Effects of ionic strength and surface charge on protein adsorption at PEGylated surfaces

PEGylated Nb2O5 surfaces were obtained by the adsorption of poly(L-lysine)-g-poly(ethylene glycol) (PLL-g-PEG) copolymers, allowing control of the PEG surface density, as well as the surface charge. PEG (MW 2 kDa) surface densities between 0 and 0.5 nm(-2) were obtained by changing the PEG to lysine-mer ratio in the PLL-g-PEG polymer, resulting in net positive, negative and neutral surfaces. Colloid probe atomic force microscopy (AFM) was used to characterize the interfacial forces associated with the different surfaces. The AFM force analysis revealed interplay between electrical double layer and steric interactions, thus providing information on the surface charge and on the PEG layer thickness as a function of copolymer architecture. Adsorption of the model proteins lysozyme, alpha-lactalbumin, and myoglobin onto the various PEGylated surfaces was performed to investigate the effect of protein charge. In addition, adsorption experiments were performed over a range of ionic strengths, to study the role of electrostatic forces between surface charges and proteins acting through the PEG layer. The adsorbed mass of protein, measured by optical waveguide lightmode spectroscopy (OWLS), was shown to depend on a combination of surface charge, protein charge, PEG thickness, and grafting density. At high grafting density and high ionic strength, the steric barrier properties of PEG determine the net interfacial force. At low ionic strength, however, the electrical double layer thickness exceeds the thickness of the PEG layer, and surface charges "shining through" the PEG layer contribute to protein interactions with PLL-g-PEG coated surfaces. The combination of AFM surface force measurements and protein adsorption experiments provides insights into the interfacial forces associated with various PEGylated surfaces and the mechanisms of protein resistance.     

Detergent-Induced Removal of β-Lactoglobulin from Stainless Steel Surfaces as Influenced by Surface Pretreatment

The adsorption of β-lactoglobulin to stainless steel and its subsequent removal were followed using in situ null ellipsometry. The influence of the surface pretreatment on the protein removal by the surfactant SDS and by sodium hydroxide was studied. All surfaces were precleaned in strongly alkaline solution. Some surfaces received no further pretreatment, while others were either passivated in nitric acid or plasma-cleaned prior to experiments. Stainless steel surfaces subjected to different surface pretreatments showed considerable differences in cleaning behavior. Cleaning, using NaOH, of surfaces which had been precleaned with alkali only or with plasma resulted in practically complete β-lactoglobulin removal. In contrast, appreciable amounts of protein remained on passivated stainless steel. Protein removal by SDS was limited and comparable for all three surface pretreatments investigated. Only minor effects on the protein adsorption tendency were observed. The amounts of β-lactoglobulin adsorbed tended to be somewhat lower on the passivated surfaces.     

Fibronectin adsorption onto polyelectrolyte multilayer films

The Layer-by-layer deposition of positively and negatively charged macromolecular species is an ideal method for constructing thin films incorporating biological molecules. We investigate the adsorption of fibronectin onto polyelectrolyte multilayer (PEM) films using optical waveguide lightmode spectroscopy (OWLS) and atomic force microscopy (AFM). PEM films are formed by adsorption onto Si(Ti)O2 from alternately introduced flowing solutions of anionic poly(sodium 4-styrenesulfonate) (PSS) and cationic poly(allylamine hydrochloride) (PAH). Using OWLS, we find the initial rate and overall extent offibronectin adsorption to be greatest on PEM films terminated with a PAH layer. The polarizability density of the adsorbed protein layer, as measured by its refractive index, is virtually identical on both PAH- and PSS-terminated films; the higher adsorbed density on the PAH-terminated film is due to an adsorbed layer of roughly twice the thickness. The binding of monoclonal antibodies specific to the protein's cell binding site is considerably enhanced to fibronectin adsorbed to the PSS layer, indicating a more accessible adsorbed layer. With increased salt concentration, we find thicker PEM films but considerably thinner adsorbed fibronectin layers, owing to increased electrostatic screening. Using AFM, we find adsorbed fibronectin layers to contain clusters; these are more numerous and symmetric on the PSS-terminated film. By considering the electrostatic binding of a segmental model fibronectin molecule, we propose a picture of fibronectin adsorbed primarily in an end-on-oriented monolayer on a PAH-terminated film and as clusters plus side-on-oriented isolated molecules onto a PSS-terminated film.     

