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A Universal Biomolecular Concentrator To Enhance Biomolecular Surface Binding Based on Acoustic NEMS Resonator


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In designing bioassay systems for low-abundance biomolecule detection, most research focuses on improving transduction mechanisms while ignoring the intrinsically fundamental limitations in solution: mass transfer and binding affinity. We demonstrate enhanced biomolecular surface binding using an acoustic nano-electromechanical system (NEMS) resonator, as an on-chip biomolecular concentrator which breaks both mass transfer and binding affinity limitations. As a result, a concentration factor of 10⁵ has been obtained for various biomolecules. The resultantly enhanced surface binding between probes on the absorption surface and analytes in solution enables us to lower the limit of detection for representative proteins. We also integrated the biomolecular concentrator into an optoelectronic bioassay platform to demonstrate delivery of proteins from buffer/serum to the absorption surface. Since the manufacture of the resonator is CMOS-compatible, we expect it to be readily applied to further analysis of biomolecular interactions in molecular diagnostics.
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A Universal Biomolecular Concentrator To Enhance Biomolecular
Surface Binding Based on Acoustic NEMS Resonator
Wenpeng Liu,
Shuting Pan,
Hongxiang Zhang,
Zifan Tang,
Ji Liang,
Yanyan Wang,
Menglun Zhang,
Xiaodong Hu,
Wei Pang,*
and Xuexin Duan*
State Key Laboratory of Precision Measuring Technology & Instruments, Tianjin University, Tianjin 300072, China
College of Precision Instrument and Optoelectronics Engineering, Tianjin University, Tianjin 300072, China
SSupporting Information
ABSTRACT: In designing bioassay systems for low-abundance biomolecule
detection, most research focuses on improving transduction mechanisms
while ignoring the intrinsically fundamental limitations in solution: mass
transfer and binding anity. We demonstrate enhanced biomolecular surface
binding using an acoustic nano-electromechanical system (NEMS) resonator,
as an on-chip biomolecular concentrator which breaks both mass transfer and
binding anity limitations. As a result, a concentration factor of 105has been
obtained for various biomolecules. The resultantly enhanced surface binding
between probes on the absorption surface and analytes in solution enables us
to lower the limit of detection for representative proteins. We also integrated
the biomolecular concentrator into an optoelectronic bioassay platform to
demonstrate delivery of proteins from buer/serum to the absorption surface.
Since the manufacture of the resonator is CMOS-compatible, we expect it to
be readily applied to further analysis of biomolecular interactions in
molecular diagnostics.
Highly sensitive detection of biomolecular interactions at
ultralow concentration is crucial for continued progress in
applications ranging fromclinicaldiagnostics
to drug
and fundamental research such as intra-/extrac-
ellular tracking,
cell signaling,
neuronal impulse trans-
articial implant technology,
and gene regulatory
The recent trend focuses on exploring miniaturized
analytical systems for scarce biomolecule detections which has
the advantages of reduced reagent consumption, high
sensitivity, rapid detection, and multiplexed analysis, etc.
Many micro-/nanoscale biosensors with novel transduction
mechanisms have been developed to this end, including
plasmonic enzyme-linked immunosorbent assay (ELISA),
nanowire-based sensors,
and micro-/nano-electromechan-
ical sensors.
Since miniaturization of sensors often increases
their signal-to-noise ratio which is largely attributed to the high
demonstrated the capability of specic biomolecule detections
of only a few thousand (or even a few hundred) analyte
molecules in the sample volume.
Despite the impressive advances in signal transduction
technology, the mass transfer and binding anity limitations
are fundamentally hindering the improvement of the limit of
detection (LOD) of micro-/nanoscale biosensors.
surface-based biosensors require analytes in solution to react
with probes immobilized on a solid surface. Being heteroge-
neous, this process depends on numerous dierent parameters.
In solution, the rate depends on the convection and diusion
of the biomolecules (mass transfer limitation). At the interface,
the rate relies on the biomolecular interaction forces between
the analytes and the probes (anity limitation). To solve these
issues, researchers have developed techniques to enhance the
total ux of the solution or actively deliver the analyte
molecules toward the sensor surface. Examples include
electrokinetics-assisted binding that brings biomolecules
toward the absorption surface by electrostatic elds, aiming
to overcome the diusion and binding barriers.
theless, this technique is restricted to targets that are inherently
charged, and solutions having low ionic strength, which limits
its applicability to many practical assays. Other active methods
include magnetically-assisted
and optically-assisted
binding, which require extra labeling steps or complicated
setups which limit their throughput. Acoustic approaches have
emerged as useful tools to manipulate microscale objects with
many distinct advantages including simplicity, biocompati-
bility, and low power consumption.
However, due to the
competition between the acoustic radiation force and frictional
force induced by Stokeslaw, a critical radius exists below
which the acoustic radiation force becomes too small to
overwhelm the frictional or streaming forces in the medium,
resulting in inecient direct manipulation of biomole-
Hydrodynamic approaches use designed micro-
Received: May 11, 2018
Research Article
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uidic chips to generate microvortices and demonstrate the
direct trapping of biomolecules.
However, such methods
are limited by the use of closed microuidic chips which
cannot apply to other biosensing techniques. Consequently,
universal, noninvasive, and highly ecient methods to directly
manipulate and trap biomolecules are imperatively preferred to
enhance biomolecular surface binding and realize highly
sensitive bioassays.
In this work, we propose a novel molecular manipulation
system to trap biomolecules in open space and enhance their
surface binding using an on-chip designed acoustic nano-
electromechanical system (NEMS) resonator. The resonator
works as a biomolecular concentrator to precisely control the
hydrodynamic collection and accumulations of biomolecules
into a three-dimensional (3D) virtual micropocket, thus
fundamentally breaking both mass transfer and binding anity
limitations. This system is compatible to any biological
solutions, and the trapping of biomolecules in open space
allows the integration of the device with other biosensing
techniques, thus achieving a real universal biomolecular
concentrator and sensing system.
