Separation of proteins on surface-modified poly(dimethylsiloxane) microfluidic devices

Separation and detection of proteins have been realized on nonionic surfactant-modified poly(dimethylsiloxane) (PDMS) microfabricated devices with end-column amperometric detection. The hydrophobic PDMS channels are turned into hydrophilic ones after being modified with Brij35 and facilitate the separation of proteins. The coating can remarkably reduce the adsorption of large protein molecules and is stable in the range of pH 6-12. The detection of proteins in such channels needs less rinsing time and thus efficiency is raised. Even large molecules of proteins can also be detected with better reproducibility and enhanced plate numbers. The relative standard deviation (RSD) of the migration time for glucose oxidase (GOD) is 2.2% (n = 19). Separation of GOD and myoglobin has been developed in modified channels. Predominant operational variables, such as the coating conditions, the concentration of surfactant and buffer, are studied in detail.     

Integrated microfluidic devices

“With the fundamentals of microscale flow and species transport well developed, the recent trend in microfluidics has been to work towards the development of integrated devices which incorporate multiple fluidic, electronic and mechanical components or chemical processes onto a single chip sized substrate. Along with this has been a major push towards portability and therefore a decreased reliance on external infrastructure (such as detection sensors, heaters or voltage sources).” In this review we provide an in-depth look at the “state-of-the-art” in integrated microfludic devices for a broad range of application areas from on-chip DNA analysis, immunoassays and cytometry to advances in integrated detection technologies for and miniaturized fuel processing devices. In each area a few representative devices are examined with the intent of introducing the operating procedure, construction materials and manufacturing technique, as well as any unique and interesting features.     

Polymer microfluidic devices

Since the introduction of lab-on-a-chip devices in the early 1990s, glass has been the dominant substrate material for their fabrication (J. Chromatogr. 593 (1992) 253; Science 261 (1993) 895). This is primarily driven by the fact that fabrication methods were well established by the semiconductor industry, and surface properties and derivatization methods were well characterized and developed by the chromatography industry among others. Several material properties of glass make it a very attractive material for use in microfluidic systems; however, the cost of producing systems in glass is driving commercial producers to seek other materials. Commercial manufacturers of microfluidic devices see many benefits in employing plastics that include reduced cost and simplified manufacturing procedures, particularly when compared to glass and silicon. An additional benefit that is extremely attractive is the wide range of available plastic materials which allows the manufacturer to choose materials' properties suitable for their specific application. In this article, we present a review of polymer-based microfluidic systems including their material properties, fabrication methods, device applications, and finally an analysis of the market that drives their development.     

Polyimide-based microfluidic devices

This paper describes the development of polyimide-based microfluidic devices. A layer transfer and lamination technique is used to fabricate flexible microfluidic channels in various shapes and with a wide range of dimensions. High bond strengths can be achieved by cure cycle adaptation and surface treatment of the polyimide layers prior to bonding. The polyimide microchannels can be combined with metallization layers to fabricate electrodes inside and outside channels. The resulting devices can be used for flexible fluidic and electrical connectors, implantable fluid delivery devices, microelectrodes with embedded fluidic channels, chip-based flow cytometry and for a great variety of other applications in medical, chemical or biological research.     

Genotyping with microfluidic devices

In the past few years, electrophoresis microchips have been increasingly utilized to interrogate genetic variations in the human and other genomes. Microfluidic devices can be readily applied to speed up existing genotyping protocols, in particular the ones that require electric field-mediated separations in conjunction with restriction fragment analysis, DNA sequencing, hybridization-based techniques, allele-specific amplification, heteroduplex analysis, just to list the most important ones. As a result of recent developments, microfabricated electrophoresis devices offer several advantages over conventional slab-gel electrophoresis, such as small sample volume requirement, low reagent consumption, the option of system integration and easy multiplexing. The analysis speed of microchip electrophoresis is significantly higher than that of any other electric field-mediated separation techniques. State-of-the-art microfluidic bioanalytical devices already claim their place in most molecular biology laboratories. This review summarizes the recent developments in microchip electrophoresis methods of nucleic acids, particularly for rapid genotyping, that will most likely play a significant role in the future of clinical diagnostics.     

Bioanalysis in microfluidic devices

Microfabricated bioanalytical devices (also referred to as laboratory-on-a-chip or micro-TAS) offer highly efficient platforms for simultaneous analysis of a large number of biologically important molecules, possessing great potential for genome, proteome and metabolome studies. Development and implementation of microfluidic-based bioanalytical tools involves both established and evolving technologies, including microlithography, micromachining, micro-electromechanical systems technology and nanotechnology. This article provides an overview of the latest developments in the key device subject areas and the basic interdisciplinary technologies. Important aspects of DNA and protein analysis, interfacing issues and system integration are all thoroughly discussed, along with applications for this novel "synergized" technology in high-throughput separations of biologically important molecules. This review also gives a better understanding of how to utilize these technologies as well as to provide appropriate technical solutions to problems perceived as being more fundamental.     

