Physical Adsorption Of Proteins Research Articles

Real-Time Dynamic Adsorption Processes of Cytochrome c on an Electrode Observed through Electrochemical High-Speed Atomic Force Microscopy

An understanding of dynamic processes of proteins on the electrode surface could enhance the efficiency of bioelectronics development and therefore it is crucial to gain information regarding both physical adsorption of proteins onto the electrode and its electrochemical property in real-time. We combined high-speed atomic force microscopy (HS-AFM) with electrochemical device for simultaneous observation of the surface topography and electron transfer of redox proteins on an electrode. Direct electron transfer of cytochrome c (cyt c) adsorbed on a self-assembled monolayers (SAMs) formed electrode is very attractive subject in bioelectrochemistry. This paper reports a real-time visualization of cyt c adsorption processes on an 11-mercaptoundecanoic acid-modified Au electrode together with simultaneous electrochemical measurements. Adsorbing cyt c molecules were observed on a subsecond time resolution simultaneously with increasing redox currents from cyt c using EC-HS-AFM. The root mean square roughness (R RMS) from the AFM images and the number of the electrochemically active cyt c molecules adsorbed onto the electrode (Γ) simultaneously increased in positive cooperativity. Cyt c molecules were fully adsorbed on the electrode in the AFM images when the peak currents were steady. This use of electrochemical HS-AFM significantly facilitates understanding of dynamic behavior of biomolecules on the electrode interface and contributes to the further development of bioelectronics.

Protein Covalently Conjugated SU-8 Surface for the Enhancement of Mesenchymal Stem Cell Adhesion and Proliferation

Cell growing behavior is significantly dependent on the surface chemistry of materials. SU-8 as an epoxy-based negative photoresist is commonly used for fabricating patterned layers in lab-on-a-chip devices. As a hydrophobic material, SU-8 substrate is not favorable for cell culture, and cell attachment on native SU-8 is limited attributed to poor surface biocompatibility. Although physical adsorption of proteins could enhance the cell adhesion, the effect is not durable. In this work, SU-8 surface chemistry is modified by immobilizing fibronectin (FN) and collagen type I (COL I) covalently using (3-aminopropyl)triethoxysilane (APTES) and cross-linker glutaraldehyde (GA) to increase surface biofunctionality. The effectiveness of this surface treatment to improve the adhesion and viability of mesenchymal stem cells (MSCs) is investigated. It is found that the wettability of SU-8 surface can be significantly increased by this chemical modification. In addition, the spreading area of MSCs increases on the SU-8 surfaces with covalently conjugated matrix proteins, as compared to other unmodified SU-8 surface or those coated with proteins simply by physical adsorption. Furthermore, cell proliferation is dramatically enhanced on the SU-8 surfaces modified under the proposed scheme. Therefore, SU-8 surface modification with covalently bound matrix proteins assisted by APTES+GA provides a highly biocompatible interface for the enhanced adhesion, spreading, and proliferation of MSCs.

Attachment of hydrogel microstructures and proteins to glass via thiol-terminated silanes

Micropatterning strategies often call for attachment of non-fouling biomaterials and immobilization of proteins in order to create biosensing surfaces or to control cell–surface interactions. Our laboratory has made frequent use of hydrogel photolithography – a micropatterning process for immobilizing poly(ethylene glycol) (PEG) hydrogel microstructures on glass surfaces. In the present study we explored the use of thiolsilane as a coupling layer for both covalent anchoring of hydrogel microstructures and covalent immobilization of proteins on glass. These new surfaces were compared to acryl-silane functionalized glass slides that allowed covalent attachment of gels but only physical adsorption of proteins as well as surfaces containing a mixture of both functional groups. We observed comparable attachment and retention of hydrogel microstructures on acryl and thiol-terminated silanes. Ellipsometry studies revealed presence of significantly higher level of proteins on thiol-functionalized glass. Overall, our studies demonstrate that thiol-silane functionalized glass surfaces may be used to create complex micropatterned surfaces comprised of covalently attached hydrogels and proteins. This simple and effective surface modification strategy will be broadly applicable in cellular engineering and biosensing studies employing hydrogel micropatterns.

Latex Immunoagglutination Assays*

Latex immunoagglutination assays continue to be widely used in biology and medicine for the detection of small quantities of an antibody or antigen of interest in fluid test samples. Main characteristics of preparation and use of these assays are examined here. Physical adsorption of proteins onto latex particles surface, with special relevance to immunoglobulins, is analyzed with major attention to those factors that influence adsorption: medium conditions such as pH and ionic strength, surface characteristics as type and amount of charge, or hydrophobicity. Different functionalized latexes for covalent linking are also presented, as well as the corresponding chemical reactions. Techniques for the detection and quantification of the immunoreaction are briefly summarized, including visual observation, light scattering, turbidimetry, nephelometry, and angular anisotropy. Finally, some problems of colloidal stability of these latex assays are analyzed, as well as the different solutions applied by scientists to solve them. *Reprinted with permission from Colloidal Biomolecules, Biomaterials, and Biomedical Applications, A. Elaissari, Ed., Marcel Dekker, Inc, New York, 2004, pp. 53–101

Hydrophilic stimuli-responsive particles for biomedical applications

Hydrophilic and stimuli-responsive submicronic latex particles based on polyalkyl(meth)acrylamide can be prepared owing to simple radical-initiated polymerizations in heterogeneous media using a water-soluble initiator and a crosslinker (methylenebisacrylamide). The paper aims at reviewing the synthesis and properties of functionalized polystyrene-poly-N-isopropylacrylamide core-shell particles or polyN-isopropylmethacrylamide microgel particles. Particle size of analysis showed that a short nucleation period afforded the synthesis of highly monodispersed latexes. The dramatic change of the colloidal properties (particle size, electrophoretic mobility) was found to reflect the thermal sensitivity of such particles. The hydrophilic nature of the particles below the volume phase transition temperature was found to drastically reduce the physical adsorption of proteins. Some examples of biomedical applications of these stimuli-responsive particles are briefly reported.

Thick polystyrene-coated quartz crystal microbalance as a basis of a cost effective immunosensor

The use of a commercial quartz crystal microbalance for a disposable low cost and reliable immunosensor is presented. Thick polystyrene coating provides a physical adsorption scheme to immobilise antigens or antibodies and avoids the oxidation of the commercial quartz crystal electrodes. The coating does not contribute significantly to the damping of quartz crystal. The polystyrene-coated quartz also shows a smoother surface than the non-coated one. This is important for applications in liquids. Using the polystyrene-coated quartz, the physical adsorption of protein is well-achieved. The frequency decrease caused by specific reactions between insulin, anti-insulin antibody, and anti-mouse IgG antibody is presented as an example. A frequency shift caused by non-specific reactions is not detected.