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aFlinders Centre for Nanoscale Science and

2100, Adelaide 5001, AustraliabSchool of Chemical and Physical Science

Adelaide 5001, Australia. E-mail: claire.lene

† Electronic supplementary informa10.1039/c3an00870c

Cite this: Analyst, 2013, 138, 4275

Received 29th April 2013Accepted 3rd June 2013

DOI: 10.1039/c3an00870c

www.rsc.org/analyst

This journal is ª The Royal Society of

On-chip capacitively coupled contactless conductivitydetection using “injected” metal electrodes†

Leigh D. Thredgold,a Dmitriy A. Khodakov,a Amanda V. Ellisa and Claire E. Lenehan*b

We demonstrate the use of injected gallium electrodes for capaci-

tively coupled contactless conductivity detection (C4D) within a

microchip electrophoresis device. Evaluation of the electrodes for

quantitative detection of electrophoretically separated lithium,

sodium and potassium ions showed the system offers competitive

detection limits of 6.1 � 10�6 M, 6.7 � 10�6 M and 8.5 � 10�6 M,

respectively. The fabrication process is fast, highly reproducible, and

eliminates difficulties with electrode alignment. Using this approach

C4D can be readily achieved in any microchip by simply adding extra

‘electrode’ channels to the microchip design.

Introduction

Lab-on-a-Chip (LOC) production has been signicantlypropelled by the development of simple, low cost fabricationtechniques. One of the most prominent of these techniques isthe use of poly(dimethylsiloxane) (PDMS) for producingmicrochannel networks from structured templates usingphotolithography for device fabrication.1 Despite signicantresearch over the last decade into such devices, very few havetranscended to commercial production.2–4 This is a result of anumber of limiting factors in producing commercially viableLOC systems. These arise from the complex fabricationprocesses required to integrate commonly utilised detectionsystems, such as uorescence and electrochemical techniques,with the microchip.4–6

Capacitively coupled contactless conductivity detection(C4D) and its integration with microchip technologies has beenthe subject of multiple reviews.4,7,8 The detection techniqueutilises integrated conductive electrodes, most commonly in the

Technology, Flinders University, GPO Box

s, Flinders University, GPO Box 2100,

han@inders.edu.au

tion (ESI) available. See DOI:

Chemistry 2013

form of metal, to continuously monitor the conductivity of asolution moving through a microchannel. A number ofapproaches for integrated contactless electrodes in LOC deviceshave been reported. These include the fabrication of micro-uidic chips on top of printed circuit boards9–12 and glasswafers13 containing microscale electrodes. Alternate methodshave mounted the electrodes in external housings14,15 andothers have reported the deposition of metal layers onto previ-ously etched glass wafer surfaces to form electrodes in the sameplane as the analysis channels.16,17 Each of these methods havepractical limitations including multi-step patterning/fabrica-tion processes oen requiring specialized equipment and/ordifficulty in consistently aligning the detection electrodes withthe microchannel. Further, these fabrication techniques areoen time consuming, for example Henderson et al. reportedthe production of laser-patterned polymer electrodes thatrequired a 20 h timeframe to complete.18 All of these limitationshave the potential to impact on analytical performance due tofabrication reproducibility or increase the overall time and costassociated with device production.19

An alternative approach for the fabrication of LOC deviceswith integrated contactless electrodes is to use injected moltenmetal. Injected metal electrodes have been demonstrated inmicrouidic applications for the production of in-plane on-chipelectromagnets20 and on-chip microplasma patterning of PDMSmicrochannels.21 More recently, this approach has been used toproduce electrodes which are in direct contact with uid inmicrochannels for mixing22 and neural stimulation23 applica-tions. Injected molten metal electrodes have the advantage thatboth electrode and analysis microchannels are patterned in thesame lithographic process, whereby the electrode design isincorporated directly onto the mask template. This allowsprecise micron-scale electrode designs that can be simply andeasily tailored to the microuidic device under construction.Once the device has been assembled, the molten metal is theninjected into the required microchannel where it either remainsmolten or solidies to form a solid electrode. Substrates formolten metal electrodes include relatively low melting point

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alloys such as molten solder (In 52%, Sn 48% m.p. z 117 �C),20

eutectic gallium indium (Ga 75%, In 25% by weight m.p. z15.7 �C)22,23 and indium alloy (In 51%, Bi 32.5%, Sn 16.5% byweight m.p. z 60 �C).22 The use of pure gallium (m.p. z29.8 �C) as an electrode material has been shown to be prefer-able over low melting point alloys due to its low toxicity, rela-tively low melting point and ease of handling.21–23 Despite itsconrmed simplicity, the use of injected metal electrodes forC4D detection on LOC devices has yet to be reported.