Theoretical Considerations and System Design. For
surface-based biosensors, the two-compartment model has
been widely used to describe the biomolecular surface binding
process, such as the convection, diusion, and reactions.
shown in Figure 1B, the model handles the variation in analyte
concentration by dividing the solution into two compartments:
concentrations of analytes in bulk solution [A]0and at
absorption surface [A]s.Adiusion layer where the ow
velocity approaches zero exists between these two compart-
ments. Theoretically, the two-compartment reaction can be
described with eq 1:
][]+[]=== [
on off (1)
where kmis the diusion or convection controlled rate
constant, kon and koff are the association and dissociation rate
constants for the biomolecular interactions, respectively, and
[B] and [AB] represent the density of probes and absorbed
analytes at the absorption surface, respectively. Because of zero
ow in the diusion layer, transfer of analytes from the bulk
solution to the absorption surface over the diusion layer
occurs primarily by diusion which normally takes a long
period of time. Such a mass transfer limitation results in an
unmatched analyte concentration between bulk solution and
absorption surface ([A]s[A]0). Active stirring is generally
recommended to increase [A]sby accelerating the mass
yet the ability is inherently limited given that [A]s
cannot exceed [A]0, leading to a corresponding binding anity
To break these two limitations, our system is designed by
the integration of an acoustic NEMS resonator in an open
liquid system to generate special hydrodynamic conditions
which can trap and accumulate the biomolecules in a predicted
virtual micropocket. The resonator is fabricated through a
standard CMOS process (Supporting Information, Figure S1).
Figure 1C,D shows the top-view scanning electron microscope
(SEM) image and schematic cross-section structure of the
resonator. It is composed of a freestanding aluminum nitride
(AlN) piezoelectric nanoplate (450 nm) sandwiched by
periodic molybdenum (Mo) electrodes as interdigital trans-
ducer (IDT) which is isolated from the silicon substrate with
an air cavity to avoid dissipation of energy into the silicon
substrate. After a power is applied to the electrodes, acoustic
waves are generated in the piezoelectric nanoplate via the
converse piezoelectric eect.
When the resonator works in
liquid, the acoustic streaming eect is generated because of the
dissipation of acoustic energy into liquid, which is
Figure 1. Enhanced biomolecular interactions using an acoustic NEMS resonator as biomolecular concentrator. (A) Diagram depicting the
conguration of the experimental setup. The resonator is fabricated on silicon substrate and mounted by a polydimethylsiloxane (PDMS) chamber.
Sinusoidal signals at the resonant frequency are generated by a vector network analyzer (VNA) and sent to the device. (B) Two-compartment
model for describing analyte transfer from the bulk solution to the absorption surface, as well as the reaction with the surface-immobilized probes.
(C) Top-view SEM image of the acoustic NEMS resonator. Scale bar, 50 μm. (D) Schematic illustrating the cross-section view of the acoustic
NEMS resonator structure. (E) Top-view schematic diagram illustrating biomolecule trapping and accumulation using the acoustic NEMS
resonator in the open space. The black solid curves represent the ow prole.
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experimentally veried by the decrease of quality factor (Q)
from 1175 to 56 (Supporting Information, Figure S2).
Consequently, four symmetric counter-owing Rankine micro-
vortices (SCRMVs) are formed close to the resonators central
region, and a virtual micropocket is further generated, where
biomolecules are massively trapped and concentrated; hence,
the concentration of the target biomolecules at the surface is
larger than the bulk concentration ([A]s> [A]0), and the
molecule diusion is enhanced by the hydrodynamic ow
(Figure 1E). Since the trapping of molecules is realized in the
open space, and the position of the micropocket can be well
predicted, this technique can be easily combined with other
biosensors by locating the transducer at or close to the virtual
micropocket to achieve a real universal biomolecular
Flow Prole. A numerical simulation is performed to
understand the resonant behaviors of the acoustic NEMS
resonator and the hydrodynamic behaviors of the ow using
the 3D nite element method (FEM). As illustrated in Figure
2A, after a power of 0.01 mW is applied on the resonator in air,
acoustic waves conned in the piezoelectric nanoplate
propagate along the transverse direction (yaxis) with the
maximum vibration amplitude of 0.09 nm. When resonating in
liquid, the resonator leads to the generation of SCRMVs, each
of which is composed of a forced microvortex core surrounded
by a free vortex zone,
as shown in the normalized simulation
results of the 2D ow prole in the xyplane that is 50 μm
above the resonator surface (z=50μm) in Figure 2Bi. The
forced microvortex core is rotational, and its ow velocity
scales with the radius and decays to zero at the microvortex
center, while the free microvortex zone is irrotational, and its
ow velocity varies inversely with the radius to satisfy the
boundary condition of no motion at innity. The formation of
the four SCRMVs results from the acoustic streaming which is
generated by spatial attenuation of acoustic waves. In brief,
acoustic waves attenuate and create a pressure gradient during
the propagation along the yaxis, resulting in the formation of
two uid jets at both sides of the resonator. Because of the ow
continuity, recirculated ows are correspondingly generated
along the xaxis. Figure 2Bii shows the zoom-in 2D ow prole
Figure 2. Numerical and experimental results of resonant behaviors and induced ow prole. (A) 3D FEM simulations of the resonators vibration
amplitude in air. The power is 0.01 mW. (B) Simulation results of the 2D ow prole in the xyplane (z=50μm) showing (i) the four SCRMVs
and (ii) the stagnation point. Note that the resonator is located at the origin of coordinates, and the velocity is normalized for simplicity. (C)
Virtual micropocket extracted from the ow velocity distribution inside the rectangle region in part Bii. (D) Illustration of the azimuthal
recirculation of the SCRMVs. Simulation results of 2D ow prole (i) in the xzplane (y=0μm) and (ii) in the yzplane (x=0μm). (iii)
Cartoon showing 3D azimuthal recirculation of the SCRMVs at the angle of θ. (E) Optical image showing the four experimental SCRMVs (in red)
in the xyplane. Scale bar, 150 μm. (F) Experimental (i) ow prole and (ii) velocity prole in the xyplane showing the stagnation point as a
virtual micropocket. Scale bar, 50 μm.
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which consists of inlets along the xaxis and outlets along the y
axis at the center of the resonator. As a consequence of the
superposition of component velocities of the four SCRMVs, a
stagnation point is created at the junction of the four SCRMVs
where the ow velocity approaches to zero. Ultimately, the
primary result of this ow velocity distribution inside the
rectangle region in Figure 2Bii is the generation of a virtual
micropocket in the open space, as shown in Figure 2C.
It is worth mentioning that an azimuthal angle exists for
SCRMVs. As revealed in Figure 2Di,ii, the uid ows toward
the bottom of the virtual micropocket along the xaxis and
departs along the yaxis at an angle downward. This azimuthal
angle originates from the fact that the mechanical vibration of
the resonator simultaneously induces the propagations of
attenuated acoustic waves along the longitudinal direction.
Namely, the velocity of the uid jets is a superposition of
component velocities along both the transverse and longi-
tudinal direction, therefore leading to the 3D azimuthal
recirculation of the SCRMVs at the angle of θ, as shown in
Figure 2Diii.
