U.S. patent application number 10/749529 was filed with the patent office on 2005-07-07 for biosensor utilizing a resonator having a functionalized surface.
This patent application is currently assigned to Intel Corporation. Invention is credited to Berlin, Andrew A., Ma, Qing, Rao, Valluri, Wang, Li-Peng, Yamakawa, Mineo, Zhang, Yuegang.
Application Number | 20050148065 10/749529 |
Document ID | / |
Family ID | 34711091 |
Filed Date | 2005-07-07 |
United States Patent
Application |
20050148065 |
Kind Code |
A1 |
Zhang, Yuegang ; et
al. |
July 7, 2005 |
Biosensor utilizing a resonator having a functionalized surface
Abstract
Systems and methods for detecting the presence of biomolecules
in a sample using biosensors that incorporate resonators which have
functionalized surfaces for reacting with target biomolecules. In
one embodiment, a device includes a piezoelectric resonator having
a functionalized surface configured to react with target molecules,
thereby changing the mass and/or charge of the resonator which
consequently changes the frequency response of the resonator. The
resonator's frequency response after exposure to a sample is
compared to a reference, such as the frequency response before
exposure to the sample, a stored baseline frequency response or a
control resonator's frequency response.
Inventors: |
Zhang, Yuegang; (Cupertino,
CA) ; Berlin, Andrew A.; (San Jose, CA) ; Ma,
Qing; (San Jose, CA) ; Wang, Li-Peng; (San
Jose, CA) ; Rao, Valluri; (Saratoga, CA) ;
Yamakawa, Mineo; (Campbell, CA) |
Correspondence
Address: |
DLA PIPER RUDNICK GRAY CARY US, LLP
4365 EXECUTIVE DRIVE
SUITE 1100
SAN DIEGO
CA
92121-2133
US
|
Assignee: |
Intel Corporation
Santa Clara
CA
|
Family ID: |
34711091 |
Appl. No.: |
10/749529 |
Filed: |
December 30, 2003 |
Current U.S.
Class: |
435/287.2 ;
422/79 |
Current CPC
Class: |
Y10T 436/25375 20150115;
B82Y 30/00 20130101; G01N 29/348 20130101; B82Y 15/00 20130101;
Y10T 436/144444 20150115; G01N 27/126 20130101; G01N 2291/0255
20130101; G01N 2291/0256 20130101; Y10T 436/143333 20150115; G01N
29/2437 20130101; G01N 29/022 20130101; G01N 29/036 20130101; G01N
2291/02466 20130101; Y10T 436/11 20150115; G01N 33/0031 20130101;
G01N 33/54373 20130101; G01N 2291/0426 20130101 |
Class at
Publication: |
435/287.2 ;
422/079 |
International
Class: |
C12M 001/34 |
Claims
What is claimed is:
1. A device comprising: a resonator; wherein the resonator has at
least one functionalized surface, wherein the functionalized
surface is configured to react with target molecules.
2. The device of claim 1, wherein the resonator comprises a
piezoelectric resonator, wherein the device further comprises a
pair of electrodes coupled to the piezoelectric resonator and
control circuitry configured to apply an excitation signal to the
pair of electrodes and to determine a frequency response for the
layer of piezoelectric material.
3. The device of claim 2, further comprising a second piezoelectric
resonator having a non-functionalized surface and an additional
pair of electrodes coupled to the second piezoelectric resonator,
wherein the control circuitry is configured to apply the excitation
signal to the additional pair of electrodes and to determine a
frequency response for the second piezoelectric resonator.
4. The device of claim 3, wherein the piezoelectric resonators
comprise film bulk acoustic resonators (FBARs).
5. The device of claim 2, wherein the excitation signal comprises
an in-phase signal.
6. The device of claim 2, wherein the excitation signal comprises
an out-of-phase signal.
7. The device of claim 2, wherein the excitation signal comprises a
single frequency signal.
8. The device of claim 2, wherein the excitation signal comprises a
mixed frequency signal.
9. The device of claim 2, wherein the excitation signal comprises a
time-variant signal.
10. The device of claim 1, wherein the functionalized surface
comprises one or more biomolecules configured to bind with the
target molecules.
11. The device of claim 10, wherein the biomolecules comprise
biologically active molecules.
12. The device of claim 10, wherein the biomolecules comprise
biologically derivatized molecules.
13. The device of claim 1, wherein the functionalized surface is
functionalized by immobilization of biomolecules on a self-assembly
monolayer.
