Adsorbed Protein Layer Research Articles

Utilizing a hydrophobic primary container surface to reduce the formation of subvisible particles in monoclonal antibody solution caused by fluid shear

The exposure of protein molecules to interfaces may cause protein aggregation and particle formation in protein formulations, especially hydrophobic interfaces, which may promote protein aggregation in solution. In this study, we found that modification of the surface properties by application of a hydrophobic Octadecyltrichlorosilane (OTS) could reduce the generation of protein aggregates and particles in protein solution induced by fluid shear. A stable protein adsorption layer was formed at the hydrophobic interface through the strong hydrophobic interaction between the protein and hydrophobic surface, which could prevent the aggregated protein from falling off into the bulk solution to form subvisible particles and insoluble protein aggregates. In addition, human complement enzyme linked immunosorbent assay results showed that the particles that were generated in the OTS-coated container did not activate human complement which indicated the OTS-coated container could be used as primary containers for certain types of monoclonal antibody formulation.

Tannin-on-protein adlayers: Secondary adsorption quantified by quartz crystal microbalance with dissipation monitoring (QCM-D)

The quartz crystal microbalance with dissipation monitoring (QCM-D) technique allows to detect adsorption and mechanical characteristics of protein adsorption layers at solid surfaces, including subsequent secondary adsorption of additional species interacting with the proteins. Here, we study the secondary adsorption of two different classes of tannins - hydrolysable vs. condensed - onto a β-lactoglobulin base layer using QCM-D in a protein/tannin adlayer configuration. Sequential flow experiments of protein solutions, tannin solutions and neat solvent or buffer are used to distinguish reversible from desorption-resistant layer formation, evaluate the viscoelastic characteristics the layers, and map protein/tannin interactions in view of their structural properties and origin. We find that for the majority of tannins, secondary layer structural changes (detected via a change in the dissipation factor) occur before significant mass adsorption (detected via the resonance frequency shift). In contrast, simultaneous onset of structural change and adsorbed mass within an observation window of 100 ms was only found for certain hydrolysable tannins. We use these data to build adsorption maps and group the tannins according to their adlayer properties, providing a physical basis for an improved understanding of the organoleptic perception of tannins and polyphenols, in particular with respect to attributes such as ‘astringency’ and ‘lingering’. Moreover, these maps provide insight into the capacity of different tannins to modify the mechanical properties of protein adsorption layers.

Interaction of Serum and Plasma Proteins with Polyelectrolyte Microparticles with Core/Shell and Shell-Only Structures.

Polyelectrolyte microparticles (MPs) synthesized on calcium carbonate cores are considered a promising basis for new drug delivery systems. It is known that microparticles entering a physiological environment absorb proteins on their surface, which can change the properties of the microparticles and alter their functional activity. This study aimed to compare the compositions of the adsorbed protein layer formed on microparticles with the core/shell and shell structures obtained by layer-by-layer deposition. The difference in the microparticle structure was associated with changes in their surface topography and ζ-potential. These microparticles were incubated with human serum or plasma at 37°C for 24 h. The adsorbed proteins were eluted and analyzed by means of SDS-PAGE. The protein composition of the eluates was determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS); a total of 357 proteins were identified, and 183 of them were detected in all samples. Our results demonstrate that the relative abundance of proteins of different functional groups (immunoglobulins, complement proteins, and apolipoproteins) varied depending on the structure and surface characteristics of the polyelectrolyte microparticles and the incubation medium. Our findings expand the understanding of the influence of the physicochemical properties of the microparticles on their interaction with proteins, which can help to improve the design of microparticles for drug delivery.

