Faster and improved microchip electrophoresis using a capillary bundle

Joule heating generated in CE microchips is known to affect temperature gradient, electrophoretic mobility, diffusion of analytes, and ultimately the efficiency and reproducibility of the separation. One way of reducing the effect of Joule heating is to decrease the cross-section area of microchannels. Currently, due to the limit of fabrication technique and detection apparatus, the typical dimensions of CE microchannels are in the range of 50-200 microm. In this paper, we propose a novel approach of performing microchip CE in a bundle of extremely narrow channels by using photonic crystal fiber (PCF) as separation column. The PCF was simply encapsulated in a poly(methyl methacrylate) (PMMA) microchannel right after a T-shaped injector. CE was simultaneously but independently carried out in 54 narrow capillaries, each capillary with diameter of 3.7 microm. The capillary bundle could sustain high electric field strength up to 1000 V/cm due to efficient heat dissipation, thus faster and enhanced separation was attained.     

Noninvasive picoliter volume thermometry based on backscatter interferometry

Using the on-chip refractive index (RI) detector based on backscatter interferometry, sensitive, small volume, noninvasive thermometry can be performed. The current optical configuration for the on-chip interferometric backscatter detector (OCIBD) is quite simple and consists of an unfocused laser, an unaltered chip with a hemispherical channel and a photodetector. Alignment is straightforward with the only requirement being that the beam fully fills the channel. The interaction of an unfocused laser beam with the uncoated etched channel with a curvature within the silica plate (chip) produces fringes whose positional changes scale with respect to the refractive index (RI), n, of the fluid in the channel. Due to the inherently high value of dn/dT for most fluids and the high sensitivity of OCIBD to RI changes, the measurement of small temperature variations in sub-nanoliter volumes is possible. Performing OCIBD with a 75 microm diameter laser beam on a silica chip that contains an etched channel with a 40 microm radius facilitates noninvasive thermometry on a N-(2-hydroxyethyl)piperazine-(2-ethanesulfonic acid) (HEPES) solution in a 188 x 10(-12) L probe volume with a temperature resolution of 9.9 x 10(-4) degrees C, at the 99% confidence level.     

Joule heating and determination of temperature in capillary electrophoresis and capillary electrochromatography columns

This article reviews the progress that has taken place in the past decade on the topic of estimation of Joule heating and temperature inside an open or packed capillary in electro-driven separation techniques of capillary electrophoresis (CE) and capillary electrochromatography (CEC), respectively. Developments in theoretical modeling of the heat transfer in the capillary systems have focused on attempts to apply the existing models on newer techniques such as CEC and chip-based CE. However, the advent of novel analytical tools such as pulsed magnetic field gradient nuclear magnetic resonance (NMR), NMR thermometry, and Raman spectroscopy, have led to a revolution in the area of experimental estimation of Joule heating and temperature inside the capillary via the various noninvasive techniques. This review attempts to capture the major findings that have been reported in the past decade.     

Influence of moderate Joule heating on electroosmotic flow velocity, retention, and efficiency in capillary electrochromatography

The influence of Joule heating on electroosmotic flow velocity, the retention factor of neutral analytes, and separation efficiency in capillary electrochromatography was investigated theoretically and experimentally. A plot of electrical current against the applied electrical field strength was used to evaluate the Joule heating effect. When the mobile phase concentration of Tris buffer exceeded 5.0 mM in the studied capillary electrochromatography systems using particulate and monolithic columns (with an accompanying power level of heat dissipation higher than 0.35 W/m), the Joule heating effect became clearly noticeable. Theoretical models for describing the variation of electroosmotic flow velocity with increasing applied field strength and the change of retention factors for neutral analytes with electrical field strength at higher Tris buffer concentrations were analyzed to explain consequences of Joule heating in capillary electrochromatography. Qualitative agreement between experimental data and implications of the theoretical model analysis was observed. The decrease of separation efficiency in capillary electrochromatography with macroporous octadecylsilica particles at high buffer concentration can be also attributed to Joule heating mainly via the increased axial diffusion of the analyte molecules and dispersion of solute bands by a nonuniform electroosmotic flow profile over the column cross-section. However, within a moderate temperature range, the contribution of the macroscopic velocity profile in the column arising from radial temperature gradients is insignificant.     