14. The device of claim 1, wherein the functionalized surface is
functionalized by immobilization of biomolecules on an organic
membrane.
15. The device of claim 14, wherein the organic membrane is
pre-coated onto the functionalized surface.
16. The device of claim 14, wherein the organic membrane is
chemically derivatized on the functionalized surface.
17. The device of claim 16, wherein the organic membrane is
chemically derivatized on the functionalized surface by
silylation.
18. The device of claim 16, wherein the organic membrane is
chemically derivatized on the functionalized surface by
acylation.
19. The device of claim 16, wherein the organic membrane is
chemically derivatized on the functionalized surface by
esterification.
20. The device of claim 16, wherein the organic membrane is
chemically derivatized on the functionalized surface by
alkylation.
21. The device of claim 1, wherein the functionalized surface is
functionalized by direct immobilization of biomolecules on
metal.
22. The device of claim 1, wherein the functionalized surface is
functionalized by direct immobilization of biomolecules on a
non-metallic inorganic film.
23. The device of claim 1, wherein the functionalized surface is
functionalized by self-assembling biomolecular layers on the
functionalized surface.
24. The device of claim 23, wherein the assembling biomolecular
layers comprise amino acid derivatized fatty acids or lipids.
25. A. system comprising: a layer of piezoelectric material,
wherein the layer of piezoelectric material has at least one
surface that is functionalized to bind with target molecules a pair
of electrodes coupled to the layer of piezoelectric material
control circuitry configured to apply an excitation signal to the
pair of electrodes and to determine a frequency response for the
layer of piezoelectric material
26. A system comprising: a pair of film bulk acoustic resonators
(FBARs), including a test FBAR and a reference FBAR, wherein each
FBAR includes a layer of piezoelectric material a pair of
electrodes coupled to opposite sides of the layer of piezoelectric
material; wherein an exposed surface of one of the electrodes of
the test FBAR is functionalized with biomolecules; further
comprising control circuitry coupled to the pair of FBARs and
configured to determine frequency responses for the test FBAR and
reference FBAR.
27. A method for detecting target molecules comprising: providing a
first resonator, wherein the first resonator has a first surface
functionalized with a first type of biomolecules, wherein the
presence of target molecules causes the first type of biomolecules
to change the frequency response of the first resonator; exposing
the first surface of the first resonator to a test fluid;
determining a frequency response of the first resonator after the
first surface has been exposed to the test fluid; and determining,
based upon the frequency response of the first resonator, whether
the test fluid contained target molecules.
28. The method of claim 27, further comprising: providing a second
resonator, wherein the second resonator has a second surface that
is not functionalized with the first type of biomolecules; exposing
the second surface of the second resonator to the test fluid;
determining a frequency response of the second resonator after the
second surface has been exposed to the test fluid; and wherein
determining, based upon the frequency response of the first
resonator, whether the test fluid contained target molecules.
29. The method of claim 28, further comprising, after exposing the
first surface of the first resonator and the second surface of the
second resonator to the test fluid, removing at least a portion of
the test fluid from the first surface of the first resonator and
the second surface of the second resonator before determining the
frequency responses of the first and second resonators.
30. The method of claim 28, further comprising, after exposing the
first surface of the first resonator and the second surface of the
second resonator to the test fluid, removing substantially all of
the test fluid from the first surface of the first resonator and
the second surface of the second resonator before determining the
frequency responses of the first and second resonators.
Description
BACKGROUND OF THE INVENTION
[0001] 1. Field of the Invention
[0002] The invention relates generally to biosensors, and more
particularly to biosensors that incorporate resonators having
functionalized surfaces for binding or otherwise reacting with
target biomolecules in a manner that changes the frequency
responses of the resonators.
[0003] 2. Background Information
[0004] Biosensors are used to detect the presence and/or levels of
biomolecules, typically in a fluid sample. For instance, biosensors
may be used to determine the levels of particular chemicals in
biological fluids, such as blood. Specific sensors can therefore be
used to determine the levels of glucose, potassium, calcium, carbon
dioxide, and other substances in blood samples.
[0005] Biosensors such as these often use an electrochemical system
to detect a particular substance of interest. The electrochemical
system includes substances such as enzymes and redox mediators to
react with the substance of interest (the target substance) and to
thereby produce ions that can carry a current. A set of electrodes
are used to generate an electrical potential that attracts the ions
to the electrodes, creating a circuit that can be used to measure
the resulting current.
