Lab on a Chip for

Live-Cell Manipulation
Gianni Medoro
Silicon Biosystems
Roberto Guerrieri
University of Bologna
Nicolo` Manaresi
Silicon Biosystems
Claudio Nastruzzi
University of Perugia
Roberto Gambari
University of Ferrara
&RECENT ADVANCES IN the life sciences stem from
a convergence of IT and new tools and machines for
large-scale analysis of the structure of DNA, proteins,
and cells. Projects such as the Human Genome Project
are striking examples of the power of this conver-
gence. Although these projects have demonstrated the
possibility of obtaining information directly from DNA,
its clear that the complexity of this task is enormous
and far from achievable with our limited understand-
ing of the basic processes that translate DNA into
structures. Fortunately, IT can provide a bridge be-
tween DNA and cells, considered here as the basic
building blocks of any complex living being.
In this article, we examine the new microelectronic
technology that gives scientists the ability to monitor,
sort, and analyze vast populations of cells and interact
with each cell individually. A microelectronic platform
called a lab on a chip (LoC) allows precise
manipulation of cells with no effect on their pheno-
types. The motivation for developing this technology is
that investigations in recent years have shown that
a few cells changing their behavior unexpectedly can
induce deadly diseases such as cancer. Current LoC
design and manufacturing techniques are spawning
new biotechnology methods with potential for re-
search, diagnosis, and therapy. Whether
the new techniques will meet the
challenge remains to be seen, but the
urgent need to find these answers is
clear.
Dielectrophoresis principles
Several approaches to cell manipu-
lation in LoCs use dielectrophoresis (DEP), which in
some cases can substitute for or integrate with fluidic
technologies and performcomplex analytical-protocol
steps in miniaturized devices. Pioneered by Pohl,
dielectrophoresis is the physical phenomenon where-
by neutral particles, in response to a spatially non-
uniform electric field (E), experience a net force
directed toward locations with increasing or decreas-
ing field intensity according to the physical properties
of the particles and the medium.
13
When the force is
directed toward locations with increasing field in-
tensity, it is called positive dielectrophoresis (pDEP);
in the second case, it is called negative dielectrophor-
esis (nDEP). Figure 1 shows the basic principle
supporting the motion of neutral particles by dielec-
trophoresis. The theory states that the DEP forces
direction is independent of the sign of the voltages that
energize the electrodes. This is a fundamental property
of dielectrophoresis because it allows the use of time-
varying (AC) signals to avoid the drawbacks con-
nected with DC or low-frequency signals (for example,
electrolysis).
Switching the frequency of the electrical stimuli
causes the transition from negative to positive dielec-
trophoresis. In fact, there is a relationship between
Editors note:
Precisely manipulating and sorting live cells on a lab on a chip is still a major
challenge. This article shows how to use dielectrophoresis for cell sorting. The
authors also describe a prototype CMOS chip with a sensor-actuator array,
row-column addressing logic, and readout circuitry.
Krishnendu Chakrabarty, Duke University
Biochips
0740-7475/07/$25.00 G 2007 IEEE Copublished by the IEEE CS and the IEEE CASS IEEE Design & Test of Computers
26
dielectrophoretic force and stimulus frequency for any
specific kind of particle or cell.
4
Graphs called
dielectrophoretic spectra plot such relationships.
Scientists have exploited this property to separate
different cell species.
5,6
For spherical geometries, the first-order expression
of time-averaged DEP forces is
~
FF
DEP
x, y, z, v ~2pe
0
e
m
R
3
< f
CM
v f g
~
++E
2
rms
where e
0
is the vacuum dielectric constant, e
m
is the
medium dielectric constant, R is the particle radius,
E
rms
is the root-mean-square value of the electric
field, v is the angular frequency, Ris the real part, and
f
CM
(v) is the Clausius-Mossotti factor. The latter is
a function of the complex permittivity of the particle
and medium, and is defined as
f
CM
v ~
e