Exploring proteins at soft interfaces and in thin liquid films – From classical methods to advanced applications of reflectometry

The history of the topic of proteins at soft interfaces dates back to the 19th century, and until the present day, it has continuously attracted great scientific interest. A multitude of experimental methods and theoretical approaches have been developed to serve the research progress in this large domain of colloid and interface science, including the area of soft colloids such as foams and emulsions. From classical methods like surface tension adsorption isotherms, surface pressure-area measurements for spread layers, and surface rheology probing the dynamics of adsorption, nowadays, advanced surface-sensitive techniques based on spectroscopy, microscopy, and the reflection of light, X-rays and neutrons at liquid/fluid interfaces offers important complementary sources of information. Apart from the fundamental characteristics of protein adsorption layers, i.e., surface tension and surface excess, the nanoscale structure of such layers and the interfacial protein conformations and morphologies are of pivotal importance for extending the depth of understanding on the topic. In this review article, we provide an extensive overview of the application of three methods, namely, ellipsometry, X-ray reflectometry and neutron reflectometry, for adsorption and structural studies on proteins at water/air and water/oil interfaces. The main attention is placed on the development of experimental approaches and on a discussion of the relevant achievements in terms of notable experimental results. We have attempted to cover the whole history of protein studies with these techniques, and thus, we believe the review should serve as a valuable reference to fuel ideas for a wide spectrum of researchers in different scientific fields where proteins at soft interface may be of relevance.

Physicochemical Quantitative Analysis of the Oil-Water Interface as Affected by the Mutual Interactions between Pea Protein Isolate and Mono- and Diglycerides.

As a commercially available ingredient, the mono- and diglycerides (MDG) were widely used in a plant protein-based emulsion to provide effective, functional, emulsifying properties. The simultaneous addition of the MDG and pea protein isolate (PPI) was investigated by the methods of interfacial rheology and quantitative protein proteomics. The physicochemical quantitative analysis of the oil-water interface revealed an interfacial stability mechanism for the protein adsorption layer. For a low MDG concentration, the interfacial quantities of vicilin and albumin were increased, which could be attributed to the adsorption rate. For a high MDG concentration, both vicilin and albumin were displaced by MDG and desorbed from the interface, while legumin was more difficult to displace due to its slow adsorption and the complex structure of protein molecules. The protein molecules with the structural rearrangement interacted with MDG, exhibiting potential effects on the interfacial film structure. Combined with some nanotechnologies, the new comprehension of protein-emulsifier interactions may promote food delivery systems. The research aims to develop an in-depth analysis of interfacial proteins, and provide more innovative and tailored functionalities for the application of the plant protein emulsion.

Biomimetic affinity membrane roll column for rapid purification of C-reactive protein

To in-depth explore the action mechanism of C-reactive protein (CRP) and precisely study its signaling pathways, it is essential to acquire high-purity CRP while preserving its intact structure and functionality. In this study, we propose and fabricate a high-density 2-methacryloyloxyethyl phosphorylcholine (MPC)-modified membrane roll column (MPC-MRC) using a surface-initiated atom transfer radical polymerization (SI-ATRP) approach, which can overcome these limitations (long incubation time and low adsorption capacity) of conventional enrichment materials. The MPC-MRC incorporates a high-density 2-hydroxyethyl methacrylate polymer brush to prevent non-specific protein adsorption and multiple MPC polymer brush layers for high-performance enrichment of CRP in the company of calcium ions. Furthermore, the MPC-MRC exhibits high permeability, hydrophilicity, and mechanical strength. Compared to previous technologies, this novel material demonstrates significantly higher CRP binding capacity (310.3 mg/g), shorter processing time (only 15 min), and lower cost (only 12 USD/column). Notably, the MPC-MRC enables fast and effective purification of CRP from both human and rat serum, exhibiting good selectivity, recovery (> 91.3 %), and purity (> 95.2 %). Thus, this proposed purification approach based on MPC-MRC holds great potential for target protein enrichment from complex samples, as well as facilitating in-depth studies of its biological functions.

ATR FTIR spectroscopy of protein and surfactant adsorption to the oil-water interface

An oil-infused polyelectrolyte multilayer (PEM) immersed in aqueous solution is a valid mimic for an oil-water interface, that can be used to probe adsorption of surfactants (including co-/competitive adsorption) using attenuated total reflection Fourier transform infrared spectroscopy (ATR FTIR). A PEM has been formed on an ATR internal reflection element, dried and then re-solvated with oil, then exposed to a flowing solution of protein, and subsequently a solution of surfactant. The PEM had undergone prior characterization with atomic force microscopy (AFM). The experiment was replicated using profile analysis tensiometry (PAT). The PEM displays a high degree of solvation with oil; it retains most of this oil in the presence of flowing aqueous solution. FTIR spectra confirmed adsorption of a dairy protein (β-lactoglobulin) to the medium chain triglyceride (MCT)-water interface, and the response of the adsorbed protein layer to a common food formulation surfactant (Tween 60) in the aqueous phase above the interface. The protein is observed to adsorb strongly and irreversibly to the MCT oil-water interface, and the Tween 60 molecules are seen to co-adsorb rather than displace the protein. This observation is supported by measurements of sequential co-adsorption undertaken with PAT.

