A polymeric microchip with integrated tips and in situ polymerized monolith for electrospray mass spectrometry

We describe the integration of a cyclo-olefin polymer based microchip with a sheathless capillary tip for electrospray ionization-mass spectrometry (ESI-MS). The microchip was fabricated by hot embossing and thermal bonding. Its design includes a side channel for adjusting the composition of the electrospray solution so that analytes in 100% water can be analyzed. The fused silica capillaries, used for sample introduction, and the electrospray tips for MS coupling were directly inserted into the microchannel before thermal bonding of the device. A microfabricated on-chip gold microelectrode was used to apply the electrospray voltage. Annealing the device after thermal bonding increased the pressure resistance of the microchip. The cross section of the microchannel was imaged by scanning electron microscopy to estimate the effects of the annealing step. The relationship between the applied electrospray voltages and MS signal was measured at different flow rates by coupling the device to an ion trap mass spectrometer. The performance of the microchip was evaluated by MS analysis of imipramine in ammonium acetate buffer solution by direct infusion. An alkylacrylate based monolith polymer bed for on-chip sample pretreatment and separation was polymerized in the microchannel and tested for ESI-MS applications.     

Microchip-based electrochromatography: designs and applications

Different techniques and methods of electrochromatography on “lab on a chip” devices are reviewed. Described approaches include open-channel microchip electrochromatography relying on C₈, C₁₈ and novel gold nanoparticle (GNP) coating of microchannel wall; packed-channel microchip electrochromatography with new ways of automated loading and unloading of conventional octadecylsilica beads; monolith-based microchip electrochromatography with tailored casting of stationary phase at the specific places of microfluidic network and novel photolitographically fabricated collocated monolithic structures. Specific issues related to the microchip electrochromatography, i.e. importance of high aspect ratio of the microchannels in the open-channel electrochromatography or approaches eliminating the wall effect in the monolith-based electrochromatography, are discussed. Various applications for environmental, pharmacological, genomic and proteomic analysis are described. The operation parameters of reviewed microsystems are summarized in easy-to-read tables.     

A simplified method for capillary embedment into microfluidic devices - exemplified by sol-gel-based preconcentration

We here describe an alternative method of embedding functionalized capillaries into microdevices fabricated in PDMS. The capillaries have square-shaped outer dimensions and fit into elastic PDMS channel networks of similar dimensions. By modifying the capillary off-chip, the technique makes it possible to integrate a new chip function without risking contamination of already existing chemically patterned areas because of new reagent solutions. Leak-free insertion of these capillaries has earlier been reported, where a thin layer of uncured PDMS bonded the capillary to the microchannel and the lid structure. In this new approach, oxygen plasma is used to bond the square capillary to the PDMS, eliminating the risk of clogging the microsystem with uncured prepolymer. The new embedding technique was exemplified and evaluated by inserting a square capillary piece containing monolithic sol-gel for sample preconcentration application. The assembled microdevice was run with mass spectrometric detection, showing that peptides were preconcentrated without leakage from either the sol-gel itself or around the inserted capillary. Repeated preconcentration runs showed migration times better than 3% for all peptides. We believe that the presented microchip assembling technique greatly simplifies the insertion of functionalized capillary pieces, e.g., an initial preconcentrator to a PDMS device containing other downstream modules.     

Coupling on-chip solid-phase extraction to electrospray mass spectrometry through an integrated electrospray tip

We report the integration of solid-phase extraction (SPE) with mass spectrometry (MS) through an on-chip electrospray tip for sample precleaning and preconcentration. An in situ polymerized alkylacrylate-based monolithic column was used as the stationary phase for the on-chip SPE. Each microchip consists of two sets of microchannels and their respective integrated electrospray tips, with a common gold electrode. After the microchip was fabricated from cycloolefin polymer by hot embossing, thermal bonding, and annealing steps, a mixture of monomers and porogenic solvents was pumped into the microchannels and certain areas of the main microchannels were exposed to UV irradiation through a mask. The resulting porous monolithic beds that were polymerized from different compositions of the mixture were characterized by scanning electron microscopy. The microchip containing the monolithic column was then interfaced to an ion trap (IT) mass spectrometer by modifying a commercially available interfacing system. Makeup solution from the side channel was infused concurrently with the solution flowing into the main channel, and the mixture of these two solutions was sprayed into the MS orifice. Both the adsorption and elution of a pharmaceutical test compound, imipramine, to and from the on-chip SPE columns were monitored by MS. The potential application of this device for sample cleanup was demonstrated by pretreatment of urine samples spiked with imipramine.     