p
v e

m
v
e

p
v z2e

m
v
where e

p
v and e

m
v are the complex permittivity of
particle and medium, defined as
e

p
v ~e
p
z s
p
=jv

and
e

m
v ~e
m
z s
m
=jv
where e
p/m
is the dielectric permittivity, and s
p/m
is the
conductivity. For nDEP, R{f
CM
(v)} , 0; for pDEP,
R{f
CM
(v)} . 0. The electric fields minima corre-
spond to the attraction regions of nDEP, and the
maxima correspond to the attraction regions of pDEP.
The tiny dimensions of electrodes available
through microlithography make it possible to realize
very high field gradients with lowvoltages (compatible
with microelectronic circuits). As a result, dielectro-
phoresis can manipulate microscopic neutral particles
such as cells or bacteria, whose dynamics are
governed by the balance between the DEP force and
the effects of viscous friction. To estimate the effects of
this force on cells, we define the approximate
instantaneous velocity of a particle in a fluidic
medium as
~vv
DEP
~
~
FF
DEP
= 6pgR
where g is the medium viscosity. We can also define
a dielectrophoretic mobility:
m
DEP
~(2e
0
e
m
<ff
CM
g j j R
2
)=6g
which describes the dependence of particle dy-
namics on fluid viscosity, particle size, and the
Clausius-Mossotti factor.
7
Particle speed is therefore
ruled by the following expression:
~vv
DEP
j j ~m
DEP
+
!
E
2
rms

which indicates the existence of a linear relation-

ship between particle speed and dielectrophoretic
Figure 1. Polarization of particles subjected to negative (a) and positive (b) dielectrophoresis (e
p
is the
particle, and e
m
is the medium dielectric constant).
JanuaryFebruary 2007
27
mobility, and a squared relationship with the ap-
plied potentials.
From these equations, it follows that we can
improve the dynamics simply by increasing the field
gradient (by reducing the electrodes feature size);
decreasing the particle size strongly reduces the
dynamics, thereby limiting the dimension of the
smallest particles that can be manipulated.
Researchers have exploited this property to de-
velop an effective approach to separation and size
characterization of bioparticles.
8
The novelty of the
approach is that it doesnt require fluid flow, using
moving dielectrophoretic cages instead. A dielectro-
phoretic cage is generated when an electric-field
configuration induces dielectrophoretic forces that
capture particles. By sequencing a set of cage patterns
in a microfabricated chip, we can selectively impart
a movement to bioparticles, depending on the ratio
between dielectrophoretic mobility and cage speed.
Dielectrophoresis-based LoCs
Studies of DEP action on biological or inorganic
matter suggest the use of dielectrophoresis not only to
sort cells but also to characterize them by differences
in physical properties, or, in general, to manipulate
them.
9
For example, Suehiro and Pethig use both
pDEP and nDEP to precisely displace cells in
a microchamber formed between two facing glass
chips with elongated electrodes.
10
The main drawback
is that cells contact device surfaces and usually stick to
them. Levitating the cells during manipulation can
overcome this problem. Because the electric fields
maxima cannot be established away from the elec-
trodes, stable levitation is possible only with nDEP
force. Hence, researchers have proposed using closed
nDEP cages. Muller et al. use 3D structures of elec-
trodes located at the vertexes of a cube for this
purpose.
11
This techniques main limitation is that it
requires fluid flowto lead cells into and out of the DEP
cage. As an alternative to fluid flow, Huang et al. use
traveling waves combined with nDEP to move cells in
a microchamber.
12
However, its difficult to position
cells as precisely as required by multistep experimen-
tal protocols, because cell speed depends on cell type.
Another approach to cell manipulation is the
moving cages mentioned earlier.
13
This approach
combines the advantages of stable cell levitation and
the use of planar technologies. It relies on the
possibility of creating large arrays of electrodes
separated from the biological liquid by a thin (a few
nanometers) dielectric layer that avoids electrolytic
effects and contamination caused by metals. These
electrodes are polarized to create dielectrophoretic
cages that effectively trap particles. Because the
applied potentials are generated under software
control, cages can be flexibly generated within the
medium. Several studies have proved the effectiveness
of this approach for implementing complex applica-
tion protocols.
14,15
The devices studied used standard
technologies ranging from a PCB to a CMOS de-
vice.
6,16,17
In the last case, small electrodes, comparable
in size to cells, trap single particles, keep them
levitated, and drag themacross the entire chip surface.
In general terms, a dielectrophoresis-based LoC
consists of two juxtaposed main modules. The first
module contains a plurality of electrodes regularly
arranged on a substrate (if this module is realized in
the CMOS process, it can include memory elements
and sensors). The second module consists of a single
large electrode fabricated in a conductive, preferably
optically transparent material. Figure 2 shows a CMOS
chip for single-cell manipulation.
A spacer inserted between the upper and lower
modules creates a chamber that contains the sample to
be analyzed or manipulated. The same spacer can also
establish separation walls in the device to create multiple
chambers optionally connected by dedicated ports.
8.1 mm
7
.
8