[0006] In one type of system, a biosensor includes an enzyme which
is immobilized by a membrane. The target substance in a fluid
sample migrates through the membrane and reacts with the enzyme.
This forms ions within the fluid sample. These ions then migrate
through the fluid sample to the system's electrodes. The migration
of the ions to the electrodes generates an electrical current that
is measured. Because the current depends upon the concentration of
the target substance in the sample, the measured current is then
translated to a concentration of the target substance.
[0007] There are a number of problems with these conventional
biosensors. For example, they are relatively slow. This is, at
least in part, a result of the fact that it is necessary in
electrochemical biosensors to allow a certain amount of time to
pass before the current resulting from the ionization of the target
substance in the sample is established. Only after this current is
allowed to establish itself can it be measured to provide a
reasonably accurate estimate of the concentration of the target
substance.
[0008] Even after the current resulting from the ionization of the
target substance is established and measured, the resulting
estimation of the target substance concentration typically is not
as accurate as would be desirable. This is a result, at least in
part, of the fact that the sample being tested typically contains
various other substances, some of which may interfere in the
process. For instance, some of these other substances may ionize in
the sample and thereby increase the measured current, leading to an
overestimation of the target substance concentration.
Alternatively, some chemicals may react with the ions of the target
substance, thereby reducing the measured current and causing an
underestimation of the target substance concentration.
[0009] It would therefore be desirable to provide systems and
methods that enable the testing of samples to determine the
presence of target substances more quickly and more accurately than
is typically possible using prior art systems and methods.
BRIEF DESCRIPTION OF THE DRAWINGS
[0010] Other objects and advantages of the invention may become
apparent upon reading the following detailed description and upon
reference to the accompanying drawings.
[0011] FIG. 1 is a diagram illustrating the structure of an
exemplary resonator in accordance with one embodiment.
[0012] FIG. 2 is a functional block diagram illustrating a
biosensor system in accordance with one embodiment.
[0013] FIGS. 3A-3C are a set of diagrams illustrating the binding
of target molecules to the functionalized surface of a biosensor in
accordance with one embodiment.
[0014] FIG. 4 is a flow diagram illustrating a method for detecting
the presence of target molecules in a sample in accordance with one
embodiment.
[0015] FIG. 5 is a flow diagram illustrating a method for detecting
the presence of target molecules in a sample in accordance with an
alternative embodiment.
[0016] While the invention is subject to various modifications and
alternative forms, specific embodiments thereof are shown by way of
example in the drawings and the accompanying detailed description.
It should be understood, however, that the drawings and detailed
description are not intended to limit the invention to the
particular embodiments which are described. This disclosure is
instead intended to cover all modifications, equivalents and
alternatives falling within the scope of the present invention as
defined by the appended claims.
DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS
[0017] One or more of the problems outlined above may be solved by
the various embodiments of the invention. Broadly speaking, the
invention comprises systems and methods for detecting the presence
of molecules (e.g., biomolecules) in a sample using sensors that
incorporate resonators which have functionalized surfaces for
binding or otherwise reacting with target molecules in a manner
that changes the frequency responses of the resonators.
[0018] In one embodiment of the invention, a device includes a
resonator, where the resonator has at least one functionalized
surface which is configured to react with target molecules. The
reaction of the target molecules with the functionalized surface
causes changes in the mass and/or charge of the resonator,
stress/strain, surface energy/tension and the like, which cause
changes in the vibrational characteristics of the resonator.
Changes in the vibrational characteristics of the resonator may be
manifested through corresponding changes in electrical
characteristics of the resonator.
[0019] In one embodiment, the resonator consists of a layer of
piezoelectric material and a pair of electrodes that are coupled to
opposite sides of the layer of piezoelectric material. One of the
electrodes forms the functionalized surface of the resonator. When
an excitation signal is applied across the electrodes, the
frequency response of the resonator can be determined. When target
biomolecules come into contact with the functionalized surface, the
target biomolecules react (e.g., bind). with the functionalized
surface and cause changes in the mass and/or electrostatic charge
of the resonator. By determining the frequency responses of the
resonator before and after exposure to a sample that may contain
target biomolecules, changes in the frequency response correlated
to the changed mass and/or electrostatic charge can be determined,
indicating the detection of the target biomolecules.
[0020] In one embodiment, a pair of resonators is used. Each of the
resonators is essentially as described above, except that one of
the resonators has a functionalized surface and the other does not.
