Analytical system with photonic crystal sensor

A system for determining whether interaction occurs between a trial substance and a target substance. The system includes a photonic crystal sensor having a photonic crystal structure and a defect member disposed adjacent the photonic crystal structure. The defect member defines an operative surface able to receive the target substance and the trial substance. The system further includes a light source that inputs a light signal to the photonic crystal structure and the defect member. The light signal is internally reflected, and a resultant output signal is outputted. The output signal relates to whether the trial substance interacts with the target substance at the operative surface. Furthermore, the system includes an identity detector that identifies the trial substance that interacts with the target substance.

FIELD

The present disclosure relates to an analytical system and, more particularly, relates to an analytical system with a photonic crystal sensor.

BACKGROUND

Photonic crystals (PC) are periodic dielectric structures with a band gap forbidding propagation of a certain frequency range of light. It is understood that controlling photon modes by PC structures can be useful. For example, electromagnetic waves can be bent with high efficiency around 90-degree corners within radii smaller than a wavelength with two-dimensional (2D) PC structures. Furthermore, a 3D PC can be used to control the timing of light emitted by a quantum dot. Moreover, a dual core photonic crystal fiber can be used for enhancing two-photon fluorescence biosensing sensitivity.

Selection from various types of PCs depends on specific applications. For example, 3D PCs are used in order to control spontaneous emission. In the case of controlling laser beams which are close to plane waves, a 1D PC is sufficient and preferable due to its simplicity in fabrication. Additionally, 1D PCs are used in various applications, such as an omnidirectional reflector, low-threshold optical switching, and nonlinear optical diodes.

Furthermore, a technique has been developed for enhancing fluorescence by sandwiching the sample of interest (e.g., chromophores) between two pieces of 1D PCs. When a structural defect is introduced in the PCs, a photon-localized state can be created in the photonic band gap and the electric field around the defect member is enhanced. In one embodiment, two-photon fluorescence emission from 2-aminopurine in the PC structure was enhanced by a factor of 120. However, the application of this approach is limited because the thick substrate of the PC structure inhibits the sample from being observed with an optical microscope or from interacting with other biomolecules.

SUMMARY

A system for determining whether interaction occurs between a trial substance and a target substance is disclosed. The system includes a photonic crystal sensor having a photonic crystal structure and a defect member disposed adjacent the photonic crystal structure. The defect member defines an operative surface able to receive the target substance and the trial substance. The system further includes a light source that inputs a light signal to the photonic crystal structure and the defect member. The light signal is internally reflected, and a resultant output signal is outputted. The output signal relates to whether the trial substance interacts with the target substance at the operative surface. Furthermore, the system includes an identity detector that identifies the trial substance that interacts with the target substance.

Furthermore, a method of determining whether interaction occurs between a trial substance and a target substance is disclosed. The method includes delivering the target substance relative to an operative surface of a photonic crystal sensor. The photonic crystal sensor includes a photonic crystal structure and a defect member disposed adjacent the photonic crystal structure. The defect member defines the operative surface. The method further includes delivering the trial substance relative to the operative surface. Moreover, the method includes inputting an input light signal from a light source to the photonic crystal structure and the defect member such that the input light signal is internally reflected and such that an output signal is outputted. The method additionally includes detecting the output signal to determine whether the trial substance interacts with the target substance. Furthermore, the method includes identifying the trial substance that interacts with the target substance.

In addition, a system for determining whether a trial substance binds with a target substance is disclosed. The system includes a fluidics system having a common channel, a target branch that branches from the common channel, and a control branch that branches from the common channel. The system also includes a target photonic crystal sensor and a control photonic crystal sensor. Each of the target and control photonic crystal sensors include a respective photonic crystal structure and a respective defect member. The defect members define a respective operative surface. The target photonic crystal sensor is fluidly connected to the target branch, and the control photonic crystal sensor is fluidly connected to the control branch. Moreover, the fluidics system delivers the target substance relative to the operative surface of the target photonic crystal sensor and withholds the target substance from the operative surface of the control photonic crystal sensor. The fluidics system also delivers a plurality of different trial substances to the target and control photonic crystal sensors. Additionally, the system includes a light source that inputs a light signal to the photonic crystal structure of the target photonic crystal sensor. The light signal is internally reflected, and a resultant output signal is outputted. The output signal relates to whether any of the plurality of trial substances binds with the target substance. Furthermore, the system includes a target mass spectrometer that receives a downstream flow of the plurality of trial substances from the target photonic crystal sensor and that measures a change in an amount of the trial substances received over time. The system further includes a control mass spectrometer that receives a downstream flow of the plurality of trial substances from the control photonic crystal sensor and that measures a change in an amount of the trial substances received over time. The system additionally includes a controller with a processor that compares the change in amount of the trial substances received by the target mass spectrometer with the change in amount of the trial substances received by the control mass spectrometer in order to identify which of the plurality of trial substances binds with the target substance and to determine how long the identified trial substance binds with the target substance.