m
m
Figure 2. CMOS chip for single-cell manipulation
by dielectrophoresis (Source: Manaresi et al.
16
).
An electronic circuit is associated with each
element of an array of 100,000 electrodes to
program the location of up to 10,000 closed
DEP traps.
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IEEE Design & Test of Computers
28
CMOS design constraints and specifications
The CMOS device mentioned earlier is optimized
for handling eukaryotic cells (such as the lymphocytes
in blood) in the range of 10 to 20 microns in
diameter.
16,17
A design guideline derived from an
analysis of simulated horizontal DEP forces as
a function of particle size with respect to the
electrodes suggests that the electrode pitch should
be similar to the cell size. Increasing the cages width
to encompass more than one electrode enables the
device to handle larger particles. Increasing the
number of electrodes in the array increases the
devices capacity (number of cells in the input
sample) and selectivity (ability to select a smaller
percentage of cells).
However, silicon cost increases with chip size.
Accordingly, the designers of the device chose the
total number of cages to be on the order of 10,000. This
is sufficient to recover a significant number of cells (10
to 100) that may be present in a low percentage (0.1%
to 1%) in the starting sample. At the same time, the
chip size (8 3 8 mm
2
) is reasonable.
The time constants for cell motion caused by DEP
forces are relatively slow(about one secondor more for
a 20-micron step). This relaxes timing constraints for
array programming, as well as for thesensing framerate.
Fabrication and technology constraints
To choose the most appropriate CMOS technology,
the device designers took the following considerations
into account. Because the DEP force is proportional to
the square of the voltages applied, the supply voltage
should be as large as possible, to limit actuation
voltages. In contrast to conventional IC designs, the
lower the technology resolution, the lower the chip
cost, because die size is set from other specifications
related to cell size. Scaling beyond the point at which
the required number of transistors fits in the microsite
area doesnt improve cost or performance. Scaling is
required only for manipulating smaller cells, such as
individual bacteria (which are typically 1,000 to
3,000 nm in diameter) or viruses (100 to 300 nm in
diameter).
In addition, LoC designers must consider some
constraints that are unusual for electronic designers.
For example, they must pay attention to the effects
of contact of the liquid sample with the top metal
electrodes. Excessive chip passivation thickness can
limit the electric fields capability to penetrate the
liquid at the desired operation frequency, especially
when biological science specifies the liquids con-
ductivity at rather high levels (for example, the
physiological solution has a conductivity of greater
than 1 siemens/meter. Fortunately, process options
that reduce this thickness to a few nanometers are
available.
Other typically biological aspects such as auto-
fluorescence of materials used are also involved. In
fact, background fluorescence from the chip surface
can impair the chips ability to detect a weak signal
for example, from fluorescently labeled antibodies
typically used to identify key cells in many biological
protocols.
Another limitation of CMOS technology is the
silicon dies opaqueness. To overcome this limitation,
researchers have proposed an approach to the
massively parallel individual manipulation of biopar-
ticles that is compatible with transistorless and trans-
parent chips.
18
They have built a prototype micro-
system using gold and indium tin oxide (ITO) on glass
to implement a 2D array of microsites, establishing
hundreds of dielectrophoretic microtraps. Each trap
can gain control over an individual cell (or mi-
crobead) and can be reconfigured by software to
transfer the cell to any adjacent trap. This approach
allows digital control of the movement of many
individual cells along different paths. Figure 3 shows
a diagram of the transparent chip.
Unlike solutions using optical or optoelectronic
tweezers,
19
this approach doesnt require external
bulky instruments, and it manipulates cells without
contactwhich is impossible with transistorless tech-
niques, such as those based on positive dielectrophor-
esis.
10
Its main limitation is the high number of signal
controls, which is comparable to the square root of the
maximum number of traps.
Other cell manipulation techniques, including
optical tweezers,
20
magnetic tweezers,
21
acoustic traps,
and hydrodynamic flows, have severe limitations if
both high throughput and high resolution are required.
Researchers have proposed another cell manipulation
technique, which combines optical images and
dielectrophoresis for parallel manipulation of many
individual cells.
22
This technique requires a bulky
external instrument and is therefore not suitable for
point-of-care applications.
Packaging LoC devices
Integrating microelectronics and biology requires
the construction of a hybrid technology with adequate
JanuaryFebruary 2007
29
fluid transport capabilities. Thus, researchers have
introduced the concept of microfluidics, the building
block of integrated LoCs. Vulto et al. describe an
example of this process integration.
23
They propose
using Ordyl SY300/550 dry film resist (DFR) from Elga
Europe to build microfluidic structures. DFR, originally
developed for PCB fabrication, offers many advan-
tages over liquid resist: conformability, adhesion to