The resonator that does not have a functionalized surface is used
as a control against which the other resonator can be compared.
Thus, when both resonators are exposed to a sample, any target
biomolecules will affect the frequency response of the
functionalized-surface resonator, but not the resonator without the
functionalized surface. Any non-target molecules will equally
affect both resonators and the corresponding frequency responses,
so a comparison of the two resonators will effectively cancel out
any effects resulting from non-target molecules.
[0021] In one embodiment of the invention, a method includes the
steps of providing a resonator having a surface functionalized with
a type of biomolecules, where the presence of target molecules
causes the biomolecules of the functionalized surface to change the
frequency response of the resonator, exposing the functionalized
surface of the resonator to a test fluid, determining a frequency
response of the resonator after the functionalized surface has been
exposed to the test fluid, and determining whether the test fluid
contains target molecules based upon the frequency response of the
resonator.
[0022] In one embodiment, the method includes the additional steps
of providing a second resonator that does not have a functionalized
surface, exposing the second resonator to the test fluid,
determining a frequency response of the second resonator after the
second resonator has been exposed to the test fluid, and comparing
the frequency response of the second resonator to the frequency
response of the first (functionalized) resonator to determine the
effect of target molecules on the frequency response of the first
resonator.
[0023] Numerous additional embodiments are also possible.
[0024] One or more embodiments of the invention are described
below. It should be noted that these and any other embodiments
described below are exemplary and are intended to be illustrative
of the invention rather than limiting.
[0025] As described herein, various embodiments of the invention
comprise systems and methods for detecting the presence of
molecules in a sample using sensors that incorporate resonators
which have functionalized surfaces for binding or otherwise
reacting with target molecules in a manner that changes the
frequency responses of the resonators.
[0026] In one embodiment, a biosensor includes a piezoelectric
resonator that has a surface which is functionalized to react with
target biomolecules. The resonator consists in one embodiment of a
layer of piezoelectric material that has a pair of electrodes that
are coupled to opposite sides of the layer of piezoelectric
material. One of the electrodes forms the functionalized surface of
the resonator. When an excitation signal is applied across the
electrodes, the frequency response of the resonator can be
determined. When target biomolecules come into contact with the
functionalized surface, the target biomolecules react (e.g., bind)
with the functionalized surface and cause changes in the mass
and/or electrostatic charge of the resonator. By determining the
frequency responses of the resonator before and after exposure to a
sample that may contain target biomolecules, changes in the
frequency response correlated to the changed mass and/or
electrostatic charge can be determined, indicating the detection of
the target biomolecules.
[0027] In one embodiment, a pair of resonators is used. Each of the
resonators is essentially as described above, except that one of
the resonators has a functionalized surface and the other does not.
The resonator that does not have a functionalized surface is used
as a control against which the other resonator can be compared.
Thus, when both resonators are exposed to a sample, any target
biomolecules will affect the frequency response of the
functionalized-surface resonator, but not the resonator without the
functionalized surface. Any non-target molecules will equally
affect both resonators and the corresponding frequency responses,
so a comparison of the two resonators will effectively cancel out
any effects resulting from non-target molecules.
[0028] Referring to FIG. 1, a diagram illustrating the structure of
an exemplary resonator in accordance with one embodiment is shown.
The resonator illustrated in this figure comprises a film bulk
acoustic resonator (FBAR) device. Device 100 includes a layer of
piezoelectric material 110 sandwiched between electrodes 121 and
122 to form a resonator component. This resonator component is
positioned with its edges on a silicon/silicon dioxide substrate
130 (a layer of silicon dioxide 131 deposited on a layer of silicon
132) in order to allow the resonator component to vibrate. The
exposed surface of electrode 121 is functionalized with a layer 140
of biologically active or derivatized material. The biologically
active or derivatized material interacts with the target
biomolecules by, for example, binding the biomolecules to layer 140
and thereby changing the mass or electrostatic charge of this layer
and, consequently, the resonator component.
[0029] The FBAR resonator may be constructed using techniques that
are known to persons of skill in the art. For example, in one
embodiment, a FBAR resonator may be constructed according to the
following process.
[0030] First, a substrate is provided. In one embodiment, a silicon
wafer is used as the substrate, although other substrate materials
used in semiconductor processing (e.g., gallium arsenide) can also
be used. A layer of sacrificial material is then deposited on the
substrate. The sacrificial layer may consist of a variety of
materials, such as Al, Cu, NiFe, ZnO, or other suitable materials
that are known in the art. The sacrificial layer may be deposited
using any suitable process, such as sputtering or vapor
deposition.
