complexes and identification of the associated biomolecules can help in understanding its biological behaviour. Established methods for the isolation of the NP-HC complexes are time-demanding and can lead to different results based on the isolation method applied. Herein, we have developed a fast and simple method using ferromagnetic beads isolated from commercial MACS column and used for the isolation of superparamagnetic NP following exposure to different types of biological milieu. We first demonstrated the ability to easily isolate superparamagnetic iron oxide NPs (IONPs) from different concentrations of human blood plasma, and also tested the method on the corona isolation using more complex biological matrices, such as culture medium containing pulmonary mucus where the ordinary corona methods cannot be applied. Our developed method showed less than 20% difference in plasma corona composition when compared with centrifugation. It also showed effective isolation of NP-HC complexes from mucus-containing culture media upon comparing with centrifugation and MACS columns, which failed to wash out the unbound proteins. Our study was supported with a full characterisation profile including dynamic light scattering, nanoparticle tracking analysis, analytical disk centrifuge, and zeta potentials. The biomolecules/ proteins composing the HC were separated by vertical gel electrophoresis and subsequently analysed by liquid chromatography-tandem mass spectrometry. In addition to our achievements in comparing different isolation methods to separate IONPs with corona from human plasma, this is the first study that provides a complete characterisation profile of particle protein corona after exposure in vitro to pulmonary mucus-containing culture media.

Surface Chemistry of Biologically Active Reducible Oxide Nanozymes.

Reducible metal oxide nanozymes (rNZs) are a subject of intense recent interest due to their catalytic nature, ease of synthesis, and complex surface character. Such materials contain surface sites which facilitate enzyme-mimetic reactions via substrate coordination and redox cycling. Further, these surface reactive sites are shown to be highly sensitive to stresses within the nanomaterial lattice, the physicochemical environment, and to processing conditions occurring as part of their syntheses. When administered in vivo, a complex protein corona binds to the surface, redefining its biological identity and subsequent interactions within the biological system. Catalytic activities of rNZs each deliver a differing impact on protein corona formation, its composition, and in turn, their recognition, and internalization by host cells. Improving the understanding of the precise principles that dominate rNZ surface-biomolecule adsorption raises the question of whether designer rNZs can be engineered to prevent corona formation, or indeed to produce "custom" protein coronas applied either in vitro, and preadministration, or formed immediately upon their exposure to body fluids. Here, fundamental surface chemistry processes and their implications in rNZ material performance are considered. In particular, material structures which inform component adsorption from the application environment, including substrates for enzyme-mimetic reactions are discussed.

Biomolecular Corona of Gold Nanoparticles: The Urgent Need for Strong Roots to Grow Strong Branches.

Gold nanoparticles (GNPs) are largely employed in diagnostics/biosensors and are among the most investigated nanomaterials in biology/medicine. However, few GNP-based nanoformulations have received FDA approval to date, and promising in vitro studies have failed to translate to in vivo efficacy. One key factor is that biological fluids contain high concentrations of proteins, lipids, sugars, and metabolites, which can adsorb/interact with the GNP's surface, forming a layer called biomolecular corona (BMC). The BMC can mask prepared functionalities and target moieties, creating new surface chemistry and determining GNPs' biological fate. Here, the current knowledge is summarized on GNP-BMCs, analyzing the factors driving these interactions and the biological consequences. A partial fingerprint of GNP-BMC analyzing common patterns of composition in the literature is extrapolated. However, a red flag is also risen concerning the current lack of data availability and regulated form of knowledge on BMC. Nanomedicine is still in its infancy, and relying on recently developed analytical and informatic tools offers an unprecedented opportunity to make a leap forward. However, a restart through robust shared protocols and data sharing is necessary to obtain "stronger roots". This will create a path to exploiting BMC for human benefit, promoting the clinical translation of biomedical nanotools.

Protein Corona Sensor Array Nanosystem for Detection of Coronary Artery Disease.