Integration of ground aerogel particles as chromatographic stationary phase into microchip

C16 modified and ground monolithic silica aerogel particles in submicrometer size, as a new type of stationary phase was prepared and integrated in polydimethylsiloxane (PDMS) microchip. The aerogel particles were packed into the microfluidic channel using a simple procedure, which does not require any special frit or fabrication step to retain the particles. The subnanoliter volume of samples can be transported through the porous, short length of packing with low pressure (< 3 bar). Food dyes as test components could be separated using low pressure within 6s. A 50-fold preconcentration could be achieved by retaining 100 nL volume of sample on the packing and elution with methanol.     

DNA sequencing in a monolithic microchannel device

We present 50 cm long microchannels in a monolithic device for high resolution, long read-length DNA sequencing. These devices were fabricated and bonded in borofloat glass using unconventional photolithography techniques with 48-188 independent, straight microchannels. The microchannel DNA separation was tested with POP-6 polymer and a DNA sequencing ladder separated at room temperature and 200 V/cm. Single-base resolution greater than 600 bases was achieved and the sequence base called to 640 bases with 98% accuracy. Under the same experimental conditions, the performance of the microchip was identical to a fused-silica capillary with similar cross-sectional area.     

Rapid amplification of genetically modified organisms using a circular ferrofluid-driven PCR microchip

The use of genetically modified organisms (GMOs) as food and in food products is becoming more and more widespread. Polymerase chain reaction (PCR) technology is extensively used for the detection of GMOs in food products in order to verify compliance with labeling requirements. In this paper, we present a novel close-loop ferrofluid-driven PCR microchip for rapid amplification of GMOs. The microchip was fabricated in polymethyl methacrylate by CO₂ laser ablation and was integrated with three temperature zones. PCR solution was contained in a circular closed microchannel and was driven by magnetic force generated by an external magnet through a small oil-based ferrofluid plug. Successful amplification of genetically modified soya and maize were achieved in less than 13 min. This PCR microchip combines advantages of cycling flexibility and quick temperature transitions associated with two existing microchip PCR techniques, and it provides a cost saving and less time-consuming way to conduct preliminary screening of GMOs. Figure Schematic of the circular ferrofluid-driven PCR microchip     

Effects of microchannel geometry on preconcentration intensity in microfluidic chips with straight or convergent-divergent microchannels

Preconcentration microfluidic devices are fabricated incorporating straight or convergent-divergent microchannels and hydrogel or Nafion membranes. Sample preconcentration is achieved utilizing concentration-polarization effects. The effects of the microchannel geometry on the preconcentration intensity are systematically examined. It is shown that for the preconcentrator with the straight microchannel, the time required to achieve a satisfactory preconcentration intensity increases with an increasing channel depth. For the convergent-divergent microchannel, the preconcentration intensity increases with a reducing convergent channel width. Comparing the preconcentration performance of the two different microchannel configurations, it is found that for an equivalent width of the main microchannel, the concentration effect in the convergent-divergent microchannel is faster than that in the straight microchannel.     

Hybrid organic-inorganic phenyl monolithic column for capillary electrochromatography

A novel hybrid organic-inorganic silica-based monolithic column possessing phenyl ligands for reversed-phase (RP) capillary electrochromatography (CEC) is described. The monolithic stationary phase was prepared by in situ co-condensation of tetraethoxysilane (TEOS) with phenyltriethoxysilane (PTES) via a two-step catalytic sol-gel procedure to introduce phenyl groups distributed throughout the silica matrix for chromatographic interaction. The hydrolysis and condensation reactions of precursors were chemically controlled through pH variation by adding hydrochloric acid and dodecylamine, respectively. The structural property of the monolithic column can be easily tailored through adjusting the composition of starting sol solution. The effect of PTES/TEOS ratios on the morphology of the created stationary phases was investigated. A variety of neutral and basic analytes were used to evaluate the column performance. The CEC columns exhibited typical RP chromatographic retention mechanism for neutral compounds and had improved peak shape for basic solutes.     

