1. 1
Dielectrophoresis of Particles in Microfluidics
Paulo Emanuel Luzio de Melo
1
plmelo@mit.edu
Abstract
This work presents the design, fabrication and testing of a quadrupole electrode chip with integrated microfluidics
prototype for manipulation, separation and concentration of microparticles, through dielectrophoresis (DEP). The built
prototype allowed successfully the characterization of silica microspheres behavior under the action of DEP forces, for
a wide range of frequencies and applied voltages. A method based on positive DEP (pDEP) and negative DEP
(nDEP) sequences was developed and led to high amounts of particles concentrated within the quadrupole structure
area.
Keywords: Dielectrophoresis, Microfluidics, Microparticles, Manipulation, Quadrupole and Comb geometries
electrodes.
I. Introduction..............................................................................................................................1
II. Working Principle and Device Design .....................................................................................1
III. Experimental.............................................................................................................................3
V. Conclusion ................................................................................................................................8
VI. Acknowledgment ......................................................................................................................9
VII. References.................................................................................................................................9
I. Introduction
Recent emerging alternative solutions in
nanotechnologies to manipulate biological particles
have opened a new reality of possibilities at the micro
and nanoscale. The dielectrophoresis is an AC
electrokinetic technique, which enables the
manipulation of particles due to the interaction of
induced dipoles with non uniform electric fields. For
some time now, dielectrophoresis (DEP) has been
continuously used to handle neutral particles (Zou,
Lee, et. al, 2008). Different approaches to manipulate
particles through DEP are continuously being
investigated (Hughes, 2000). Besides the
manipulation of particles, cell and biological molecules
manipulation is also becoming more frequent
(Albrecht et al., 2005; Mittal et al, 2007; Gray et al.,
2004; Chou et al., 2002).
The time averaged DEP force is given by the
following equation (Gascoyne et al, 2002),
𝐹𝐷𝐸𝑃 = 2𝜋𝜀 𝑚 𝑟3
𝑅𝑒[𝑓𝐶𝑀]𝐸2
Where εm is the permittivity of the medium, r the
radius of the particles, E is the root mean square of
the electric field and fCM the Clausius-Mossoti factor
than can be defined as (Gascoyne et al, 2002),
𝑓𝐶𝑀 =
𝜀 𝑝
∗
− 𝜀 𝑚
∗
𝜀 𝑝
∗
+ 2𝜀 𝑚
∗
Where 𝜀∗
is the complex permittivity 𝜀∗
= 𝜀 − 𝑗𝜎/𝜔,
𝜎 is the conductivity, and ω is the electric field
frequency. The p and m correspond to the particles
and the medium, respectively. If the real part of the
Clausius-Mossoti is greater than zero, the particles will
experience positive DEP, and if it is lower than zero
they will experience negative DEP. Particles
experiencing positive DEP, will be attracted to the
maximum electric fields regions, and particles
experience negative DEP will be repelled from the
higher electric field regions.
The subject of this paper is to apply this powerful
technique to manipulate and concentrate a high
amount of particles in a specific designed prototype
device.
II. Working Principle and
Device Design
A. Finite element modeling and
determination of system behavior
To analyze the electrical field distribution in an
aqueous solution around the electrodes that were
designed, finite-element analysis (FEA) simulations
were performed. The simulations were carried out
using COMSOL Multiphysics 3.5 software. From
Figure 1 it can be seen that the generated electric
field energy is nonuniform and that near the vertexes
of the electrodes, the maximum electric field regions
1
Supervisor – Prof. João Pedro Conde

2. 2
are expected. For the quadrupole geometry (Figure 1,
left), the minimum electric field regions are expected
to occur in the middle of the quadrupole. Regarding
the comb geometry (Figure 1, right), the minimum
electric field region occurs between each pair of
electrode fingers. Therefore, particles experiencing
positive DEP (pDEP) will be attracted to the electrode
edges and when negative DEP (nDEP) occurs some
particles will be trapped in the middle of the
quadrupole geometry and trapped in between
electrode fingers (in the case of the comb geometry).
A variation in the geometries used was also
tested, as can be seen in Figure 1. The use of round
shaped edges might allow higher concentration of
particles during dielectrophoresis, since a larger
region of maximum field intensity will occur and the
minimum field regions will be better defined.
