Fibrinogen adsorption on blocked surface of albumin

We have investigated the adsorption of albumin and fibrinogen onto PET (polyethylene terephthalate) and glass surfaces and how pre-adsorption of albumin onto these surfaces can affect the adsorption of later added fibrinogen. For materials and devices being exposed to blood, adsorption of fibrinogen is often a non-wanted event, since fibrinogen is part of the clotting cascade and unspecific adsorption of fibrinogen can have an influence on the activation of platelets. Albumin is often used as blocking agent for avoiding unspecific protein adsorption onto surfaces in devices designed to handle biological samples, including protein solutions. It is based on the assumption that proteins adsorbs as a monolayer on surfaces and that proteins do not adsorb on top of each other. By labelling albumin and fibrinogen with two different radioactive iodine isotopes that emit gamma radiation with different energies, the adsorption of both albumin and fibrinogen has been monitored simultaneously on the same sample. Information about topography and coverage of adsorbed protein layers has been obtained using AFM (Atomic Force Microscopy) analysis in liquid. Our studies show that albumin adsorbs in a multilayer fashion on PET and that fibrinogen adsorbs on top of albumin when albumin is pre-adsorbed on the surfaces.     

Platelet adhesion to photodynamic therapy-treated extracellular matrix proteins

Photodynamic therapy (PDT) produces reactive species that alter vascular wall biology and vessel wall proteins. In this study, we examined platelet adhesion to PDT-treated (photosensitizer = Photofrin; fluence 100 J/cm2; lambda = 630 nm) extracellular matrix (ECM), fibrinogen, von Willebrand factor (vWF) and collagen Types I and III, under flow conditions in a recirculating perfusion chamber. Platelet adhesion was quantified by image analysis. The effect of PDT on vWF was assessed by measuring the binding of domain-specific antibodies to treated vWF. PDT significantly decreased platelet adhesion to the ECM, fibrinogen and vWF. However, PDT of collagen resulted in significantly increased platelet adhesion, with large aggregate formation. PDT affected mostly the A1 (glycoprotein [GP]-Ib-binding site), A2 and A3 (collagen-binding site) domains of vWF but not the D'-D3 (factor VIII-binding site) and B-C1 (GP-IIb/IIIa-binding site) domains. In conclusion, PDT can alter the ECM, resulting in decreased platelet adhesion. However, vessels with high collagen content, such as veins and small arteries, may become increasingly prone to thrombosis. The results of this study may thus play a role in understanding the thrombogenic properties and mechanisms of vascular PDT.     

Colloid probe AFM investigation of interactions between fibrinogen and PEG-like plasma polymer surfaces

Interaction forces between surfaces designed to be protein resistant and fibrinogen (Fg) were investigated in phosphate-buffered saline with colloid probe atomic force microscopy. The surfaces of the silica probes were coated with a layer of fibrinogen molecules by adsorption from the buffer. The technique of low-power, pulsed AC plasma polymerization was used to make poly(ethylene glycol) (PEG)-like coatings on poly(ethylene teraphthalate) by using diethylene glycol vinyl ether as the monomer gas. The degree of PEG-like nature of the films was controlled by use of a different effective plasma power in the chamber for each coating, ranging from 0.6 to 3.6 W. This produced a series of thin films with a different number of ether carbons, as assessed by X-ray photoelectron spectroscopy. The interaction force measurements are discussed in relation to trends observed in the reduction of fibrinogen adsorption, as determined quantitatively by (125)I radio-labeling. The plasma polymer coatings with the greatest protein-repelling properties were the most PEG-like in nature and showed the strongest repulsion in interaction force measurements with the fibrinogen-coated probe. Once forced into contact, all the surfaces showed increased adhesion with the protein layer on the probe, and the strength and extension length of adhesion was dependent on both the applied load and the plasma polymer surface chemistry. When the medium was changed from buffer to water, the adhesion after contact was eliminated and only appeared at much higher loads. This indicates that the structure of the fibrinogen molecules on the probe is changed from an extended conformation in buffer to a flat conformation in water, with the former state allowing for stronger interaction with the polymer chains on the surface. These experiments underline the utility of aqueous surface force measurements toward understanding protein-surface interactions, and developing nonfouling surfaces that confer a steric barrier against protein adsorption.     