Nonlithographic fabrication of microfluidic devices

A facile nonlithographic method for expedient fabrication of microfluidic devices of poly(dimethylsiloxane) is described. Positive-relief masters for the molds are directly printed on smooth substrates. For the formation of connecting channels and chambers inside the polymer components of the microfluidic devices, cavity-forming elements are adhered to the surfaces of the masters. Using this nonlithographic approach, we fabricated microfluidic devices for detection of bacterial spores on the basis of enhancement of the emission of terbium (III) ions.     

Surface-imprinted polymers in microfluidic devices

Molecularly imprinted polymers are generated by curing a cross-linked polymer in the presence of a template. During the curing process, noncovalent bonds form between the polymer and the template. The interaction sites for the noncovalent bonds become "frozen" in the cross-linking polymer and maintain their shape even after the template is removed. The resulting cavities reproduce the size and shape of the template and can selectively reincorporate the template when a mixture containing it flows over the imprinted surface. In the last few decades the field of molecular imprinting has evolved from being able to selectively capture only small molecules to dealing with all kinds of samples. Molecularly imprinted polymers (MIPs) have been generated for analytes as diverse as metal ions, drug molecules, environmental pollutants, proteins and viruses to entire cells. We review here the relatively new field of surface imprinting, which creates imprints of large, biologically relevant templates. The traditional bulk imprinting, where a template is simply added to a prepolymer before curing, cannot be applied if the analyte is too large to diffuse from the cured polymer. Special methods must be used to generate binding sites only on a surface. Those techniques have solved crucial problems in separation science as well as chemical and biochemical sensing. The implementation of imprinted polymers into microfluidic chips has greatly improved the applicability of microfluidics. We present the latest advances and different approaches of surface imprinting and their applications for microfluidic devices.     

Urine analysis in microfluidic devices

Microfluidics has attracted considerable attention since its early development in the 1980s and has experienced rapid growth in the past three decades due to advantages associated with miniaturization, integration and automation. Urine analysis is a common, fast and inexpensive clinical diagnostic tool in health care. In this article, we will be reviewing recent works starting from 2005 to the present for urine analysis using microfluidic devices or systems and to provide in-depth commentary about these techniques. Moreover, commercial strips that are often treated as chips and their readers for urine analysis will also be briefly discussed. We start with an introduction to the physiological significance of various components or measurement standards in urine analysis, followed by a brief introduction to enabling microfluidic technologies. Then, microfluidic devices or systems for sample pretreatments and for sensing urinary macromolecules, micromolecules, as well as multiplexed analysis are reviewed, in this sequence. Moreover, a microfluidic chip for urinary proteome profiling is also discussed, followed by a section discussing commercial products. Finally, the authors' perspectives on microfluidic-based urine analysis are provided. These advancements in microfluidic techniques for urine analysis may improve current routine clinical practices, particularly for point-of-care (POC) applications.     

Model-controlled hydrodynamic focusing to generate multiple overlapping gradients of surface-immobilized proteins in microfluidic devices

Historically, it has been difficult to generate accurate and reproducible protein gradients for studies of interactions between cells and extracellular matrix. Here we demonstrate a method for rapid patterning of protein gradients using computer-driven hydrodynamic focusing in a simple microfluidic device. In contrast to published work, we are moving the complexity of gradient creation from the microfluidic hardware to dynamic computer control. Using our method, switching from one gradient profile to another requires only a few hours to devise a new control file, not days or weeks to design and build a new microfluidic device. Fitting existing protein deposition models to our data, we can extract key parameters needed for controlling protein deposition. Several protein deposition models were evaluated under microfluidic flow conditions. A mathematical model for our deposition method allows us to determine the parameters for a protein adsorption model and then predict the final shape of the surface density gradient. Simple and non-monotonic single and multi-protein gradient profiles were designed and deposited using the same device.     