This paper presents a novel in-plane C4D detection systemfor LOC utilising injected metal electrodes. The system wasfabricated by injecting liquid gallium into pre-designed elec-trode microchannels prepared during microchip fabrication.The optimal electrolyte concentration, excitation voltage andexcitation frequency are discussed. Quantitative on-chip detec-tion using the injected electrode system was demonstratedusing a model electrophoretic system comprising of aqueouslithium, sodium and potassium cations.

Experimental

Unless otherwise specied, all chemicals were of analyticalgrade, purchased from Sigma-Aldrich (Australia) and used asreceived. Background electrolyte (BGE) solutions were preparedby dissolving 2-(N-morpholino)ethanesulfonic acid (MES) (10–20 mM) and histidine (His) (10–20 mM) in Milli-Q water(18.2 MU cm). Sample stock solutions of aqueous Li+, Na+ andK+ (100 mM) were prepared from their chloride salts. Workingstandards of Li+, Na+ and K+ were prepared by diluting stocksolutions in BGE. All solutions were degassed by ultrasonicationprior to use.

A schematic of the microchip design is shown in Fig. 1 andconsisted of a standard elongated cross-conguration with a57 mm separation channel and an 18 mm injection channel.The electrode arrangement was positioned 37 mm from theseparation/injection channel cross-over and consisted oftransmitter and receiver electrodes in anti-parallel congura-tion. The detection electrodes were offset by 1 mm along theseparation channel with two ground plane shield plates situ-ated between them to reduce direct coupling. The channel

Fig. 1 Schematic of microchip design with a separation channel connected toBGE (B) and BGE waste (BW) reservoirs and cross-injection channel connected tosample (S) and sample waste (SW) reservoirs. C4D electrodes are also shown(inset) with transmitter electrode (TE) and receiver electrode (RE) in anti-parallelconfiguration separated by grounded shield plate microchannels (G).

4276 | Analyst, 2013, 138, 4275–4279

dimensions were 100 mm wide by 40 mm deep for both analysisand electrode channels and 200 mm wide and 40 mm deep forshielding plate channels. The detection electrode and shieldingplate channels were situated 75 mm and 50 mm from the sepa-ration channel respectively. Reservoirs were placed at the endsof each channel to facilitate uid access and allow connection ofthe electrode channels to external electronic circuitry. This wasachieved using a custom designed printed circuit board(developed in house) that facilitated connection through tinplated copper electrode pins (see ESI† for photographs).

C4D is inherently susceptible to external electrical noisewhich can lead to a loss in detector sensitivity.14 In order tominimise this, the printed circuit board electronics, highvoltage electrodes and microchip were housed in a die cast box.A removable lid facilitated external access, sample introductionand contained an additional recessed ground plane to furtherminimise direct coupling between electrodes. The housing wasgrounded to eliminate interferences from external noisesources.

PDMSmicrouidic chips were fabricated using standard solithography techniques.1 A master mold of the patternedmicrochip electrophoresis and electrode channels was preparedon a silicon wafer (non-porous, polished Si (100) wafers (p++

type, boron doped, 0.0008–0.0012 U cm), Siltronix, France) withSU-8 photoresist (MicroChem Corp., USA). PDMS replicas of themaster mold were made using a Sylgard� 184 silicone elas-tomer kit (Dow Corning, USA) by pouring a mix of PDMS baseand curing agent in a ratio of 10 : 1 (%w/w). Aer curing,replicas were removed, holes punched for later uid introduc-tion and cleaned in iso-propanol. PDMS replicas, along with aat sheet of PDMS were then oxygen plasma treated (PDC-32G-2Plasma Cleaner, Harrick Plasma, USA) at 0.2 Torr of O2 at 18 Wfor 45 s. The replica and at sheet of PDMS were then broughtinto contact with each other within 10 s of plasma treatmentwhich resulted in irreversible covalent bonding,24 creating asealed microuidic chip.

Molten gallium (99.99%) was injected into the electrodemicrochannels according to Priest et al.21 Gallium was warmedto 50 �C and subsequently drawn into plastic laboratory tubingusing a 1 mL syringe. A plastic pipette tip, attached to the end ofthe tubing, was placed in the electrode channel inlet andgallium was injected until it protruded from the opposite end ofthe channel. The gallium remained molten until a seed crystalwas added, solidifying the metal.