any substrate, flatness, no liquid handling, no edge
bead formation, uniform distribution, low exposure
energy, low cost, short processing time, and near-
vertical sidewalls.
24
DFR is an excellent material for fast
prototyping of fluidic structures. Its low number of
simple processing steps make DFR ideal for industrial
applications as well, particularly because the process-
ing steps are fully compatible with CMOS silicon
technology. Moreover, manufacturers can perform
DFR processing on a full wafer scale with great
advantages for mass production over conventional
microfabrication techniques. Figure 4 shows photos of
resist structures on a silicon substrate. (These struc-
tures are examples of building blocks used in several
common situations.) The smallest reproducible
features for freestanding
structures are 20 microns
wide for a resist thickness
of 54 microns. The smal-
lest gap dimension for the
same parameters is 40
microns wide.
An alternative tech-
nique for creating micro-
fluidic prototypes is poly-
dimethylsiloxane (PDMS)
molding. PDMS allows
easy sealing and fast fa-
brication of complicated
structures.
25
Other com-
monly used fabrication
techniques are glass wet
etching,
26
silicon etching,
27
and polymer molding us-
ing the LIGA technique
(LIGA is an acronym for
the German words for x-
ray lithography, electro-
forming, and molding).
28
Fluidic structures can also
be sandwiched between
two processed substrates.
The most common method of making such hybrid
chips is to pattern the fluidic structures in SU-8
photoresist.
29,30
UV-laser-photoablated polymers,
31
CO
2
-
laser-engraved polymers, and even double sticky
tape
32
have also been reported suitable for creating
fluidic gaskets. SU-8 has advantages over the other
techniques in resolution and in requiring no additional
adhesives for double-bonding the substrate.
Whatever the technology used to realize an LoC,
the device must move a biological sample through the
channels of a microfluidic circuit to load the sample or
recover selected cells. One mechanism proposed for
this liquid transport is the micropump, a device that
can move volumes of liquid in the micro-to-nanoliter
range. So far, the use of this mechanism is limited,
mainly because micropumps with the right combina-
tion of cost and performance are unavailable.
33
In most cases, scientists achieve liquid transport in
microfluidic chips through manual pipetting with
external pneumatic sources or through electro-osmot-
ic flow. To selectively recover separated particles, they
use a double pipette that establishes a laminar flow in
the separation chamber and recovers two halves of the
Figure 3. Fully transparent chip using a glass substrate for single-cell manipulation by
dielectrophoresis (Source: Medoro et al.
18
Reproduced with permission). The cell
manipulation approach is based on the changing phases of the stimuli applied to rows
and columns. With N + M control signals, the array can control up to N 3 M cells.
Biochips
IEEE Design & Test of Computers
30
sample liquid.
34
In this
case, interface hole dia-
meters are adapted to the
pipette tips size, enabling
liquid actuation by pipette
pressure. Because the sep-
aration mechanism is very
sensitive to unwanted liq-
uid flow, scientists prevent
hydrostatic flow by direct-
ly inserting the sample
into the chamber. They
control the resulting liq-
uid-air interface by intro-
ducing phase guides (reg-
ular spatial structures
characterized by different
wettability that can create
a precise movement di-
rection of the fluid). This
technique is excellent for
recovering spatially separated particles and requires
minimal surface area for microfluidics. This is
especially important with expensive silicon chip
technology. Figure 5 shows an example of a working
LoC designed as just described.
Biotechnological LoC applications
LoC devices offer a unique opportunity for in-
tegrating complex protocols on single miniaturized
platforms. The platforms combine sophisticated
sensing and computational technologies in miniatur-
ized analytical instruments for manipulating single
living cells or cell populations. Reproducing a com-
plete biological experiment on a single cell, the
smallest living element, can provide scientists with
exclusive deterministic information rather than the
statistical results so far available with conventional
laboratory procedures. The ability to manipulate cell
populations enables scientists to performprogrammed
interactions between cells, or between cells and
microspheres, without requiring complex laboratory
toolsenabling new experimental plans as well as
diagnostic procedures. This explains scientists grow-
ing interest in exploiting microelectronics in combi-
nation with live biological matter.
Cell isolation
Researchers have reported on using DEP-based LoC
platforms to isolate cell populations or single cells.
15,35
Isolating rare cells from biological fluids, such as
whole blood and bone marrow, would provide new
scope for cell-based diagnosis, cell-based therapy, and
phenotypic characterization of cell population sub-
sets. For instance, characterizing a few metastatic
cancer cells for the purpose of further molecular
analysis can play an important role in the diagnosis
and prognosis of cancer patients. Isolating so-called
Figure 5. A working LoC (a CMOS biosensor-
actuator for cell analysis). The LoC developers
implemented the fluidic microchamber
packaging by double-bonding the ITO-coated
glass, patterned with dry-resist film, to a CMOS
chip. (ITO is indium tin oxide.)
Figure 4. Resist structures on a silicon substrate, with exposure energy

