Surface amination of poly(acrylonitrile)

The surface amination of poly (acrylonitrile) by ammonia plasma treatment has been studied. Furthermore, two other surface modification techniques have been investigated, the plasma chemical decomposition of an amino group containing chemical (tris-(2-aminoethyl)amine) onto the polymer surface and the surface reduction by lithium aluminium hydride. The three different methods are compared with respect to the adhesion improvement of the coatings onto the modified surfaces.The number of groups introduced on the surfaces has been determined by a wet chemical method.     

Determination of amine and aldehyde surface densities: application to the study of aged plasma treated polyethylene films

The aim of this work was to test and to compare different methods reported in the literature to quantify amine and aldehyde functions on the surface of polyethylene (PE) films treated by ammonia plasma and to specify their stability against time. A low pressure ammonia plasma reactor was used to functionalize PE films with amine groups, which could be subsequently used for further immobilization of biomolecules. In order to determine the density of amine groups on the surface of treated films, various molecule probes and spectrophotometric analytical methods have been investigated. Two methods using (i) sulfosuccinimidyl 6-[3'-(2-pyridyldithio)-propionamido] hexanoate (sulfo-LC-SPDP) and (ii) 2-iminothiolane (ITL) associated with bicinchoninic acid (BCA) have been proved to be reliable and sensitive enough to estimate the surface concentration of primary amine functions. The amount of primary amino groups on the functionalized polyethylene films was found to be between 1.2 and 1.4 molecules/nm2. In a second step, the surface concentration of glutaraldehyde (GA), which is currently used as a spacer arm before immobilization of biomolecules, has been assessed: two methods were used to determine the surface density of available aldehyde functions, after the reaction of GA with the aminated polyethylene film. The concentration of GA was found to be in the same range as primary amine concentration. The influence of aging time on the density of available amino and aldehyde groups on the surfaces were evaluated under different storage conditions. The results showed that 50% of the initial density of primary amine functions remained available after storage during 6 days of the PE samples in PBS (pH 7.6) at 4 degrees C. In the case of aldehyde groups, the same percentage of the initial density (50%) remained active after storage in air at RT over a longer period, i.e., 15 days.     

基于金属离子螯合的压电免疫传感器新型固定化方法

提出了一种基于金属离子螯合作用的压电免疫传感器新型固定化方法. 先在压电石英晶振表面沉积正丁胺等离子体聚合膜(BA-PPF), 再在BA-PPF表面修饰可与金属离子螯合的氨三乙酸基团, 用金属铜离子活化后, 修饰了二乙三胺五乙酸基团的IgG抗体蛋白质分子即可螯合固定于BA-PPF上. 将固定了抗体的压电石英传感器用于正常人免疫球蛋白IgG (NHIgG)的测定, 其频率响应与NHIgG浓度在0.36～63.8 μg/mL范围内存在良好的线性关系. 这种新型压电免疫传感器固定化方法简单快速, 具有良好的通用性.     

Solvent-induced porosity in ultrathin amine plasma polymer coatings

Plasma polymers deposited from n-heptylamine onto silicon wafers have been found to form a porous microstructure when immersed in water and other solvents, with pores of dimensions and densities that vary considerably between coatings deposited under different plasma conditions. This solvent-induced pore formation was found to correlate with the observed percentage of extractable material. With low radio frequency (rf) power inputs, the resultant softer coatings possess considerably more extractable material than coatings deposited at higher applied power levels. The porosity is thus proposed to result from the formation of voids created by the extraction of soluble low-molecular-weight polymeric material, which produces shrinkage stress that the coating, firmly attached to the substrate, cannot relieve by macroscopic contraction. The microscopic contraction of plasma polymer volume creates voids that appear to span the entire film thickness. The effect of aging plasma polymers in air was also investigated. For films deposited at low power it led to reduced extraction of soluble material and different pore morphology, whereas for films deposited at higher rf power levels, the extracted amounts and pore formation were the same for aged coatings. It was also found that the density of surface amine groups was lower for films deposited under the two lowest power settings, in contrast to the commonly held belief that the use of minimal applied rf power aids retention of functional groups. These porous plasma polymer coatings with surface groups suitable for further interfacial chemical immobilization reactions may be useful for various membrane and biotechnology applications.     