We employed micro-PIV to experimentally visualize the
PS particles of 5 μm were used to track the motion
of the microvortices by the actuation of the resonator
(Supporting Information, Movie S1). Figure 2E shows the
typical captured uorescence image of PS particle motion
which clearly shows four SCRMVs at a power of 4 mW
(Supporting Information, Movie S2). Figure 2F shows the 2D
ow prole and velocity of the SCRMVs at the xyplane
analyzed by Diatrack 3.04 (Supporting Information, Movie
S3). It is characterized by an obvious stagnation point which is
surrounded by a high-velocity region with a maximum ow
velocity at 220 μm/s.
It is also noted that, from Figure 2E, the uorescence
intensity of the vortex zone is remarkably enhanced compared
to the background, indicating that the particles were dragged
into the vortices and trapped inside. We further compared the
size and the trapping eciency of the vortex by applying
dierent powers. The location and the size of the microvortices
stay constant at dierent applied powers (Supporting
Information, Figures S4 and S5). However, the number of
the particles being trapped in the vortex increased by using
higher power, as indicated by the uorescence intensity results
in Figure S4. This is due to the fact that the higher power will
induce a stronger vortex and will generate larger drag forces
which will in turn trap more particles in the vortex. This is also
veried by the simulation. The amplitude of the resonator
vibration at the resonant frequency increases with higher
power. As a consequence, larger resonant amplitude will
induce more vigorous microvortices via the acoustic streaming
eect; thus, more particles are brought into the vortices zone.
Since the volume and the location of the vortex zone can be
predesigned with the device dimension, it will benet the
application to use such trapping for dierent applications. In
addition, the power can be readily adjusted to tune the amount
of the trapped particles or molecules, which will be described
in the following section.
Biomolecular Concentrator. As reported by hydro-
dynamic trapping in microuidics, biomolecules in vortex
Figure 3. Analysis of biomolecular concentration using the acoustic NEMS resonator. (A) Time-lapse uorescence images of concentrated FITC-
SAV. Scale bar, 50 μm. (B) 3D prole of concentrated FITC-SAV inside the virtual micropocket by DIPHM measurement. Scale bar, 20 μm. (C,
D) Growth of the area and the average height of concentrated FITC-SAV biomolecules. (E) Cartoon showing the concentration of biomolecules
inside the 3D ow eld structure.
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ow prefer to migrate from regions of high uid velocity
downward to regions where the uid velocity becomes
hence, they will depart from the streamlines
of the vortex and are further collected into the virtual
micropocket. In our case, the molecular trapping is achieved in
an open space which is above the resonator surface. For a
demonstration of the molecular trapping and concentration
eect by the acoustic NEMS resonator, uorescent isothio-
cyanate-labeled streptavidin (FITC-SAV) is used as the model
protein. FITC-SAVs are rst dissolved in 100 μL of PBS buer
(200 nM) which covers the resonator in an open chamber.
Because of the low protein concentration, no uorescence can
be observed at this condition (Figure 3A, 0 min). Under the
actuation of the resonator, FITC-SAVs are gradually trapped
into the virtual micropocket, which can be clearly observed
from the uorescence images (Figure 3A). After 5 min, the
uorescence intensity (Int) is remarkably enhanced from
0.0004 to 4.0744. These results suggest that a trapping force
has been successfully applied on biomolecules toward the
center of SCRMVs. It is worth mentioning that the
concentration eect occurs only within several seconds, and
the trapping area extends until saturation as long as the
resonator actuation persists (Supporting Information, Movie
For further analysis of the 3D spatial distributions of the
trapped FITC-SAVs, reection digital image-plane holographic
microscopy (DIPHM) is introduced to record the time-lapse
of the growth of the virtual micropocket (Supporting
Information, Figure S3). Static features in the images are
eliminated by calculating the phase shift between the real-time
images and the initial image; thus, only the gradually
concentrated FITC-SAVs are recorded, as shown in Figure
3B. Figure 3C,D quanties the area and the average height of
the concentration region, respectively. The active area reaches
up to 2700 μm2with the actuation of the resonator for 15 min,
approaching saturation. A similar trend is revealed in terms of
the average height which shows an average saturated value
around 80 nm with the maximum height as high as 260 nm.
This height restriction is resulted by the azimuthal
recirculation of the SCRMVs at an angle of θas shown in
Figure 3E. Namely, biomolecules move toward the resonator
from inlets at an angle downward and tend to be collected at
the bottom of the virtual micropocket. It should be noted that
the position and the size of the virtual micropocket are rather
repeatable by using the same power. Thus, this method can be
directly applied to enhance protein detections by locating the
probe-functionalized sensor substrate at this region which can
be signicantly benecial from the markedly enhanced analyte
concentration during biomolecular interaction assays where
the response time and surface-absorbed proteins are inherently
limited by diusion and anity, especially for measurement of
biomolecules at ultralow concentrations.
It is also noted that
since the acoustic trapping is noninvasive, the enhanced
molecular assay can be applied to any surface-based biosensors.
Enhanced Immunoassay. The clearly demonstrated
protein concentration eect will markedly lower the LOD of
the specic protein detection, such as in an immunoassay. To
prove this, we performed the detection of the human
immunoglobulin G (IgG) through specic antibodyantigen
interactions using a resonator-enhanced immunoassay. Figure
4Adepictstheconguration of the detection system.
Antihuman IgGs were immobilized on the resonator surface
through a PLLPEGbiotinSAV linker.
The main reason
that we employed PLLPEGbiotinSAV for anti-IgG
Figure 4. Enhanced immunoassays of human IgG using the acoustic NEMS resonator as biomolecular concentrator. (A) Cartoon showing the
process of functionalization on the resonator surface. (B) Time-lapse uorescence images of Cy3-labeled human IgG (i) with and (ii) without the
actuation of the resonator in solution. Scale bar, 25 μm. (C) Extraction of time-lapse uorescence intensity from part B. The data are calculated
from the region of highest intensity of uorescence signals (with an area of 5 μm×30 μm at the center of the resonator). (D) Fluorescence
intensity after buer rinsing and drying. Scale bar, 10 μm. The inset shows the uorescence images of each sample. (E) Time-lapse uorescence
intensity under the actuation of the resonator with the power changing from 0 to 4 mW. (F) Time-lapse uorescence intensity under the actuation
of the resonator with dierent Qvalue.
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immobilization instead of direct conjugation of anti-IgG is to
test the concentration eect on the same device by
regenerating the sensor surface. Since the assemblies of the
PLLPEGbiotinSAV on the resonator are driven by
electrostatic interactions between the positively charged
polymer and negatively charged surface, such assemblies can
be regenerated easily by tuning the pH value of the buer.