[0031] A photoresist layer is then formed on top of the sacrificial
layer. A pattern is then formed in the photoresist using
conventional methods. The patterned photoresist forms a mask which
is used to selectively etch the sacrificial layer. More
specifically, the photoresist mask covers an area of the
sacrificial layer that will later form an air gap beneath the
piezoelectric resonator component. After the sacrificial layer is
etched, an insulator layer is deposited on the substrate,
effectively replacing the sacrificial layer that was previously
etched away. The insulator layer can be deposited or otherwise
formed using conventional means.
[0032] After the insulator layer is deposited, the photoresist is
removed using, for example, a lift-off process. The portion of the
insulator layer that is on top of the photoresist is also removed.
This results in a patterned layer of insulator and sacrificial
materials on top of the substrate. In other words, the sacrificial
material is inset within the insulator material (or vice versa) to
form a pattern within this layer. This layer will form the
supporting structure for the resonator component after the
sacrificial material is removed in a later step.
[0033] A membrane layer may optionally be formed on top of the
patterned insulator/sacrificial layer. References below to
formation of structures on top of the layer of
insulator/sacrificial material should be construed as formation of
the structures on the membrane layer if the membrane layer is
used.
[0034] A conductive layer is then formed on the layer of
insulator/sacrificial material. This conductive layer may consist
of any conductive material such as a metal. Suitable metals may
include Al, Au, W, Pt or Mo. This conductive layer is patterned to
form a lower electrode of the resonator component. A layer of
piezoelectric material, such as AlN or ZnO, is then formed on top
of the conductive layer. This piezoelectric layer is patterned to
form the body of the piezoelectric resonator. A second conductive
layer is then formed on top of the piezoelectric layer. This
conductive layer is patterned to form the upper electrode of the
resonator component. After the resonator component is formed in
this manner, the sacrificial material on the substrate below the
resonator component is removed using, for example, a wet etch
process (it may be necessary to form a via to the sacrificial layer
in order to effect the removal of this material).
[0035] The upper electrode of the resonator component has a lower
side which is bound to the piezoelectric layer and an upper side
which is exposed. This exposed to surface is then functionalized so
that it will react (e.g., bound) with target molecules. In one
embodiment, the electric surface is functionalized with antibody or
DNA molecules. This may be accomplished by forming self-assembling
monolayers of various thiols or sulfides on the electrode surface
using a chemisorption process. The antibody or DNA molecules can
then be covalently linked to the self-assembled monolayer using an
activation process.
[0036] Various alternative means for functionalizing the surface of
the resonator are also possible. For example, the functionalization
of the FBAR device may be achieved by immobilization of
biomolecules on an organic membrane that is pre-coated on the
surface of the device, or chemically derivatized such as
silylation, esterification, alkylation, or similar processes that
are known in the art. The functionalization of the FBAR device may
also be achieved by direct immobilization of biomolecules on a
metal or other inorganic film on the surface of the device, or
self-assembled biomolecular layers such as amino acid-derivatized
fatty acids/lipids on the surface of the device.
[0037] Because the biosensor may be used in a "wet" environment, it
may be necessary to protect portions of the biosensor other than
the exposed resonator surfaces. In other words, the sample which is
being tested to determine the presence of the target biomolecules
may be a liquid sample. For some of the components of the
biosensor, exposure to a liquid sample may cause problems that
could prevent proper operation of the biosensor. For example, the
liquid sample could cause a short circuit between the electrodes of
the resonator component, thereby preventing measurement of changes
in the frequency response of the resonator. Some embodiments of the
invention may therefore also include a protective layer that covers
the components of the biosensor, except for the functionalized
surface of the resonator component (and possibly the
non-functionalized electrode surface of a corresponding control
biosensor). This protective layer may be provided by forming a
polymer membrane over the components that require protection.
[0038] It should be noted that the foregoing description pertains
to a single, exemplary embodiment. Alternative embodiments may be
formed using slightly different steps in the described process. A
number of these variations are included in the description of the
foregoing process, and additional such variations will be apparent
to those of skill in the art upon reading the present disclosure.
For example, in one alternative embodiment, the exposed electrode
surface of the resonator component may be functionalized by coating
the surface with an ion-selective membrane. The ion-selective
membrane can then be functionalized with enzymes such as glucose
oxidase. Alternatively, the ion-selective membrane can be
functionalized with a functional membrane that provides transport
mechanisms such as carrier molecules, ion pores or channels
extracted from biological materials or synthetic biochemicals.