Immobilized streptavidin gradients as bioconjugation platforms

Surface density gradients of streptavidin (SAV) were created on solid surfaces and demonstrated functionality as a bioconjugation platform. The surface density of immobilized streptavidin steadily increased in one dimension from 0 to 235 ng cm(-2) over a distance of 10 mm. The density of coupled protein was controlled by its immobilization onto a polymer surface bearing a gradient of aldehyde group density, onto which SAV was covalently linked using spontaneous imine bond formation between surface aldehyde functional groups and primary amine groups on the protein. As a control, human serum albumin was immobilized in the same manner. The gradient density of aldehyde groups was created using a method of simultaneous plasma copolymerization of ethanol and propionaldehyde. Control over the surface density of aldehyde groups was achieved by manipulating the flow rates of these vapors while moving a mask across substrates during plasma discharge. Immobilized SAV was able to bind biotinylated probes, indicating that the protein retained its functionality after being immobilized. This plasma polymerization technique conveniently allows virtually any substrate to be equipped with tunable protein gradients and provides a widely applicable method for bioconjugation to study effects arising from controllable surface densities of proteins.     

Photochemical functionalization of polymer surfaces for microfabricated devices

Herein we report the topochemical modification of polymer surfaces with perfluorinated aromatic azides. The aryl azides, which have quaternary amine or aldehyde functional groups, were linked to the surface of the polymer by UV irradiation. The polymer substrates used in this study were cyclic olefin copolymer and poly(methyl methacrylate). These substrates were characterized before and after modification using reflection-absorption infrared spectroscopy, sessile water contact angle measurements, and X-ray photoelectron spectroscopy. Analysis of the surface confirmed the presence of aromatic groups with aldehyde or quaternary amine functionality. Enzyme immobilization and patterning onto polymer surfaces were studied using confocal microscopy. Enzymatic digests of protein were carried out on modified probes manufactured from thermoplastic substrates, and the resulting peptide analysis was completed using matrix-assisted laser desorption/ionization mass spectrometry. The use of functionalized perfluorinated aromatic azides allows the surface chemistry of thermoplastics to be tailored for specific lab-on-a-chip applications.     

Amination of Poly(ethylene-terephthalate) polymer surface for biochemical applications

Amine functionalization of Poly(ethylene-terephthalate) (PET) films for covalent binding of peptides is described. Ammonia plasma treatments have been used to graft nitrogen-containing functional groups onto the PET surface. The samples were then analyzed by X-ray photoelectron spectroscopy (XPS) and a parametric study was performed to define the best plasma grafting conditions. For biological tests, samples were sterilized by steam autoclaving: this induces a four to fivefold loss of the nitrogen functional groups on the polymer surface. XPS does not differentiate easily between the various nitrogen groups present on the surface so it is difficult to estimate the amount of surface amine groups available for direct coupling of bioactive molecules (proteins, peptides, nucleic acids, ...). To obtain a direct measurement of the amines present, we assayed for cysteine fixation through its carboxylic group by detection of the thiolaminoacid by XPS. We obtained cysteine fixation, showing the presence of grafted primary amine functions on PET surface after ammonia plasma treatment. Radiochemical assays were also made to quantify the amount of amine groups on plasma treated PET. XPS, cysteine fixation and radiochemical assays all show the presence of amine functions on ammonia plasma treated PET.     

Magnetically activated micromixers for separation membranes

Presented here is a radically novel approach to reduce concentration polarization and, potentially, also fouling by colloids present in aqueous feeds: magnetically responsive micromixing membranes. Hydrophilic polymer chains, poly(2-hydroxyethyl methacrylate) (PHEMA), were grafted via controlled surface-initiated atom transfer radical polymerization (SI-ATRP) on the surface of polyamide composite nanofiltration (NF) membranes and then end-capped with superparamagnetic iron oxide magnetite (Fe(3)O(4)) nanoparticles. The results of all functionalization steps, that is, bromide ATRP initiator immobilization, SI-ATRP, conversion of PHEMA end groups from bromide to amine, and carboxyl-functional Fe(3)O(4) nanoparticle immobilization via peptide coupling, have been confirmed by X-ray photoelectron spectroscopy (XPS) and field emission scanning electron microscopy (FESEM). These nanoparticles experience a magnetic force as well as a torque under an oscillating external magnetic field. It has been shown, using particle image velocimetry (PIV), that the resulting movement of the polymer brushes at certain magnetic field frequencies induces mixing directly above the membrane surface. Furthermore, it was demonstrated that with such membranes the NF performance could significantly be improved (increase of flux and salt rejection) by an oscillating magnetic field, which can be explained by a reduced concentration polarization in the boundary layer. However, the proof-of-concept presented here for the active alteration of macroscopic flow via surface-anchored micromixers based on polymer-nanoparticle conjugates has much broader implications.     