Thus, the device can be reused many times which facilitated
the comparison of the concentration eect. In addition, the
PEG chains were grafted in the polymer to prevent nonspecic
protein bindings.
Dierent concentrations of Cy3-labeled human IgGs were
then introduced into the solution chamber. After reaction with
the immobilized anti-IgGs, the uorescence intensity can be
used to quantify the amount of the surface-absorbed proteins.
The concentration eect is compared with and without the
resonator actuation. As the trapped proteins were saturated
after 15 min of actuation of the resonator in the DIPHM
measurement, we kept all the actuation experiments no longer
than 15 min. Figure 4B,C shows the time-lapse uorescence
images and corresponding uorescence intensities by exposing
various concentrations of the Cy3-labeled human IgG to
antihuman IgG coated resonator. This clearly shows the
enhancement of the uorescence intensity with the resonator
actuation. The uorescence intensity at 50 pM with the
resonator-induced concentration for 15 min is comparable to
that at 5 μM without the resonator operation, indicating that
the concentration factor Areaches up to 105. As calculated
from the uorescence intensity of 50 pM IgG solutions
(Supporting Information), the total number of IgG molecules
in solution is 5.88 ×108, and the total number of concentrated
molecules is 4.50 ×108; hence, the concentration eciency for
50 pM IgG solution ηis 4.50 ×108/5.88 ×108= 76%.
Similarly, the total number of IgG molecules in 500 pM IgG
solution is 5.88 ×109, and the total number of concentrated
molecules is 1.33 ×109; hence, the concentration eciency for
500 pM IgG solution ηis 1.33 ×109/5.88 ×109= 23%. It is
worth noting that the concentration factor and the total
number of the molecules to saturate the trap may vary case-by-
case considering the practical dependence on a series of
conditions such as protein interactions, the concentration of
the analytes, the viscosity of the solution, and the intrinsic
properties of the protein molecules.
It is also worth mentioning that the uorescence intensities
from Figure 4C are derived from the analytes both absorbed at
the resonator surface and collected in 3D space. To specically
quantify the proteins absorbed at the resonator surface, we
removed the protein solutions after 15 min of incubation and
actuation by the resonator. The devices were then thoroughly
rinsed with buer to remove the physical absorbed proteins.
After drying, the uorescence intensity of each concentration
was recorded (Figure 4D). The results again clearly
demonstrate the signicantly enhanced amount of the
surface-absorbed proteins by the resonator actuation. This
indicates that the resonator plays a critical role in the
enhancement of biorecognition events via the controlled
hydrodynamic trapping. Namely, analytes are eciently
trapped, concentrated, and specically bound to surface-
Figure 5. Biomolecular detection using optoelectronic bioassay platform. (A) Measurement for SAV in buer: (i) thermodynamic measurement for
SAV with and without the resonator-induced concentration eect (the inset shows the zoom-in view at ultralow concentration), and (ii) binding
enhancement factor (Be)atdierent SAV concentrations (the inset illustrates the schematic diagram of optoelectronic bioassay platform which
shows the resonator actuation in the open space and BLI sensing for biomolecules). (B) Thermodynamic measurement for PSA in buer with and
without the resonator-induced concentration eect. The inset shows the binding enhancement factor (Be)atdierent PSA concentrations. (C)
Thermodynamic measurement for IgG in serum with and without the resonator-induced concentration eect.
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immobilized probes under the actuation of the resonator by
breaking the mass transfer and binding anity limitations.
As predicted in theory analysis and simulations, the
hydrodynamic manipulations are related with the power
applied to the device and the Qof the resonator. The
power-dependent and Q-dependent characteristics of the
resonator-induced concentration eect are characterized in
detail by using the same antibodyantigen interactions. Figure
4E shows the results of power-dependent concentration eects
(according to uorescence intensity) with the power changing
from 0 to 4 mW. No uorescence is observed in the control
experiment (0 mW) after 15 min of incubation, indicating that
the analyte concentration is below the LOD of the
uorescence microscope. With the same incubation time and
power increasing from 0.25 to 4 mW, the uorescence
intensity enhances signicantly. The simulation analysis
explicitly indicates that the amplitude of resonator vibration
increases to 0.68 nm with the power growing up to 4 mW
(Supporting Information, Figure S4). As a consequence, higher
resonant amplitude will induce more vigorous microvortices,
whereby more target molecules are brought into the vortices
and nally trapped at the virtual micropocket. The power-
dependent characteristics of the resonator-induced concen-
tration eect render the resonator as a regulator to control the
biomolecular interaction rate via adjusting the power, which
plays a key role in enzymology and other biological research.
Except for the power, the resonator with higher Qvalue is
characterized by larger vibration amplitude and has a stronger
streaming eect in solution,
which will lead to a more
ecient molecular concentration eect. As shown in Figure 4F,
after incubation and actuation of the resonator for 15 min, the
resulting uorescence intensity for the resonator with Q=41is
3.7 times larger than that for the resonator with Q= 33.
Optoelectronic Bioassay System. A key advantage of the
resonator-induced enhancement is that it can concentrate
biomolecules at a 3D collection zone in an open space, which
can directly benet any type of surface-based biosensor by
locating the transducer at the virtual micropocket. In such a
case, the resonator is used as an active uid delivery and
molecular manipulation component. Such a combination will
achieve a real universal biomolecular concentrator and sensing
To explore this assumption, we developed an optoelectronic
bioassay system by integrating resonator actuation into a
biolayer interferometry (BLI) biosensor for protein binding
analysis (Supporting Information, Figure S5).
The BLI is a
label-free technique for measuring biomolecular interactions
using a ber-optic probe approach. Any changes in the number
of molecules bound to the probe surface would induce a
wavelength shift in the interference pattern between the
incident and reected light. Thus, it can provide real-time
measurement for biomolecular surface absorptions. Our
optoelectronic bioassay platform uses a resonator as an
actuator to provide analyte accumulations at the BLI optical
probe interface, thus enhancing the amount of the surface-
absorbed molecules.
The BLI optical probe was functionalized with PLLPEG
biotin; then, the resonator was integrated into the BLI system
by locating the device directly below the optical probe.
Dierent concentrations of SAV were applied into the system.