MyD88-dependent Toll-like receptor 2 signaling modulates macrophage activation on lysate-adsorbed Teflon™ AF surfaces in an in vitro biomaterial host response model.

The adsorbed protein layer on an implanted biomaterial surface is known to mediate downstream cell-material interactions that drive the host response. While the adsorption of plasma-derived proteins has been studied extensively, the adsorption of damage-associated molecular patterns (DAMPs) derived from damaged cells and matrix surrounding the implant remains poorly understood. Previously, our group developed a DAMP-adsorption model in which 3T3 fibroblast lysates were used as a complex source of cell-derived DAMPs and we demonstrated that biomaterials with adsorbed lysate potently activated RAW-Blue macrophages via Toll-like receptor 2 (TLR2). In the present study, we characterized the response of mouse bone marrow derived macrophages (BMDM) from wildtype (WT), TLR2-/- and MyD88-/- mice on Teflon™ AF surfaces pre-adsorbed with 10% plasma or lysate-spiked plasma (10% w/w total protein from 3T3 fibroblast lysate) for 24 hours. WT BMDM cultured on adsorbates derived from 10% lysate in plasma had significantly higher gene and protein expression of IL-1β, IL-6, TNF-α, IL-10, RANTES/CCL5 and CXCL1/KC, compared to 10% plasma-adsorbed surfaces. Furthermore, the upregulation of pro-inflammatory cytokine and chemokine expression in the 10% lysate in plasma condition was attenuated in TLR2-/- and MyD88-/- BMDM. Proteomic analysis of the adsorbed protein layers showed that even this relatively small addition of lysate-derived proteins within plasma (10% w/w) caused a significant change to the adsorbed protein profile. The 10% plasma condition had fibrinogen, albumin, apolipoproteins, complement, and fibronectin among the top 25 most abundant proteins. While proteins layers generated from 10% lysate in plasma retained fibrinogen and fibronectin among the top 25 proteins, there was a disproportionate increase in intracellular proteins, including histones, tubulins, actins, and vimentin. Furthermore, we identified 7 DAMPs or DAMP-related proteins enriched in the 10% plasma condition (fibrinogen, apolipoproteins), compared to 39 DAMPs enriched in the 10% lysate in plasma condition, including high mobility group box 1 and histones. Together, these findings indicate that DAMPs and other intracellular proteins readily adsorb to biomaterial surfaces in competition with plasma proteins, and that adsorbed DAMPs induce an inflammatory response in adherent macrophages that is mediated by the MyD88-dependent TLR2 signaling pathway.

PH-induced egg white protein foaming properties enhancement: Insight into protein structure and quantitative proteomic analysis at protein adsorption layer

The foaming properties of egg white protein (EWP) after pH-induced unfolding (pH 2, pH 4, pH 10 and pH 12) and refolding treatment (pH 2–7, pH 4–7, pH 10–7 and pH 12-7 (adjusting the above pH back to 7)) were studied by structure changes and quantitative proteomic analysis. It was found that all pH treatment improved foaming capacity (FC) and foaming stability (FS) of EWP, especially at pH 2 and pH 12, where EWP was in a molten globule state with unfolded structure (especially at pH 12), high surface hydrophobicity (except for pH 12-treated EWP), more free sulfhydryl groups and up-regulated adsorption proteins. However, adjusting the pH back to 7 limited the foaming properties due to protein refolding. Meanwhile the presence of down-regulated proteins was another reason for relatively poor FC at pH 2–7. Notably, the firmest foam and high FS at pH 12-7 were attributed to the generation of aggregates and the long-term stability of adsorbed proteins. The results of quantitative proteomics implied that lysozyme, ovalbumin and protein TENP were identified as key proteins associated with excellent FC at pH 2, pH 12 and pH 12-7, while β-ovomucin contributed the most to FC at pH 2–7. Moreover, protein TENP and ovomucoid were more relevant to maintaining and reducing FS. Overall, this work elucidated the mechanism of pH treatment to enhance the foaming properties of EWP, which would provide the necessary theoretical support for the quality improvement and product development of EWP powder.