Studies of electroosmotic flow and the effects of protein adsorption in plasma‐polymerized microchannel surfaces

This paper presents a study of EOF properties of plasma‐polymerized microchannel surfaces and the effects of protein (fibrinogen and lysozyme) adsorption on the EOF behavior of the surface‐modified microchannels. Three plasma polymer surfaces, i.e. tetraglyme, acrylic acid and allylamine, are tested. Results indicate EOF suppression in all plasma‐coated channels compared with the uncoated glass microchannel surfaces. The EOF behaviors of the modified microchannels after exposure to protein solutions are also investigated and show that even low levels of protein adsorption can significantly influence EOF behavior, and in some cases, result in the reversal of flow. The results also highlight that EOF measurement can be used as a method for detecting the presence of proteins within microchannels at low surface coverage (<1 ng/cm² on glass). Critically, the results illustrate that the non‐fouling tetraglyme plasma polymer is able to sustain EOF. Comparison of the plasma‐polymerized surfaces with conventionally grafted polyelectrolyte surfaces demonstrates the stabilities of the plasma polymer films, enabling multiple EOF runs over 3 days without deterioration in performance. The results of this study clearly demonstrate that plasma polymers enable the surface chemistry of microfluidic devices to be tailored for specific applications. Critically, the deposition of the non‐fouling tetraglyme coating enables stable EOF to be induced in the presence of protein.     

聚(N-异丙基丙烯酰胺)改性硅表面与血浆蛋白质的相互作用

通过表面引发原子转移自由基聚合(ATRP)在硅表面接枝了聚(N-异丙基丙烯酰胺)(PNIPAAm)聚合物刷,并考察了PNIPAAm改性表面在单一蛋白质溶液以及血浆中与血浆蛋白质之间的相互作用.蛋白质吸附测试表明,与未改性的硅表面相比,改性后的表面对纤维蛋白原的吸附量大大降低,特别是在血浆中纤维蛋白原吸附量小于5ng/c...     

Fibrinogen adsorption on three silica-based surfaces: conformation and kinetics

Using AFM (atomic force microscopy) to probe protein conformation and arrangement, and TIRF (total internal reflectance fluorescence) to monitor kinetics, fibrinogen adsorption on three different silica-based surfaces was studied: the native oxide on silicon, acid-etched microscope slides, and acid-etched polished glass. The three are chemically similar, but the microscope slide is rougher and induces AFM tip instabilities that appear as high spots on the bare surface. Fibrinogen's conformation and transport-limited adsorption kinetics are found to be quantitatively similar on all three surfaces. Further, the number of adsorbed proteins in progressive AFM micrographs quantitatively match the coverages measured by TIRF during early adsorption. Surfaces appear full, via AFM, when adsorbed amounts are about an order of magnitude below their true saturation levels (via TIRF) because, above about 0.26 mg/m(2), individual proteins cannot be discerned. The results demonstrate how the appearance of AFM micrographs can be misleading regarding surface saturation. On all three surfaces, fibrinogen is, at most, slightly aggregated, showing limited, if any, surface mobility. The complexities of the microscope slide's surface landscape minimally impact adsorption.     

Interpretation of protein adsorption: surface-induced conformational changes

Protein adhesion plays a major role in determining the biocompatibility of materials. The first stage of implant integration is the adhesion of protein followed by cell attachment. Surface modification of implants (surface chemistry and topography) to induce and control protein and cell adhesion is currently of great interest. This communication presents data on protein adsorption (bovine serum albumin and fibrinogen) onto model hydrophobic (CH(3)) and hydrophilic (OH) surfaces, investigated using a quartz crystal microbalance (QCM) and grazing angle infrared spectroscopy. Our data suggest that albumin undergoes adsorption via a single step whereas fibrinogen adsorption is a more complex, multistage process. Albumin has a stronger affinity toward the CH(3) compared to OH terminated surface. In contrast, fibrinogen adheres more rapidly to both surfaces, having a slightly higher affinity toward the hydrophobic surface. Conformational assessment of the adsorbed proteins by grazing angle infrared spectroscopy (GA-FTIR) shows that after an initial 1 h incubation few further time-dependent changes are observed. Both proteins exhibited a less organized secondary structure upon adsorption onto a hydrophobic surface than onto a hydrophilic surface, with the effect observed greatest for albumin. This study demonstrates the ability of simple tailor-made monochemical surfaces to influence binding rates and conformation of bound proteins through protein-surface interactions. Current interest in biocompatible materials has focused on surface modifications to induce rapid healing, both of implants and for wound care products. This effect may also be of significance at the next stage of implant integration, as cell adhesion occurs through the surface protein layer.     