Highly efficient dynamic modification of plastic microfluidic devices using proteins in microchip capillary electrophoresis

New dynamic coating agents were investigated for the manipulation of electroosmotic flow (EOF) in poly(methylmethacrylate) (PMMA) microchips. Blocking proteins designed for enzyme-linked immunosorbent assay (ELISA) applications (e.g. Block Ace and UltraBlock), and egg-white lysozyme were proposed in this study. The EOF could be enhanced, suppressed or its direction could be reversed, depending on the buffer pH and the charge on the proteins. The coating procedure is simple, requiring only filling of the microchannels with a coating solution, followed by a rinse with a running buffer solution prior to analysis. One major advantage of this method is that it is not necessary to add the coating agent to the running buffer solution. Block Ace and UltraBlock coatings were stable for at least five runs in a given microchannel without the need to condition the coating between runs other than replenishing the buffer solution after each run, i.e. the RSD values of EOF (n=5) were less than 4.3%, and there was no significant change in the EOF after 5 runs. The reproducibility of the coating procedures was found from the channel-to-channel RSD values of the EOF, and were less than 5.0% when using HEPES-Na buffer (pH 7.4) as the running buffer. Several examples of electrophoretic separations of amino acids and biogenic amines derivatized with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F) are demonstrated in this paper. The dynamic coating method has the potential for a broad range of applications in microchip capillary electrophoresis (microchip CE) separations.     

Macro-to-micro interfaces for microfluidic devices

Since the concept of miniaturized total analysis systems (microTAS) was invented, a great number of microfluidic devices have been demonstrated for a variety of applications. However, an important hurdle that still needs to be cleared is the connection of a microfluidic device with the rest of the world, which is often referred to as the macro-to-micro interface, interconnect, or world-to-chip interface. In this review, we will examine the methods used by pioneers in the field and other investigators, review the approaches for capillary electrophoresis-based devices and those using pneumatic pumping, and present additional discussion on interface standardization and choosing and designing interconnects for your applications.     

表面印迹聚合物及其在微流控器件中的应用

分子印迹聚合物是通过在模板存在下固化交联的聚合物制备的.在固化过程中,聚合物和模板间形成非共价键.这些非共价结合位点被"冻结"在交联的聚合物中,即使移去模板后也依然维持他们的形状.余下的空穴与模板的尺寸和形状一致,并且可以选择性地从流过的混合物中俘获模板物质.在近几十年中,分子印迹的领域由选择性俘获小分子扩展到处理各种类型的样品.分子印迹聚合物(MIP)被用于分析种类繁多的样品,比如金属离子、药物分子、环境污染物、蛋白、病毒以至整个细胞.本文中我们综述相对较新的领域——表面印迹,这是一种可以用来生成相对较大的生物相关模板的印迹方法.传统的整体印迹法是直接在固化前将模板加入预聚体中,因而不适用于那些大到无法从固化后的聚合物中扩散出来的物质.要仅在表面上生成结合位点,必须要使用特别的方法,由此产生的表面印迹技术解决了分离科学以及化学和生物化学监测的重要问题.将印迹聚合物植入微流控芯片,大大扩展了微流体技术的适用性.本文叙述表面印迹最新的进展以及不同的实施手段,以及它们在微流控器件中的应用.     

Bulk modification of polymeric microfluidic devices

The surface properties of microfluidic devices play an important role in their flow behavior. We report here on an effective control of the surface chemistry and performance of polymeric microchips through a bulk modification route during the fabrication process. The new protocol is based on modification of the bulk microchip material by tailored copolymerization of monomers during atmospheric-pressure molding. A judicious addition of a modifier to the primary monomer solution thus imparts attractive properties to the plastic microchip substrate, including significant enhancement and/or modulation of the EOF (with flow velocities comparable to those of glass), a strong pH sensitivity and high stability. Carboxy, sulfo, and amino moieties have thus been introduced (through the incorporation of methylacrylic acid, 2-sulfoethyl-methacrylate and 2-aminoethyl-methacrylate monomers, respectively). A strong increase in the electroosmotic pumping compared to the native poly(methylmethacrylate)(PMMA) microchip (ca. electroosmotic mobility increases from 2.12 to 4.30 x 10(-4) cm(2) V(-1) s(-1)) is observed using a 6% methylacrylate (MAA) modified PMMA microchip. A 3% aminoethyl modified PMMA microchip exhibits a reversal of the electroosmotic mobility (for example, -5.6 x 10(-4) cm(2) V(-1) s(-1) at pH 3.0). The effects of the modifier loading and the pH on the EOF have been investigated for the MAA-modified PMMA chips. The bulk-modified devices exhibit reproducible and stable EOF behavior. The one step fabrication/modification protocol should further facilitate the widespread production of high-performance plastic microchip devices.     