Photosensitive Azopolymer Brushes via Atom Transfer Radical Polymerization for Protein Sensing

Novel photosensitive azopolymer brushes were synthesized via surface initiated atom transfer radical polymerization using initiator self‐assembled on Au surface. The chemical structures of azobenzene derivatives were confirmed by Fourier transform infrared spectroscopy (FTIR) and nuclear magnetic resonance spectroscopy (NMR). The surface morphology of azopolymers via atom transfer radical polymerization (ATRP) for different time was investigated by atomic force microscopy (AFM). Additionally, the photoisomerization of azopolymer was measured by ultraviolet‐visible spectroscopy (UV‐Vis). The results indicate that such azopolymers can undergo trans‐cis‐trans photoisomerization efficiently by photo‐irradiation with UV light. Furthermore, this photoisomerization property could also induce the reversible adsorption of bovine serum albumin (BSA) adsorption on azopolymer brush surfaces. This adsorption kinetics of the reversible process can be measured by surface plasmon resonance (SPR) spectroscopy in situ. It suggests that the protein biochips could be regenerated safely by UV irradiation rather than by being rinsed with chemical reagents.     

Protein resistance of titanium oxide surfaces modified by biologically inspired mPEG-DOPA

In the present study, we have utilized X-ray photoelectron spectroscopy (XPS), spectroscopic ellipsometry (ELM), and optical waveguide lightmode spectroscopy (OWLS) to examine the surface adsorption and protein resistance behavior of bio-inspired polymers consisting of poly(ethylene glycol) (PEG) conjugated to peptide mimics of mussel adhesive proteins. Peptides containing up to three residues of 3,4-dihydroxyphenylalanine (DOPA), a key component of mussel adhesive proteins, were conjugated to monomethoxy-terminated PEG polymers. These mPEG-DOPA polymers were found to be highly adhesive to TiO2 surfaces, with quantitative XPS analysis providing useful insight into the binding mechanism. Additionally, the antifouling properties of immobilized PEG were reflected in the excellent resistance of mPEG-DOPA-modified TiO2 surfaces to protein adsorption. Measurements of mPEG-DOPA and human serum adsorption were related in terms of ethylene glycol (EG) surface density and serum mass adsorbed and demonstrated a threshold of approximately 15-20 EG/nm2, above which substantially little protein adsorbs. With respect to surface density of adsorbed PEG and the associated nonfouling behavior of the adlayers, strong parallels exist between the nonfouling properties of the surface-bound mPEG-DOPA polymers and PEG polymers immobilized to surfaces using other approaches. Peptide anchors containing three DOPA residues resulted in PEG surface densities higher than those achieved using several existing PEG immobilization strategies, suggesting that peptide mimics of mussel adhesive proteins may be useful for achieving high densities of protein-resistant polymers on surfaces.     

An aqueous-based surface modification of poly(dimethylsiloxane) with poly(ethylene glycol) to prevent biofouling

We report a simple modification of poly(dimethylsiloxane) (PDMS) surfaces with poly(ethylene glycol) (PEG) through the adsorption of a graft copolymer, poly(l-lysine)-graft-poly(ethylene glycol) (PLL-g-PEG) from aqueous solution. In this approach, the PDMS surface was treated with oxygen plasma, followed by immersion into aqueous solution containing PLL-g-PEG copolymers. Due to the hydroxyl/carboxylic groups generated on the PDMS surface after oxygen plasma, the polycationic PLL backbone is attracted to the negatively charged surface and PEG side chains exhibit an extended structure. The PEG/aqueous interface generated in this way revealed a near-perfect resistance to nonspecific protein adsorption as monitored by means of optical waveguide lightmode spectroscopy (OWLS) and fluorescence microscopy.     

Adsorption of trypsin on hydrophilic and hydrophobic surfaces

The adsorption of trypsin onto polystyrene and silica surfaces was investigated by reflectometry, spectroscopic methods, and atomic force microscopy (AFM). The affinity of trypsin for the hydrophobic polystyrene surface was higher than that for the hydrophilic silica surface, but steady-state adsorbed amounts were about the same at both surfaces. The conformational characteristics of trypsin immobilized on silica and polystyrene nanospheres were analyzed in situ by circular dichroism and fluorescence spectroscopy. Upon adsorption the trypsin molecules underwent structural changes at the secondary and tertiary level, although the nature of the structural alterations was different for silica and polystyrene surfaces. AFM imaging of trypsin adsorbed on silica showed clustering of enzyme molecules. Rinsing the silica surface resulted in 20% desorption of the originally adsorbed enzyme molecules. Adsorption of trypsin on the surface of polystyrene was almost irreversible with respect to dilution. After adsorption on silica the enzymatic activity of trypsin was 10 times lower, and adsorbed on polystyrene the activity was completely suppressed. The trypsin molecules that were desorbed from the sorbent surfaces by dilution with buffer regained full enzymatic activity.     