Investigation on Human Serum Protein Depositions Inside Polyvinylidene Fluoride-Based Dialysis Membrane Layers Using Synchrotron Radiation Micro-Computed Tomography (SR-μCT).

Hemodialysis (HD) membrane fouling with human serum proteins is a highly undesirable process that results in blood activations with further severe consequences for HD patients. Polyvinylidene fluoride (PVDF) membranes possess a great extent of protein adsorption due to hydrophobic interaction between the membrane surface and non-polar regions of proteins. In this study, a PVDF membrane was modified with a zwitterionic (ZW) polymeric structure based on a poly (maleic anhydride-alt-1-decene), 3-(dimethylamino)-1-propylamine derivative and 1,3-propanesultone. Fourier transform infrared spectroscopy (FTIR), scanning electron microscope (SEM), and zeta potential analyses were used to determine the membrane's characteristics. Membrane fouling with human serum proteins (human serum albumin (HSA), fibrinogen (FB), and transferrin (TRF)) was investigated with synchrotron radiation micro-computed tomography (SR-μCT), which allowed us to trace the protein location layer by layer inside the membrane. Both membranes (PVDF and modified PVDF) were detected to possess the preferred FB adsorption due to the Vroman effect, resulting in an increase in FB content in the adsorbed protein compared to FB content in the protein mixture solution. Moreover, FB was shown to only replace HSA, and no significant role of TRF in the Vroman effect was detected; i.e., TRF content was nearly the same both in the adsorbed protein layer and in the protein mixture solution. Surface modification of the PVDF membrane resulted in increased FB adsorption from both the protein mixture and the FB single solution, which is supposed to be due to the presence of an uncompensated negative charge that is located at the COOH group in the ZW polymer.

Producing mixed-soy protein adsorption layers on alginate microgels to controlled-release β-carotene

In this study, the effects of soy protein isolate (SPI) on the morphology, encapsulation efficiency, storage stability, swelling behavior, and in vitro digestion behavior of calcium alginate (CA) microgels were investigated. CA and calcium alginate-SPI (CAS) microgels with encapsulated β-carotene were prepared by extruding a mixture of alginate and SPI using a co-extrusion technique, followed by cross-linking with Ca2+. All microgels exhibited homogeneous sizes and spherical shapes, and CAS microgels showed high levels of protein loading efficiency. The encapsulation efficiency and storage stability of β-carotene within CAS microgels were higher than those within CA microgels. The introduction of SPI into CAS microgels resulted in a higher degree of gel size shrinkage in gastric fluid and a lower degree of swelling in intestinal fluid compared to CA microgels. In vitro digestion was conducted to investigate the effects of the addition of SPI on the release behavior of CA and CAS microgels. Results obtained showed that CAS microgels were more resistant to simulated gastric fluid than CA microgels. Cryo-scanning electron microscopy (cryo-SEM) and confocal laser scanning microscopy (CLSM) observations indicated that the release behavior was dependent on the porosity of the CA and CAS microgels, and the porosity was influenced by the concentration of SPI. This study showed that the introduction of SPI to CA microgels can lead to the development of an effective controlled release delivery system.

Exploiting the size exclusion effect of protein adsorption layers for electrochemical detection of microRNA: A new mechanism for design of E-DNA sensor