Figure 5Ai shows the results of SAV bindings in HEPES buer
with and without resonator actuations. After a power of 4 mW
is applied, the LOD of SAV detection extends to 50 fM, which
is 1000-fold lower than the results without resonator
enhancement (50 pM). The improvement of LOD proves
the overcoming of the surface limitation due to the
accumulation of SAV molecules around the optical probe
surface. Binding enhancement factor, Be, which is dened as
the ratio of response with resonator to response without
resonator, is calculated as well. As shown in the inset of Figure
5Aii, Beis as high as 145 at the concentration of 50 pM while it
gradually decreases to 1.6 at the concentration of 5 nM. The
decrease of Bewith the increasing SAV concentration results
from the depletion of limited binding sites (biotin) at the
absorption surface which indicates the same saturation
response for the two measurements process. In other words,
the binding enhancement under the actuation of the resonator
is more prominent for biomolecular interactions at extremely
diluted conditions. We believe this method would be useful
not only in clinic diagnosis but also in the anity measurement
of biomolecular interactions such as for drug screening where
the protein binding is usually applied in buer conditions.
The resonator was also used to concentrate PSA molecules
during the PSA measurement. The BLI optical probe was
functionalized with anti-PSA through a similar approach as
shown in Figure 4A (Supporting Information, Figure S6).
Figure 5B shows the results of PSA bindings in HEPES buer
with and without resonator actuations. After a power of 4 mW
is applied, the LOD of PSA detection extends to 50 pM, which
is 200-fold lower than the results without resonator enhance-
ment (10 nM). As shown in the inset of Figure 5B, Beis 93 at
the concentration of 10 nM while it decreases to 8 at the
concentration of 50 nM. The results are consistent with the
response for SAV measurement.
To further demonstrate the truly meaningful sensing
enhancement of this optoelectronic bioassay system, we also
conducted the protein detection in serum where background
signals from nonspecic binding are considered to be a non-
negligible factor to the sensing results. Alternatively, the PSA
binding pairs are replaced by IgG binding pairs to prove the
universality of the optoelectronic bioassay system for a diverse
range of biomolecules. Figure 5C shows the measurement
results of IgG in serum with and without resonator actuations.
As revealed, the LOD of IgG measurement extends to 2 nM,
which is 10-fold lower than the results without the resonator-
induced concentration eect (20 nM), and Bereaches 20.6 at
the concentration of 20 nM while it decreases to 9.6 at the
concentration of 200 nM. The optoelectronic bioassay does
not show very high sensitivity in serum since the hydro-
dynamic trapping is not selective, and other proteins or
molecules will be concentrated by the acoustic devices as well.
Thus, the nonspecic bindings will be strongly inuenced their
performance in serum. Further studies are denitely required
to improve the binding enhancement in serum or other
complicated conditions where clinic diagnoses are usually run.
One solution could be using a preltration chip, where specic
probe-functionalized micropillars or nanoparticles could be
used to purify the serum samples.
The successfully demonstrated optoelectronic bioassays
prove the practical feasibility of the integration of the device
with other biosensing techniques, which is attributed to their
merit of open-space trapping. Meanwhile, the results also
indicate that the hydrodynamic trapping of biomolecules using
the NEMS resonator is a noninvasive approach without
denaturing their bioactivities, which is rather important to
develop an enhanced biosensing platform.
ACS Central Science Research Article
DOI: 10.1021/acscentsci.8b00301
Overall, we have demonstrated a universal approach to
enhance biomolecular surface binding for biosensing applica-
tions in the open space with an acoustic NEMS resonator,
which is featured by the concentration factor of 105. This
approach bridges a major gap between signal transduction
technology and the uidic system in the investigation of
biomolecular interactions. It oers several competitive
advantages: First, it is a noninvasive and biocompatible
approach which works for a diverse range of biomolecules,
regardless of their physical and chemical properties. Second,
the concentration process is highly ecient, requiring only a
few minutes, which is benecial for rapid biomarker detections.
Third, the trapping of biomolecules in open space without
microuidic channels allows the combination of the device
with many surface-based biosensing techniques (e.g., surface
plasma resonance, quartz crystal microbalance, electrochemis-
try, and ELISA, etc.) to achieve a real universal biomolecular
concentrator and sensing system. Fourthly, the concentration
process is achieved using a simple, miniaturized, low-cost,
CMOS-compatible device; thus, it can be readily applied to an
established system for biomolecule analysis. Given the above
advantages, our approach is valuable in the eld of biomedical
engineering such as molecular diagnostics and drug discoveries.
Synthesis of PLLPEGBiotin. PLL was dissolved in 50
mM sodium carbonate buer (pH = 8.5) at a concentration of
40 mg/mL. The solution was then ltered through a 220 nm
pore syringe lter. NHS-PEGbiotin was added to the
dissolved PLL solution under vigorous stirring. The reaction
was allowed to proceed for 5 h under room temperature,
followed by the dialysis against PBS at pH = 7.4 and deionized
water for 24 h using a centrifugal lter device (molecular
weight cuto8 kDa). The dialyzed solution was lyophilized for
24 h and stored in a 25 °C freezer.
Device Fabrication. The acoustic NEMS resonator was
fabricated using a CMOS-compatible process (Supporting
Information, Figure S1). It was started by etching an air cavity
on silicon substrate by reactive ion etching, followed by
deposition of phosphosilicate glass (PSG) using chemical
vapor deposition (CVD). After that, the surface was planarized
using chemical mechanical polish (CMP). Then, 200 nm Mo
lm was deposited and patterned as the bottom electrode.
After that, T= 450 nm AlN lm was employed as piezoelectric
layer by RF reactive magnetron sputtering. Next, 200 nm Mo
lm was deposited and patterned as the top electrode. Then,
AlN was etched by a combination of Cl2-based plasma etching
and potassium hydroxide wet etching. After the AlN etch, Au
was then evaporated and patterned by lift-o, serving as
electrical connection and pads. Finally, the silicon wafer was
immersed in diluted hydrouoric acid solution to release PSG
in the cavity. Here, the top electrode was patterned with the
same shape of the bottom electrode to form IDT. As a
consequence, a 350 MHz resonator, with an aperture p=15
μm, width w=10μm, length l= 150 μm, and number n=12
IDT ngers was fabricated. The resonance characteristic is
presented by Smith chart in air and liquid (Supporting
Information, Figure S2). When working in solution, mechan-
ical resonance generated from the resonator is partially coupled
into liquid that can be seen by the shrink of Smith chart which
is an indicator of energy losses in the system.
Flow Prole Quantication. We tracked the movement
of 5 μm PS particles from which the ow prole and velocity
are quantitatively analyzed. With the assumption that PS
particles are small enough to have no signicant eects on the
ow prole, they are simply dragged along with behaviors
similar to the ow prole. PS particles were tracked using a
video camera (Olympus DP73, Tokyo, Japan) attached to an
optical microscope (Olympus BX53, Tokyo, Japan), and their
movements were analyzed with commercially available
software, Diatrack 3.04.