The simulations performed were done mainly
to assess the distribution of the electric field for the
different geometries, however further analysis could
combine these simulations with microfluidics and DEP
force simulations, which would allow to estimate the
drag force to remove a particle in continuous flow or
even estimate the maximum/minimum number of
particles that can be trapped.
Conductivity
(S·m-1
)
Relative
Dielectric
constant
Particles
(Silica)
10-16
2,5
Medium (DI
water)
8,5×10-5
80
Figure 1. Electrical behavior close to the quadrupole and comb geometry electrodes (dark blue). Simulation of the
electric field energy distribution (in V/m) around the quadrupole geometry (left) and around the comb geometry (right)
with normal edges (a and b) and round shaped edges (c and d). The distance between electrodes is 5 µm (FEA
simulations, COMSOL Multiphysics 3.5).
a) b)
c) d)
Table 1. Parameters used in calculations of the
Clausius-Mossoti factor (note: the vacuum
dielectric constant considered was 8,8×10-12
)

3. 3
The prediction of the particle behavior for this
particular set of experiments was also carried out, by
obtaining the relation between the frequency and the
real part of the Clausius-Mossoti factor (Figure 2).
. Observing the figure, it can be seen that for all range
of frequencies, the real part of the Clausius-Mossoti
factor is always negative. This means that
theoretically, for the given conditions, the particles will
always experience negative dielectrophoresis
independent of the applied frequency, and for that
they will be repelled from the higher electric field
regions to the lower electric field regions.
10
0
10
1
10
2
10
3
10
4
10
5
10
6
10
7
10
8
10
9
10
10
10
11
10
12
10
13
10
14
10
15
-0,505
-0,500
-0,495
-0,490
-0,485
-0,480
-0,475
Real(fCM)
Frequency (Hz)
Figure 2. Predicting the particles behavior. Relation
between the real part of Clausius-Mossoti factor and the
electric field frequency.
B. Device design and experiment
planning
The prototypes fabricated in this experiment were
designed with AutoCAD 2009. The design of the
devices had to have in account specific issues related
to equipment constraints. The thickness of the PDMS
had to be controlled since it should be minimized in
order to increase the visibility of the aluminum
structures, but also to have in account the focus
distance of the objective. For that reason, the PDMS
thickness was always below 0,3 cm. Another issue
was related to the distance inlet-outlet in the
microchannels and the diameter of the objective,
because this distance should be sufficient for the
objective to move freely around the quadrupole and
comb structures. As the diameter of the objective was
around 2,5 cm, the distance inlet-outlet chosen was
3,6 cm, having in account the distance between
structures. The design of the devices built is depicted
in Figure 4. Observing this figure, it can be seen that
both devices were designed with 16 aluminum pads,
which means that in the quadrupole design, there are
four sets of structures (4 pads for each), and for the
comb design there are 8 sets of structures (2 pads for
each). There are two microchannels in each design,
each of these with half the total structures of the
design (2 for the quadrupole and 4 for the comb). The
difference between the two microchannels is in the
quadrupole (or comb) geometries, meaning that in
one of the channels there was only electrode
geometries with normal edges (squared) and on the
other there was only round-shaped electrode
geometries.
In the modeling previously shown, the distance
between electrode structures was 5 µm, but since the
beads used had 1 µm of diameter, distance of 2 µm
was also designed in attempt to achieve single bead
entrapment.
The experiment design setup is depicted in Figure 3.
The objective was to use a syringe pump to inject the
beads solution in the microchannel, then apply an AC
signal in the electrodes to excite the electric field
inside the channel and observe the dielectrophoresis
under a fluorescence microscope.
III. Experimental
A. Device Fabrication
The microfabrication process used to produce the
device is summarized in Figure 5. Essentially, the two
chips were produced separately (the DEP electrodes
chip and the microfluidics chip) and then bonded to
each other. The detailed protocols for the
microfabrication processes are in appendix.
To address the two types of geometries modeled, two
different devices were fabricated, the first one based
on the quadrupole geometry electrodes and the
second one based on the comb geometry. The
fabrication processed was the same for both, except
in the step to pattern the geometries on the aluminum,
where different masks were loaded.