Efficiency studies in nonaqueous capillary electrophoresis

Nonaqueous capillary electrophoresis (NACE) is a relatively new area with several advantages that include enhanced efficiency and improved detection sensitivity. The goal of this study was to investigate the influence of NACE compared to aqueous CE on the separation efficiency of oligosaccharides. The applied voltage and buffer concentration were optimized for the aqueous and nonaqueous buffer media to minimize the band broadening effects of Joule heating and electrophoretic dispersion. At the optimized conditions a 1.5-fold enhancement in efficiency was obtained with the nonaqueous buffer medium.     

Peak dispersion and contributions to plate height in nonaqueous capillary electrophoresis at high electric field strengths: ethanol as background electrolyte solvent

Nonaqueous capillary electrophoretic separations were performed under high electric field strengths (up to 2000 Vcm(-1)) in ethanolic background electrolyte solution and the contributions of different band broadening effects to plate height were evaluated. Under optimum conditions, increasing the field strength will provide faster separations and increased separation efficiency. Decrease in the separation efficiency at high field strengths was, however, observed in a previous study and now in the present paper an attempt is made to quantify various band broadening effects by applying a plate height model, which included the contributions of the injection plug length, diffusion, electromigration dispersion, Joule heating, analyte adsorption to the capillary wall, and detector slit aperture length. Of special interest were the contributions of Joule heating and analyte adsorption to the capillary wall. Poly(glycidylmethacrylate-co-N-vinylpyrrolidone)-coated fused-silica capillaries were used with internal diameters (ID) ranging from 30 to 75 microm. The separation efficiencies obtained experimentally were compared with the theoretically calculated efficiencies and fairly good agreement was observed for the 30 microm ID capillary. Relatively large deviation from the predictions of the model was found for the other capillary diameters especially at higher field strengths. The possible reasons for the deviation were discussed.     

Assessment of Joule heating and its effects on electroosmotic flow and electrophoretic transport of solutes in microfluidic channels

Joule heating is inevitable when an electric field is applied across a conducting medium. It would impose limitations on the performance of electrokinetic microfluidic devices. This article presents a 3-D mathematical model for Joule heating and its effects on the EOF and electrophoretic transport of solutes in microfluidic channels. The governing equations were numerically solved using the finite-volume method. Experiments were carried out to investigate the Joule heating associated phenomena and to verify the numerical models. A rhodamine B-based thermometry technique was employed to measure the solution temperature distributions in microfluidic channels. The microparticle image velocimetry technique was used to measure the velocity profiles of EOF under the influence of Joule heating. The numerical solutions were compared with experimental results, and reasonable agreement was found. It is found that with the presence of Joule heating, the EOF velocity deviates from its normal "plug-like" profile. The numerical simulations show that Joule heating not only accelerates the sample transport but also distorts the shape of the sample band.     

Joule heating in packed capillaries used in capillary electrochromatography

Effective heat dissipation is critical for reproducible and efficient separations in electrically driven separation systems. Flow rate, retention kinetics, and analyte diffusion rates are some of the characteristics that are affected by variation in the temperature of the mobile phase inside the column. In this study, we examine the issue of Joule heating in packed capillary columns used in capillary electrochromatography (CEC). As almost all commonly used CEC packings are poor thermal conductors, it is assumed that the packing particles do not conduct heat and heat transfer is solely through the mobile phase flowing through the system. The electrical conductivity of various mobile phases was measured at different temperatures by a conductivity meter and the temperature coefficient for each mobile phase was calculated. This was followed by measurement of the electrical current at several applied voltages to calculate the conductivity of the solution within the column as a function of the applied voltage. An overall increase in the conductivity is attributed to Joule heating within the column, while a constant conductivity means good heat dissipation. A plot of conductivity versus applied voltage was used as the indicator of poor heat dissipation. Using theories that have been proposed earlier for modeling of Joule heating effects in capillary electrophoresis (CE), we estimated the temperature within CEC columns. Under mobile and stationary phase conditions typically used in CEC, heat dissipation was found to be not always efficient. Elevated temperatures within the columns in excess of 23 degrees C above ambient temperature were calculated for packed columns, and about 35 degrees C for an open column, under a given set of conditions. The results agree with recently published experimental findings with nuclear magnetic resonance (NMR) thermometry, and Raman spectroscopic measurements.     