150 mJ/cm
2
,
85uC/(5 min): castellated channels with 50-micron dimensions (a); square and round
pillars with dimensions of 50 and 20 microns (b). Insets showdetailed top views (Source:
Vulto et al.
23
Reproduced by permission of The Royal Society of Chemistry).
JanuaryFebruary 2007
31
cancer stem cells might lead to diagnostic and
therapeutic innovations.
Yu et al. describe an application of DEP-based
protocols to cell isolation.
35
They have developed an
efficient method for trapping neurons and construct-
ing ordered neuronal networks on bioelectronic chips
using arrayed nDEP forces. According to their pro-
tocol, neurons adhere and are then cultured directly
onto the bioelectronic chip, exhibiting good neuron
viability and neurite development.
Cell population separation
Huang et al. describe a dielectrophoretic field-flow-
fractionation method for purging human breast cancer
cells from hematopoietic stem cells.
36
The same group
also demonstrated high performance in separating
human blood cells: T or B lymphocytes from mono-
cytes, T or B lymphocytes from granulocytes, and
monocytes from granulocytes.
Another interesting application is using dielectro-
phoresis to separate viable from nonviable yeast
cells. Researchers used known mixtures of viable
and heat-treated cells of Saccharomyces cerevisiae
(bakers yeast) with 60% nonviable cells present; the
results demonstrated that the DEP-separated non-
viable portion contained only 3% viable cells, and
the viable portion contained 8% dead cells. Impor-
tantly, the separation procedure didnt affect cell
viability.
Another example is using DEP-based LoCs to
isolate infected cells from noninfected counterparts.
LoC platforms carrying spiral electrodes can isolate
malaria-infected cells from blood. Parasitized cells
cluster at the center of a spiral electrode array
because during development of Plasmodium falci-
parum (the malaria pathogen), the ionic permeabil-
ity of the plasma membrane of infected erythrocytes
increases.
Borgatti et al. have demonstrated the application
of a PCB-based chip to generating DEP-based
cylinder-shaped cages for separation and recovery
of white blood cells from erythrocytes.
37
This work is
an important contribution to developing low-cost
LoC devices for diagnostic purposes. The researchers
showed that white blood cells recovered from their
LoC are suitable for polymerase-chain-reaction-based
molecular-diagnosis procedures using DNA sequenc-
ing, or biospecific-interaction analysis based on
surface plasmon resonance and biosensor technol-
ogy.
Marker-specific rare-cell sorting
Most of the available high-speed cell-sorting tech-
niques, especially for isolating rare cells, are limited by
parameters such as throughput, purity, and rare-cell
recovery. One approach for efficiently isolating
marker-specific rare cells from complex mixtures is
an electrokinetic sorting methodology exploiting DEP
in microfluidic channels.
38
This approach modulates
the dielectrophoretic amplitude response of rare target
cells by labeling cells with particles that differ in
polarization response.
A second approach is DEP-based routing of
identified rare cells to selected recovery fields of the
LoC platform. In this case, the cells might be identified
through fluorescence labeling using monoclonal
antibodies. Fuchs et al. provide a proof of principle
of this strategy.
39
They demonstrated the sorting and
recovering of specific live cells from samples contain-
ing less than a few thousand cells. An important
feature of this approach is that the cells maintain
viability and proliferation ability. The LoCmanipulated
cells using dynamic dielectrophoretic traps controlled
by an electronic interface.
Pharmaceutical LoC applications
Microfluidic technologies that improve the cost-
effective productionof establisheddrugs have provided
