Patricia Lisboa

pH-Dependent Immobilization of Proteins on Surfaces Functionalized by Plasma-Enhanced Chemical Vapor Deposition of Poly(acrylic acid)-and Poly(ethylene oxide)-like Films

The interaction of the proteins bovine serum albumin (BSA), lysozyme (Lys), lactoferrin (Lf), and fibronectin (Fn) with surfaces of protein-resistant poly(ethylene oxide) (PEO) and protein-adsorbing poly(acrylic acid) (PAA) fabricated by plasma-enhanced chemical vapor deposition has been studied with quartz crystal microbalance with dissipation monitoring (QCM-D). We focus on several parameters which are crucial for protein adsorption, i.e., the isoelectric point (pI) of the proteins, the pH of the solution, and the charge density of the sorbent surfaces, with the zeta-potential as a measure for the latter. The measurements reveal adsorption stages characterized by different segments in the plots of the dissipation vs frequency change. PEO remains protein-repellent for BSA, Lys, and Lf at pH 4-8.5, while weak adsorption of Fn was observed. On PAA, different stages of protein adsorption processes could be distinguished under most experimental conditions. BSA, Lys, Lf, and Fn generally exhibit a rapid initial adsorption phase on PAA, often followed by slower processes. The evaluation of the adsorption kinetics also reveals different adsorption stages, whereas the number of these stages does not always correspond to the structurally different phases as revealed by the D- f plots. The results presented here, together with information obtained in previous studies by other groups on the properties of these proteins and their interaction with surfaces, allow us to develop an adsorption scenario for each of these proteins, which takes into account electrostatic protein-surface and protein-protein interaction, but also the pH-dependent properties of the proteins, such as shape and exposure of specific domains.

Nanostructure Protein Repellant Amphiphilic Copolymer Coatings with Optimized Surface Energy by Inductively Excited Low Pressure Plasma

Statistically designed amphiphilic copolymer coatings were deposited onto Thermanox, Si wafer, and quartz crystal microbalance (QCM) substrates via Plasma Enhanced Chemical Vapor Deposition of 1H,1H,2H,2H-perfluorodecyl acrylate and diethylene glycol vinyl ether in an Inductively Excited Low Pressure Plasma reactor. Plasma deposited amphiphilic coatings were characterized by Field Emission Scanning Electron Microscopy, X-ray Photoelectron Spectroscopy, Atomic Force Microscopy, and Water Contact Angle techniques. The surface energy of the coatings can be adjusted between 12 and 70 mJ/m(2). The roughness of the coatings can be tailored depending on the plasma mode used. A very smooth coating was deposited with a CW (continuous wave) power, whereas a rougher surface with R(a) in the range of 2 to 12 nm was deposited with the PW (pulsed wave) mode. The nanometer scale roughness of amphiphilic PFDA-co-DEGVE coatings was found to be in the range of the size of the two proteins namely BSA and lysozyme used to examine for the antifouling properties of the surfaces. The results show that the statistically designed surfaces, presenting a surface energy around 25 mJ/m(2), present no adhesion with respect to both proteins measured by QCM.

Structured biotinylated poly(3,4-ethylenedioxypyrrole) electrodes for biochemical applications

The immobilization of biotin on transducer surfaces is a very important step for the fabrication of biosensors for many applications (immunoassay, DNA-hybridization assays, targeted imaging). Biotinylated polypyrroles have been studied and tested but gave rise to problems of polymerization and stability due to the intrinsic properties of pyrrole. As an alternative, biotinylated pyrroles were often used in a copolymerization with pyrrole or with an amphiphilic pyrrole derivative in a copolymerization to reduce the problems due to the pyrrole substitution. To find a new strategy, this paper presents the homopolymerization, instead of the use of a copolymerization, by replacing pyrrole by 3,4-ethylenedioxypyrrole bearing biotinylated substituent. We report the synthesis, characterization and electrochemical properties of two biotinylated 3,4-ethylenedioxypyrroles differing by the length of the alkyl spacer (ethyl or dodecyl) as well as the characterization of the corresponding polymer films. We successfully show, by cyclic voltammetry, that these monomers polymerize perfectly and give relatively stable polymer films. The increase of the alkyl spacer improves the polymerization and increases the polymer stability. For the first time, we also studied the surface morphology of an electrodeposited biotinylated polymer. The electrodeposition of these biotinylated derivatives gave rise to the ability to modulate the surface microstructuration, which consists of microspheres or cauliflower-like microstructures according to the length of the alkyl spacer.

Nanopatterned Surfaces for Bio-Detection

The development of advanced biosensor devices for very sensitive detection is highly required for many applications. The careful design of the bio-interface on the transducer surface is known to be one of the major bottlenecks for the development of high performance sensing devices. This paper reviews the emerging role of nanopatterned surfaces as alternative bio interface in the field of bio-detection. The different material used to functionalize this type of surfaces and the fabrication methodologies are described. Finally, the application of these surfaces in bio-detection using different biological systems and detection techniques is presented. In particular, a recent and very promising approach based on the optical interaction of nanoarrays with Surface Plasmon Resonance detection is described.

Fabrication of Bio-Functionalised Polypyrrole Nanoarrays for Bio-Molecular Recognition

The present study demonstrates that nanosphere lithography and electro-polymerization can be successfully combined to produce bioactive protein nanoarrays. In particular, we describe a method to produce well-defined nanoarrays of polypyrrole functionalized with biomolecules. The nanoarrayed surfaces were fabricated on gold coated surface plasmon resonance prisms by first creating silicon oxide or polyethylene oxide nanotemplate using nanosphere lithography. The nanotemplate was subsequently used to grow bio-functionalized polypyrrole nanoarrays by electrocopolymerization. Atomic force microscopy analysis showed that the fabricated surfaces have a well-organized 2D hexagonal geometry with nanoscale dimensions. The biological activity of the bio-functionalized polypyrrole was assessed by surface plasmon resonance detection. The results showed that the immobilized biomolecules within the nanoarrayed polypyrrole films had the necessary bioactivity for successful molecular recognition. Moreover the detection signals normalized to the bioactive area were increased by a factor 5 as compared to non-structured bio-functionalized polypyrrole in the nanoarrayed surfaces using polyethylene oxide.