DETAILED DESCRIPTION

Referring initially toFIG. 1, one embodiment of a photonic crystal (PC) sensor10is illustrated. In the embodiment shown, the PC sensor10includes a PC structure12, a defect member14(e.g., defect layer) disposed adjacent the PC structure12on one side, and a prism16disposed adjacent the PC structure12on an opposite side. The defect member14defines an operative surface15. Also, a substrate18is interposed between the PC structure12and the prism16. The input light has total internal reflection (TIR) at the sensor top surface. Owing to the TIR, an imaginary PC structure exists, which results in a unique open microcavity.

As will be discussed in greater detail below, only one PC is used in a total internal reflection (TIR) geometry instead of a prior art closed cavity with two pieces of PCs. Thus, the PC sensor10is open and exposed, and can be easily influenced by other molecules or signals to be detected. The reflection spectrum of the PC structure is extremely sensitive to changes in thickness of the defect member14and/or to the refractive index of biomolecules bound to the defect member14. Thus, using the PC sensor10in this unique configuration opens up a wide range of applications and forms the basic detection mechanism for constructing highly sensitive sensors for various situations.

As shown, input light from a light source (not shown) is inputted to the prism16, the input light passes through the prism16, the substrate18, the PC structure12, and the defect member14if a resonance condition is satisfied in the defect member14. Then, the input light is totally internally reflected at the operative surface15. If a resonance condition is not satisfied in the defect member14, the input light merely passes through the prism16and the substrate18, while it gets highly reflected by the PC structure. A light signal is outputted from the PC sensor10. As will be discussed in greater detail below, the output light signal is monitored in order to detect a condition at the operative surface15. For instance, the PC sensor10can be used for biomolecular assay or as an ultrasonic sensor.

The PC structure12includes a plurality of alternating pairs of layers of different dielectric materials. In one embodiment, each dielectric material has a different refractive index. In one embodiment, there are seven alternating layers of silica and hafnia with the top layer (i.e., the layer on which the defect member14is provided) being hafnia. Also, in this embodiment, the substrate18is made of glass. Moreover, the defect member14is made of silica and has a thickness that is different than the thickness of any of the layers within the PC structure12. In another embodiment, the defect member14includes a plurality of layers of different materials. In addition, the defect member14includes an absorbing material in one embodiment. It will be appreciated that the materials used to make the PC structure12, the substrate18, and/or the defect member14could be of any suitable type without departing from the scope of the present disclosure.

Furthermore, in one embodiment, the thickness of each dielectric layers in the PC structure12is determined according to the following considerations. It is assumed that the incident angle at the substrate layer18is θi. The refraction angles in the substrate18, the silica layers of the PC structure12, the hafnia layers of the PC structure12, and the defect member14are θs, θA, θB, and θX, respectively. Also, ns, nA, nB, and nXare the refractive index of the substrate18, the silica layers of the PC structure12, the hafnia layers of the PC structure12, and the defect member14, respectively. Therefore:
nssin θs=nAsin θA, nXsin θs=nBsin θB, nSsin θs=nxsin θX(1)

The thickness of the dielectric layers in the PC structure12satisfies the following relation:

nA⁢dA⁢cos⁢⁢θA=nB⁢dB⁢cos⁢⁢θB=λ4(2)
where λ is the center wavelength of the photonic band gap, and dAand dBare the thickness for the silica layers and hafnia layers, respectively.

In order to obtain local field enhancement in the defect member14, the thickness of the defect member14satisfies the following resonant condition,

2⁢nX⁢dX⁢cos⁢⁢θX+α⁢⁢λd2⁢π=λd2(4)
where α represents the Goos Hänchen phase shift, and λdis the wavelength of the photonic defect state, which should be within the photonic band gap. The factor of 2 in the first term on the left hand side of equation 4 is due to the fact that the light double passes the defect member due to the TIR. From Eqs. (1) and (4), we have,

For s-polarization of incident light, the Goos Hänchen phase shift is given by the following expression:

For p-polarization of incident light, the Goos Hänchen phase shift is given by the following expression:

In order to test fluorescence enhancement of the PC sensor10, the center wavelength is chosen to be 600 nm. A 100 μL of 1 μM ethanol solution of Nile blue, a known dye molecule, is spin coated on the operative surface15of the defect member14. A reference sample (not shown) is also constructed having a substrate deposited with only one silica layer. The same solution Nile blue solution is spin-coated on the reference sample. The reference sample and the PC sensor10are placed on a prism with index-matching fluid and excited by a laser output from an optical parametric amplifier tuned to 600 nm in wavelength. The emitted fluorescence is collected with a 20× objective lens and passes through a sharp-cut long-pass filter. A spectrometer coupled with a photon-counting streak scope is used to observe the wavelength- and time-resolved fluorescence.

The excitation power dependence of the fluorescence intensity is measured. In one embodiment represented inFIG. 2, a saturation of absorption occurs at approximately 0.24 mW excitation power for the PC sensor10. Before the saturation, the observed fluorescence intensity from the PC sensor10was enhanced by 20-fold compared to that from the reference sample without a PC structure.