Polyester microchannel chip for electrophoresis - incorporation of a blue LED as light source

A blue-light-emitting diode was incorporated as a fluorescence-excitation light source into a polyester microchannel chip fabricated by in situ polymerization. Placing the light-emitting facet of the diode close to the microchannel obviated any need for an additional optical arrangement. Fluorescence from the sample was transmitted by an optical fiber incorporated into the microchip perpendicular to the LED. FITC labeled amino acids were separated in the presence of 5 mM SDS by using the microchip and were detected by LED-induced fluorescence.     

On-chip solid phase extraction coupled with electrophoresis using modified magnetic microspheres as stationary phase

A poly(dimethylsiloxane)(PDMS)/glass hybrid microchip for on-line solid phase extraction (SPE) and electrophoresis separation has been developed and evaluated. The SPE microchannel was crossed to the electrophoresis microchannel. All the microfluidic channels were etched on the glass substrate. The magnetic microspheres were coated with hydroxyl-terminated poly-dimethylsiloxane (PDMS-OH) serving as extraction phase, which could be conveniently immobilized into the sample pretreatment channel by magnetic field. The PDMS-OH microspheres were mobilized into and out of the pretreatment channel by injection flow. The 0.1 μmol/L solution of fluorescence isothiocyanate (FITC)-labeled phenylalanine (Phe) was electrically injected into the SPE channel and extracted onto the PDMS-OH microspheres bed. The enriched FITC-labeled Phe was electrically eluted by 9 mmol/L sodium acetate containing 10% acetonitrile and electrically driven into the electrophoresis channel and then separated. The preconcentration factor could reach 87.5 after sufficient extraction. A linear preconcentration curve was obtained with the initial FITC-labeled Phe concentration ranging from 6 nmol/L to 300 nmol/L (R ²=0.9922) with 200 s loading time. The detection limit (S/N=3) for the FITC-labeled Phe was 3 nmol/L.     

Electrochemical Behavior of Parallel Opposed Dual Electrode in a Microchannel

The electrochemical behavior dependent on the microchannel depth is discussed using a parallel opposed dual electrode in the microchip. The microchannel depth was controlled easily by the thickness of the photoresist. Cyclic voltammetry (CV) was carried out with conventional mode and the generation‐collection mode. High collection efficiency (max: 98%) and high current amplification (max: 5.2) were achieved without miniaturizing the dual electrode by micromachining techniques, when the microchip with the microchannel depth of 30 μm was used in the generation‐collection mode at low sweep rates. Quantitative analysis was applied to electrochemically reversible species with a quicker response time of around a few seconds for the same microchannel depth on chronoamperometry (CA). We observed good linearity on the calibration plot of dopamine (2×10^?6 – ca. 1×10^?3 mol dm^?3 ).     

10,000-fold concentration increase in proteins in a cascade microchip using anionic ITP by a 3-D numerical simulation with experimental results

This paper describes both the experimental application and 3-D numerical simulation of isotachophoresis (ITP) in a 3.2 cm long "cascade" poly(methyl methacrylate) (PMMA) microfluidic chip. The microchip includes 10 × reductions in both the width and depth of the microchannel, which decreases the overall cross-sectional area by a factor of 100 between the inlet (cathode) and outlet (anode). A 3-D numerical simulation of ITP is outlined and is a first example of an ITP simulation in three dimensions. The 3-D numerical simulation uses COMSOL Multiphysics v4.0a to concentrate two generic proteins and monitor protein migration through the microchannel. In performing an ITP simulation on this microchip platform, we observe an increase in concentration by over a factor of more than 10,000 due to the combination of ITP stacking and the reduction in cross-sectional area. Two fluorescent proteins, green fluorescent protein and R-phycoerythrin, were used to experimentally visualize ITP through the fabricated microfluidic chip. The initial concentration of each protein in the sample was 1.995 μg/mL and, after preconcentration by ITP, the final concentrations of the two fluorescent proteins were 32.57 ± 3.63 and 22.81 ± 4.61 mg/mL, respectively. Thus, experimentally the two fluorescent proteins were concentrated by over a factor of 10,000 and show good qualitative agreement with our simulation results.     