DEP Electrodes chip
The DEP electrode chip was fabricated on a glass
slide where a 100 nm thick layer of aluminum was
deposited (Nordiko 7000). A 5 µm thick layer of
negative photoresist was first spin-coated above the
aluminum and then patterned in a Direct Write Laser
system (DWL), with the desired DEP electrodes
geometry. After, aluminum wet etch was performed in
Figure 3. Schematics of the experiments setup (adapted
from Albrecht et al., 2005).

4. 4
the chip to remove the aluminum that was not
protected by the photoresist. The residual photoresist
on the chip was then removed by placing the chip in
microstrip.
Microfluidic chip
The microfluidic chip was fabricated in polymeric
compound named polydimethylsiloxane (PDMS), by
using a mold. To accomplish this, a physical mask was
first fabricated to carry out the patterning of another
substrate, which would be the microfluidics mold that
would allow the fabrication of the microfluidic chip.
The fabrication of the physical mask was performed in
a quartz slide, where a 100 nm thick layer of
aluminum was deposited (Nordiko 7000). A 5 µm thick
layer of negative photoresist was first spin-coated
above the aluminum and then patterned in a DWL
system, with the desired microfluidic geometry.
Subsequently, aluminum wet etch was performed in
the mask to remove the aluminum that was not
protected by the photoresist. The residual photoresist
on the mask was then removed by placing the mask in
microstrip. The microfluidic mold was fabricated on a
Silicon substrate, where a 20 µm thick layer of SU-8
photoresist was spin-coated and baked. Afterwards,
the SU-8 was patterned in a ultra-violet chamber, by
means of the quartz physical mask previously
fabricated and then it was developed. Having the
microfluidics mold, the PDMS was prepared,
dispensed on top of the mold and then cured at 60ºC
for 1h30. The processed PDMS was fabricated to
have a height inferior to 0,25 cm, in order that a
proper focus of the structures could be achieved.
Holes at the inlets and outlets were drilled for fluidic
interconnection, with a 20-gauge flat tip needle. In
order to achieve a permanent hydrophilization of the
fabricated PDMS, a treatment based in triethylamine,
ethyl acetate and acetone was applied to the PDMS.
The bonding of the microfluidic chip to the DEP
electrodes chip was performed by applying a corona
discharge to the PDMS surface that is supposed to
DEP electrodes Chip
Figure 5. Simplified illustration of the microfabrication process used to produce the device (adapted from Zou et al.,
2008).
Figure 4. Design of the devices built, the quadrupole geometries (left) and the comb geometries (right) chips (images from
AutoCAD 2009)

5. 5
bond, and then bringing it in contact with the
electrodes surface. This bonding was performed
under the optical microscope with some drops of DI
water to allow the alignment of the structures. Metal
connectors were introduced at the inlets and outlets to
strengthen the fluidic interface connection. Silicone
had to be placed in the connectors, near the inlets and
outlets, due to the low thickness of the fabricated
PDMS.
The final device can be seen in Figure 7. The chip
size is 4,2 cm x 1,6 cm. It has two microfluidics
channels, each with 20 µm height, 100 µm wide and
3,2 cm long. The inlets and outlets are squares of 2
mm, also with 20 µm of height. The total volume of the
each channel is 0,224 µL. Regarding the aluminum,
the current lines are 200 µm wide and the distance
between adjacent current lines is always 200 µm. The
difference between the two microchannels is in the
quadrupole (or comb) geometries, meaning that in
one of the channels there was only electrode
geometries with normal edges (squared) and on the
other there was only round-shaped electrode
geometries. Within each channel, differences exist in
each set of electrodes, but only regarding the distance
between electrodes of the quadrupole (or each finger
for the comb geometry case). For these distances, 2
µm and 5 µm were used.
One important issue that occurred during the
fabrication process was related to the PDMS
dimensions, since the microfluidics mask was made
without having in account differences in the PDMS
dimensions after the cure, which led to inability to
align correctly both channels. However, this problem
was solved by cutting the PDMS in two pieces (each
with a channel) and aligning them separately. The
alternative to avoid this problem could be designing
the mask to have in account the shrinkage percentage
of the PDMS, through the control of parameters such
as cure time and temperature. The relation between
shrinkage, cure time and temperature for PDMS can
be found elsewhere (Krogh & Asberg, 2003).