Immunosensing based on site-directed immobilization of antibody fragments and polymers that reduce nonspecific binding

Antibody Fab'-fragments can be directly coupled onto gold, and the space between the fragments can be filled with protein repellent disulfide bearing polymers. Coupling of the antibody Fab'-fragments, and thus both the amount of nonspecific binding and antigen binding but also the ability to regenerate the layer, is dependent on the immobilization procedure. First, the immobilization has taken place by coupling the Fab'-fragments to the surface and thereafter attaching the polymer in the remaining space between the antibodies. Second, the Fab'-fragments have been added after the surface has been coated by polymer. Third, the Fab'-fragments and polymer have been added onto the surface from the same solution. Up to 80% of the antigen could be removed during regeneration, if proper concentrations of polymer and Fab'-fragments were immobilized onto the gold surface. Only about 60% of the antigen could be removed, when the fragments were coupled directly onto a clean Au surface before the polymer or if low concentrations of polymer were attached onto gold before the Fab'-fragments. The first immobilization method, however, showed the highest response to antigen.     

Chemical modifications of inert organic monolayers with oxygen plasma for biosensor applications

In this paper we present a study of using oxygen plasma for chemically modifying inert hydrocarbon self-assembled monolayers of octadecyltrichlorosilane (OTS-SAMs) and rendering active surfaces for protein immobilization. Detailed surface modification and protein immobilization were characterized by using ellipsometry, X-ray photoelectron spectroscopy (XPS), Fourier transform infrared-attenuated total reflectance spectroscopy, and fluorescence microscopy. Our XPS results showed that the surface reaction between OTS-SAMs and oxygen plasma can generate new surface functional groups such as alcohol (C-O), aldehyde (C=O), and carboxylic acid (O-C=O), and their compositions can be controlled by using different treatment times and powers. A short treatment time ( approximately 1 s) and high power (10 W) can lead to a higher density of aldehyde groups, which can serve as linker groups for protein immobilization through the formation of Schiff bases with the amine groups of proteins. By using the fluorescence immunostaining method, we confirmed that human immunoglobulin (IgG) can be immobilized on a glass slide, only if the surface was decorated with OTS-SAMs and if the OTS-SAMs were pretreated with oxygen plasma. The protein immobilized on the oxygen-plasma-treated surface can only be recognized by using a highly specific antibody, FITC-anti-IgG, but not FITC-anti-biotin.     

Plasma modified polymers as a support for enzyme immobilization 1.: Allyl alcohol plasma

The paper describes deposition of plasma polymerized allyl alcohol on polysulfone film. It is shown that film surface becomes more hydrophilic after plasma treatment independently on presence of argon in a reaction mixture. The chemistry of the new surface layer was established by FTIR-ATR and ESCA spectroscopy. The substrate placed close to the plasma edge was the most hydrophilic but the amount of hydoxyl groups was not the highest there. Presence of argon stabilized the plasma but the deposited layer contained relatively less oxygen-bearing functionalities. The plasma treated polymer was subjected to xylose isomerase immobilization. For this purpose the divinylsulfone method was adapted. The studies revealed no correlation between the surface hydrophilicity and efficiency of immobilization.     