Holographic Microscopy. Reection DIPHM was utilized
to obtain the 3D surface prole of analytes during the
concentration (Supporting Information, Figure S3). The
illumination source was a tunable diode laser at λ= 690 nm
(Nanobase, Xperay-TL-STD, 639697 nm) which was split
into the object beam and the reference beam. The expanded
object beam illuminated and reected from the resonator
surface to create the object wavefront, which interfered with
the reference wavefront and formed the surface prole in 3D
perspective images. Here, the resonator was used to
concentrate 10 μg/mL SAV in 10 mM HEPES buer with a
power of 1 mW. The 3D prole was extracted from the phase
shift between real-time images and initial images, which gave
quantiable information about the optical thickness of
concentrated analytes.
Protein Concentration. The resonator was exposed to air
plasma for 5 min to form a clean and negatively charged
surface, and then immersed in the aqueous solution of PLL
PEGbiotin (1 mg/mL) at room temperature for 30 min,
followed by rinsing with HEPES buer (10 mM). SAV (200
nM) was then attached onto PLLPEGbiotin linker via
biotinSAV binding for 30 min. Afterward, 100 μg/mL biotin-
labeled antihuman IgGs were immobilized. After surface
functionalization, the chip is wire-bound to a chip holder,
and a PDMS channel is mounted on top of the device to
facilitate the protein sensing experiments. All the analytes used
in the experiments were dissolved in HEPES (pH = 7.4) buer.
Fluorescence intensity was recorded by uorescence micros-
copy (Olympus BX53).
Measurement of PSA/IgG Using Optoelectronic Bio-
assay Platform. The resonator was integrated into an
optoelectronic bioassay platform (Supporting Information,
Figure S5). The ber-optic probe was rst immerged in
piranha solution for cleaning and generation of the negatively
charged hydroxyl group. Then, 1 mg/mL PLLPEGbiotin,
200 nM SAV, and 200 μg/mL biotin-labeled anti-PSA/anti-
IgG were immobilized sequentially on the probe surface by
immersion in their solution for 15 min (Supporting
Information, Figure S6). After that, the ber-optic probe was
precisely positioned around the stagnation point by a
positioning stage, and a continuous set of PSA/IgG samples
dissolved in buer/serum were introduced.
Caution. Piranha solution reacts violently with organic
solvents and should be handled with great care. For more
information, please see
SSupporting Information
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acscents-
ACS Central Science Research Article
DOI: 10.1021/acscentsci.8b00301
Additional theoretical analysis, gures, materials, fab-
rication procedures, characterization data, and exper-
imental results (PDF)
Movie S1: movement of 1 μm PS particles under the
actuation of the acoustic NEMS resonator with a power
of 4 mW (AVI)
Movie S2: movement of uorescence 5 μm PS particles
under the actuation of the acoustic NEMS resonator
with a power of 4 mW (AVI)
Movie S3: movement of 5 μm PS particles under the
actuation of the acoustic NEMS resonator with a power
of 0.01 mW (AVI)
Movie S4: concentration of FITC-SAV molecules under
the actuation of the acoustic NEMS resonator with a
power of 4 mW (AVI)
Corresponding Authors
Menglun Zhang: 0000-0002-5174-7812
Xuexin Duan: 0000-0002-7550-3951
Author Contributions
W.L. and S.P. contributed equally. W.L. and X.D. conceived
and designed the experiments. W.L., S.P., and Z.T. performed
the experiments and data analysis. H.Z. and J.L. fabricated
LWR devices; H.Z. performed the simulations. W.L. wrote the
manuscript. X.D., Y.W., M.Z., X.H. and W.P. revised the
manuscript. All the authors contributed to the scientic
The authors declare no competing nancial interest.
This work was supported by the National Natural Science
Foundation of China (NSFC 61674114, 91743110,
21861132001), National Key R&D Program of China
(2017YFF0204600), and the 111 Project (B07014). We are
grateful to our colleague Yang Yang for help on the
characterization of the ow prole. We are grateful to our
colleague Ziyu Han for help on the surface modications of the
sensors. We are grateful to our colleague Xinyu Chang for help
on the holographic measurement.
(1) Mahshid, S. S.; Camiré
, S.; Ricci, F.; Vallé
lisle, A. A highly
selective electrochemical DNA-based sensor that employs steric
hindrance effects to detect proteins directly in whole blood. J. Am.
Chem. Soc. 2015,137, 1559615599.
(2) Kosaka, P. M.; Pini, V.; Ruz, J. J.; da Silva, R.; Gonzá
lez, M.;
Ramos, D.; Calleja, M.; Tamayo, J. Detection of cancer biomarkers in
serum using a hybrid mechanical and optoplasmonic nanosensor. Nat.
Nanotechnol. 2014,9, 10471053.
(3) Meister, S.; Plouffe, D. M.; Kuhen, K. L.; Bonamy, G. M.; Wu,
T.; Barnes, S. W.; Bopp, S. E.; Borboa, R.; Bright, A. T.; Che, J.
Imaging of plasmodium liver stages to drive next-generation
antimalarial drug discovery. Science 2011,334, 13721377.
(4) Mellman, I.; Warren, G. The road taken: past and future
foundations of membrane traffic. Cell 2000,100,99112.
Lauffenburger, D. A.; Yaffe, M. B. A systems model of signaling
identifies a molecular basis set for cytokine-induced apoptosis. Science
2005,310, 16461653.
(6) Neiditch, M. B.; Federle, M. J.; Pompeani, A. J.; Kelly, R. C.;
Swem, D. L.; Jeffrey, P. D.; Bassler, B. L.; Hughson, F. M. Ligand-
induced asymmetry in histidine sensor kinase complex regulates
quorum sensing. Cell 2006,126, 10951108.
(7) Pugh, J. R.; Raman, I. M. GABAAreceptor kinetics in the
cerebellar nuclei: evidence for detection of transmitter from distant
release sites. Biophys. J. 2005,88, 17401754.
(8) Abraham, S.; Brahim, S.; Ishihara, K.; Guiseppi-Elie, A.
Molecularly engineered p(HEMA)-based hydrogels for implant
biochip biocompatibility. Biomaterials 2005,26, 47674778.
(9) Davidson, E. H.; Erwin, D. H. Gene regulatory networks and the
evolution of animal body plans. Science 2006,311, 796800.
(10) Sameenoi, Y.; Koehler, K.; Shapiro, J.; Boonsong, K.; Sun, Y.;
Collett, J., Jr; Volckens, J.; Henry, C. S. Microfluidic electrochemical
sensor for on-line monitoring of aerosol oxidative activity. J. Am.
Chem. Soc. 2012,134, 1056210568.
(11) Lee, J. H.; Daugharthy, E. R.; Scheiman, J.; Kalhor, R.; Yang, J.
