In-channel atom-transfer radical polymerization of thermoset polyester microfluidic devices for bioanalytical applications

A new technique for polymer microchannel surface modification, called in-channel atom-transfer radical polymerization, has been developed and applied in the surface derivatization of thermoset polyester (TPE) microdevices with poly(ethylene glycol) (PEG). X-ray photoelectron spectroscopy, electroosmotic flow (EOF), and contact angle measurements indicate that PEG has been grafted on the TPE surface. Moreover, PEG-modified microchannels have much lower and more pH-stable EOF, more hydrophilic surfaces and reduced nonspecific protein adsorption. Capillary electrophoresis separation of amino acid and peptide mixtures in these PEG-modified TPE microchips had good reproducibility. Phosducin-like protein and phosphorylated phosducin-like protein were also separated to measure the phosphorylation efficiency. Our results indicate that PEG-grafted TPE microchips have broad potential application in biomolecular analysis.     

Microfluidic flow control on charged phospholipid polymer interface

A type of charged phospholipid polymer biointerface was constructed on a quartz microfluidic chip to control the electroosmotic flow (EOF) and to suppress non-specific protein adsorption through one-step modification. A negatively charged phospholipid copolymer containing 2-methacryloyloxyethyl phosphorylcholine (MPC), n-butyl methacrylate (BMA), potassium 3-methacryloyloxypropyl sulfonate (PMPS) and 3-methacryloxypropyl trimethoxysilane (MPTMSi) moieties (referred to as PMBSSi) was synthesized to introduce such phosphorylcholine segments as well as surface charges onto the silica-based microchannels via chemical bonding. At neutral pH, the homogenous microchannel surface modified with 0.3 wt% PMBSSi in alcoholic solution, retained a significant cathodic EOF ((1.0 +/- 0.1) x 10(-4) cm(2) V(-1) s(-1)) with approximately one-half of the EOF of the unmodified microchannel ((1.9 +/- 0.1) x 10(-4) cm(2) V(-1) s(-1)). Along with another non-charged copolymer (poly(MPC-co-MPTMSi), PMSi), the regulation of the surface charge density can be realized by adjusting the concentration of PMBSSi or PMSi initial solutions for modification. Coincidently, the zeta-potential and the EOF mobility at neutral pH showed a monotonically descending trend with the decrease in the charge densities on the surfaces. This provides a simple but feasible approach to controlling the EOF, especially with regard to satisfying the requisites of miniaturized systems for biological applications requiring neutral buffer conditions. In addition, the EOF in microchannels modified with PMBSSi and PMSi could maintain stability for a long time at neutral pH. In contrast to the EOF in the unmodified microchannel, the EOF in the modified microchannel was only slightly affected by the change in pH (from 1 to 10). Most importantly, although PMBSSi possesses negative charges, the non-specific adsorptions of both anionic and cationic proteins (considering albumin and cytochrome c, respectively, as examples) were effectively suppressed to a level of 0.15 microg cm(-2) and lesser in the case of the 0.3 wt% PMBSSi modification. Consequently, the variation in the EOF mobility resulting from the protein adsorption was also suppressed simultaneously. To facilitate easy EOF control with compatibility to biomolecules delivered in the microfluidic devices, the charged interface described could provide a promising option.     

Non-fouling microfluidic chip produced by radio frequency tetraglyme plasma deposition

This Technical Note presents the direct surface modification of a glass/PTFE hybrid microfluidic chip, via radio frequency glow discharge plasma polymerisation of tetraethlylene glycol dimethylether (tetraglyme), to produce hydrophilic, non-fouling, PEO-like surfaces. We use several techniques including X-ray photoelectron spectroscopy (XPS), direct enzyme-linked immunosorbent assays (ELISA) and immunofluorescent imaging to investigate the channel coatings. Our results indicate the successful deposition of a PEO-like coating onto microchannel surfaces that has both solution and shelf stability (>3 months) and is capable of preventing fibrinogen adsorption to the microchannel surfaces.     

Microchannel wall coatings for protein separations by capillary and chip electrophoresis

The necessity for microchannel wall coatings in capillary and chip-based electrophoretic analysis of biomolecules is well understood. The regulation or elimination of EOF and the prevention of analyte adsorption is essential for the rapid, efficient separation of proteins and DNA within microchannels. Microchannel wall coatings and other wall modifications are especially critical for protein separations, both in fused-silica capillaries, and in glass or polymeric microfluidic devices. In this review, we present a discussion of recent advances in microchannel wall coatings of three major classes--covalently linked polymeric coatings, physically adsorbed polymeric coatings, and small molecule additives. We also briefly review modifications useful for polymeric microfluidic devices. Within each category of wall coatings, we discuss those used to eliminate EOF, to tune EOF, to prevent analyte adsorption, or to perform multiple functions. The knowledgeable application of the most promising recent developments in this area will allow for the separation of complex protein mixtures and for the development of novel microchannel wall modifications.     