significant benefitssuchasimprovedsafetyandefficacy,
increased patient compliance, greater ease of use, and
expanded indications. Pharmaceutical scientists use
LoC platforms to control single biological objects,
including liposomes or microspheres immersed in
a liquid overhanging and in contact with the chip.
Two important classes of microparticles are micro-
spheresandliposomes. Thefirst classcomprisesparticles
characterized by a rigid structure made of biocompat-
ible materials. A common use of microspheres is the
activation of their surface with compounds that have
a well-defined biological activity when in contact with
cell membranes. Liposomes are lipidic shells that
contain chemical compounds, such as drugs. A useful
property of liposomes is that when properly prepared
they release their content into the cell cytoplasm.
Microparticles for LoC applications
We classify microparticles by size: Large micro-
particles are more than 100 microns in diameter,
medium microparticles are from 10 to 100 microns in
diameter, and small microparticles are less than 10
microns in diameter.
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IEEE Design & Test of Computers
32
Several types of microparticles are suitable for LoC
preparation, including experimental laboratory-made
microparticles specifically tailored to selected applica-
tions. Many types of special microparticles are
commercially available. They consist of materials such
as polystyrene, silica, melamine, glass, magnetite,
carboxymethylcellulose, biodegradable polymers, tri-
glycerides, partial glycerides, fatty acids, steroids, and
waxes. To produce microparticles, pharmaceutical
companies use experimental approaches such as
solvent extraction,
40
melt dispersion and solvent
evaporation,
41
ultrasound, high-shear and high-
pressure homogenization,
42
air jet milling,
43
coacerva-
tion,
44
salting out,
45
fluid bed coating (hot air
coating),
46
and spray drying.
47
In addition, many
companies supply fluorescent or dye-loaded micro-
particles. Most microparticles are supplied in deio-
nized water and have a shelf life of several years at 4uC.
Biodegradable microparticles, such as starch-based
microparticles, have a shorter shelf life.
Researchers have tailored some classes of micro-
particles, including lipospheres and cellulose acetate
microspheres, for specific LoC applications.
48,49
They
found that the dimensions of microparticles play a very
important role. Accordingly, the researchers tested
stabilizers and plasticizers to obtain useful micropar-
ticle characteristics.
Microparticle-cell interactions
The current literature contains a few examples of
LoC applications of microparticles. For example,
Borgatti et al. showed that an LoC can achieve
software-guided interactions between microspheres
and target cells.
37
The DEP-based device used parallel
electrodes to force interactions between microspheres
and K562 (human leukemia) cells, after moving them
to the central electrode in the corresponding DEP
cage.
Figure 6 shows an experiment demonstrating that
an LoC is suitable for directing single microspheres to
a single identified target cell.
50
The figure shows three
cationic microspheres (M1, M2, and M3) and two
K562 cells, entrapped in five independent spherical
DEP cages. Only one of the two cells was the three
Figure 6. Programmed sequential interactions (a-c) between three cationic microspheres (M1, M2, and M3) and one
target K562 cell. Moving M1 produced the K562-M1 complex (d). Moving M2 and M3 produced the K562-M1M2 (e)
and K562-M1M2M3 (f) complexes (Source: Borgatti et al.
50
Reprinted with permission).
JanuaryFebruary 2007
33
microspheres cellular target. The device first moved
microsphere M1, generating the cell-microsphere
complex shown in Figure 6d; then it moved micro-