Coronary artery disease (CAD) is the most common type of heart disease and represents the leading cause of death in both men and women worldwide. Early detection of CAD is crucial for decreasing mortality, prolonging survival, and improving patient quality of life. Herein, a non-invasive is described, nanoparticle-based diagnostic technology which takes advantages of proteomic changes in the nano-bio interface for CAD detection. Nanoparticles (NPs) exposed to biological fluids adsorb on their surface a layer of proteins, the "protein corona" (PC). Pathological changes that alter the plasma proteome can directly result in changes in the PC. By forming disease-specific PCs on six NPs with varying physicochemical properties, a PC-based sensor array is developed for detection of CAD using specific PC pattern recognition. While the PC of a single NP may not provide the required specificity, it is reasoned that multivariate PCs across NPs with different surface chemistries, can provide the desirable information to selectively discriminate the condition under investigation. The results suggest that such an approach can detect CAD with an accuracy of 92.84%, a sensitivity of 87.5%, and a specificity of 82.5%. These new findings demonstrate the potential of PC-based sensor array detection systems for clinical use.

Understanding protein-nanoparticle interactions leading to protein corona formation: In vitro - in vivo correlation study.

Nanoparticles (NPs) in contact with biological fluids form a biomolecular corona through interactions with proteins, lipids, and sugars, acquiring new physicochemical properties. This work explores the interaction between selected proteins (hemoglobin and fetuin-A) that may alter NP circulation time and NPs of different surface charges (neutral, positive, and negative). The interaction with key proteins albumin and transferrin, the two of the most abundant proteins in plasma was also studied. Binding affinity was investigated using quartz crystal microbalance and fluorescence quenching, while circular dichroism assessed potential conformational changes. The data obtained from in vitro experiments were compared to in vivo protein corona data. The results indicate that electrostatic interactions primarily drive protein-NP interactions, and higher binding affinity does not necessarily translate into more significant structural changes. In vitro and single protein-NP studies provide valuable insights that can be correlated with in vivo observations, opening exciting possibilities for future protein corona studies.

Effect of the Protein Corona Formation on Antibody Functionalized Liquid Lipid Nanocarriers.

The main aim of this study is to report basic knowledge on how a protein corona (PC) could affect or modify the way in which multifunctionalized nanoparticles interact with cells. With this purpose, we have firstly optimized the development of a target-specific nanocarrier by coupling a specific fluorescent antibody on the surface of functionalized lipid liquid nanocapsules (LLNCs). Thus, an anti-HER2-FITC antibody (αHER2) has been used, HER2 being a surface receptor that is overexpressed in several tumor cells. Subsequently, the in vitro formation of a PC has been developed using fetal bovine serum supplemented with human fibrinogen. Dynamic Light Scattering (DLS), Nanoparticle Tracking Analysis (NTA), Laser Doppler Electrophoresis (LDE), and Gel Chromatography techniques have been used to assure a complete physico-chemical characterization of the nano-complexes with (LLNCs-αHER2-PC) and without (LLNCs-αHER2) the surrounding PC. In addition, cellular assays were performed to study the cellular uptake and the specific cellular-nanocarrier interactions using the SKBR3 (high expression of HER2) breast cancer cell line and human dermal fibroblasts (HDFa) (healthy cell line without expression of HER2 receptors as control), showing that the SKBR3 cell line had a higher transport rate (50-fold) than HDFa at 60 min with LLNCs-αHER2. Moreover, the SKBR3 cell line incubated with LLNCs-αHER2-PC suffered a significant reduction (40%) in the uptake. These results suggest that the formation of a PC onto LLNCs does not prevent specific cell targeting, although it does have an important influence on cell uptake.

The In Vivo Fate of Polycatecholamine Coated Nanoparticles Is Determined by a Fibrinogen Enriched Protein Corona.

Polycatecholamine coatings have attracted significant attention in the past 10 years owing to their ability to functionalize a wide range of materials. Here we apply the use of such coatings to drug nanocrystals, made from a poorly soluble drug compound, to postfunctionalize the nanocrystal surface with the aim of providing steric stabilization and extending their circulation time after intravenous injection. We show that both polydopamine and polynorepinephrine can be used to successfully modify drug nanocrystals and subsequently incorporate end-functionalized PEG to the surface. Even though high grafting densities of PEG were achieved, we observed rapid clearance and increased liver uptake for polycatecholamine functionalized drug nanocrystals. Using both surface sensitive model systems and protein corona profiling, we determine that the rapid clearance was correlated with an increase in adsorption of proteins involved in coagulation to the polycatecholamine surface, with fibrinogen being the most abundant. Further analysis of the most abundant proteins revealed a significant increase in thiol-rich proteins on polycatecholamine coated surfaces. The observed interaction with coagulation proteins highlights one of the current challenges using polycatecholamines for drug delivery but might also provide insights to the growing use of these materials in hemostatic applications.