Single DNA molecules as probes for interrogating silica surfaces after various chemical treatments

We examined the adsorption of single YOYO-1-labeled λ-DNA molecules at glass surfaces after treatment with various chemical cleaning methods by using total internal reflection fluorescence microscopy (TIRFM). The characteristics of these surfaces were further assessed using contact angle (CA) measurements and atomic force microscopy (AFM). By recording the real-time dynamic motion of DNA molecules at the liquid/solid interface, subtle differences in adsorption affinities were revealed. The results indicate that the driving force for adsorption of DNA molecules on glass surfaces is mainly hydrophobic interaction. We also found that surface topography plays a role in the adsorption dynamics.     

Cleaning and hydrophilization of atomic force microscopy silicon probes

The silicon surface of commercial atomic force microscopy (AFM) probes loses its hydrophilicity by adsorption of airborne and package-released hydrophobic organic contaminants. Cleaning of the probes by acid piranha solution or discharge plasma removes the contaminants and renders very hydrophilic probe surfaces. Time-of-flight secondary-ion mass spectroscopy and X-ray photoelectron spectroscopy investigations showed that the native silicon oxide films on the AFM probe surfaces are completely covered by organic contaminants for the as-received AFM probes, while the cleaning methods effectively remove much of the hydrocarbons and silicon oils to reveal the underlying oxidized silicon of the probes. Cleaning procedures drastically affect the results of adhesive force measurements in water and air. Thus, cleaning of silicon surfaces of the AFM probe and sample cancelled the adhesive force in deionized water. The significant adhesive force values observed before cleaning can be attributed to formation of a bridge of hydrophobic material at the AFM tip-sample contact in water. On the other hand, cleaning of the AFM tip and sample surfaces results in a significant increase of the adhesive force in air. The presence of water soluble contaminants at the tip-sample contact lowers the capillary pressure in the water bridge formed by capillary condensation at the AFM tip-sample contact, and this consequently lowers the adhesive force.     

High salt stability and protein resistance of poly(L-lysine)-g-poly(ethylene glycol) copolymers covalently immobilized via aldehyde plasma polymer interlayers on inorganic and polymeric substrates

The electrostatic adsorption onto charged surfaces of comb copolymers comprising a polyelectrolyte backbone and pendent PEG side chains, such as poly(l-lysine)-g-poly(ethylene glycol) (PLL-g-PEG), has in previous studies provided protein-repellent thin coatings, particularly on metal oxide surfaces. A drawback of this approach is, however, the instability of such adsorbed layers under extreme pH values or high ionic strength. We have overcome this limitation in the present study by covalently immobilizing PLL-g-PEG copolymers onto aldehyde plasma-modified substrates. Silicon wafers, optical waveguide chips, and perfluorinated ethylene-co-propylene (FEP) polymer substrates were first coated with a thin plasma polymer layer using a propionaldehyde plasma, followed by covalent immobilization of PLL-g-PEG via reductive amination between amine groups of the PLL backbone with aldehyde groups on the plasma-deposited interlayer. The stability in high salt media and the protein resistance of different molecular architectures of immobilized PLL-g-PEG layers were quantitatively investigated by XPS, an optical waveguide technique (OWLS), and ToF-SIMS. The adsorption of bovine serum albumin was found to be below the detection limit (<2 ng/cm(2)), as for electrostatically adsorbed PLL-g-PEG layers. However, after 24 h of exposure of covalently immobilized layers of PLL-g-PEG to high ionic strength buffer (2400 mM NaCl), no significant change in the protein resistance was observed, whereas under the same conditions electrostatically adsorbed PLL-g-PEG coatings lost their protein resistance. Moreover, covalent immobilization via an aldehyde plasma interlayer enabled the application of PLL-g-PEG layers onto substrates such as FEP onto which electrostatic binding is not possible. These findings create a generic platform for the covalent immobilization of PLL-g-PEG onto a wide variety of substrates.     