Covalent attachment of trypsin on plasma polymerized allylamine

This paper focuses on the immobilization of a proteolytic enzyme, trypsin, on plasma polymerized allylamine (ppAA) films. The later have been deposited onto silicon substrate by means of radiofrequency glow discharge. The covalent attachment of the enzyme was achieved in three steps: (i) activation of the polymer surface with glutaraldehyde (GA) as a linker, (ii) immobilization of trypsin and (iii) imino groups reduction treatment. The effects and efficiency of each step were investigated by X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM). Fluorescent spectroscopy was used to evaluate the change of the biological activity following the immobilization steps. The results showed that enzyme immobilization on GA-modified substrate increases the enzyme activity by 50% comparing to adsorbed enzymes, while the imino reduction treatment improves the enzyme retention by about 30% comparing to untreated samples. In agreement with XPS and AFM data, UV–vis absorption spectroscopy, used to quantify the amount of immobilized enzyme, showed that allylamine plasma polymer presents a high adsorption yield of trypsin. Although the adsorbed enzymes exhibit a lower activity than that measured for enzymes grafted through GA linkers, the highest catalytic activity obtained was for the enzymes that underwent the three steps of the immobilization process.     

Formation of Polymer Nanolayers with Special Properties at Polymer Surfaces

A principal possibility of the utilization of polymer peroxides for the modification of polymer surfaces that provides an immobilization of the tailored quantity of peroxide groups at them has been established. The processes of polymer surface activation with following “grafting from” or “grafting to” utilizing immobilized peroxide groups has been studied using a FTIR-ATR spectroscopy, ellipsometry and measurements of contact angles. The proposed technique of polymer surface activation is based on the universal ability of carbon chain polymer to participate the free radical reactions.     

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.     

一种新的基于正丁胺等离子体聚合膜的免疫传感器抗体(抗原)固定化方法

等离子体聚合膜 ( Plasma- polymerized film)是由有机蒸气在辉光放电下聚合而成 ,这种高度交联的膜具有均匀、超薄、附着力强、化学稳定、机械性能良好、基质类型多样以及成膜有机物品种多样等优点 ,因此已引起了传感器工作者的兴趣 ,目前 ,所研制的传感器已用于有机气体的测定 [1 ,2 ] .Karube小组报道了乙烯二胺等离子体聚合膜在免疫传感器方面的应用[3,4] .但由于抗体直接共价键合于等离子体聚合膜上 ,无法洗脱 ,使等离子体聚合膜修饰的传感器不能再生 ,而不同批次沉积的等离子体聚合膜其交联度、活性基团含量等又难以一致 ,严重影响了…     

Plasma polymerization and deposition of glycidyl methacrylate on Si(100) surface for adhesion improvement with polyimide

Thin polymer films were deposited on Si(100) surfaces by plasma polymerization of glycidyl methacrylate (GMA) under different glow discharge conditions. The FT‐IR, X‐ray photoelectron spectroscopy (XPS), and amine treatment results suggested that the epoxide functional groups of the deposited films had been preserved to various extents, depending on the plasma deposition conditions. The use of a low radio frequency power (～ 5 W) and a relatively high system pressure (100–400 Pa) readily resulted in the deposition of thin films having nearly the same composition of the epoxide functional groups as that of the GMA homopolymer. The plasma‐polymerized GMA (PP‐GMA) thin films deposited on the Ar plasma‐pretreated Si(100) surfaces were retained to a large extent after acetone extraction, suggesting the presence of covalent bonding between the PP‐GMA layer and the Si surface. Thermal imidization of the poly(amic acid) precursor of polyimide on the GMA plasma‐polymerized Si(100) surface resulted in a strongly adhered polyimide film. The adhesion results further suggested that the GMA polymer had been grafted on the Si(100) surface and the epoxide functional groups had undergone reactive interaction (curing) with the carboxylic and amine groups of the poly(amic acid) during thermal imidization. Copyright © 2001 John Wiley & Sons, Ltd.     

Development of an efficient amine-functionalized glass platform by additional silanization treatment with alkylsilane

Aminosilane-treated molecular layers on glass surfaces are frequently used as functional platforms for biosensor preparation. All the amino groups present on the surface are not available in reactive forms, because surface amino groups interact with remaining unreacted surface silanol groups. Such nonspecific interactions might reduce the efficiency of chemical immobilization of biomolecules such as DNA, enzymes, antibodies, etc., in biosensor fabrication. To improve immobilization efficiency we have used additional surface silanization with alkylsilane (capping) to convert the remaining silanol groups into Si–O–Si linkages, thereby liberating the amino groups from nonspecific interaction with the silanol groups. We prepared different types of capped amine surface and evaluated the effect of capping on immobilization efficiency by investigating the fluorescence intensity of Cy3-NHS (N-hydroxysuccinimide) dye that reacted with amino groups. The results indicate that most of the capped amine surfaces resulted in enhanced efficiency of immobilization of Cy3-NHS compared with the untreated control amine surface. We found a trend that trialkoxysilanes had greater capping effects on immobilization efficiency than monoalkoxysilanes. It was also found that the aliphatic chain of alkylsilane, which does not participate in the capping of the silanol, had an important function in enhancing immobilization efficiency. These results would be useful for preparation of an amine-modified surface platform, with enhanced immobilization efficiency, which is essential for developing many kinds of biosensors on a silica matrix. Enhancement of amine funtionality by capping with alkylsilane     