Silicate glass coated microchannels through a phase conversion process for glass-like electrokinetic performance

The surface modified polydimethylsiloxane (PDMS) microchannels show a much more inferior performance to the durable and reproducible glass chip. In this paper, a facile approach to preparing a silicate glass modified PDMS microchannel for glass-like performance is presented. This glass-like performance is made possible by a phase conversion of a preceramic polymer--allylhydridopolycarbosilane (AHPCS). The, several hundred nanometer thick, polymer that coats the PDMS channel is hydrolyzed to form hydrophilic silicate glass via phase conversion under an aqueous alkali condition. It is characterized by XPS, FTIR-ATR, AFM, and contact angle measurements. The silicate glass coated PDMS channel from AHPCS has an excellent solvent resistance, delivers a high electroosmotic flow (EOF) that is stable in the long-term (4.9±0.1×10(-4) cm(2) V(-1) s(-1)) and a reliable capillary electrophoresis (CE), which are comparable to those of native glass channels. Moreover, the silicate glass PDMS channel allows easy regeneration of the electrokinetic behavior, just as in a glass channel, by a simple treatment with alkali solution. This coating approach can be applied to other polymer substrates such as polyimide (PI).     

Surface grafted sulfobetaine polymers via atom transfer radical polymerization as superlow fouling coatings

One of the sulfobetaine methacrylate (SBMA) monomers, N-(3-sulfopropyl)-N-(methacryloxyethyl)-N,N-dimethylammonium betaine, was polymerized onto initiator-covered gold surfaces using atom transfer radical polymerization (ATRP) to form uniform polymer brushes. Self-assembled monolayers (SAMs) with ATRP initiators were characterized by X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM). The thickness of grafted poly(SBMA) films was measured by ellipsometry. Fibrinogen adsorption on poly(SBMA) grafted surfaces was measured with a surface plasmon resonance (SPR) sensor. Two approaches were compared to graft ATRP initiators onto gold surfaces for surface polymerization and subsequent protein adsorption on these polymer grafted surfaces. The first was to prepare a SAM from omega-mercaptoundecyl bromoisobutyrate onto a gold surface. Superlow fouling surfaces with well-controlled poly(SBMA) brushes were achieved using this approach (e.g., fibrinogen adsorption <0.3 ng/cm2). The second approach was to react bromoisobutyryl bromide with a hydroxyl-terminated SAM on a gold surface. Although protein adsorption decreased as the density of surface initiators increased, the surface prepared using the second approach was not able to achieve as low protein adsorption as the first approach. Key parameters to achieve superlow fouling surfaces were studied and discussed.     

Surface modification of polymer microfluidic devices using in-channel atom transfer radical polymerization

In-channel atom transfer radical polymerization (ATRP) was used to graft a PEG layer on the surface of microchannels formed in poly(glycidyl methacrylate)-co-(methyl methacrylate) (PGMAMMA) microfluidic devices. The patterned and cover plates were first anchored with ATRP initiator and then thermally bonded together, followed by pumping a solution containing monomer, catalyst, and ligand into the channel to perform ATRP. A PEG-functionalized layer was grafted on the microchannel wall, which resists protein adsorption. X-ray photoelectron spectroscopy (XPS) was used to investigate the initiator-bound surface, and EOF was measured to evaluate the PEG-grafted PGMAMMA microchannel. Fast, efficient, and reproducible separations of amino acids, peptides, and proteins were obtained using the resultant microdevices. Separation efficiencies were higher than 1.0x10(4) plates for a 3.5 cm separation microchannel. Compared with microdevices modified using a previously reported ATRP technique, these in-channel modified microdevices demonstrated better long-term stability.     

