2009 LabChip Shah EWOD Conduit

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Fluidic conduits for highly efficient purification of target species inEWOD-driven droplet microfluidics†

Gaurav J. Shah* and Chang-Jin ‘‘CJ’’ Kim

Received 7th January 2009, Accepted 14th May 2009

First published as an Advance Article on the web 27th May 2009

DOI: 10.1039/b823541d

Due to the lack of continuous flows that would wash unwanted specifies and impurities off from

a target location, droplet microfluidics commonly employs a long serial dilution process to purify target

species. In this work, we achieve high-purity separation for the case of electrowetting-on-dielectric

(EWOD) based droplet microfluidics by introducing a ‘‘fluidic conduit’’ between a sample droplet and

a buffer droplet. The long and slender fluidic path minimizes the diffusion and fluidic mixing between

the two droplets (thus eliminating non-specific transport) but provides a conduit between them for

actively transported particles (thus allowing the specific transport). The conduit is purely fluidic,

stabilized chemically (e.g. using surfactants) and controlled by EWOD. The effectiveness of the

technique is demonstrated by eliminating$97% non-magnetic beads in just one purification step, while

maintaining high collection efficiency (>99%) of magnetic beads.

Background and motivation

Electrowetting operations of biochemical liquids

Due to its simple design, low power consumption and reprog-

rammable fluid paths, droplet-based or digital microfluidics

driven by electrowetting-on-dielectric (EWOD)1–3 is an attractive

technology to develop microfluidic devices and systems on for

many applications. Unlike continuous flows through micro-

channels, fluids are handled in the form of droplets driven by

sequential actuation of electric potential. Although more difficult

than pure water, there have been increasing reports of using

EWOD for biochemical fluids. One approach is to immerse the

biochemical droplets in an oil2,4 environment, preventing the

intimate contact between the droplet liquid and the hydrophobic

solid surface.5 Although the operation is more challenging in an

air environment,1,6 where the direct contact is allowed, consid-

erable progress has been made.7–9

Non-specific transport undermines purification

In many biochemical assays, high-purity concentration or sepa-

ration of the target species (‘‘TS’’) from the non-target species

(‘‘nonTS’’) is critical. Unlike continuous microfluidics, where the

TS is immobilized while wash-buffer is flowed through the

channels to remove impurities (i.e. nonTS),10 purification in

droplet microfluidics (e.g. by EWOD) typically involves serial

(i.e., repeated steps of) dilution of the nonTS,11,12†. In each step,

a wash-buffer droplet is added and the nonTS are removed (as

‘‘depleted’’ droplets) by splitting the droplet, while collecting the

TS (in ‘‘collected’’ droplet). The distribution of the nonTS

between the two daughter droplets is governed by their initial

distribution in the parent droplet and non-specific transport

phenomena that occur during purification. Two important

mechanisms for the non-specific transport are: (a) diffusion and

(b) fluidic movement. The diffusion of a species inside a fluid is

given by Fick’s first law13 in the one-dimensional case:

J dif ¼ ÀDvf

vx(1)

where J dif is the diffusion flux of a species across a fluidic section,

D is the diffusion coefficient determined by the particle radius,

the temperature and the viscosity of the medium, and f is the

chemical concentration.The second mechanism for contamination is fluidics-driven

transport, i.e. species transported due to the viscous forces

during fluidic movement. According to Stokes’ law, the viscous

drag force F v on a spherical particle of radius r inside a fluid of

viscosity m is proportional to the fluid velocity v:

F v ¼ 6prmv (2)

Although a well-designed electrode layout and droplet actua-

tion sequence can reduce the flow into the collected droplet,14

some flow is inevitable during neck creation and pinch-off

required for droplet splitting. As the droplet is stretched, the

ensuing flow drags the nonTS along with it. For EWOD-drivenmicrofluidics in air, fluidic transport is particularly pronounced

along the free droplet interfaces along its two sides, where flow

velocity is much higher.15

Fluidic conduits: Idea and implementation

NonTS contamination into the collected droplet due to both the

mechanisms described above could be reduced if a slender neck

was created in the buffer droplet prior to sample introduction, as

schematically illustrated in Fig. 1.16 While this neck could be an

effective ‘‘conduit’’ for active TS transport, its slender (long and

UCLA, 420 Westwood Plaza, Engineering IV Bldg, Los Angeles, CA, USA90095. E-mail: [email protected]; [email protected]; Fax: +1-310-206-2302;Tel: +1-310-825-3977

† Electronic supplementary information (ESI) available: 1. Knownpurification techniques in droplet microfluidics; 2. Materials andmethods; 3. Supplementary experimental results. See DOI:10.1039/b823541d

2402 | Lab Chip, 2009, 9, 2402–2405 This journal is ª The Royal Society of Chemistry 2009

TECHNICAL NOTE www.rsc.org/loc | Lab on a Chip



narrow) structure would make it an excellent barrier against

diffusion. Moreover, since little fluidic movement would be

required to cut the slender conduit, fluidics-driven nonTS

transport would be minimized.