[0039] As noted above, the FBAR biosensor described herein is used
by detecting changes in the frequency response of the resonator
that result from the exposure of the biosensor to a sample and
subsequent reaction of the functionalized surface with the target
molecules. In order to determine the frequency response of the
resonator, control circuitry is provided. An exemplary embodiment
of a system including an FBAR biosensor and corresponding control
electronics is shown in FIG. 2.
[0040] Referring to FIG. 2, a functional block diagram illustrating
a biosensor system in accordance with one embodiment is shown. In
this embodiment, system 200 includes a resonator 210 and control
the circuitry 220. In one embodiment, resonator 210 is as described
above. Resonator 210 is coupled to control circuitry 220 by the
electrodes of the resonator. The electrodes are coupled to signal
generation circuitry 221 and processing circuitry 222 components of
control circuitry 220.
[0041] Signal generator circuitry 221 is configured to produce an
excitation signal that is applied to the electrodes of resonator
210. The excitation signal has an AC (alternating current)
component that causes the piezoelectric material of the resonator
to vibrate. Because of the physical characteristics of resonator
210, the resonator has a characteristic frequency response. The
frequency response of resonator 210 manifests itself in the
variability of the electrical characteristics of the resonator
(e.g., the impedance of the resonator). These electrical
characteristics can be measured by processing circuitry 222.
[0042] The frequency response of resonator 210 has a fundamental
resonance at a frequency at which the corresponding wavelength is
twice the thickness of the resonator. The wavelength is equal to
the acoustic velocity of the piezoelectric material, divided by the
frequency. The acoustic velocity of the piezoelectric material
depends upon the specific material that is used. For instance, AlN
has an acoustic velocity of about 10,400 meters per second, while
ZnO has an acoustic velocity of about 6,330 meters per second.
Thus, for a resonator using AlN, if the thickness of the resonator
is about 2.5 micrometers, the resonant frequency is about 2 GHz. If
it is desired to adjust the resonant frequency to a different
frequency, this can be achieved by, for example, changing the
piezoelectric material or changing the thickness of the
resonator.
[0043] It should be noted that, because the technology used in the
manufacturing of FBAR devices is compatible with both Si and GaAs
wafer processing techniques, it is possible to combine these
technologies to make all-in-one biosensors. In other words, it is
possible to manufacture both the resonator and the control
circuitry on a single chip. This may provide additional advantages
over the prior art in terms of simplification of the design of the
respective components of the biosensors, improved power efficiency,
and so on.
[0044] Thus, in one embodiment, biosensor system 200 operates by
generating an excitation signal that includes a plurality of
frequencies (not necessarily at the same time), applying this
excitation signal to resonator 210 and then measuring the
electrical characteristics of the resonator corresponding to each
of the frequencies. For example, signal generator circuitry 221 may
generate an excitation signal that includes a single frequency
which varies as a function of time. In other words, signal
generator circuitry 221 scans through a range of frequencies.
Processing circuitry 222 may then measure, for example, the
impedance across resonator 210 as a function of frequency (which is
a function of time). This frequency response (i.e., the impedance
of resonator 210 as a function of frequency) may be digitized,
stored and compared to a baseline response, or in may be compared
to the response of a control resonator, which would be operated in
the same manner.
[0045] It should be noted that the excitation signal applied to the
resonator may include various components (e.g., single or mixed
frequencies, or time-variant components). Similarly, the frequency
response may be measured in terms of various response components
(e.g., in-phase and out-of-phase components) or other response
characteristics. Such response characteristics may include the
steady-state frequency shifts of the resonators due to changes of
mass or electrostatic charge resulting from the specific binding of
the target molecules with the immobilized biomolecules on the
resontaors' surfaces (e.g., antibody-antigen, DNA hybridization,
molecular receptor binding, molecular configurational changes).
[0046] The biosensor is useful in the detection of target molecules
because, in reacting with the functionalized surface of the
resonator, the target molecules change the mass and/or
electrostatic charge of the resonator, both of which affect the
resonance of the resonator. In other words, these characteristics
change the vibrational characteristics of the resonator. For
example, if the target molecules bind with the functionalized
surface and thereby effectively increase the mass of the resonator,
the resonator will tend to respond less quickly to the forces
generated by the applied excitation signal. The resonant frequency
will therefore be lower. Thus, if, prior to the binding of target
molecules to the functionalized surface, a resonator resonates at a
frequency f, the additional mass of the target molecules that are
bound to the functionalized surface will cause the resonator to
resonate at a frequency f-.DELTA.f. If the frequency response of
the resonator is viewed as a function of frequency, this
corresponds to a shift of the peak response to the left (the lower
frequencies).