Biofouling-resistance expanded poly(tetrafluoroethylene) membrane with a hydrogel-like layer of surface-immobilized poly(ethylene glycol) methacrylate for human plasma protein repulsions

In general, it is a challenge to control the highly polar material grafting from the chemically inert Teflon-based membrane surface. This work describes the surface modification and characterization of expanded poly(tetrafluoroethylene) (ePTFE) membranes grafted with poly(ethylene glycol) methacrylate (PEGMA) macromonomer via surface-activated plasma treatment and thermally induced graft copolymerization. The chemical composition and microstructure of the surface-modified ePTFE membranes were characterized by Fourier transform infrared spectroscopy (FT-IR), contact angle, and bio-atomic force microscopy (bio-AFM) measurements. Biofouling property of the modified membranes was evaluated by the measurements of the plasma protein (γ-globulin, fibrinogen, or albumin) adsorption determined using an enzyme-linked immunosorbent assay (ELISA). In general, the hydrophilicity of the surface of ePTFE membranes increases with increasing the grafting degree of the copolymerized PEGMA. The highly hydrated PEGMA chain on the resulting ePTFE membranes was found to form a surface hydrogel-like layer with regulated coverage in aqueous state, which can be controlled by the content of PEGMA macromonomer in the reaction solution. The relative protein adsorption was effectively reduced with increasing capacity of the hydration for the PEGMA chain grafted on the ePTFE membrane surface. From both results of protein adsorption and platelet adhesion test in vitro, it is concluded that the PEGMA-grafted hydrophilic ePTFE membranes could provide good biofouling resistance to substantially reduce plasma protein and blood platelet fouling on the membrane surface in human body temperature.     

Protein resistant polyurethane surfaces by chemical grafting of PEO: amino-terminated PEO as grafting reagent

The objective of this work was to gain a better understanding of the mechanism of resistance to protein adsorption of surfaces grafted with poly(ethylene oxide) (PEO). A polyurethane-urea was used as a substrate to which PEO was grafted. Grafting was carried out by introducing isocyanate groups into the surface followed by reaction with amino-terminated PEO. Surfaces grafted with PEO of various chain lengths (PUU-NPEO) were prepared and characterized by water contact angle and X-ray photoelectron spectroscopy (XPS). XPS data indicated higher graft densities on the PUU-NPEO surfaces than on analogous surfaces prepared using hydroxy-PEO (PUU-OPEO) as reported previously [J.G. Archambault, J.L. Brash, Colloids Surf. B: Biointerf. 33 (2004) 111-120]. Protein adsorption experiments using radiolabeled myoglobin, concanavalin A, albumin, fibrinogen and ferritin as single proteins in buffer showed that adsorption was reduced on the PEO-grafted surfaces by up to 95% compared to the control. Adsorption decreased with increasing PEO chain length and reached a minimum at a PEO MW of 2000. Adsorption levels on surfaces with 5000 and 2000 MW grafts were similar. There was no clear effect of protein size on resistance to protein adsorption. Adsorption on the PUU-NPEO surfaces was significantly lower than on the corresponding PUU-OPEO surfaces, again suggesting higher graft densities on the former. Adsorption of fibrinogen from plasma was also greatly reduced on the grafted surfaces. From analysis (SDS-PAGE, immunoblotting) of the proteins eluted after plasma exposure, it was found that the grafted surfaces and the unmodified substrate adsorbed the same proteins in roughly the same proportions, suggesting that adsorption to the PEO surfaces occurs on patches of bare substrate. The PEO grafts did not apparently cause differential access to the substrate based on protein size.     

Doubly Biomimetic Catecholic Phosphorylcholine Copolymer: A Platform Strategy for Fabricating Antifouling Surfaces

A doubly biomimetic PMNC polymer bearing cell antifouling phosphorylcholine and mussel adhesive protein catechol groups is synthesized. The polymer can be deposited onto a variety of substrates by dip‐coating in an aqueous solution, adhering to surfaces via the catechol functional group while at the same time forming a cell outer membrane mimetic antifouling surface. Contact angle, ATR‐FTIR and XPS measurements confirm polymer coating formation on a variety of inorganic and organic substrates. BSA and bovine plasma fibrinogen protein adsorption on PMNC coated surfaces are reduced significantly compared to unmodified substrates, and platelet adhesion from human serum onto the PMNC coated substrate surfaces is highly suppressed in this study.