L.; Ferrante, T. C.; Terry, R.; Jeanty, S. S.; Li, C.; Amamoto, R. Highly
multiplexed subcellular RNA sequencing in situ. Science 2014,343,
(12) Stern, E.; Vacic, A.; Rajan, N. K.; Criscione, J. M.; Park, J.; Ilic,
B. R.; Mooney, D. J.; Reed, M. A.; Fahmy, T. M. Label-free biomarker
detection from whole blood. Nat. Nanotechnol. 2010,5, 138142.
(13) Liu, W.; Zhang, H.; Zhao, H.; Tang, Z.; Wang, Y.; Sun, C.;
Pang, W.; Duan, X. Comparative analysis of static and non-static
assays for biochemical sensing using on-chip integrated field effect
transistors and solidly mounted resonators. Sens. Actuators, B 2017,
243, 775783.
(14) De La Rica, R.; Stevens, M. M. Plasmonic ELISA for the
ultrasensitive detection of disease biomarkers with the naked eye. Nat.
Nanotechnol. 2012,7, 821824.
(15) Tian, B.; Cohen-Karni, T.; Qing, Q.; Duan, X.; Xie, P.; Lieber,
C. M. Three-dimensional, flexible nanoscale field-effect transistors as
localized bioprobes. Science 2010,329, 830834.
(16) Stern, E.; Klemic, J. F.; Routenberg, D. A.; Wyrembak, P. N.;
Turner-Evans, D. B.; Hamilton, A. D.; LaVan, D. A.; Fahmy, T. M.;
Reed, M. A. Label-free immunodetection with CMOS-compatible
semiconducting nanowires. Nature 2007,445, 519522.
(17) Zheng, G.; Patolsky, F.; Cui, Y.; Wang, W. U.; Lieber, C. M.
Multiplexed electrical detection of cancer markers with nanowire
sensor arrays. Nat. Biotechnol. 2005,23, 12941301.
(18) Li, M.; Tang, H. X.; Roukes, M. L. Ultra-sensitive NEMS-based
cantilevers for sensing, scanned probe and very high-frequency
applications. Nat. Nanotechnol. 2007,2, 114120.
(19) Kim, J.; Junkin, M.; Kim, D.-H.; Kwon, S.; Shin, Y. S.; Wong, P.
K.; Gale, B. K. Applications, techniques, and microfluidic interfacing
for nanoscale biosensing. Microfluid. Nanofluid. 2009,7, 149167.
(20) Kim, D. R.; Zheng, X. Numerical characterization and
optimization of the microfluidics for nanowire biosensors. Nano
Lett. 2008,8, 32333237.
(21) Liao, K.-T.; Chou, C.-F. Nanoscale molecular traps and dams
for ultrafast protein enrichment in high-conductivity buffers. J. Am.
Chem. Soc. 2012,134, 87428745.
(22) Eijkel, J. C.; van den Berg, A. Nanofluidics: tiny electrostatic
traps. Nature 2010,467, 666667.
(23) Lien, K.-Y.; Lee, W.-C.; Lei, H.-Y.; Lee, G.-B. Integrated reverse
transcription polymerase chain reaction systems for virus detection.
Biosens. Bioelectron. 2007,22, 17391748.
(24) Lee, S.; Hohng, S. An optical trap combined with three-color
FRET. J. Am. Chem. Soc. 2013,135, 1826018263.
(25) Grier, D. G. A revolution in optical manipulation. Nature 2003,
424, 810816.
(26) Wang, K.; Schonbrun, E.; Steinvurzel, P.; Crozier, K. B.
Trapping and rotating nanoparticles using a plasmonic nano-tweezer
with an integrated heat sink. Nat. Commun. 2011,2, 469.
ACS Central Science Research Article
DOI: 10.1021/acscentsci.8b00301
(27) Xin, H.; Li, Y.; Li, B. Controllable patterning of different cells
via optical assembly of 1D periodic cell structures. Adv. Funct. Mater.
2015,25, 28162823.
(28) Mao, Z.; Li, P.; Wu, M.; Bachman, H.; Mesyngier, N.; Guo, X.;
Liu, S.; Costanzo, F.; Huang, T. J. Enriching Nanoparticles via
Acoustofluidics. ACS Nano 2017,11, 603612.
(29) Ahmed, D.; Ozcelik, A.; Bojanala, N.; Nama, N.; Upadhyay, A.;
Chen, Y.; Hanna-Rose, W.; Huang, T. J. Rotational manipulation of
single cells and organisms using acoustic waves. Nat. Commun. 2016,
7, 11085.
(30) Xu, T.; Soto, F.; Gao, W.; Dong, R.; Garcia-Gradilla, V.;
Magaña, E.; Zhang, X.; Wang, J. Reversible swarming and separation
of self-propelled chemically powered nanomotors under acoustic
fields. J. Am. Chem. Soc. 2015,137, 21632166.
(31) Bruus, H. Acoustofluidics 10: scaling laws in acoustophoresis.
Lab Chip 2012,12, 15781586.
(32) Hammarström, B.; Laurell, T.; Nilsson, J. Seed particle-enabled
acoustic trapping of bacteria and nanoparticles in continuous flow
systems. Lab Chip 2012,12, 42964304.
(33) Collins, D. J.; Ma, Z.; Han, J.; Ai, Y. Continuous micro-vortex-
based nanoparticle manipulation via focused surface acoustic waves.
Lab Chip 2017,17,91103.
(34) Tanyeri, M.; Ranka, M.; Sittipolkul, N.; Schroeder, C. M. A
microfluidic-based hydrodynamic trap: design and implementation.
Lab Chip 2011,11, 17861794.
(35) Shenoy, A.; Tanyeri, M.; Schroeder, C. M. Characterizing the
performance of the hydrodynamic trap using a control-based
approach. Microfluid. Nanofluid. 2015,18, 10551066.
(36) Schasfoort, R. B.; Tudos, A. J. Handbook of surface plasmon
resonance; Royal Society of Chemistry: Cambridge, 2008; pp 9098.
(37) Brandhoff, L.; Zirath, H.; Salas, M.; Haller, A.; Peham, J.;
Wiesinger-Mayr, H.; Spittler, A.; Schnetz, G.; Lang, W.; Vellekoop, M.
J. A multi-purpose ultrasonic streaming mixer for integrated magnetic
bead ELISAs. J. Micromech. Microeng. 2015,25, 104001.
(38) Wiklund, M.; Green, R.; Ohlin, M. Acoustofluidics 14:
Applications of acoustic streaming in microfluidic devices. Lab Chip
2012,12, 24382451.