Differential cellular responses to FDA-approved nanomedicines: an exploration of albumin-based nanocarriers and liposomes in protein corona formation.

Albumin nanoparticles (NPs) and PEGylated liposomes have garnered tremendous interest as therapeutic drug carriers due to their unique physicochemical properties. These unique properties also have significant effects on the composition and structure of the protein corona formed around these NPs in a biological environment. Herein, protein corona formation on albumin NPs and liposomes was simultaneously evaluated through in vitro and simulation studies. The sizes of both types of NPs increased with more negatively charged interfaces upon being introduced into fetal bovine serum. Gel electrophoresis and label-free quantitative proteomics were performed to identify proteins recruited to the hard corona, and fewer proteins were found in albumin NPs than in liposomes, which is in accordance with isothermal titration calorimetry. The cellular uptake efficiency of the two NPs significantly differed in different serum concentrations, which was further scrutinized by loading an anticancer compound into albumin NPs. The presence of the hard protein corona increased the cellular uptake of albumin NPs in comparison with liposomes. In our simulation study, a specific receptor present in the membrane was greatly attracted to the albumin-apolipoprotein E complex. Overall, this study not only evaluated protein corona formation on albumin NPs, but also made promising advancements toward albumin- and liposome-based therapeutic systems.

Real-Time Optical Tracking of Protein Corona Formation on Single Nanoparticles in Serum.

The formation of a protein corona, where proteins spontaneously adhere to the surface of nanomaterials in biological environments, leads to changes in their physicochemical properties and subsequently affects their intended biomedical functionalities. Most current methods to study protein corona formation are ensemble-averaging and either require fluorescent labeling, washing steps, or are only applicable to specific types of particles. Here we introduce real-time all-optical nanoparticle analysis by scattering microscopy (RONAS) to track the formation of protein corona in full serum, at the single-particle level, without any labeling. RONAS uses optical scattering microscopy and enables real-time and in situ tracking of protein adsorption on metallic and dielectric nanoparticles with different geometries directly in blood serum. We analyzed the adsorbed protein mass, the affinity, and the kinetics of the protein adsorption at the single particle level. While there is a high degree of heterogeneity from particle to particle, the predominant factor in protein adsorption is surface chemistry rather than the underlying nanoparticle material or size. RONAS offers an in-depth understanding of the mechanisms related to protein coronas and, thus, enables the development of strategies to engineer efficient bionanomaterials.

Plasma Parameters During Nanoparticle-Enhanced Laser-Induced Breakdown Spectroscopy (NELIBS) in the Presence of Nanoparticle-Protein Conjugates.

Nanoparticle-enhanced laser-induced breakdown spectroscopy (NELIBS) is an optical emission technique based on the laser-induced plasma (LIP) on a sample after the deposition of plasmonic nanoparticles (NPs) on its surface. The employment of the NPs allows an enhancement of the signal with respect to the one obtained with the conventional laser-induced breakdown spectroscopy (LIBS) enabling an extremely high sensitivity and very low limits of detection compared with the LIBS performance. Recently, NELIBS was used for monitoring the NP protein corona formation. As a matter of fact, the NPs in the presence of proteins adsorbed on the surface change their surface properties, therefore the sensing of protein corona formation was possible because of the strong dependence of NELIBS effects on the NP organization on the substrate, which in turn is deeply affected by the surface properties of the NPs. A correlation was found between NELIBS enhancement and the structure of the NP-protein conjugate in terms of protein content absorbed on the NP surface. An interesting question that was not yet exploited regards the role of LIP during the NELIBS when the NPs are covered with proteins. Since the presence of organic matter can strongly quench the LIP emission, the study of the LIP properties during protein corona sensing by NELIBS is of interest for two main reasons: (i) to understand whether the plasma parameters can vary in the presence of proteins adsorbed on the NP surface and (ii) to investigate how and if the plasma parameters themselves can influence the NELIBS processes. With this aim, the study of plasma parameters, i.e., electron densities and temperatures, during the sensing of NP protein corona by NELIBS is presented and discussed. The NPs used during these experiments were ultrapure gold NPs (AuNPs) produced by pulsed laser ablation in liquid, which are stable without any stabilizer. The human serum albumin protein is used to form AuNP-protein conjugates further deposited on a titanium target in NELIBS measurements. Dynamic light scattering, surface plasmon resonance spectroscopy, and laser Doppler electrophoresis for ζ-potential determination were employed to monitor the protein coverage of NP surface in the conjugate solutions before the NELIBS measurements.