[0047] Referring to FIGS. 3A-3C, a set of diagrams illustrating the
binding of target molecules to the functionalized surface of a
biosensor in accordance with one embodiment shown. Referring first
to FIG. 3A, a diagram illustrating a biosensor prior to exposure to
a sample is shown. The biosensor has essentially the same structure
shown in FIG. 1, including a support structure formed by substrate
331 and insulator layer 332, and a resonator component formed by
piezoelectric layer 310 sandwiched between electrodes 321 and 322.
Electrode 322 of the resonator component is functionalized by
antibodies 340, which can be bound to the electrode, for example,
by a self-assembling monolayer of a thiol. A protective polymer
layer 350 (not shown in FIG. 1) covers the resonator component and
support structure, except for the functionalized surface of the
resonator component.
[0048] Referring next to FIG. 3B, a diagram illustrating the
biosensor of FIG. 3A during exposure to a sample is shown. As
depicted in this figure, sample 360 contains a variety of different
biomolecules, including antigen molecules (target molecules,
represented in the figure by triangles) and various other molecules
(non-target molecules, represented in the figure by circles and
squares). The biomolecules are distributed throughout sample 360,
so that some of the biomolecules come into contact with the
functionalized surface of the biosensor. As the biomolecules come
into contact with the functionalized surface, they may or may not
become bound to the functionalized surface. More specifically, if a
biomolecule that comes into contact with the functionalized surface
is an antigen corresponding to the antibodies of the functionalized
surface, it will be bound to one of the antibodies. If the
biomolecule is not the specific antigen corresponding to the
antibodies, it will not be bound to the antibodies of the
functionalized surface.
[0049] Referring next to FIG. 3C, a diagram illustrating the
biosensor of FIGS. 3A and 3B after the sample is removed is shown.
It can be seen from this figure that, when the sample (e.g., a
biological fluid) is removed from the biosensor, the non-antigen
biomolecules contained in the sample are also removed. The antigen
biomolecules bound to the antibodies remain. These antigens affect
the mass and/or electrostatic charge of the resonator component and
will therefore change the frequency response of the resonator
component. Thus, the presence of the antigen biomolecules is
detected by determining whether the frequency response of the
resonator component has changed. This is accomplished as described
above.
[0050] It should be noted that, in some cases, removal of the
sample from the biosensor may not ensure that all of the non-target
biomolecules have been removed from the functionalized surface of
the resonator component. These non-target molecules may affect the
frequency response of the resonator component and consequently
affect the determination of whether target molecules were present
in the sample. In one embodiment, the effect of these non-target
molecules is compensated for through the use of a control biosensor
in addition to the test biosensor. The control biosensor is
essentially identical to the test biosensor, except that the
surface of the resonator component is not functionalized to react
with the target biomolecules. When the sample is tested, both the
test biosensor and control biosensor are exposed to the sample. Any
non-target molecules that are not removed from the test biosensor
should likewise not be removed from the control biosensor. Because
the control biosensor does not bind the target biomolecules, any
change in the frequency response of the resonator component of the
control biosensor should be due to the presence of these non-target
biomolecules. Since the effect of the non-target biomolecules is
known from the frequency response of the control biosensor, this
effect can effectively be "subtracted out" of the changes in the
frequency response of the test biosensor.
[0051] It should be noted that, while the foregoing example
describes the use of two biosensors (a test biosensor and a control
biosensor), the two biosensors may be considered either separate
units, or parts of the same biosensor. The two biosensors may each
have their own control circuitry, or they may share all or part of
the control circuitry. In the latter instance, the example may be
more easily understood if the term "biosensor" is replaced with the
term "resonator." Both instances are within the scope of the
invention.