Surface modification of polymer-based microfluidic devices

We report the chemical modification of poly(methyl methacrylate) (PMMA), and poly(carbonate) (PC) surfaces for applications in microfluidic systems. For PMMA, a reaction of the surface methyl ester groups with a monoanion of α,ω-diaminoalkanes (aminolysis reaction) to yield amine-terminated PMMA surfaces will be described. Furthermore, it was found that the amine functionalities were tethered to the PMMA backbone through an alkane bridge to amide bonds formed during the aminolysis of the surface ester functionalities. The electro-osmotic flow (EOF) in aminated-PMMA microchannels was reversed when compared to that in unmodified channels. Finally, the availability of the surface amine groups was further demonstrated by their reaction with n-octadecane-1-isocyanate to form PMMA surfaces terminated with well ordered and highly crystalline octadecane chains, appropriate for performing reverse-phase separations. Examples of reverse-phase separations of ion-paired double-stranded DNAs in electric fields (capillary electrochromatography (CEC)) will be demonstrated using a PMMA-based fluidic chip. For PC, sulfonation of the surface with SO₃ will be described; this sulfonation makes the surface very hydrophilic. EOF studies of the sulfonated-PC surfaces indicated changes in the pH-dependent profile when compared to unmodified PC.     

Commercialization of microfluidic point-of-care diagnostic devices

A large part of the excitement behind microfluidics is in its potential for producing practical devices, but surprisingly few lab-on-a-chip based technologies have been successfully introduced into the market. Here, we review current work in commercializing microfluidic technologies, with a focus on point-of-care diagnostics applications. We will also identify challenges to commercialization, including lessons drawn from our experience in Claros Diagnostics. Moving forward, we discuss the need to strike a balance between achieving real-world impact with integrated devices versus design of novel single microfluidic components.     

Micromolding of solvent resistant microfluidic devices

We demonstrate a rapid fabrication procedure for solvent-resistant microfluidic devices based on the perfluoropolyether (PFPE) SIFEL. We carefully modified the poly-dimethylsiloxane (PDMS) micromolding procedure, such that it can still be executed using the standard facilities for PDMS devices. Most importantly, devices with a thin SIFEL layer for the patterned channels and a PDMS support layer on top offered the best of two worlds in terms of chemical and mechanical stability during fabrication and use. Tests revealed that these devices overcome two important drawbacks of PDMS devices: (i) incompatibility with almost all non-aqueous solvents, and (ii) leaching of oligomer into solution. The potential of our device is shown by performing a relevant organic synthesis reaction with aggressive reactants and solvents. PFPE-PDMS devices will greatly expand the application window of micromolded devices.     

Surface modification for PDMS-based microfluidic devices

This review focuses on advances reported from April 2009 to May 2011 in PDMS surface modifications for the application in microfluidic devices. PDMS surface modification techniques presented here include improved plasma and graft polymer coating, dynamic surfactant treatment, hydrosilylation-based surface modification and surface modification with nanomaterials such as carbon nanotubes and metal nanoparticles. Recent efforts to generate topographical and chemical patterns on PDMS are also discussed. The described surface modifications not only increase PDMS wettability, inhibit or reduce non-specific adsorption of hydrophobic species onto the surfaces in the act, but also result in the display of desired functional groups useful for molecular separations, biomolecular detection via immunoassays, cell culture and emulsion formation.     

Continuous flow separations in microfluidic devices

Biochemical sample mixtures are commonly separated in batch processes, such as filtration, centrifugation, chromatography or electrophoresis. In recent years, however, many research groups have demonstrated continuous flow separation methods in microfluidic devices. Such separation methods are characterised by continuous injection, real-time monitoring, as well as continuous collection, which makes them ideal for combination with upstream and downstream applications. Importantly, in continuous flow separation the sample components are deflected from the main direction of flow, either by means of a force field (electric, magnetic, acoustic, optical etc.), or by intelligent positioning of obstacles in combination with laminar flow profiles. Sample components susceptible to deflection can be spatially separated. A large variety of methods has been reported, some of these are miniaturised versions of larger scale methods, others are only possible in microfluidic regimes. Researchers now have a diverse toolbox to choose from and it is likely that continuous flow methods will play an important role in future point-of-care or in-the-field analysis devices.     

Diffusion coefficient measurements in microfluidic devices

A glassy carbon electrode (GCE) modified with Pd/IrO(2) provides excellent electrocatalytic oxidation of hydrogen peroxide. Glucose oxidase (GOD) and xanthine oxidase (XOD) were co-immobilized on the modified electrode with a thin film Nafion coated on the enzyme layer to form a glucose (Glu)/hypoxanthine (Hx) sensor, without interference from electroactive species such as ascorbic acid (AA) and uric acid (UA). Its response was evaluated with respect to the enzyme amount on the electrode, pH and temperature of the electrolyte. The prepared bienzymic biosensor, used as the detector of HPLC gave a detection limit of 1.0x10(-6) mol l(-1) Glu and 2.0x10(-7) mol l(-1) Hx (Hx) with a linear concentration range of 5.0x10(-6)-2.5x10(-3) mol l(-1) and 1.0x10(-6)-5.0x10(-4) mol l(-1), respectively. Coupled with microdialysis, it was used to monitor the concentrations of Glu and Hx in rat brain.