(39) Liang, J.; Zhang, H.; Zhang, D.; Duan, X.; Zhang, H.; Pang, W.
Design and fabrication of aluminum nitride Lamb wave resonators
towards high figure of merit for intermediate frequency filter
applications. J. Micromech. Microeng. 2015,25, 035016.
(40) Loiseleux, T.; Chomaz, J.; Huerre, P. The effect of swirl on jets
and wakes: linear instability of the Rankine vortex with axial flow.
Phys. Fluids 1998,10, 11201134.
(41) Monnier, N.; Barry, Z.; Park, H. Y.; Su, K.-C.; Katz, Z.; English,
B. P.; Dey, A.; Pan, K.; Cheeseman, I. M.; Singer, R. H. Inferring
transient particle transport dynamics in live cells. Nat. Methods 2015,
12, 838840.
(42) Armani, A. M.; Kulkarni, R. P.; Fraser, S. E.; Flagan, R. C.;
Vahala, K. J. Label-free, single-molecule detection with optical
microcavities. Science 2007,317, 783787.
(43) Duan, X.; Mu, L.; Sawtelle, S. D.; Rajan, N. K.; Han, Z.; Wang,
Y.; Qu, H.; Reed, M. A. Functionalized polyelectrolytes assembling on
nano-bioFETs for biosensing applications. Adv. Funct. Mater. 2015,
25, 22792286.
(44) Liu, W.; Wang, J.; Yu, Y.; Chang, Y.; Tang, N.; Qu, H.; Wang,
Y.; Pang, W.; Zhang, H.; Zhang, D. Tuning the resonant frequency of
resonators using molecular surface self-assembly approach. ACS Appl.
Mater. Interfaces 2015,7, 950958.
(45) Whitehead, T. A.; Chevalier, A.; Song, Y.; Dreyfus, C.;
Fleishman, S. J.; De Mattos, C.; Myers, C. A.; Kamisetty, H.; Blair, P.;
Wilson, I. A. Optimization of affinity, specificity and function of
designed influenza inhibitors using deep sequencing. Nat. Biotechnol.
2012,30, 543548.
ACS Central Science Research Article
DOI: 10.1021/acscentsci.8b00301
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The advancement in micro/nanotechnologies has been continuously providing possibilities for inventing novel biochemical sensors. However, variations in the transducer type can cause different sensing results due to the differences in their mechanisms of analyzing biomolecular interactions. In this work, we focused on the comparative analysis of static and non-static assays for molecular interactions using on-chip integrated extended-gate field effect transistor (EGFET) as a static sensing interface and solidly mounted resonator (SMR) as a non-static sensing interface. Analysis of polyelectrolytes (PETs) surface assembly and antigen-antibody interaction using the two types of biochemical sensors presented consistent and complementary sets of information. Meanwhile, due to the difference in their operating mechanisms, variations on the detection efficiency, kinetics and thermodynamics were observed. Our results highlighted the critical dependence of signal detection on biochemical sensors’ operating mechanisms and provided a valuable guidance for static and non-static assays for biomolecular detections.
We present an ultrasonic streaming mixer for disposable and on-chip magnetic bead ELISAs. The ultrasonic transducer is placed at system-level to keep cost per chip as low as possible, and is coupled to the chip by means of a solid ultrasonic horn. The system provides mixing of liquids, as well as dispersion of the superparamagnetic beads in the ELISA. Additionally it can be used clean the chamber surface from nonspecifically bound proteins during the washing steps in the ELISA protocol. Using our system the time for the ELISA protocol has been greatly reduced down to 30 min.
The development of rapid, low cost, easy-to-use approaches for the quantitative detection of multiple biomarkers would drastically impact global health by enabling medical diagnosis at the point-of-care. Unfortunately, current multiplexed methods for the quantitative detection of antibodies or other disease markers (e.g., enzyme-linked immunosorbent assays (ELISAs), Western blots, fluorescence polarization assays) remain complex multiple-step assays that rely on well-trained technicians and expensive instrumentations. As a promising alternative, we describe here a novel signal transduction mechanism for the one-step detection of any large macromolecules (e.g. antibodies) that utilizes steric hindrance effects at the nanoscale. More specifically, this homogenous assay takes advantage of the large dimension of macromolecules in comparison to the diameter of a double stranded DNA helix: upon binding to a signaling DNA strand that contain a small recognition element, the relatively large macromolecule will reduce the number of signaling strands that can hybridize on its complementary sequence immobilized on a surface. Here we demonstrate this steric hindrance hybridization assay using an electrochemical readout (eSHHA) and show that it enables the quantitative, one-step detection of four different macromolecules in the low nanomolar range in less than 10 minutes directly in whole blood. This inexpensive eSHHA sensor can be readily multiplexed and used for the point-of-care detection of any proteins for which we possess a small recognition element that we can attach to DNA.
Flexible patterning of different cells into designated locations with direct cell–cell contact at single-cell patterning precision and control is of great importance, however challenging, for cell patterning. Here, an optical assembly method for patterning of different types of cells via direct cell–cell contact at single-cell patterning precision and control is demonstrated. Using Escherichia coli and Chlorella cells as examples, different cells are flexibly patterned into 1D periodic cell structures (PCSs) with controllable configurations and lengths, by periodically connecting one type of cells with another by optical force. The patterned PCSs can be flexibly moved and show good light propagation ability. The propagating light signals can be detected in real-time, providing new opportunities for the detection of transduction signals among patterned cells. This patterning method is also applicable for cells of other kinds, including mammalian/human cells.
A new surface functionalization scheme for nano-Bio field effect transistors (FETs) using biocompatible polyelectrolyte thin films (PET) is developed. PET assemblies on Si nanowires (Si-NWs) are driven by electrostatic interactions between the positively charged polymer backbone and negatively charged Si/SiO2 surface. Such assemblies can be directly coated from PET aqueous solutions and result in a uniform nanoscale thin film, which is more stable compared to the conventional amine silanization. Short oligo-ethylene glycol chains are grafted on the PETs to prevent nonspecific protein binding. Moreover, the reactive groups of the polymer chains can be further functionalized to other chemical groups in specific stoichiometry for biomolecules detection. Therefore, it opens a new strategy to precisely control the functional group densities on various biosensor surfaces at the molecular level. In addition, such assemblies of the polymers together with the bound analytes can be removed with the pH stimulation resulting in regeneration of a bare sensor surface without compromising the integrity and performance of the Si-NWs. Thus, it is believed that the developed PET coating and sensing systems on Si-NW FETs represent a versatile, promising approach for regenerative biosensors which can be applied to other biosensors and will benefit real device applications, enhancing sensor lifetime, reliability, and repeatability.