B. Experimental Setup
The experiment setup used to perform the
dielectrophoresis with microfluidics is shown in Figure
6. The flows containing the microparticles solution
were injected through a syringe pumping system at
controlled rates (NE-1000 Single Syringe Pump, New
Era Pump Systems, NY, USA). Flow rates up to 5
µl·min
-1
were used to inject the desired solutions in the
4
1
2
3
5
1. Waveform generator
2. Oscilloscope
3. Syringe pump
4. Fluorescence microscope
5. Designed device
Figure 6. Experiment setup for the DEP of microparticles in Microfluidics using the designed device.
Alluminum (pad)
Inlet PMDS
SiliconeOutlets
Metal connectors
Figure 7. Photograph of the final device.

6. 6
microfluidics channels. To apply the AC signal, a
function waveform generator (Agilent 33220A function
waveform generator, Agilent Technologies, Inc., CA,
USA) was electrically connected to the aluminum
pads of the device, by using gold wires and indium to
perform the connection at the interface. An
oscilloscope was also coupled to the system setup
(Tektronix TDS 340, Tektronix, USA). A sinusoidal
voltage signal with frequencies up to 20 MHz and
amplitudes up to 10 Vpp (peak-to-peak) was applied
to the electrodes. The images and videos of the DEP
behaviors were captured by means of a fluorescence
microscope (Leica Microsystems, USA) coupled with a
video recording system and by the use of the software
Jasc Paint Shop Pro installed on a computer. Green
fluorescent silica microspheres (sicastar ® -
greenF (plain), Micromod, Germany) with 1 µm of
diameter were used in the experiments. The solution
flowed into the microchannels was 50 µl of the stock
solution (50 mg·mL
-1
in DI water) diluted in 150 µl of
DI water.
The tests performed in the device were only
done in one of the two channels, since the other
channel (with the round shaped quadrupoles) was
misaligned, due to the fact that the PDMS had shrunk
during its processing. The experiments made were
basically with no flow, meaning that the particles were
injected into the channels, and then only when all the
flow had stopped and the Brownian motions were
dominant, the experiments were performed. However,
some preliminary DEP tests with flow were also
carried out.
IV. Results and Discussion
The preliminary DEP experiments were only
performed in the quadrupole geometry chip. However,
as it was previously stated, a misalignment of the
channels occurred, because the PDMS shrunk. For
this reason, tests were only performed in one of the
channels. All the next microscope images were
captured with an amplification of 600 times.
To avoid a possible electrode corrosion (water
electrolysis), due to low frequencies or higher
voltages, the first experiments were performed starting
with low voltages (around 500 mV) and going up to 10
V, while for the frequencies, tens of megahertz were
chosen (10 MHz and below). However, the device was
used for extensive periods of time (around 3 hours
continuously) and no damage on the electrodes was
visible. In Figure 8 can be seen the dielectrophoretic
force acting on the particles. As it was supposed,
nDEP occurred for a frequency of 10 MHz (see Figure
2) that made the particles be repelled from the higher
electric field regions to the lowest ones, which led to a
trapping of the cells in the center of the quadrupole.
The experiments results show that the DEP force is
frequency and voltage dependent. In Figure 9 can be
seen the DEP force response with the variation of the
applied frequency (left) and voltage (right). The
voltage dependence in Figure 9 (right) shows that as
the applied increases the the DEP force also
increases, meaning that in this case that the particles
will be more repelled from the higher electric field
regions (nDEP). Also it is noticeable, that for 2,5 Vpp
the DEP force is not strong enough to counteract the
Brownian motions of the particles as well as the
hydrodynamic forces existent due to some residual
flow velocities, which makes the particles in the
quadrupole center to be dragged away continuously
until there are no particles (see Figure 9, bottom figure
on the right). The results showed that there is a
threshold for the particles to become trapped in the
center of the quadrupole. For an applied frequency of
10 MHz, this threshold is around 5 to 6 Vpp (peak-to-
peak). It is important to notice that if the DEP
experiment starts with low applied voltages (below the
threshold), no trapping will occur in the quadrupole
center until a higher threshold is achieved (for 10 MHz
this value is around 8-9 Vpp). Other important fact is
that the initial threshold to trap particles in the
quadrupole center is actually higher than the lower
limit to maintain them trapped in the center.
Regarding the frequency dependence, Figure 9 (left)
shows that, for an applied voltage of 10 Vpp, the
nDEP force increases as the frequency decreases.