sphere M2, generating the K562-M1M2 complex shown
in Figure 6e. Finally, it moved microsphere M3 for
a further targeting of the K562 cell, obtaining the K562-
M1M2M3 complex shown in Figure 6f. The buffer
employed was 280 milliMolar (mM) mannitol and
6.5 mM potassium chloride. The experimental condi-
tions were 50 kHz for the AC electric field used to
create dielectrophoretic patterns, and 37uC for the
supernatant. The passivation layer provides protection
against metal contamination and inhibits electrolysis.
THE BENEFITS OF CARRYING out experiments based on
single cells open new horizons in several key fields,
although the implications of this novel source of
information are still poorly understood. In general,
variability has always been a hallmark of living beings,
and scientists have tended to manage it with statistical
techniques. We hope that understanding the deviant
behavior of unique renegade cells will provide the
key to diagnosis and therapy for pathologies that still
lack adequate treatments. &
Acknowledgments
Our work is supported by FIRB-2001 (Italian
Fund for Basic Research, of the Italian Ministry of
Education), Development of a Lab on a Chip Based
on Microelectronic Technologies and Its Biotech-
nological Validation, to Roberto Guerrieri, Roberto
Gambari, and Claudio Nastruzzi. Our research is
also supported by the Italian Foundation for Cystic
Fibrosis, the Italian Association for Cancer Re-
search, and the Veneta Association for the Fight
against Thalassaemia. Funding from the European
Commission for the eInfrastructure for Thalassae-
mia Research Network is gratefully acknowledged.
&References
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Gianni Medoro is cofounder and
chief scientific officer of Silicon Bio-
systems, in Bologna, Italy. His re-
search interests include lab-on-a-chip
modeling and design. Medoro has
a PhD in electrical engineering and computer science
from the University of Bologna.
Roberto Guerrieri is a full pro-
fessor and teaches design of in-
tegrated systems at the University of
Bologna. His research interests in-
clude IC modeling and design, in-
cluding digital systems and biometric sensors, and
applications of microelectronics to biotechnology.
Guerrieri has a PhD in electrical engineering from
the University of Bologna.
Nicolo` Manaresi is cofounder and
chief technology officer of Silicon
Biosystems. His research interests
include analog design, fuzzy systems,
integrated sensors, and microelec-
tronic devices for analysis and manipulation of
biological particles. Manaresi has a PhD in electrical
engineering from the University of Bologna.
Claudio Nastruzzi is an associate
professor in the Department of Chem-
istry and Pharmaceutical Technology
at the University of Perugia, Italy. He
also heads the universitys Laboratory
of Biomaterials and Bioencapsulation. His research
interests include drug delivery microsystems, nerve
repair conduits, microparticle application to lab-on-a-
chip systems, lipospheres, and multifunctional micro-
capsules for cell entrapment. Nastruzzi has a PhD in
pharmaceutical sciences from the University of
Ferrara, Italy.
Roberto Gambari is a full professor
of biochemistry, chair of the PhD
program in biotechnology, and direc-
tor of the Biotechnology Center at the
University of Ferrara. His research
interests include pharmacogenomic and gene thera-
py of thalassaemia, molecular diagnosis, regulation
of gene expression, and transcription alteration using
decoy molecules and peptide nucleic acids. Gambari
has a PhD in biological sciences from the University
of Rome.
&Direct questions or comments about this article to
Roberto Guerrieri, ARCES University of Bologna, Viale
Pepoli 3-2, 40136 Bologna, Italy; roberto.guerrieri@
unibo.it.
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