Slender physical channels or other filter-like structures have

been used to achieve high purity separation.17 However, such

a permanent provision adds complexity to the fabrication andlacks re-configurability in hardware and operation, defeating an

important advantage of the EWOD platform. Instead, we

propose a purely fluidic conduit generated and ruptured by

EWOD actuation. To overcome the fundamental challenge of

the hydrodynamic instability of such a thin liquid column for

pure water or buffer media, surfactants are added to stabilize the

purely fluidic conduit between the droplets.

Recent reports have shown surfactants to be useful in pre-

venting the irreversible adsorption of biological species like

proteins and cells7,9 on the hydrophobic EWOD surface.However, addition of surfactants to the solution tends to impede

droplet splitting by stabilizing the neck,18 often leading to long

necks, particularly at higher concentrations. Controlling the

neck’s dimensions, location and stability purely chemically is

difficult though. Here, we report achieving controllability using

a combination of surfactants and electrode modification. Specif-

ically, nonionic surfactant pluronic F68 was added to stabilize

neck (conduit) formation, while a stabilizing electrode (‘‘SE’’) was

used to toggle between sustaining and severing the conduit.

Demonstration on EWOD: Results and discussion

Fig. 1 illustrates how the proposed idea is implemented on the

EWOD chip. The typical EWOD electrode layout is modified to

incorporate a slender line electrode (i.e. SE) through the center of

the square EWOD electrodes. The conduit-forming buffer

droplet (left) is stretched towards the sample droplet containing

TS and nonTS. Keeping the SE on, a slender fluidic conduit is

created from the conduit-forming droplet (Fig. 1(a,b)), whose

width is defined by the SE. On merging with the sample droplet,

the TS are transported across the conduit using an active trans-

port mechanism (Fig. 1(c)) such as magnetic.9 However, very few

nonTS can cross the diffusion barrier presented by the fluidic

conduit. After TS transport, the SE is turned off, and the droplet

is stretched further (Fig. 1(d)), breaking the conduit and

completing the droplet split (Fig. 1(e)). Since the neck was

already formed with the buffer droplet before merging with the

Fig. 1 Schematic representation of proposed technique for high-purity

rare TS separation using fluidic conduit. (a) TS are transported to left

edge of sample, while nonTS are randomly distributed in the droplet.With stabilizing electrode (SE) on, the conduit-forming droplet is

stretched, (b) forming a slender ‘‘conduit’’. On merging with sample, (c)

conduit allows active TS transport but restricts nonTS transport. (d,e)

When droplet is stretched with SE off, the droplet splits with minimal

fluidically-driven nonTS transport for high-purity TS collection.

Fig. 2 Image sequence for high purity magnetic separation using droplet conduit structures: All droplets contain 0.15% pluronic F68 in PBS. (a)

Magnet is positioned to the left of the sample and the conduit-forming buffer droplet, so that MBs (dark) are attracted to the left edge of sample (see

inset). (b) A long and thin fluidic conduit is formed by stretching the buffer droplet while the SE is on. (c) Sample is merged with the conduit-forming

droplet, (d) allowing MBs to pass through. After transport, very little fluidic movement is involved as (e) the droplet further stretched with SE turned off,

(f) cutting it into collected (MBs collected) and depleted droplets (depleted of MBs). Satellite droplets can be cleaned up by depleted droplet.

This journal is ª The Royal Society of Chemistry 2009 Lab Chip, 2009, 9, 2402–2405 | 2403



sample, much less fluidic movement is involved during splitting.

As such, the fluidically driven nonTS transport is much reduced.In Fig. 1(e), the collected droplet (left) contains the TS and very

few nonTS, most of which are left in the depleted droplet (right).