A model for Joule heating-induced dispersion in microchip electrophoresis

This paper presents an analytical and parameterized model for analyzing the effects of Joule heating on analyte dispersion in electrophoretic separation microchannels. We first obtain non-uniform temperature distributions in the channel resulting from Joule heating, and then determine variations in electrophoretic velocity, based on the fact that the analyte's electrophoretic mobility depends on the buffer viscosity and hence temperature. The convection-diffusion equation is then formulated and solved in terms of spatial moments of the analyte concentration. The resulting model is validated by both numerical simulations and experimental data, and holds for all mass transfer regimes, including unsteady dispersion processes that commonly occur in microchip electrophoresis. This model, which is given in terms of analytical expressions and fully parameterized with channel dimensions and material properties, applies to dispersion of analyte bands of general initial shape in straight and constant-radius-turn channels. As such, the model can be used to represent analyte dispersion in microchannels of more general shape, such as serpentine- or spiral-shaped channels.     

Effect of Joule heating on efficiency and performance for microchip-based and capillary-based electrophoretic separation systems: a closer look

An attempt is made to revisit the main theoretical considerations concerning temperature effects ("Joule heating") in electro-driven separation systems, in particular lab-on-a-chip systems. Measurements of efficiencies in microfabricated devices under different Joule heating conditions are evaluated and compared to both theoretical models and measurements performed on conventional capillary systems. The widely accepted notion that planar microdevices are less susceptible to Joule heating effects is largely confirmed. The heat dissipation from a nonthermostatically controlled glass microdevice was found to be comparable to that from a liquid-cooled-fused silica capillary. Using typically dimensioned glass and glass/silicon microdevices, the experimental results indicate that 5-10 times higher electric field strengths can be applied than on conventional capillaries, before detrimental effects on the separation efficiency occur. The main influence of Joule heating on efficiency is via the establishment of a radial temperature profile across the lumen of the capillary or channel. An overall temperature increase of the buffer solution has only little influence on the quality of the separation. Still, active temperature control (cooling, thermostatting) can help prevent boiling of the buffer and increase the reproducibility of the results.     

Nonaqueous capillary electrophoresis with alcoholic background electrolytes: separation efficiency under high electrical field strengths

The effect of high voltage on capillary electrophoresis (CE) separations of anionic analytes in nonaqueous separation media was investigated. Methanol, ethanol, 1-propanol, and 1-butanol were tested as background electrolyte (BGE) solvents. Experiments were carried out with a laboratory-built CE instrument suitable for high-voltage separations. Potentials up to 60 kV were applied with reversed polarity to generate unusually high field strengths (e.g. 2000 Vcm-1) and so achieve fast and efficient separations. Highest separation efficiencies were obtained with propanol as BGE solvent, and the dependency of the efficiency on the separation voltage was more or less linear. With the other alcohols, separation efficiency decreased or remained roughly constant with increasing absolute voltage. The separation efficiencies are discussed in terms of longitudinal diffusion, Joule heating, and analyte interaction with the capillary wall. Capillary preconditioning had a varied effect on the separations in the different BGEs as the BGE and the conditioning process affected the electroosmotic flow (EOF) velocity and direction.     