This makes the particles be more repelled from the
higher electric field regions (corners and edges of the
quadrupole) as the frequency decreases. However,
the experiment tests show that this only occurs until
around 10 kHz. At this frequency, both pDEP and
nDEP start to occur, and below only pDEP occurs
(see Figure 10). This frequency corresponds to the
case where the real part of the Clausius-Mossoti
factor is equal to 0. In the frequencies of 10 kHz and
below, the particles are attracted to the high electric
field regions (corners and edges). The conducted
tests for pDEP ranged from 10 Hz to 10kHz, and even
at these lower frequencies no damage to the
electrodes occurred.
Having the possibility to use pDEP forces, several
tests were performed. A high concentration of particles
in the quadrupole center was achieved through the
combination of pDEP and nDEP in a particular
sequence. Observing Figure 11, two scenarios are
presented, the first one applying a signal of
Figure 8. Dielectrophoresis force experiment in the
quadrupole geometry (5 µm), within the microfluidic
channel with no flow. Negative DEP occurred for a
sinusoidal signal of 10 V @ 10 MHz.

7. 7
10V@1kHz and then changing the frequency to
10MHz, and the second one applying a 10V@1kHz
signal and then changing it to 10kHz. What happens
is that by applying a pDEP force first (10V@1kHz) and
then changing the frequency to 10 kHz, which is the
frequency where K(w) equals zero (meaning pDEP
and nDEP) a great amount of particles was possible
to be trapped in a large area of the quadrupole.
Until now, all the presented results were obtained with
the syringe pump turned off. Though, some
preliminary DEP tests at different flow velocities were
performed. Flow velocities from 0.5 to 5 µL/min were
used combined with strong DEP forces
(10V@10MHz), however the particles were unable to
be trapped at the center of the quadrupoles. This
probably happened because the flow velocities used
were still too high for the dimensions of the channel,
which had a volume of approximately 0,22 µL.
Nevertheless further testing should be done to assess
these issues.
Figure 9. Variation of the DEP force with the applied frequency (left), maintaining the voltage constant at an amplitude of 10 V.
Variation of the DEP force with voltage (right), maintaining the frequency constant at 10 MHz.Quadrupole geometry was used
with a distance between electrodes of 5 µm.
100 kHz
10 MHz
1 MHz
10 V
2,5 V
5 V

8. 8
V. Conclusion
In this work, a quadrupole electrode chip with
integrated microfluidics prototype was designed,
fabricated and characterized to manipulate, separate
and concentrate microparticles using
dielectrophoresis. Silica microspheres of 1µm of
diameter were successfully manipulated, through
DEP, to accumulate in the quadrupole center, without
being in a continuous flow (within a microfluidic
channel).Regarding the dimensions of the microfluidic
channels, it became clear that in the future the depth
and width of microfluidic channels can be further
reduced, forcing the particles to flow more close to the
electrodes, and probably increasing the separation
and concentration of particles. Besides the
quadrupole geometry, a different geometry to perform
DEP was also designed and fabricated, the comb
geometry. However, testing of this type of structure is
yet to be performed. Several tests in the quadrupole
geometry were performed to determine the frequency
and voltage dependencies of this type of device. A
10kHz
Figure 10. DEP force for an applied sinusoidal signal of 10
V@ 10 kHz. nDEP and pDEP occur simultaneously.
1kHz 10 MHz
10 kHz1kHz
Figure 11. Method to concentrate a high amount of particles at the quadrupole center. Two scenarios: A 10 Vpp @ 1kHz
signal is applied to quadrupole and then changed to 10 MHz (top row) and to 10 kHz.

9. 9
method to concentrate a high amount of particles in
the quadrupole center was also successfully
developed. This type of device can be further tested
and optimized for the particles used, as well as other
types of beads, but also for cells, making it useful for
biological applications, such as lab-on-a-chip.
VI. Acknowledgment
The author gratefully thanks all the INESC-MN staff
for their support, and a special thanks to Prof. João
Conde, Milene Santos, Ana Teresa Pereira, Diogo
Martins, Agnieszka Joskowiak and Riccardo Scipinotti,
for their technical support and discussions.
VII. References
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Zou, Z., Lee, S., & Ahn, C. (2008). A Polymer
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