Surface modification on microfluidic devices with 2-methacryloyloxyethyl phosphorylcholine polymers for reducing unfavorable protein adsorption

Surface modification of polymer materials for preparing microfluidic devices including poly(dimethyl siloxane) (PDMS) was investigated with phospholipids polymers such as poly(2-methacryloyloxylethyl phosphorylcholine(MPC)-co-n-butyl methacrylate) (PMB) and poly(MPC-co-2-ethylhexyl methacrylate-co-2-(N,N-dimethylamino)ethyl methacrylate) (PMED). The hydrophilicity of every surface on the polymer materials modified with these MPC polymers increased and the value of zeta-potential became close to zero. The protein adsorption on the polymer materials with and without the surface modification was evaluated using a protein mixture of human plasma fibrinogen and serum albumin. Amount of proteins adsorbed on these polymeric materials showed significant reduction by the surface modification with the MPC polymers compared to the uncoated surfaces ranging from 56 to 90%. Furthermore, we successfully prepared PDMS-based microchannel which was modified by simple coating with the PMB and PMED. The modified microchannel also revealed a significant reduction of adsorption of serum albumin. We conclude that the MPC polymers are useful for reducing unfavorable protein adsorption on microfluidic devices.     

Grafting epoxy-modified hydrophilic polymers onto poly(dimethylsiloxane) microfluidic chip to resist nonspecific protein adsorption

In order to achieve a simple covalent hydrophilic polymer coating on poly(dimethylsiloxane) (PDMS) microfluidic chip, epoxy modified hydrophilic polymers were synthesized in aqueous solution with a persulfate radical initiation system, and crosslinked onto PDMS pretreated by oxygen plasma and silanized with 3-aminopropyl-triethoxysilanes (APTES). Glycidyl methacrylate (GMA) was copolymerized with acrylamide (poly(AAM-co-GMA)) or dimethylacrylamide (poly(DAM-co-GMA)), and graft polymerized with polyvinylpyrrolidone (PVP-g-GMA) or polyvinylalcohol (PVA-g-GMA). The epoxy groups in the polymers were determined by UV spectra after derivation with benzylamine. Reflection absorption infrared spectroscopy (RAIRS) confirmed covalent grafting of GMA-modified polymers onto PDMS surface. Electroosmotic flow (EOF) in the polymer grafted microchannel was strongly suppressed within the range pH 3-11. Surface adsorption of lysozyme and bovine serum albumin (BSA) was reduced to less than 10% relative to that on the native PDMS surface. On the GMA-modified polymer coated PDMS microchip, basic proteins, peptides, and sodium dodecyl sulfate (SDS) denatured proteins were separated successfully.     

Polyurethane-based microfluidic devices for blood contacting applications

Protein adsorption on PDMS surfaces poses a significant challenge in microfluidic devices that come into contact with biofluids such as blood. Polyurethane (PU) is often used for the construction of medical devices, but despite having several attractive properties for biointerfacing, it has not been widely used in microfluidic devices. In this work we developed two new fabrication processes for making thin, transparent and flexible PU-based microfluidic devices. Methods for the fabrication and bonding of microchannels, the integration of fluidic interconnections and surface modification with hydrophilic polyethylene oxide (PEO) to reduce protein adsorption are detailed. Using these processes, microchannels were produced having high transparency (96% that of glass in visible light), high bond strength (326.4 kPa) and low protein adsorption (80% reduction in fibrinogen adsorption vs. unmodified PDMS), which is critical for prevention of fouling. Our findings indicate that PEO modified PU could serve as an effective alternative to PDMS in blood contacting microfluidic applications.     

Dynamic coating using methylcellulose and polysorbate 20 for nondenaturing electrophoresis of proteins on plastic microchips

A dynamic coating using methylcellulose (MC) and a nonionic detergent (polysorbate 20) was developed, which controlled protein adsorption onto the surface of microchannels on a microchip made of poly(methyl methacrylate) (PMMA). Optimum concentration of polysorbate 20 in combination with the range of MC concentrations controlled the protein adsorption onto the microchannel surface, and increased the solubility of the protein samples while facilitating the injection of high concentrations of MC solutions into the microchannels. Higher concentrations of nonionic detergent increased the EOF mobility as opposed to the electrophoretic mobility and caused the electrophoresis to fail. Nondenaturing microchip electrophoresis of protein samples with molecular masses ranging from 20 to 100 kDa were completed in 100 s. Also, successful separation of a BSA sample and its complex with anti-BSA mAb ( 220 kDa) was achieved on a PMMA microchip. The separation exhibited high reproducibility in both migration time (RSD = 1%) and peak area (RSD = 10-15%).     