The assay performance of electrochemical DNA (E-DNA) sensors is deeply influenced by the state of DNA probes immobilized on electrode. Moreover, the immobilization procedures for DNA probes are tedious and vary according to the probes and analytes. In this work, we find that the adsorption layers of bovine serum albumin (BSA) on gold electrode (AuE) possess a size exclusion effect to distinguish between single-stranded (-ss) DNA probes and the DNA fragments generated from enzymatic digestion of ssDNA probes. In detail, the BSA layers act as a gatekeeper that hinders the adsorption of a ssDNA probe on AuE but permits the DNA fragments with much smaller sizes to pass through the adsorption layers and adsorb on AuE. This finding is developed into a novel E-DNA sensor for microRNA (miRNA) detection by coupling with duplex-specific nuclease (DSN)-assisted target recycling strategy. The ssDNA probe in solution phase is enzymatically digested during the DSN-assisted target recycling process initiated by target miRNA-21, generating plenty of DNA fragments. The adsorption of these DNA fragment on BSA/AuE is permitted, which arouses electrochemical signals after binding with [Ru(NH3)6]3+ to indicate the recognition of miRNA-21. The developed E-DNA sensor possesses a wide calibration range from 0.001 to 100 pM and a low detection limit of 0.48 fM. Significantly, accurate evaluation of miRNA-21 expression levels in cancer cell lines and non-small-cell lung carcinomas (NSCLC) serum samples are successfully achieved using the developed method. This work provides a new mechanism for constructing sensitive E-DNA sensor without tedious probe immobilization procedures.

Optimizing the Porohyperelastic Response of a Layered Compliance Matched Vascular Graft to Promote Luminal Self-Cleaning.

Thrombosis and intimal hyperplasia have remained the major failure mechanisms of small-diameter vascular grafts used in bypass procedures. While most efforts to reduce thrombogenicity have used a biochemical surface modification approach, the use of local mechanical phenomena to aid in this goal has received somewhat less attention. In this work, the mechanical, fluid transport, and geometrical properties of a layered and porous vascular graft are optimized within a porohyperelastic finite element framework to maximize self-cleaning via luminal reversal fluid velocity (into the lumen). This is expected to repel platelets as well as inhibit the formation of and/or destabilize adsorbed protein layers thereby reducing thrombogenic potential. A particle swarm optimization algorithm was utilized to maximize luminal reversal fluid velocity while also compliance matching our graft to a target artery (rat aorta). The maximum achievable luminal reversal fluid velocity was approximately 246 μm/s without simultaneously optimizing for host compliance. Simultaneous optimization of reversal flow and compliance resulted in a luminal reversal fluid velocity of 59 μm/s. Results indicate that a thick highly permeable compressible inner layer and a thin low permeability incompressible outer layer promote intraluminal reversal fluid velocity. Future research is needed to determine the feasibility of fabricating such a layered and optimized graft and verify its ability to improve hemocompatibility.

Highly Luminescent Positively Charged Quantum Dots Interacting with Proteins and Cells†

Comprehensive SummaryWe have studied interactions between positively charged MUTAB‐stabilized quantum dots (QDs) and model proteins, serum and live cells using fluorescence correlation spectroscopy (FCS), dynamic light scattering (DLS), time‐resolved photoluminescence (PL) and live‐cell fluorescence imaging. Using human serum albumin (HSA) as a model protein, we measured the growth of a protein adsorption layer (“protein corona”) via time‐resolved FCS. Corona formation was characterized by an apparent equilibrium dissociation coefficient, KD ≈ 10 μM. HSA adlayer growth was surprisingly slow (timescale ca. 30 min), in stark contrast to many similar measurements with HSA and other proteins and different NPs. Time‐resolved PL data revealed a characteristic quenching behavior depending on the QD surface coverage with HSA. Taken together, we found that MUTAB‐QDs initially bind HSA molecules weakly (KD ≈ 700 μM); however, the affinity is enhanced over time, presumably due to proton injection into the MUTAB layer by HSA triggering ligand dissociation. This process was also observed with human blood serum, showing equal kinetics for comparable HSA concentration. Moreover, imaging experiments with cultured human cells (HeLa) revealed that MUTAB‐QDs bind to the cell membrane and perforate it. This process is reduced upon pre‐adsorption of proteins on the MUTAB‐QD surfaces.

Poly(ethylene glycol)-Based Surfactant Reduces the Conformational Change of Adsorbed Proteins on Nanoparticles.

When in contact with a biological medium, the surfaces of nanoparticles are usually covered by proteins. In this regard, it was found that poly(ethylene glycol) (PEG) promotes the “stealth effect”. This implies a reduction of unspecific protein adsorption and cellular uptake. Although information about the PEG–protein interaction was reported, more accurate and sophisticated structure and dynamics analyses are needed to understand the interaction processes in detail. This work studies the PEG–protein interaction using model nanoparticles stabilized either by the PEG-based surfactant Lutensol AT50 or sodium dodecyl sulfate. The interaction with human serum albumin was studied using neutron scattering techniques. The parameters obtained by small-angle neutron scattering yielded information about the adsorbed protein layer thickness. Protein structure changes were detected via differential scanning fluorimetry and elastic neutron scattering. This combination gives a better insight into the PEG–protein interaction, contributing to the design of nanomaterials for medical applications.