     

Fibrinogen Adsorption on Biomaterials – A Numerical Study

A model for the adsorption of fibrinogen or, in general, non‐globular shaped proteins on solid surfaces are presented. Two‐dimensional cellular automata simulations of the adsorption of fibrinogen on two different surfaces were performed. The model includes mass transfer toward the surface, adsorption of fibrinogen molecules, and surface diffusion mechanisms for both fibrinogen molecules and clusters. We show that the major physical processes are represented in the recent model. Particularly, the influence of the surface hydrophobicity on the behavior of fibrinogen. Atomic force microscopy images of fibrinogen adsorption on Si model surfaces with different hydrophobicity are compared to the results.

     

Modification of polyurethane surface with an antithrombin-heparin complex for blood contact: influence of molecular weight of polyethylene oxide used as a linker/spacer

Polyurethane (PU) was modified using isocyanate chemistry to graft polyethylene oxide (PEO) of various molecular weights (range 300-4600). An antithrombin-heparin (ATH) covalent complex was subsequently attached to the free PEO chain ends, which had been functionalized with N-hydroxysuccinimide (NHS) groups. Surfaces were characterized by water contact angle and X-ray photoelectron spectroscopy (XPS) to confirm the modifications. Adsorption of fibrinogen from buffer was found to decrease by ~80% for the PEO-modified surfaces compared to the unmodified PU. The surfaces with ATH attached to the distal chain end of the grafted PEO were equally protein resistant, and when the data were normalized to the ATH surface density, PEO in the lower MW range showed greater protein resistance. Western blots of proteins eluted from the surfaces after plasma contact confirmed these trends. The uptake of ATH on the PEO-modified surfaces was greatest for the PEO of lower MW (300 and 600), and antithrombin binding from plasma (an indicator of heparin anticoagulant activity) was highest for these same surfaces. The PEO-ATH- and PEO-modified surfaces also showed low platelet adhesion from flowing whole blood. It is concluded that for the PEO-ATH surfaces, PEO in the low MW range, specifically MW 600, may be optimal for achieving an appropriate balance between resistance to nonspecific protein adsorption and the ability to take up ATH and bind antithrombin in subsequent blood contact.     

白蛋白原位复合的生物医用功能材料的研究(Ⅱ)──白蛋白的选择性吸附和血液相容性研究

采用¹²⁵I放射标记技术研究了血浆白蛋白和纤维蛋白原在聚甲基丙烯酸甲酯-接枝-十八烷基聚氧乙烯(PMMA-g-SPEO)、聚甲基丙烯酸甲酯-接枝-乙基聚氧乙烯(PMMA-g-EPEO)和聚甲基丙烯酸甲酯-甲基丙烯酸十八酯共聚物(PMMA-co-SMA)表面的竞争吸附行为.结果表明,十八烷基聚氧乙烯复合修饰的PMMA-g-SPEO可高选择性地形成白蛋白可逆吸附层,有效地阻抗血小板的粘附,延长材料的凝血时间,是一种理想的白蛋白原位复合的生物医用功能材料.     

Platelet adhesion and protein adsorption on silicone rubber surface by ozone-induced grafted polymerization with carboxybetaine monomer

Platelet adhesion and protein adsorption on the silicone rubber film grafted with N,N'-dimethyl-N-methacryloyloxyethyl-N-(2-carboxyethyl) ammonium (DMMCA) was studied. The grafting was carried out by means of ozone-induced method and was confirmed by ATR-FTIR and XPS investigations. The grafted films possessed relatively hydrophilic surface revealed by contact angle measurement. The blood compatibility of the grafted film was evaluated in vitro by platelet adhesion in platelet-rich plasma (PRP) and protein absorption in bovine fibrinogen (BFG) using silicone film as the reference. No substantial platelet adhesion was observed for the grafted films incubated in PRP for 60 and 180 min. The protein absorption was also significantly reduced after incubated in bovine fibrinogen for 60 min. Both the results indicated that the blood compatibility of silicone rubber was greatly improved by ozone-induced grafting of carboxybetaine zwitterionic polymer onto its surface.     