Variations in metabolite profiles of serum coronas produced around PEGylated liposomal drugs by surface property.

Understanding biomolecular coronas that spontaneously occur around nanocarriers (NCs) in biological fluids is critical to nanomedicine as the coronas influence the behaviors of NCs in biological systems. In contrast to extensive investigations of protein coronas over the past decades, understanding of the coronas of biomolecules beyond proteins, e.g., metabolites, has been rather limited despite such biochemicals being ubiquitously involved in the coronas, which may influence the bio-nano interactions and thus exert certain biological impacts. In this study, serum biomolecular coronas, in particular the coronas of metabolites including lipids, around PEGylated doxorubicin-loaded liposomes with different surface property were investigated. The surface properties of liposomal drugs varied in terms of surface charge and PEGylation density by employing different ionic lipids such as DOTAP and DOPS and different concentrations of PEGylation lipids in liposome formulation. Using the liposomal drugs, the influence of the surface property on the serum metabolite profiles in the coronas was traced for target molecules of 220 lipids and 88 hydrophilic metabolites. From the results, it was found that metabolites rather than proteins mainly constitute the serum coronas on the liposomal drugs. Most of the serum metabolites were found to be retained in the coronas but with altered abundances. Depending on their class, lipids exhibited a different dependence on the surface property. However, overall, lipids appeared to favor corona formation on more negatively charged and PEGylated surfaces. Hydrophilic metabolites also exhibited a similar propensity for corona formation. This study on the surface dependence of metabolite corona formation provides a fundamental contribution toward attaining a comprehensive understanding of biomolecular coronas, which will be critical to the development of efficient nanomedicine.

Towards a simple in vitro surface chemistry pre-screening method for nanoparticles to be used for drug delivery to solid tumours.

An efficient nanoparticulate drug carrier intended for chemotherapy based on intravenous administration must exhibit a long enough blood circulation time, a good penetrability into the tumour volume, as well as an efficient uptake by cancer cells. Limiting factors for the therapeutic outcome in vivo are recognition of the nanoparticles as foreign objects, which triggers nanoparticle uptake by defence organs rich in macrophages, e.g. liver and spleen, on the time-scale of accumulation and uptake in/by the tumour. However, the development of nanomedicine towards efficient nanoparticle-based delivery to solid tumours is hampered by the lack of simple, reproducible, cheap, and predictive means for early identification of promising nanoparticle formulations. The surface chemistry of nanoparticles is known to be the most important determinant for the biological fate of nanoparticles, as it influences the extent of serum protein adsorption, and also the relative composition of the protein corona. Here we preliminarily evaluate an extremely simple screening method for nanoparticle surface chemistry pre-optimization based on nanoparticle uptake in vitro by PC-3 cancer cells and THP-1 macrophages. Only when both selectivity for the cancer cells as well as the extent of nanoparticle uptake are taken into consideration do the in vitro results mirror literature results obtained for small animal models. Furthermore, although not investigated here, the screening method does also lend itself to the study of actively targeted nanoparticles.

An In Situ Investigation of the Protein Corona Formation Kinetics of Single Nanomedicine Carriers by Self-Regulated Electrochemiluminescence Microscopy.

Protein coronas are present extensively at the bio-nano interface due to the natural adsorption of proteins onto nanomaterials in biological fluids. Aside from the robust property of nanoparticles, the dynamics of the protein corona shell largely define their chemical identity by altering interface properties. However, the soft coronas are normally complex and rapidly changing. To real-time monitor the entire formation, we report here a self-regulated electrochemiluminescence (ECL) microscopy based on the interaction of the Ru(bpy)3 3+ with the nanoparticle surface. Thus, the heterogeneity of the protein corona is in situ observed in single nanoparticle "cores" before and after loading drugs in nanomedicine carriers. The label-free, optical stable and dynamic ECL microscopy minimize misinterpretations caused by the variation of nanoparticle size and polydispersity. Accordingly, the synergetic actions of proteins and nanoparticles properties are uncovered by chemically engineered protein corona. After comparing the protein corona formation kinetics in different complex systems and different nanomedicine carriers, the universality and accuracy of this technique were well demonstrated via the protein corona formation kinetics curves regulated by competitive adsorption of Ru(bpy)3 3+ and multiple proteins on surface of various carriers. The work is of great significance for studying bio-nano interface in drug delivery and targeted cancer treatment.