A preliminary experimental investigation on the biotribocorrosion of a metal-on-polyethylene hip prosthesis in a hip simulator

Corrosion at the taper/trunnion interface of total hip replacement (THR) often results in severe complications. However, the underlying mechanisms of biotribocorrosion at the taper/trunnion interface during the long-term walking gait cycles remain to be fully understood. In this study, a hip joint simulator was therefore instrumented with an electrochemical cell for in-situ monitoring of the tribocorrosion evolution in a metal-on-polyethylene (MoP) THR during a typical long-term walking gait. In addition, the biotribocorrosion mechanism was investigated via surface and chemical characterizations. The experimental results confirmed that the taper/trunnion interface dominated the contemporary MoP hip joint corrosion. Three cyclic variations in the open circuit potential (OCP) were observed throughout the long-term electrochemical measurements, attributed to the formation and disruption of the adsorbed protein layer. The corrosion exhibited an initial increase at each period, peaking at approximately 0.125 million cycles, followed by a subsequent gradual reduction. Surface and chemical analyses revealed the formation of a tribochemical reaction layer (tribolayer) on the worn surface of the taper/trunnion interface. The surface/chemical characterizations and the electrochemical measurements indicated that the adhesion force of the adsorbed protein layer was weaker than that of the tribolayer. In contrast, the opposite was true for the corrosion resistance. Based on the observations from this study, the tribocorrosion mechanism of the taper/trunnion interface under the long-term walking gait cycles is deduced.

The extended effect of adsorbed damage-associated molecular patterns and Toll-like receptor 2 signaling on macrophage-material interactions.

Implanted biomaterials elicit an immune-mediated foreign body reaction (FBR) that results in the fibrous encapsulation of the implant and can critically impact the performance of some implants. Consequently, understanding the molecular mechanisms that underpin cell-materials interactions that initiate biomaterial-induced inflammation and fibrosis is critical to improving the performance of biomaterial implants negatively impacted by the FBR. Damage-associated molecular patterns (DAMPs) are endogenous mediators of inflammation that are released upon tissue injury and induce sterile inflammation via Toll-like receptors (TLRs). However, the prevalence of DAMPs within the adsorbed protein layer on material surfaces and their role mediating cell-material interactions is unclear. Previously, our group demonstrated that molecules in fibroblast lysates adsorbed to various biomaterials and induced a potent TLR2-dependent inflammatory response in macrophages at 24 h. In this study, we examined the extended response of RAW-Blue reporter macrophages on lysate or serum-adsorbed Teflon™ AF surfaces to understand the potential role of adsorbed DAMPs in macrophage-material interactions at later time points. Lysate-conditioned surfaces maintained increased nuclear factor kappa B (NF-κB) and activator protein 1 (AP-1) transcription factor activity and increased expression Regulated upon Activation, Normal T Cell Expressed and Presumably Secreted (RANTES/CCL5) at 72 h and 120 h, compared to FBS-conditioned surfaces. In contrast, monocyte chemoattractant protein 1 (MCP-1/CCL2) was only elevated at 72 h in lysate conditions. Transforming growth factor beta 1 (TGF-β1) secretion was significantly increased on lysate-conditioned surfaces, while conditioned media from macrophages on lysate-conditioned surfaces induced alpha smooth muscle actin (αSMA) expression in 3T3 fibroblasts. TLR2 neutralizing antibody treatment significantly decreased NF-κB/AP-1 activity and attenuated TGF-β1 expression at both time points, and MCP-1 and RANTES at 72 h. Finally, multinucleated cells were observed on lysate-conditioned surfaces at 72 h, indicating adsorbed DAMPs induced a fusion permissive environment for adherent macrophages. This study demonstrates that adsorbed DAMPs continue to influence macrophage-material responses beyond the initial 24-h period and maintain a pro-inflammatory and fibrotic response that models aspects of the early FBR. Furthermore, the transient inhibition of TLR2 continued to exert an effect at these later time points, suggesting TLR2 may be a target for therapeutic interventions in FBR.