Mechanism of the Immobilization of Surfactants on Polymer Surfaces by Means of an Argon Plasma Treatment: Influence of UV Radiation

The mechanism of the immobilization of the surfactant sodium 10-undecenoate (C11(:)) on poly(ethylene) (PE) by means of an argon plasma treatment has been investigated. In particular, the influence of the vacuum ultraviolet (UV) radiation emitted by the argon plasma on the immobilization was studied. For this purpose, PE samples were coated with C11(:) (PE/C11(:) samples) and treated with an argon plasma under different conditions. PE/C11(:) samples were placed inside (glow) and outside (afterglow) the visible region of the plasma. Additionally, polymer samples that were placed in the glow of the plasma were covered with lithium fluoride or quartz crystals. These materials are transparent for electromagnetic radiation with a wavelength longer than 104 and 150 nm, respectively. Derivatization X-ray Photoelectron Spectroscopy was applied to characterize the modified polymer surfaces. It was demonstrated that vacuum UV radiation with a wavelength shorter than 150 nm made a predominant contribution to the process of immobilization. Under certain conditions it was possible to retain about 30% of the functional groups of the initially coated surfactant layer on PE. Furthermore, the UV radiation accounted for etching of PE and PE/C11(:) surfaces and initiated oxidation of the polymer surfaces.     

Facile immobilization of biomolecules onto various surfaces using epoxide-containing antibiofouling polymers

The surface modifications of plastic or glass substrate and the subsequent immobilization of biomolecules onto the surfaces has been a central feature of the fabrication of biochips. To this end, we designed and synthesized new epoxide-containing random copolymers that form stable polymer adlayers on plastic or glass surface and subsequently react with amine or sulfhydryl functional groups of biomolecules under aqueous conditions. Epoxide-containing random copolymers were synthesized by radical polymerization of three functional monomers: a monomer acting as an anchor to the surfaces, a PEG group for preventing nonspecific protein adsorption, and an epoxide group for conjugating to biomolecules. Polymer coating layers were facilely formed on cyclic olefin copolymer (COC) or glass substrate by simply dipping each substrate into a solution of each copolymer. The polymer-coated surfaces characterized by a contact angle analyzer and X-ray photoelectron spectroscopy (XPS) showed very low levels of nonspecific immunoglobulin G (IgG) adsorption compared to the uncoated bare surface (control). Using a microcontact printing (μCP) method, antibodies as representative biomolecules could be selectively attached onto the copolymers-coated glass or COC surface with high signal-to-noise ratios.     

Biofouling of dextran-derivative layers investigated by quartz crystal microbalance

This study reports the fouling of carboxymethyl dextran (CMD) layers in cell culture medium, fibronectin, and serum solutions. CMD layers were covalently immobilized onto amine groups available either on an n-heptylamine plasma polymer (HApp) layer or onto a polyethylenimine (PEI) coating grafted to an acetaldehyde plasma polymer (AApp) layer. The successful immobilization of the graft layers was demonstrated by X-ray photoelectron spectroscopy (XPS). Primary amines on HApp and AApp + PEI surfaces were quantified using a colorimetric assay. Quartz crystal microbalance (QCM) was used to investigate in real-time the fouling of the graft layers upon incubation in cell culture medium (RPMI), fibronectin, and foetal bovine serum (FBS) solutions. HApp, AApp and AApp + PEI layers exhibited large fouling in fibronectin and FBS solutions, while fouling in RPMI cell culture medium was not significant. Protein repellent properties of CMD layers in FBS and fibronectin have been demonstrated compared to the other tested surfaces. QCM has shown that both CMD layers were fouled to a small extent in RPMI medium.