Effect of interfacial proteins on osteoblast-like cell adhesion to hydroxyapatite nanocrystals

A quartz crystal microbalance with dissipation (QCM-D) technique was employed to detecting the protein adsorption and subsequent osteoblast-like cell adhesion to hydroxyapatite (HAp) nanocrystals. The interfacial phenomena with the preadsorption of three proteins (albumin (BSA), fibronectin (Fn), and collagen (Col)), the subsequent adsorption of fetal bovine serum (FBS), and the adhesion of the cells were investigated. The QCM-D measured the frequency shift (Δf) and dissipation energy shift (ΔD), and the viscoelastic properties of the adlayers were evaluated using ΔD-Δf plot and Voigt-based viscoelastic model. The Col adsorption significantly showed higher Δf, ΔD, elasticity, and viscosity values as compared to the BSA and Fn adsorption, and the subsequent FBS adsorption depended on the preadsorbed proteins. The ΔD-Δf plot of the cell adhesion also showed a different behavior depending on the surfaces, and the Fn- and Col-modified surfaces showed the rapid mass and ΔD changes by forming the viscous interfacial layers with cell adhesion, indicating that the processes were affected by the cellular reaction through the extracellular matrix (ECM) proteins. The confocal laser scanning microscope images of adherent cells showed a different morphology and pseudopod on the surfaces. The cells adhered to the surfaces modified with the Fn and Col had significantly uniaxially expanded shapes and fibrous pseudopods, and those modified with the BSA had a round shape. Therefore, the different cell-protein interactions would cause the arrangement of the ECM and the cytoskeleton changes at the interfaces, and these phenomena were successfully detected by the QCM-D and Voigt-based model.     

Ion implantation into collagen-coated surfaces for the development of small diameter artificial grafts

Ion implantation into collagen (Type I) coated inner surfaces of test tubes with a length of 50 mm and an inner diameter of 2 and 3 mm were performed to develop hybrid type small diameter artificial vascular grafts. To obtain information about the cellular response and chemical and physical structure of those collagen surfaces, several experiments such as platelets adhesion test, endothelial cell culture, analysis of amino acids and animal study were performed. He(+) ion implanted collagen coated specimen exhibited cell attachment and inhibit platelet adhesion. From these results, it was assumed that He(+) ions broke the ligands that correspond to platelet, and the ligands that correspond to endothelial cell adhesion still existed after ion implantation. It was suggested that platelets and cell attachment could be control individually by ion implantation into collagen.     

Real-Time Characterization of Fibrinogen Interaction with Modified Titanium Dioxide Film by Quartz Crystal Microbalance with Dissipation

The adsorption of fibrinogen can be used as a quick indicator of surface haemocompatibility because of its prominent role in coagulation and platelet adhesion. In this work the molecular interaction between fibrinogen and a modified titanium oxide surface/platelet has been studied by quartz crystal microbalanee with dissipation （QCM-D） in situ. In order to further characterize the conformation of adsorbed fibrinogen, αC and γ-chain antibody were used to check the orientation and denaturation of fibrinogen on solid surface. QCM-D investiga- tions revealed the fibrinogen have the trend to adsorb on hydropllilic surface in a side-on orientation by positively charged αC domains, which would reduce the exposure of platelet bonding site on γ chain and enable less platelet adhesion and be activated. These obser- vations suggest that certain conformations of adsorbed fibrinogen are less platelet adhesive than others, which opens a possibility for creating a non-platelet adhesive substrates.     

A highly stable nonbiofouling surface with well-packed grafted zwitterionic polysulfobetaine for plasma protein repulsion