Surface modification with BSA blocking based on in situ synthesized gold nanoparticles in poly(dimethylsiloxane) microchip

A stable BSA blocking poly(dimethylsiloxane) (PDMS) microchannel was prepared based on in situ synthesized PDMS–gold nanoparticles composite films. The modified microchip could successfully suppress protein adsorption. The assembly was followed by contact angle, charge-coupled device (CCD) imaging, electroosmotic flow (EOF) measurements and electrophoretic separation methods. Contact angle measurements revealed the coated surface was hydrophilic, water contact angle for coated chips was 45.2° compared to a water contact angle for native PDMS chips of 88.5°. The coated microchips exhibited reproducible and stable EOF behavior. With FITC-labeled myoglobin incubation in the coated channel, no fluorescence was observed with CCD image, and the protein exhibited good electrophoretic effect in the modified microchip.     

On the surface modification of microchannels for microcapillary electrophoresis chips

This paper presents systematic investigation of the microchannel surface properties in microCE chips. Three popular materials for microCE chips, polydimethylsiloxane (PDMS), quartz, and glass, are used. The zeta potentials of these microchannels are calculated by measuring the EOF velocity to evaluate the surface properties after surface modification. The hydrophobic PDMS is usually plasma-treated for microCE applications. In this study, a new method using a high-throughput atmospheric plasma generator is adopted to treat the PDMS surface under atmospheric conditions. In this approach, the cost and time for surface treatment can be significantly reduced compared with the conventional vacuum plasma generator method. Experimental results indicate that new functional groups could be formed on the PDMS surface after treatment, resulting in a change in the surface property. The time-dependent surface property of the plasma-treated PDMS is then measured in terms of the zeta potential. Results show that the surface property will reach a stable condition after 1 h of plasma treatment. For glass CE chips, two new methods for changing the microchannel surface properties are developed. Instead of using complicated and time-consuming chemical silanization procedures for CE channel surface modification, two simple and reliable methods utilizing organic-based spin-on-glass and water-soluble acrylic resin are reported. The proposed method provides a fast batch process for controlling the surface properties of glass-based CE channels. The proposed methods are evaluated using PhiX-174 DNA maker separation. The experimental data show that the surface property is modified and separation efficiency greatly improved. In addition, the long-term stability of both coatings is verified in this study. The methods proposed in this study show potential as an excellent solution for glass-based microCE chip surface modification.     

Water-vapor plasma-based surface activation for trichlorosilane modification of PMMA

Separation rates and resolutions within capillary electrophoretic (CE) systems can be enhanced when surface zeta potentials are uniform with minimum deviations from ideal pluglike flow. Microfluidic CE devices based on poly(methyl methacrylate) (PMMA) are being developed due to the optical clarity, availability, stability, and reproducible electroosmotic flow (EOF) rates displayed by this polymer. Control of EOF in polymer-based CE systems can be achieved by surface zeta potential alteration through chemical modification. Herein, a method will be presented for the surface functionalization of PMMA with chemistry analogous to formation of trichlorosilane self-assembled monolayers on SiO2. The current method involves two separate steps. First, surface activation with water-vapor plasma introduces surface hydroxylation. Second, treatment of the plasma-treated PMMA with a substituted trichlorosilane solution forms the functional surface layer. The modified surfaces were characterized using several analytical techniques, including water contact angle, X-ray photoelectron spectroscopy, Fourier transform infrared-attenuated total reflection, secondary ion mass spectroscopy, and measurement of EOF velocities within PMMA microchannels.     

Streaming potential and electroosmotic flow in heterogeneous circular microchannels with nonuniform zeta potentials: requirements of flow rate and current continuities

Real surfaces are typically heterogeneous, and microchannels with heterogeneous surfaces are commonly found due to fabrication defects, material impurities, and chemical adsorption from solution. Such surface heterogeneity causes a nonuniform surface potential along the microchannel. Other than surface heterogeneity, one could also pattern the various surface potentials along the microchannels. To understand how such variations affect electrokinetic flow, we proposed a model to describe its behavior in circular microchannels with nonuniform surface potentials. Unlike other models, we considered the continuities of flow rate and electric current simultaneously. These requirements cause a nonuniform electric field distribution and pressure gradient along the channel for both pressure-driven flow (streaming potential) and electric-field-driven flow (electroosmosis). The induced nonuniform pressure and electric field influence the electrokinetic flow in terms of the velocity profile, the flow rate, and the streaming potential.     