Ex Situ Chemical Determination of Free Radicals and Peroxides on Plasma Treated Surfaces

Ex situ determinations of free radicals by 2,2-Diphenyl-1-pricylhydrazyl (DPPH) method and of peroxides by the iodine method applied to plasma treated surfaces were examined. Plasma polymers and plasma-treated polymers, which are already exposed to ambient oxygen, the original free radicals could be detected as peroxides. It was found that the specificity of the both methods was not high enough for quantitative determination. Chemical cleaning of aluminum alloy surfaces, such as alkaline cleaning and deoxidizing treatment, was found to leave large amount of DPPH scavengers, which could be removed by argon or oxygen plasma treatment, implying that the reactivity of DPPH is not exclusive to free radicals. DPPH method should be viewed as an analytical tool to detect scavengers of DPPH (rather than free radicals) existing on the surface region accessible to the DPPH solution. The iodine method should be likewise viewed as an analytical tool for iodine-liberating species existing on the surface region. Plasma treatment (O₂ or Ar) of aluminum surface did not remove the species, and in some cases increased the amount. The both methods could be used as tools to assess the overall surface reactivity, particularly of metals, which could influence the adhesion of coatings.     

Cleaning of pyrolytic hexagonal boron nitride surfaces

Hexagonal boron nitride (h‐BN) has recently garnered significant interest as a substrate and dielectric for two‐dimensional materials and devices based on graphene or transition metal dichalcogenides such as molybdenum disulfide (MoS₂). As substrate surface impurities and defects can negatively impact the structure and properties of two‐dimensional materials, h‐BN surface preparation and cleaning are a critical consideration. In this regard, we have utilized X‐ray photoelectron spectroscopy to investigate the influence of several ex situ wet chemical and in situ thermal desorption cleaning procedures on pyrolytic h‐BN surfaces. Of the various wet chemistries investigated, a 10 : 1 buffered HF solution was found to produce surfaces with the lowest amount of oxygen and carbon contamination. Ultraviolet/ozone oxidation was found to be the most effective ex situ treatment for reducing carbon contamination. Annealing at 1050 °C in vacuum or 10^?5 Torr NH₃ was found to further reduce oxygen and carbon contamination to the XPS detection limits. Copyright © 2015 John Wiley & Sons, Ltd.     

Electrostatic self-assembly of PEG copolymers onto porous silica nanoparticles

A critical requirement toward the clinical use of nanocarriers in drug delivery applications is the development of optimal biointerfacial engineering procedures designed to resist biologically nonspecific adsorption events. Minimization of opsonization increases blood residence time and improves the ability to target solid tumors. We report the electrostatic self-assembly of polyethyleneimine-polyethylene glycol (PEI-PEG) copolymers onto porous silica nanoparticles. PEI-PEG copolymers were synthesized and their adsorption by self-assembly onto silica surfaces were investigated to achieve a better understanding of structure-activity relationships. Quartz-crystal microbalance (QCM) study confirmed the rapid and stable adsorption of the copolymers onto silica-coated QCM sensors driven by strong electrostatic interactions. XPS and FT-IR spectroscopy were used to analyze the coated surfaces, which indicated the presence of dense PEG layers on the silica nanoparticles. Dynamic light scattering was used to optimize the coating procedure. Monodisperse dispersions of the PEGylated nanoparticles were obtained in high yields and the thin PEG layers provided excellent colloidal stability. In vitro protein adsorption tests using 5% serum demonstrated the ability of the self-assembled copolymer layers to resist biologically nonspecific fouling and to prevent aggregation of the nanoparticles in physiological environments. These results demonstrate that the electrostatic self-assembly of PEG copolymers onto silica nanoparticles used as drug nanocarriers is a robust and efficient procedure, providing excellent control of their biointerfacial properties.     

Synthesis of poly(vinylidene fluoride) (PVdF)/silica hybrids having interpenetrating polymer network structure by using crystallization between PVdF chains

High transparent and homogeneous poly(vinylidene fluoride) (PVdF)/silica hybrids were obtained by using an in‐situ interpenetrating polymer network (IPN) method. The simultaneous formation of PVdF gel resulting from the physical cross‐linking and silica gel from sol–gel process prevented the aggregation of PVdF in silica gel matrix. To form the physical cross‐linking between PVdF chains, the cosolvent system of dimethylformaide (DMF) and γ‐butyrolactone was used. The obtained PVdF/silica hybrids had an entangled combination of physical PVdF gel and silica gel, which was called a “complete‐ IPN” structure. The physical cross‐linking between PVdF chains in silica gel matrix was confirmed by differential scanning calorimetry (DSC) measurements. The miscibility between PVdF and silica phase was examined by scanning electron microscopy (SEM) and tapping mode atomic force microscopy (TM‐AFM) measurements. © 2005 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 43: 3543–3550, 2005     