Electrophoretic separations were performed in PDMSmicrochips using the following standard procedure. The chipswere rst ushed with Milli-Q water for 2 min followed by BGEsolution (15 mM) with n-dodecyl b-D-maltoside (DDM) (0.4 mM)for a further 20 min (see ESI† for discussion). Aer ushing, allreservoirs were lled with the running buffer solution (15 mMBGE). The microchip was rinsed with both the BGE containingDDM and running buffer between analysis runs to ensurereproducibility and maintain a stable baseline. The buffer andsample solutions were dispensed into the respective reservoirsusing a xed volume of 25 mL. Sample introduction was per-formed electrokinetically using a cross-channel injection. Volt-ages of 0.8 kV were applied to the sample reservoir (S) to

This journal is ª The Royal Society of Chemistry 2013

Fig. 2 Effect of BGE concentration on the peak height and S/N ratio (inset) of aNa+ peak (5 � 10�4 M). Operating conditions: microchip 57/37 mm total/effec-tive length; injection voltage, 0.8 kV for 10 s; separation voltage, 1.4 kV. C4Ddetector: sine waveform of 220 kHz, 5 Vp-p. Error bars: �1 standard deviation.

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facilitate injection, whilst sample waste (SW) was grounded andBGE (B) and BGE waste (BW) reservoirs were oated. Separa-tions were performed by applying a voltage of 1.4 kV to the Breservoir (anode) whilst the BW (cathode) was grounded and Sand SW were oated, producing a cathodic EOF. Voltageswitching was carried out by a custom designed LabVIEW�program built in-house.

On-chip detection of analytes was achieved using the previ-ously outlined injected electrode arrangement (Fig. 1). Anexternally sourced excitation signal in the form of sine waves,with a frequency of 220 kHz and amplitude of 5.5 Vp-p, wastransferred to the transmitter electrode (TE). This signalproduced an induced alternating current that passed throughthe detection cell area and was subsequently picked up by thereceiver electrode (RE). The shielding plates between the elec-trodes remained grounded throughout (G). The current wasconverted to a voltage using a receiver operational amplierOPA827A (Texas Instruments, 2.2 MU feedback resistor). Thereceiver amplier was encased inside the microchip housing forminimal signal loss. The output signal of the receiver amplierwas transferred to the signal processing circuit encased in agrounded housing. The amplied signal was then rectied,amplied and low pass ltered, as has been described previ-ously.14 The resulting signal was then collected using a NI USB-6009 DAQ card (National Instrument) controlled by a customdesigned LabVIEW� program built in-house.

Fig. 3 Effect of excitation frequency on the peak height and S/N ratio (inset)of Li+ (–:– and –D–) Na+ (–C– and –B–) and K+ (–-– and –,–) peaks (5 �10�4 M). Operating conditions as for Fig. 2.

Results and discussionElectrode operating conditions

For reliable operation, contactless conductivity detectionrequires careful optimisation of the electrode excitation voltageand excitation frequency.25,26 These are directly affected by theBGE and as such all three are discussed below. A standardmixture of Li+, Na+ and K+ cations (5 � 10�4 M) was chosen as amodel analyte system as they are commonly used ions to eval-uate the performance of C4D detectors.18

The effect of BGE concentration (10, 15 and 20 mM) on theperformance of the C4D system was explored by separating Li+,Na+ and K+ ions using an excitation voltage of 5 Vp-p andfrequency of 220 kHz. It was observed that the data trends for allthree cationswere identical and therefore only the results forNa+

are shown (see ESI† for gures). The peak height and signal tonoise (S/N) ratio both decreased with increasing BGE concen-tration. However, the reproducibility obtained was poorer using10 mM BGE (Fig. 2). Therefore, 15 mM BGE was chosen for theoptimization of excitation voltage and frequency as it gaveexcellent signals without compromising reproducibility.

The excitation frequency of the C4D system was evaluatedbetween 100 and 400 kHz at a xed excitation voltage of 5 Vp-p.As outlined above, replicate analyses of a Li+, Na+ and K+ stan-dard were performed in 15 mM BGE. Fig. 3 depicts the effect ofaltering the excitation frequency on the peak height and S/Nratios of the Li+, Na+ and K+ peaks. As can be seen, themaximumpeak height and S/N ratio were both achieved using an excitationfrequency of 220 kHz. At lower frequencies the signal responsedecreases as the impedance of the microchannel wall is

This journal is ª The Royal Society of Chemistry 2013

relatively high resulting in only a small current passing into theelectrolyte. In contrast, at high frequencies the signal is inu-enced by both stray capacitance and poorer performance of thecurrent to voltage convertor resulting in the decreased peakheights and S/N ratios observed. It was concluded that 220 kHzbe used for further optimisation experiments due to its superiorsensitivity.