Modeling of nucleic acid adsorption on 3D prisms in microchannels

Understanding nucleic acid adsorption in microchannels is critical to improve the efficiency of purifying and extracting nucleic acid (NA) from sample solutions by microfluidic technologies. Using a microchannel with 3D prismatic silica elements on the wall can dramatically increase the surface area-to-volume ratio, and hence facilitate the nucleic acid adsorption on the wall. In this study a theoretical model for modeling adsorption in a microchannel with a designed 3D surface structure was developed, and five dimensionless numbers were found to be the key parameters in the adsorption process. Extensive numerical simulations were conducted. Two flow modes, the electroosmotic flow (EOF) and pressure-driven flow (PDF), were investigated for their effect on the adsorption. It was found that the EOF is more desirable than PDF. The 3D prismatic elements can increases the NA molecule adsorption not only by providing more surface areas, but also by the induced pressure resisting the central bulk electroosmotic flow. Finally, the effects of adsorption kinetic parameters (i.e., the kinetic association/dissociation constants, the diffusion coefficient, the total site density, the loading concentration, and the channel height), on the adsorption process were discussed in detail.     

Characterization of fibrinogen adsorption onto glass microcapillary surfaces by ELISA

Adsorption of biomolecules onto microchannel surfaces remains a critical issue in microfluidic devices. This paper investigates the adsorption of fibrinogen on glass microcapillaries using an immunoassay method (ELISA) and X-ray photoelectron spectroscopy (XPS). Various adsorption conditions such as protein concentrations and incubation times, buffer pH, buffer ionic strengths and effects of flow are presented. ELISA is successfully demonstrated as a facile and robust technique to examine these phenomena. The highest adsorption level occurs near the isoelectric point of fibrinogen (pH 5.0) and low buffer ionic strengths (0-8 mM). Microchannel surface saturation was achieved at a fibrinogen solution concentration of approximately 50 microg ml(-1). Fibrinogen adsorption under flow was always higher than that seen in static systems. The importance of diffusion phenomena in microchannels on protein adsorption was demonstrated. ELISA experiments using fused silica and PEEK have also confirmed significant adsorption on these mass spectrometer transfer line materials.     

A novel method of preparing of glass capillary columns: Low-temperature plasma etching and polymerization

A novel method is described for the preparation of stable glass capillary columns (glass open tubular columns), including the etching and formation of a polymer film on the inner glass capillary surfaces. The approach used here is based on low-temperature plasma etching and polymerization. Under the influence of a field of radio frequency discharge, low pressure gases of fluoric compounds, introduced into the glass capillary tube, generate excited fluorine radicals which etch the inner surface. The plasma of organosilicone monomer in the glass capillary yields a uniform polymerized film on the inner surface. The resultant material functions as a good stationary phase for glass capillary gas chromatography (GC²). The inner surfaces treated with such a plasma, can be studied by means of a scanning electron microscope (SEM). The flexibility of this method permits the use of various stationary phases and surface modification.     

Fouling and protein adsorption effect of low-temperature plasma treatment of membrane surfaces

Adsorption of proteins and the effect of the chemical nature of membrane surfaces on protein adsorption were investigated using¹⁴C-tagged albumin and several microporous membranes (polyvinilydene fluoride, PVDF; nylon; polypropylene, PP; and polycarbonate, PC). The membrane surfaces were modified by exposing them to low-temperature plasma of several different monomers (n-butane, oxygen, nitrogen alone or as mixtures) in a radiofrequency plasma reactor. Transients in the permeability of albumin solutions through the membranes and changes in flux of distilled water through the membranes before and after adsorption of albumin were used to investigate the role of protein adsorption on membrane fouling. The results show that the extent of adsorption of albumin on hydrophobic membranes was considerably more than that on hydrophilic membranes. The hydrophilic membranes were susceptible to electrostatic interactions and less prone to fouling. A pore-blocking model was successfully used to correlate the loss of water flux through pores of defined geometry     

Poly(oxyethylene) based surface coatings for poly(dimethylsiloxane) microchannels

Control of surface properties in microfluidic systems is an indispensable prerequisite for successful bioanalytical applications. Poly(dimethylsiloxane) (PDMS) microfluidic devices are hampered from unwanted adsorption of biomolecules and lack of methods to control electroosmotic flow (EOF). In this paper, we propose different strategies to coat PDMS surfaces with poly(oxyethylene) (POE) molecules of varying chain lengths. The native PDMS surface is pretreated by exposure to UV irradiation or to an oxygen plasma, and the covalent linkage of POE-silanes as well as physical adsorption of a triblock-copolymer (F108) are studied. Contact angle measurements and atomic force microscopy (AFM) imaging revealed homogeneous attachment of POE-silanes and F108 to the PDMS surfaces. In the case of F108, different adsorption mechanisms to hydrophilic and hydrophobic PDMS are discussed. Determination of the electroosmotic mobilities of these coatings in PDMS microchannels prove their use for electrokinetic applications in which EOF reduction is inevitable and protein adsorption has to be suppressed.