Correlation between desorption force measured by atomic force microscopy and adsorption free energy measured by surface plasmon resonance spectroscopy for peptide-surface interactions

Surface plasmon resonance (SPR) spectroscopy is a useful technique for thermodynamically characterizing peptide-surface interactions; however, its usefulness is limited to the types of surfaces that can readily be formed as thin layers on the nanometer scale on metallic biosensor substrates. Atomic force microscopy (AFM), on the other hand, can be used with any microscopically flat surface, thus making it more versatile for studying peptide-surface interactions. AFM, however, has the drawback of data interpretation due to questions regarding peptide-to-probe-tip density. This problem could be overcome if results from a standardized AFM method could be correlated with SPR results for a similar set of peptide-surface interactions so that AFM studies using the standardized method could be extended to characterize peptide-surface interactions for surfaces that are not amenable for characterization by SPR. In this article, we present the development and application of an AFM method to measure adsorption forces for host-guest peptides sequence on surfaces consisting of alkanethiol self-assembled monolayers (SAMs) with different functionality. The results from these studies show that a linear correlation exists between these data and the adsorption free energy (ΔG(o)(ads)) values associated with a similar set of peptide-surface systems available from SPR measurements. These methods will be extremely useful to characterize thermodynamically the adsorption behavior for peptides on a much broader range of surfaces than can be used with SPR to provide information related to understanding protein adsorption behavior to these surfaces and to provide an experimental database that can be used for the evaluation, modification, and validation of force field parameters that are needed to represent protein adsorption behavior accurately for molecular simulations.     

Development of a non-labeled immunosensor for the herbicide trifluralin via optical waveguide lightmode spectroscopic detection

A highly sensitive immunosensor using optical waveguide lightmode spectroscopy (OWLS) was developed for the detection of the herbicide trifluralin. OWLS as an in situ and label free method of detection, based on the measurement of the diffraction of a linearly polarized laser beam (He-Ne laser, 632.8 nm) on a diffraction grating in a thin waveguide layer (SiO₂-TiO₂), offered means to produce immunosensors utilizing immobilized antibodies raised against trifluralin allowing a non-competitive biosensor, or immobilized trifluralin conjugate allowing a competitive biosensor for this analyte. Immobilization of molecules sensitizing the sensor was undertaken on amino silanized waveguide surfaces in a two-step procedure using glutaraldehyde. Within the immobilized antibody (Ab) based immunosensor the signal measured was proportional to the trifluralin content in the samples, but the method allowed detection of trifluralin only above 100 ng ml⁻¹ due to the small molecular size of the antigen (Ag). In the immobilized antigen based immunosensor, a trifluralin-bovine serum albumin (BSA) conjugate was covalently linked to the waveguide surface. During measurements the standard solutions and samples were mixed in 1:1 ratio with antiserum, containing constant amounts of antibodies. The amount of free antibodies bound to the surface was inversely proportional to the trifluralin content of the solutions measured. The immobilized antigen based method allowed detection of trifluralin in the concentration range of 2×10⁻⁷ to 3×10⁻⁵ ng ml⁻¹. Results of trifluralin determinations were compared to those obtained in parallel enzyme-linked immunosorbent assay (ELISA) tests and in gas chromatorgraphic-mass spectrometric (GC-MS) analyses, and indicated an increase of six orders of magnitude in the limit of detection (LOD).     

Use of in‐situ atomic force microscopy to monitor the biodegradation of polyhydroxyalkanoates (PHAs)

Thin films of a co‐polymer mixture of poly 3‐hydroxybutyrate and poly 3‐hydroxyvalerate P(3HB‐3HV) were spun‐cast onto glass slides resulting in 35 nm thick layers with a spherulitic microstructure. An untyped strain of Streptomyces sp. bacteria was isolated from soil samples, and it's PHA depolymerase was used to degrade the P(3HB‐3HV) thin films. Both ex‐situ and in‐situ atomic force microscopy (AFM) biodegradation studies were performed to determine the kinetics of the biodegradation over the course of three hours at room temperature. Ex‐situ AFM was performed in Tapping Mode and in‐situ AFM was performed in the PHA depolymerase using contact mode AFM in the liquid cell, allowing for the real‐time analysis of P(3HB‐3HV) biodegradation. Biodegradation is observed uniformly throughout the surface, and can be observed within 30 min. of depolymerase exposure. In‐situ AFM analysis yields a linear degradation rate as a function of time, while the ex‐situ study suggests a more complex kinetics.     