Noninvasive fluid flow measurements in microfluidic channels with backscatter interferometry

The ability to measure fluid velocity within picoliter volumes or on-chip noninvasively, is important toward fully realizing the potential of microfluidics and micrototal analysis systems, particularly in applications such as micro-high-performance liquid chromatography (HPLC) or in metering mixing where the flow rate must be quantified. Additionally, these measurements need to be performed directly on moving fluids in a noninvasive fashion. We presented here the proof of principle experiments showing nonintrusive fluid flow measurements can be accomplished on-chip using a pump and probe configuration with backscattering interferometry. The on-chip interferometric backscatter detector (OCIBD) is based on a fiber-coupled HeNe laser that illuminates a portion of an isotropically etched 40 microm radius channel and a position sensitive transducer to measure fringe pattern shifts. An infrared laser with a mechanical shutter is used to heat a section of a flowing volume and the resulting refractive index (RI) change is detected with the OCIBD downstream as a time-dependent RI perturbation. Fluid velocity is quantified as changes in the phase difference between the shutter signal and the OCIBD detected signal in the Fourier domain. The experiments are performed in the range of 3-6 microL/h with 3sigma detection limits determined to be 0.127 nL/s. Additionally, the RI response of the system is calibrated using temperature changes as well as glycerol solutions.     

Attomole sensitivity for unlabeled proteins and polypeptides with on-chip capillary electrophoresis and universal detection by interferometric backscatter

A universal detector based on backscatter interferometry has been developed to perform nanoliter volume refractive index measurements for on-chip sodium dodecyl sulfate (SDS) gel based (polyethylene oxide gel) separations and quantification label-free proteins. The on-chip interferometric backscatter detector (OCIBD) system consists of a simple, folded optical train based on the interaction of a laser beam with an etched channel in the shape of half cylinder in a fused-silica plate. The backscattered light from the channel takes on the form of a high-contrast interference pattern that contains information related to the bulk properties of the fluid located within the probe or detection volume of 2.32 x 10(-9) L. Depending on capillary electrophoresis (CE) injection method, the positional changes of the interference pattern extrema (fringes) allow for the quantification of unlabeled proteins at levels ranging from 11 to 310 amol (2.7 x 10(-8)mol/L) with a linear dynamic range of 2.5 decades (egg albumin). Using OCIBD microchannel-based SDS capillary gel electrophoresis (SDS/CGE), separation and detection of five label-free proteins was achieved in less than 100 seconds with detection limits ranging from 0.95 pg (1.1 x 10(-16)mol or 2.5 x 10(-7)mol/L) of calmodulin to 7.0 pg (1.0 x 10(-16)mol or 2.4 x 10(-7)mol/L) for bovine serum albumin (BSA) without signal filtering or active thermal control. This development shows that a universal detector based on backscatter interferometry can be used effectively for on-chip label-free solute analysis.     

Protein tryptic digests analyzed by carrier ampholyte-based capillary electrophoresis coupled to ESI-MS

In this study, narrow pH cuts of carrier ampholytes have been used as buffers in CE for the analysis of protein tryptic digests. Their low conductivity allows very efficient separations under high electric field strength without inducing any significant Joule heating. In this study, the capabilities of narrow pH cuts of carrier ampholytes for the separation of protein tryptic digests have been assessed. Three proteins of different molecular masses have been studied: cytochrome C (horse heart), beta-lactoglobulin B (bovine) and human transferrin. Efficient, rapid and repeatable separations of the peptides resulting from the tryptic digestion have been achieved in this buffer. Moreover, the feasibility of the coupling of carrier ampholyte-based capillary electrophoresis with ESI-MS has been demonstrated through the study of the cytochrome C tryptic digest.     

Bundled capillary electrophoresis using microstructured fibres

Joule heating, arising from the electric current passing through the capillary, causes many undesired effects in CE that ultimately result in band broadening. The use of narrow‐bore capillaries helps to solve this problem as smaller cross‐sectional area results in decreased Joule heating and the rate of heat dissipation is increased by the larger surface‐to‐volume ratio. Issues arising from such small capillaries, such as poor detection sensitivity, low loading capacity and high flow‐induced backpressure (complicating capillary loading) can be avoided by using a bundle of small capillaries operating simultaneously that share buffer reservoirs. Microstructured fibres, originally designed as waveguides in the telecommunication industry, are essentially a bundle of parallel ～5 μm id channels that extend the length of a fibre having otherwise similar dimensions to conventional CE capillaries. This work presents the use of microstructured fibres for CZE, taking advantage of their relatively high surface‐to‐volume ratio and the small individual size of each channel to effect highly efficient separations, particularly for dye‐labelled peptides.     