Previous reports have found that increasing excitation volt-ages in microuidic C4D systems can result in improvedanalytical performance.16,25 To evaluate the effect of thisparameter on the performance of the developed electrodesystem a Li+, Na+ and K+ standard was analysed using varyingexcitation voltages (3–5.5 Vp-p). Fig. 4 shows the peak height andS/N ratios of all three cations obtained over the excitationvoltage range studied. The data trend demonstrates thatincreasing the excitation voltage resulted in a subsequentincrease in both the peak height and S/N ratio. Beyond 5.5 Vp-p

the high level of stray capacitance results in an increasedbackground current that overloads the feedback resistorresulting in the complete loss of signal. Therefore, themaximum peak height and S/N ratio for each ion was achieved

Analyst, 2013, 138, 4275–4279 | 4277

Fig. 4 Effect of excitation voltage on the peak height (PH) and S/N ratio of Li+

(PH: –:–, S/N: –D–) Na+ (PH: –C–, S/N: –B–) and K+ (PH: –-–, S/N: –,–) peaks(5 � 10�4 M). Operating conditions as for Fig. 2.

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with an excitation voltage of 5.5 Vp-p making it the optimumvalue for the injected gallium electrode C4D system.

Determination of inorganic cations

A sample comprising of 5 � 10�4 M each of K+, Na+ and Li+ (allas chlorides) was separated in under 32 s with an efficiency of17724 theoretical plates per m (Fig. 5). The cations migrated inthe order K+, Na+ and Li+. This is consistent with the migrationvelocity being proportional to the effective charge to solvatedradius ratio of ions in electrophoresis.27 The repeatability ofanalysis using injected electrodes was evaluated by relativestandard deviations (%RSD) of migration times and peakheights. The migration time and peak height repeatability ofeach analyte was determined by performing repeat analyses(n ¼ 6 over a single day) of the same sample using a standardprotocol. The %RSD for migration time was determined to be0.9 for K+, 1.2 for Na+ and 1.1 for Li+ and the %RSD for peakheight was determined to be 1.6 for K+, 2.1 for Na+ and 2.5 forLi+. These values reect the excellent repeatability and

Fig. 5 Electrophoretic analysis with C4D detection of K+, Na+ and Li+ cations in astandard mixture containing 5 � 10�4 M of each ion. Operating conditions:microchip 57/37 mm total/effective length; BGE solution, 15 mM Mes/His;injection voltage, 0.8 kV for 10 s; separation voltage, 1.4 kV. C4D detector: sinewaveform of 220 kHz 5.5 Vp-p.

4278 | Analyst, 2013, 138, 4275–4279

analytical performance of the injected electrode system forquantitative analysis.

Linear calibration curves were measured in the concentra-tion range 1 � 10�4 M to 7 � 10�4 M for each analyte producingR2 values of 0.9789, 0.9794 and 0.9982 for K+, Na+ and Li+

respectively. Using the linear calibrations the limits of detection(LODs) calculated as the average noise + 3.3 � standard devia-tion of the noise for the ions28 were 8.5 � 10�6 M for K+, 6.7 �10�6 M for Na+ and 6.1 � 10�6 M for Li+. LODs of around 3 �10�5 M have beenmost commonly reported with integrated C4Dusing similar conditions to those used in this study.4,18 Ourwork shows slightly superior detection limits than those previ-ously reported for similar devices. The signicant advantagehowever, is the ability to incorporate both electrode and elec-trophoresis channels into the same lithographic process, withsubsequent electrode integration through the injection of liquidgallium. The process is simple and highly reproducible align-ment of electrodes on the chip is achievable.

Conclusions

We have demonstrated the rst use of injected metal electrodesfor C4D detection in PDMS microchips. This novel approach toC4D electrode integration is considerably simpler to achievethan traditional approaches that involve multiple fabricationstages and subsequent electrode alignment using costlyequipment. The electrophoretic separation of Li+, Na+ and K+

cations was used to test the analytical performance of thedevice, showing a detection limit down to 6.1 � 10�6 M and anefficiency of 17724 theoretical plates per m. This is comparableto the performance of devices employing similar electrodedesigns using more traditional fabrication approaches.

Acknowledgements

This research was supported by the Smart Futures FundNational and International Research Alliances Program –

Australian Future Forensic Network (AFFIN), the AustralianNational Fabrication Facility Ltd (ANFF) South Australian NodeStart-Up Award and the South Australian Department of Justice.

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