Protein adsorption: A quest for a universal mechanism

Recent theoretical and experimental results pertinent to protein adsorption kinetics obtained for well-defined systems using direct experimental techniques are discussed. Attention is focused on albumins and fibrinogen, whose structure and physicochemical characteristic are well-known. It is confirmed that the experimental data obtained by AFM imaging, QCM, OWLS, XPS and electrokinetic techniques (streaming potential) are prone to a quantitative interpretation in terms of the coarse-grained and molecular dynamics modeling. This allows to derive reliable data concerning the mass transfer rates, hydration functions, maximum coverages and adsorption/desorption kinetic constants. These results confirm that the protein adsorption mechanism is governed by electrostatic interactions among heterogeneously distributed charges. The protein substrate interactions promote the molecule transfer through the surface layer, control the free energy and in consequence the residence time of the molecule on substrate surfaces. On the other hand, the interactions among adsorbed molecules control the maximum coverage and the formation of bilayer structures. As a result of this complex electrostatics, one often observes in protein adsorption studies the formation of irreversibly bound fraction of molecules that contact the substrate and a reversibly adsorbed fraction otherwise. This leads to the appearance of anomalous isotherms, characterized by considerable adsorption for negligible bulk protein concentration, which deviate from the Langmuir model.     

Optical Response of Porous Titania‐Silica Waveguides to Surface Charging in Electrolyte Filled Pores

In this work, we present a novel method for in situ investigation of surface charging and ion transport inside nanopores of titania‐silica waveguide by means of the optical‐waveguide‐lightmode spectroscopy. Porous oxide waveguides show a strong optical response when exposed to electrolyte solutions, and this response is consistent with oxide surface charging due to changes in ionic strength and pH of the solution in contact with the waveguide. The optical response to pH or electrolyte concentration change is stabilized within several minutes when the solution ionic strength is sufficiently high (0.1M ), while it takes two orders of magnitude longer to reach stable optical response at very low ionic strengths (<0.1mM ). The relaxation times at the high ionic strength are still by several orders of magnitude slower than expected from bulk diffusion coefficients of electrolytes in water. Our results indicate that diffusion of electrolytes is severely hindered (and more so with decreasing ionic strength) in charged pores inside waveguides.     

Adsorption characteristics of zwitterionic diblock copolymers at the silica/aqueous solution interface

The adsorption of a zwitterionic diblock copolymer, poly(2-(diethylamino)ethyl methacrylate)-block-poly(methacrylic acid) (PDEA59-PMAA50), at the silica/aqueous solution interface has been characterised as a function of pH. In acidic solution, this copolymer forms core-shell micelles with the neutral PMAA chains being located in the hydrophobic cores and the protonated PDEA chains forming the cationic micelle coronas. In alkaline solution, the copolymer forms the analogous inverted micelles with anionic PMAA coronas and hydrophobic PDEA cores. The morphology of the adsorbed layer was observed in situ using soft-contact atomic force microscopy (AFM): this technique suggests the formation of a thin adsorbed layer at pH 4 due to the adsorption of individual copolymer chains (unimers) rather than micelle aggregates. This is supported by the remarkably low dissipation values and the relatively low degrees of hydration for the adsorbed layers, as estimated using a combination of quartz crystal microbalance with dissipation monitoring (QCM-D) and optical reflectometry (OR). In alkaline solution, analysis of the adsorption data suggests a conformation for the adsorbed copolymers where one block projects normal to the solid/liquid interface; this layer consists of a hydrophobic PDEA anchor block adsorbed on the silica surface and an anionic PMAA buoy block extending into the solution phase. Tapping mode AFM studies were also carried out on the silica surfaces after removal from the copolymer solutions and subsequent drying. Interestingly, in these cases micelle-like surface aggregates were observed from both acidic and alkaline solutions. The lateral dimension of the aggregates seen is consistent with the corresponding hydrodynamic diameter of the copolymer micelles in bulk solution. The combination of the in situ and ex situ AFM data provides evidence that, for this copolymer, micelle aggregates are only seen in the ex situ dry state as a result of the substrate withdrawal and drying process. It remains unclear whether these aggregates are caused by micelle deposition at the surface during the substrate withdrawal from the solution or as a result of unimer rearrangements at the drying front as the liquid recedes from the surface.