An ideal nonbiofouling surface for biomedical applications requires both high-efficient antifouling characteristics in relation to biological components and long-term material stability from biological systems. In this study we demonstrate the performance and stability of an antifouling surface with grafted zwitterionic sulfobetaine methacrylate (SBMA). The SBMA was grafted from a bromide-covered gold surface via surface-initiated atom transfer radical polymerization to form well-packed polymer brushes. Plasma protein adsorption on poly(sulfobetaine methacrylate) (polySBMA) grafted surfaces was measured with a surface plasmon resonance sensor. It is revealed that an excellent stable nonbiofouling surface with grafted polySBMA can be performed with a cycling test of the adsorption of three model proteins in a wide range of various salt types, buffer compositions, solution pH levels, and temperatures. This work also demonstrates the adsorption of plasma proteins and the adhesion of platelets from human blood plasma on the polySBMA grafted surface. It was found that the polySBMA grafted surface effectively reduces the plasma protein adsorption from platelet-poor plasma solution to a level superior to that of adsorption on a surface terminated with tetra(ethylene glycol). The adhesion and activation of platelets from platelet-rich plasma solution were not observed on the polySBMA grafted surface. This work further concludes that a surface with good hemocompatibility can be achieved by the well-packed surface-grafted polySBMA brushes.     

Bioadhesive control of plasma proteins and blood cells from umbilical cord blood onto the interface grafted with zwitterionic polymer brushes

In this work, bioadhesive behavior of plasma proteins and blood cells from umbilical cord blood (UCB) onto zwitterionic poly(sulfobetaine methacrylate) (polySBMA) polymer brushes was studied. The surface coverage of polySBMA brushes on a hydrophobic polystyrene (PS) well plate with surface grafting weights ranging from 0.02 mg/cm(2) to 0.69 mg/cm(2) can be effectively controlled using the ozone pretreatment and thermal-induced radical graft-polymerization. The chemical composition, grafting structure, surface hydrophilicity, and hydration capability of prepared polySBMA brushes were determined to illustrate the correlations between grafting properties and blood compatibility of zwitterionic-grafted surfaces in contact with human UCB. The protein adsorption of fibrinogen in single-protein solutions and at complex medium of 100% UCB plasma onto different polySBMA brushes with different grafting coverage was measured by enzyme-linked immunosorbent assay (ELISA) with monoclonal antibodies. The grafting density of the zwitterionic brushes greatly affects the PS surface, thus controlling the adsorption of fibrinogen, the adhesion of platelets, and the preservation of hematopoietic stem and progenitor cells (HSPCs) in UCB. The results showed that PS surfaces grafted with polySBMA brushes possess controllable hydration properties through the binding of water molecules, regulating the bioadhesive and bioinert characteristics of plasma proteins and blood platelets in UCB. Interestingly, it was found that the polySBMA brushes with an optimized grafting weight of approximately 0.1 mg/cm(2) at physiologic temperatures show significant hydrated chain flexibility and balanced hydrophilicity to provide the best preservation capacity for HSPCs stored in 100% UCB solution for 2 weeks. This work suggests that, through controlling grafting structures, the hemocompatible nature of grafted zwitterionic polymer brushes makes them well suited to the molecular design of regulated bioadhesive interfaces for use in the preservation of HSPCs from human UCB.     

Quartz crystal microbalance with dissipation coupled to on-chip MALDI-ToF mass spectrometry as a tool for characterising proteinaceous conditioning films on functionalised surfaces

Proteinaceous conditioning films (pCFs) are thought to play a key role in microbial adhesion, leading to the fouling of technical and biomedical devices and biofilm formation, which in turn causes material damage or persistent infections, respectively. However, little is definitively known about the process of surface conditioning via proteins. Herein, we demonstrate the potential of quartz crystal microbalance with dissipation coupled to MALDI-ToF mass spectrometry (QCM-D-MALDI) to investigate protein adsorption on different surfaces, enabling both the monitoring of CF formation and the determination of the molecular composition of CFs. After running QCM-D experiments, a subsequent tryptic on chip digestion step allows the identification of the proteins deposited on the sensor chip surface via MALDI-ToF mass spectrometry. Prominent blood plasma proteins, i.e., human serum albumin (HSA), fibrinogen (FG) and fibronectin (FN), were used. Chemically well defined sensor surfaces were prepared, among others, via self-assembled monolayer (SAM) technology. In cases where protein adsorption was observed by QCM-D, the adsorbed proteins were clearly detected and identified using MALDI-ToF/MS for both single-protein solutions of HSA, FG and FN as well as for protein mixtures. However, for equimolar protein mixtures on TiO₂ surfaces, only signals attributed to FG and FN were observed in the mass spectra. No signals indicating the presence of HSA could be detected. This finding leads to the assumption that only FG and FN attach to the TiO₂ sensor surface under the given experimental conditions.