To evaluate high-purity separation of TS (4.5mm dia. magnetic

beads (‘‘MBs’’)) from fluorescent nonTS (5.2 mm dia. nonMBs)

using the slender fluidic conduit, separationwith and without† the

conduit was performed. (Details on fabrication,3 materials,16 and

experimental setup19 in ESI†.) These beads were chosen instead of

smaller beads or dyes due to theease of quantification without the

need for high-sensitivity detection systems, particularly since the

nonTS concentration in the collected droplet can be quite low.

Gravity even at this size is still much smaller than other forces like

viscous drag and interfacial forces. In order to demonstrate the

utility of fluidicconduit for purification of rare species, a low ($1 :20) MB : nonMB ratio was chosen.

Fig. 2 shows the sequence of images for the case using a fluidic

conduit. The sample droplet containing MBs and nonMBs,

along with 0.15% w/v pluronic surfactant F68 (optimum

concentration for the current device geometry, to allow repeat-

ably stable neck formation during TS transport, as well as the

subsequent splitting), is placed on the right, while the conduit-

forming buffer droplet, also containing the surfactant, is intro-

duced from the left (Fig. 2(a)). The magnet is positioned at the

left, collecting MBs at the left meniscus of the sample droplet

(Fig. 2(a) inset). A stable, slender conduit is formed by stretching

the conduit-forming droplet while keeping the SE on (Fig. 2(b)).

On merging with the sample (Fig. 2(c)), MBs from the sample areactively transported across the conduit towards the magnet,

while most of the nonMBs remain behind at the right (Fig. 2(d)).

After the MBs are transported (wait time of 10–20 s), the SE is

turned off and the droplet is stretched further (Fig. 2(e)) to split it

into the collected (left) and depleted (right) droplets (Fig. 2(f)).

Since the neck was already formed prior to the merging, the

splitting operation involves much lesser fluidic movement.

Hence, not only diffusion, but also fluidics-driven transport (the

dominant non-specific transport for the present case) into the

collected droplet is drastically reduced.

To quantify the purity of the separation, MBs (dark) and

nonMBs (bright) are counted in the collected and depleted

droplets for cases without (Fig. 3(a,b)) and with (Fig. 3(c,d)) thefluidic conduit. Even though high MB collection efficiency

(>99%) is achieved in both cases, the collection of $33%

nonMBs reduces MB purity. Using the fluidic conduit, on the

other hand, dramatically improves the purity, collecting over 10

times fewer nonMBs without affecting MB collection. The results

have been summarized in Table 1.

Conclusions

In EWOD-driven droplet microfluidics, the purification effi-

ciency of TS by the simple serial-dilution washing steps is limited

Table 1 Summary of experimental results

CaseTotal MBin sample

Total nonMBin sample

MB in collecteddroplet

NonMB incollected droplet

MB indepleted droplet

NonMB indepleted droplet

% MBcollected

% NonMBremoved

No conduit $15 $313 $15 $105 0 $205 $100% 66%With conduit 16 $283 16 <10 0 $267 100% 97%

Fig. 3 Fluorescence images showing much improved purity using fluidic conduit. Some brightfield illumination is used to visualize non-fluorescent

features like droplets’ left meniscus and dark MBs (inset). (a) Collected and (b) depleted droplets without conduit: Despite efficient (>99%) MB

collection, many nonMBs are collected as well, lowering purity. (c) Collected and (d) depleted droplets using conduit: Collected droplet has <10

nonMBs, showing high ($97%) purity with high (>99%) efficiency.

2404 | Lab Chip, 2009, 9, 2402–2405 This journal is ª The Royal Society of Chemistry 2009



because of the non-specific transport of nonTS, mainly as a result

of diffusion and fluidic movement. Contamination due to both

the factors has been reduced using a fluidic conduit without

sacrificing the TS collection efficiency.

The chemically and electrically stabilized fluidic conduit not

only created a diffusion barrier between droplets under EWOD

actuation but also provided a pre-formed neck to minimize

fluidic movement during droplet splitting. By taming contami-

nation from both, diffusion and fluidic mixing, high purity aswell as high collection efficiency of TS was achieved. The

proposed technique helps bolster the purification step, a greater

challenge for droplet microfluidics as compared to the usual

continuous-flow based platforms.

Acknowledgements

This work was supported by NASA (CMISE), NIH (R01

RR020070:01A2 and Pacific Southwest RCE AI065359), and

UCLA Department of Urology.

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2009 LabChip Shah EWOD Conduit