Reversed-phase liquid chromatography on a microchip with sample injector and monolithic silica column

In micro total analysis systems, liquid chromatography (LC) works under pressure-driven flow is the essential analysis component. There were not, however, much works on microchip LC. Here we developed a microchip for reversed-phase LC using porous monolithic silica. The chip consisted of a double T-shaped injector and a approximately 40-cm serpentine separation channel. The octadecyl-modified monolithic silica was prepared in the specified part of the channel on the microchip using sol-gel process. Furthermore, the effect of geometry of turn sections on band dispersion at turns was examined under pressure-driven flow. High separation efficiencies of 15,000-18,000 plates/m for catechins were obtained using the LC chip.     

Hybrid organic-inorganic octyl monolithic column for in-tube solid-phase microextraction coupled to capillary high-performance liquid chromatography

An octyl-functionalized hybrid silica monolithic column was developed for in-tube solid-phase microextraction (SPME) to perform on-line preconcentration coupled to capillary high-performance liquid chromatography (microHPLC) analysis. A hybrid silica monolithic column functionalized with octyl groups was conveniently synthesized by a two-step acid/base-catalyzed hydrolysis/co-condensation of tetraethoxysilane (TEOS) and n-octyltriethoxysilane (C8-TEOS). The size of through-pores as well as the carbon content can be adjusted by changing the ratio of TEOS to C8-TEOS in the polymerization mixture. The extraction characteristics of the monolithic column prepared under optimized fabrication conditions were studied by using polycyclic aromatic hydrocarbons (PAHs) as the analytes. The sample volume that could be injected into the system was increased up to 1mL with simultaneous increase of column efficiency, when hybrid silica monolithic column was used as a precolumn. Good linear calibration curves (R>0.999) were obtained, and the limits of detection (signal-to-noise ratio, S/N=3) for the analytes were found to be between 2.4 and 8.1ng/mL with a UV absorbance detector, which are 299-456 times lower than those obtained without preconcentration. The column-to-column RSD values were 1.3-8.0% for recoveries of PAHs investigated.     

Preconcentration of milk proteins using octadecylated monolithic silica microchip

Sample preparation is a bottleneck in systems for chemical analysis and it is a required step in order to remove interference and preconcentrate the target analytes. Much research in recent years has focused on porous monolithic materials since they are highly permeable to liquid flow and show high mass transfer compared with common packed beds. This study has focused on the use of a glass microchip containing an inorganic silica-based monolithic material modified with octadecyl groups for preconcentration of milk proteins from skimmed cows’ milk that vary in molecular weight, hydrophobicity, and abundance. Comparison between the fabricated device and a commercial cartridge for the preconcentration of proteins in skimmed cows’ milk showed the ability of the device to successfully enrich protein mixtures from the sample. The three replicate experiments showed that the RSD of the mass to charge ratio of milk proteins ranged from 0.01 to 0.46%. In addition, it was found that there were no significant differences between the observed and reported masses of the milk proteins and the relative percentage error of the molecular masses ranged between 0.03 and 0.90%. The fact that the small amounts of sample required and short sample preparation time suggest that this new microfluidic device may be a viable alternative to existing procedures for protein extraction from real samples.     