[0052] Referring to FIG. 4, a flow diagram illustrating a method
for detecting the presence of target molecules in a sample in
accordance with one embodiment is shown. In this embodiment, a
biosensor having a single resonator is used. The resonator has a
functionalized surface that will react with target molecules in a
sample. The method of this embodiment includes the steps of
providing a resonator having a functionalized surface (block 410),
determining the frequency response of the resonator prior to
exposure to a sample (block 420), exposing the resonator to the
sample (block 430), determining the frequency response of the
resonator following exposure to the sample (block 440), comparing
the frequency responses of the resonator prior to and following
exposure to the sample (block 450), and determining the presence of
target molecules based upon changes in the frequency response of
the resonator resulting from exposure to the sample (block
460).
[0053] A single resonator is employed in this method, and it is
necessary to provide a baseline frequency response from which
changes in the frequency response can be determined. This baseline
is provided in one embodiment by determining the frequency response
of the resonator prior to exposure to the sample. In an alternative
embodiment, the baseline may be provided by testing a plurality of
resonators that are identically manufactured and establishing a
composite frequency response, or an average frequency response for
resonators having an identical design. In such embodiment, the
composite or average frequency response can be stored in a memory
coupled to the processing circuitry so that it can be retrieved and
compared to the measured frequency response of the resonator after
exposure to the sample. Various other means for providing the
baseline are also possible.
[0054] Referring to FIG. 5, a flow diagram illustrating a method
for detecting the presence of target molecules in a sample in
accordance with an alternative embodiment is shown. In this
embodiment, a biosensor having a pair of resonators is used. One of
the resonators functions as a test resonator, while the other
functions as a control resonator. As described above, the test
resonator has a functionalized surface, while the control resonator
does not. The signal generation circuitry and processing circuitry
for determining the frequency response of each of these resonators
may be common. That is, a single set of signal generation and
processing circuitry can be used in conjunction with both of the
resonators. Alternatively, each resonator may have separate signal
generation and processing circuitry.
[0055] This alternative method includes the steps of providing a
biosensor having dual (test and control) resonators (block 510),
exposing the resonator to the sample (block 520), determining the
frequency response of the test resonator and the control resonator
following exposure to the sample (block 530), comparing the
frequency responses of the test resonator and control resonator
(block 54), and determining the presence of target molecules based
upon differences in the frequency responses of the test resonator
and control resonator (block 550.
[0056] While this alternative method determines the presence of
target molecules based upon differences between the frequency
response of a test resonator and the frequency response of a
control resonator, it may also be useful to be able to compare
either or both of these frequency responses to a baseline response.
For instance, even though the resonators may be identically
designed (except for non-functionalized surface of the control
resonator), the change in frequency response due to the presence of
target molecules may not be linear. Therefore, it may be helpful to
know not only the magnitude of the difference between the test and
control frequency responses, but also the magnitude of the
frequency response changes resulting from non target molecules
(i.e., the difference between a baseline, pre-sample-exposure
frequency response and the control frequency response).
[0057] As noted above, the various embodiments of the present
invention may provide a number of advantages over the prior art.
These advantages may include greater sensitivity and faster
response time than other types of biosensors, higher resonance
frequencies that may provide higher sensitivity to changes in mass
or in stiffness (resulting from changes in charge), simpler
structures than other types of resonators (e.g., single-crystal
quartz microbalance or SAW resonators), better power handling
characteristics at high frequencies, sharper response peaks (due to
reduced parasitic effects, larger surface areas for detection of
target molecules, and the ability to perform detection of target
molecules during or after exposure to wet environments. Another
advantage of some of the embodiments of the present invention is
that they may make use of relatively mature technologies (e.g.,
relating to methods for designing and manufacturing FBAR devices
and integrated circuits, or functionalizing various surfaces) and
may therefore present fewer difficulties in the design, practice
and/or manufacture of the respective embodiments.
[0058] The benefits and advantages which may be provided by the
present invention have been described above with regard to specific
embodiments. These benefits and advantages, and any elements or
limitations that may cause them to occur or to become more
pronounced are not to be construed as critical, required, or
essential features of any or all of the claims. As used herein, the
terms "comprises," "comprising," or any other variations thereof,
are intended to be interpreted as non-exclusively including the
elements or limitations which follow those terms. Accordingly, a
system, method, or other embodiment that comprises a set of
elements is not limited to only those elements, and may include
other elements not expressly listed or inherent to the claimed
embodiment.
[0059] While the present invention has been described with
reference to particular embodiments, it should be understood that
the embodiments are illustrative and that the scope of the
invention is no limited to these embodiments. Many variations,
modifications, additions and improvements to the embodiments
described above are possible. It is contemplated that these
variations, modifications, additions and improvements fall within
the scope of the invention as within the following claims.
* * * * *