Human Vimentin Layers on Solid Substrates: Adsorption Kinetics and Corona Formation Investigations.

Adsorption kinetics of human vimentin on negatively charged substrates (mica, silica, and polymer particles) was analyzed using atomic force microscopy (AFM), quartz microbalance (QCM), and the laser doppler velocimetry (LDV) method. AFM studies realized under diffusion conditions proved that the adsorbed protein layer mainly consisted of aggregates in the form of compact tetramers and hexamers of a size equal to 11–12 nm. These results were consistent with vimentin adsorption kinetics under flow conditions investigated by QCM. It was established that vimentin aggregates efficiently adsorbed on the negatively charged silica sensor at pH 3.5 and 7.4, forming compact layers with the coverage reaching 3.5 mg m–2. Additionally, the formation of the vimentin corona at polymer particles was examined using the LDV method and interpreted in terms of the electrokinetic model. This allowed us to determine the zeta potential of the corona as a function of pH and the electrokinetic charge of aggregates, which was equal to −0.7 e nm–2 at pH 7.4 in a 10 mM NaCl solution. The anomalous adsorption of aggregates exhibiting an average negative charge on the negatively charged substrates was interpreted as a result of a heterogeneous charge distribution. These investigations confirmed that it is feasible to deposit stable vimentin layers both at planar substrates and at carrier particles with well-controlled coverage and zeta potential. They can be used for investigations of vimentin interactions with various ligands including receptors of the innate immune system, immunoglobulins, bacterial virulence factors, and spike proteins of viruses.

The effects of thermal treatment on emulsifying properties of soy protein isolates: Interfacial rheology and quantitative proteomic analysis

In this work, the interfacial rheological properties and the quantitative changes of proteins at interfacial protein layers of emulsions stabilized by soy protein isolates (SPI) and heat-treated soy protein isolates (HSPI) were investigated. The quantification results showed that the relative quantities of albumin (2S) and glycinin (11S) in SPI decreased at the oil-water interface, suggesting that they possessed lower interfacial affinities at the interface. Basic 7S globulin presented more adsorption at the oil-water interface due to the well balance of the hydrophobic and hydrophilic groups of its amino acid sequence. The HSPI (95 °C, 20 min) showed a larger apparent diffusion rate (Kdiff) and a shorter equilibrium adsorption time. The results of interfacial rheology of globulins were consistent with their interfacial quantitative changes, which demonstrated that the interfacial behavior and adsorption ability of globulin were improved by thermal treatment. In this research, the interfacial behaviors of SPI and HSPI was illustrated by their interfacial properties and quantitative results of interfacial adsorbed protein layers, would promote a profound comprehension for the interfacial behavior of the protein and the influence of thermal treatment on protein interfacial properties.

Adsorption of lysozyme and antibodies at material surfaces: Implications to material compatibility for development of biologics

The impacts of contact materials such as borosilicate glass, polycarbonate (PC) and cellulose, on the adsorption of lysozyme, along with the interaction of an immunoglobulin (IgG) with polyethersulfone (PES)-typical filter material, were evaluated using quartz crystal microbalance with dissipation (QCM-D). The adsorption and viscoelastic properties of the adsorbed protein layers, along with displacement and adsorption prevention by polysorbate 80 (PS80) and poloxamer 188, were characterized based on frequency and dissipation change. A decrease in frequency denotes a mass gain from adsorption, while an increase in dissipation indicates formation of a viscoelastic layer. The adsorption at a fixed lysozyme concentration increases in the following order: borosilicate glass > PC > cellulose. Furthermore, the adsorbed protein layers on PC surface appears to be the most viscoelastic, whereas the adsorbed layer at borosilicate seems to be the least viscoelastic. Finally, adsorption at interfaces is irreversible and cannot be completely displaced by PS80 or poloxamer 188. However, the adsorption of IgG on PES was shown to be effectively prevented when PS80 is present in the formulation. These results provide insights in understanding material compatibilities during manufacturing, storage and administration of therapeutic proteins in biopharmaceutical industry.