Convenient enantioseparation by monolithic imprinted capillary clamped in a chip with electrochemical detection

A microchip integrated with a monolithic imprinted capillary has been manufactured for performing the chip-based capillary electrochromatographic enantioseparation. The microporous monolith anchored on the inner wall of the microchannel was prepared by in situ chemical copolymerization, and characterized with scanning electron microscopy, IR spectroscopy, and solid-state UV-vis spectroscopy. The monolithic network with high porosity gave a large surface area, good permeability, low mass-transfer resistance, and thus high separation efficiency. A portable microchip was conveniently constructed by integrating an imprinted capillary with 5-cm length as the separation channel and a carbon fiber microdisk working electrode for amperometric detection. Using L-tyrosine (L-Tyr) as the template molecule, Tyr enantiomers could be baseline separated within 55 s under the optimized preparation and separation conditions. The linear ranges for online amperometric detection of both Tyr enantiomers were from 20 to 2400 μM. The microporous monolithic chip strategy exhibited excellent separation efficiency and promising analytical application in enantioseparation. It opens an avenue for high-throughput screening of chiral compounds.     

Solid Phase Extraction of Neutral Analytes through Silica Gel Coated with Layers of Au Nanoparticles Self‐Assembled with Alkanethiols

In this study, we used Au nanoparticle (NP)‐coated silica gel as a solid phase extraction sorbent for the preconcentration of neutral analytes (steroid drugs). The sorbent was fabricated using two alkanethiol self‐assembly processes: one to deposit the Au NPs onto a 3‐aminopropyltrimethoxysilane‐modified silica gel and the other to functionalize the surfaces of the Au NPs. A large volume of the steroid solution was passed through the silica gel to facilitate adsorption mediated by hydrophobic interactions between the steroids and the hydrophobic moieties on the silica gel surface. Extraction of the steroids was accomplished by flushing the silica gel with a low‐polarity solvent. In this preliminary study, we found that the particle size of the silica gel and the number of layers of Au NPs coated on the silica gel both affected the preconcentration performance for the steroids. When using six layers of Au NPs coated on 5–20‐μm silica gel, the detection limits for steroids were below 80 ng L^?1; the preconcentration efficiency was over 170‐fold higher than that of the original steroid solution. Our findings provide further evidence that nanotechnology has much to benefit analytical science.     

A simple and compact blue diode laser powered excitation source for fluorescence detection in capillary electrochromatographic microchip separation

A simple and compact fluorescence excitation source was prepared using a 405 nm blue laser diode module and characterized in capillary electrochromatographic or capillary electrophoretic microchip separation. An inexpensive blue laser diode module with a tiny focusing lens was simply mounted at the center of an aluminum block on a miniature linear motion guide for heat dissipation and position control. A slit unit has a series of fifteen laser-machined slits with 1 mm space along the direction of the separation channel of the microchip above this unit. The laser beam was focused through a slit with 50 μm width to the separation channel at the position of a desired length. Although the excitation source unit was connected to a simple current controlled power supply, it was stable with 0.1% drift per hour and 1.3% (1σ) fluctuation in intensity. This simple excitation source can be prepared easily with inexpensive minimum optical components and mounted with a microchip on the stage of an ordinary fluorescence microscope for daily separation studies using a CE or CEC microchip. The applicability of the excitation source was evaluated with FITC-amino acid derivative mixtures using a polymer based CEC microchip packed fully with submicron silica beads in its microchannel.     

Fabrication of a fiberglass-packed channel in a microchip for flow injection analysis

A novel fiberglass-packed channel in a microchip has been fabricated for flow injection analysis (FIA) based on the in situ polymerization of methyl methacrylate (MMA). MMA prepolymer molding solution containing an ultraviolet initiator was sandwiched between a poly(methyl methacrylate) (PMMA) cover plate and a PMMA base plate bearing a glycerol-permeated fiberglass bundle and exposed to UV light. During the UV-initiated polymerization, the fiberglass bundle was embedded in the PMMA substrate to form a fiberglass-packed microchannel. When the glycerol in the fiberglass bundle was flushed away with water, the obtained porous fiberglass-packed microchannel could serve as an electroosmotic pump and a FIA channel. Scanning electron micrographs offer insights into the fiber-based microchip. The analytical performance of the FIA system has been demonstrated by detecting catechol in connection with end-column amperometric detection. The fiber- based microchips can be fabricated by the new approach without the need for complicated and expensive lithography-based microfabrication techniques, indicating great promise for the low-cost production of microchips, and should find a wide range of applications.