Fast and simple sample introduction for capillary electrophoresis microsystems

A newly designed capillary electrophoresis (CE) microchip with a simple and efficient sample introduction interface is described. The sample introduction is carried out directly on the separation channel through a sharp inlet tip placed in the sample vial, without an injection cross, complex microchannel layouts or hardware modification. Alternate placement of the inlet tip in vials containing the sample and buffer solutions permits a volume defined electrokinetic sample introduction. Such fast and simple sample introduction leads to highly reproducible signals with no observable carry over between different analyte concentrations. The performance of the system was demonstrated in flow-injection and CE measurements of nitroaromatic explosives and for on-chip enzymatic assays of glucose in the presence of ascorbic acid. Employing an 8 cm long separation channel and a separation voltage of 4000 V it offers high-throughput flow-injection assays of 100 samples h(-1) with a relative standard deviation of 3.7% for TNT (n= 100). Factors influencing the analytical performance of the new microchip have been characterized and optimized. Such ability to continuously introduce discrete samples into micrometer channels indicates great promise for high-speed microchip analysis.     

Temperature measurements in microfluidic systems: heat dissipation of negative dielectrophoresis barriers

The manipulation of living biological cells in microfluidic channels by a combination of negative dielectrophoretic barriers and pressure-driven flows is widely employed in lab-on-a-chip systems. However, electric fields in conducting media induce Joule heating. This study investigates if the local temperatures reached under typical experimental conditions in miniaturized systems cause a potential risk for hyperthermic stress or cell damage. Two methods of optical in situ temperature detection have been tested and compared: (i) the exposure of the thermo-dependent fluorescent dye Rhodamine B to heat sources situated in microfluidic channels, and (ii) the use of thermoprecipitating N-alkyl-substituted acrylamide polymers as temperature threshold probes. Two-dimensional images of temperature distributions in the vicinity of active negative dielectrophoresis (nDEP)-barriers have been obtained and local temperature variations of more than 20 degrees C have been observed at the electrode edges. Heat propagation via both buffer and channel walls lead to significant temperature increases within a perimeter of 100 microm and more. These data indicate that power dissipation has to be taken into account when experiments at physiological temperatures are planned.     

Electrochromatography with a 2.7 mm inner diameter monolithic column

Monolithic columns of 2.7 mm I.D. have been prepared and used in electrochromatography (EC) separation. Although capillary electrochromatography (CEC) has higher separation efficiency, it displays some shortcomings, such as limited sample loadability and restricted concentration detectability etc. In this paper, we investigate the feasibility of EC separation with millimeter diameter monolithic columns. By using a designed preparation method of monolithic column packed with about 150 microm quartz sand, the effect of Joule heating can be reduced, and the processes of frit making and column packing can be avoided. The concentration detectability of the EC is improved comparing with that of CEC. Moreover, the separation efficiency of 52,000 plates/m was achieved with a 70 mm length and 2.7 mm I.D. monolithic column.     

Influence of flow and diffusion on protein separation in a continuous flow electrophoresis cell: computation procedure

Continuous flow zone electrophoresis is an efficient method for the non-destructive separation of biological materials in a flowing film of buffer solution in which the constituents of a sample are separated according to their electrophoretic mobilities, under the influence of an electric field. This paper deals with modeling of flow structure of the buffer solution taking into account the effect of electroosmosis, Joule heating and thermal free convection and the forced convection of axial flow. Modeling of diffusion is examined in a second part. The modeling equations and the corresponding boundary conditions are solved by finite difference methods. The various parameters affecting the quality of the fractionation are analyzed.     

Revisit of Joule heating in CE: the contribution of surface conductance

We present in this short communication the true form of Joule heating in CE which considers the contribution of surface conductance. This increased conductivity of electrolyte solution within electrical double layer has never been discussed in previous studies. The resultant intensive heat generation near the capillary wall is demonstrated using numerical simulation to produce not a locally strong temperature rise, but an additional temperature elevation in the whole solution compared to the model neglecting surface conductance. The latter effect is, however, negligible in typical CE while it might become significant in very small channels.