This result experimentally verifies that a single PC can be used in a TIR geometry to achieve local field enhancement. In addition, the fluorescence spectrum collected from top of the PC-TIR sample is not modified by the photonic band gap structure as represented inFIG. 3A. Also, the fluorescence decay curve shown inFIG. 3Bhas a decay constant of 2.3 ns, which is in agreement with the value of 2.6 ns for the reference sample within a margin of detection error. These results demonstrate that it is possible to form a microcavity by using only one piece of PC structure in a TIR geometry.

As will be discussed in greater detail below, the operative surface15of the defect member14is generally exposed so as be affected by external influences to be discussed. As such, the properties of the light reflected at the operative surface15will vary depending on the external influences at the operative surface15. Thus, the output signal from the PC sensor10can be monitored for changes in order to detect the condition at the operative surface15. Accordingly, the PC sensor10can be used for various sensing applications, such as kinetic analysis of biomolecular interactions or sensitive ultrasonic detections, and the like.

APPLICATION EXAMPLE 1

Biomolecular Assay

Specific interactions of biological molecules with various ligands, biopolymers, and membranes, such as protein:protein, protein:lipid, protein:DNA and protein:membrane binding, provide a chemical foundation for all cellular processes. The study of these biomolecular affinity and binding kinetics is of great importance for biomedical researches. For example, in order to discover a novel therapeutic antibody to treat cancer and other diseases, screening of a large repertoire of small molecules is necessary in order to identify high specificity and high affinity binders, followed by a more detailed characterization of the binding properties and determination of epitope specificity. Thus, there is an increasing demand for sensitive, accurate and high throughput analytical instruments that can provide insights on a molecular basis into critical biological processes.

One of the most widely used analytical techniques is based on surface plasmon resonance (SPR). In this known technique, a thin noble metal film is placed in an evanescent optical field, and the change of the SPR reports the binding of analytes to the ligand immobilized on the metal surface. Although this method allows label-free, real-time analysis, it has many drawbacks. For example, it is difficult to obtain accurate kinetic analysis due to the mass-transport limitation and it is not suitable for the measurement of small analytes (e.g., ˜1000 Da).

However, the PC sensor10of the present disclosure has a detection mechanism fundamentally different from the SPR and leads to a significant improvement in detection sensitivity. More specifically, the photonic defect state of the PC sensor10is extremely sensitive to the change in the thickness and/or refractive index of the defect member14, and this characteristic is employed for a sensitive biomolecule analysis tool as described below.

In order to test this tool a PC sensor10is constructed that includes a PC structure12having three alternating layers of silica and titania disposed on a substrate18, which is made of BK7 glass. The resonant wavelength is chosen to be 600 nm. The silica and titania layers of the PC structure12have a refractive index of 1.458 and 2.225, respectively, and the substrate18has a refractive index of 1.516. A defect member14with a refractive index of 1.49 is deposited on the top layer (titania) of the PC structure12. The incident angle in the substrate18is chosen to be 68°. For s-polarization input light, the thickness of the silica, titania and the defect member14are 388.32 nm, 86.98 nm, and 446.32 nm, respectively, based on Eqs. (3) and (7), assuming the refractive index of the solution in contact with the defect member is 1.333. When the incident light wavelength falls in the photonic band gap, it is reflected by the PC structure12and does not reach the defect member14. When the incident light wavelength matches the photonic defect state, the light reaches the defect member14and experiences TIR at the operative surface15.

FIG. 4Ashows the spectrum of light that reaches the defect member and experiences TIR when there is no absorption in the defect member. When the defect member14is doped with some absorbing material, the light is partially absorbed. In the calculation, we assume the extinction coefficient to be 0.0005 in the absorptive defect member14, which corresponds to about only 0.32% absorption when there is no PC structure12.FIG. 4Bshows the corresponding spectrum in contrast to the case with an absence of absorption. The result shows that the absorption is more than 90%, enhanced by the PC structure12comparing 0.32% in the absence of a PC structure12.

In practice, the portion of light reflected by the PC structure12and the portion reflected at the operative surface15are mixed and difficult to separate. Therefore, the total amount of the reflected light is used as a measurable quantity.

The defect member14is doped with absorbing materials, so that the reflected light that passes through the defect member has lower reflectance, thus resulting a reflectance dip at a specific wavelength corresponding to the defect state within the photonic band gap. When a thin layer is formed on the operative surface15(defined as an “adlayer” on the operative surface15), the wavelength of the reflectance dip varies. It is appreciated that the adlayer could be formed by biological molecules to be observed in a biomolecular assay.

Then, the change in the output signal (i.e, the reflected light) from the PC sensor10is observed in order to detect the condition at the operative surface15due to the adlayer. More specifically, in one embodiment, a change in wavelength of the output light signal is detected as shown inFIG. 5and/or a change in output signal light intensity at a wavelength of 632.8 nm is detected as shown inFIG. 6.

Specifically, a sharp and prominent dip appears in the reflection spectrum corresponding to the photonic defect state as shown inFIG. 5. This dip is relatively narrow and the resonant wavelength of this dip is sensitive to the material bound to the operative surface15. For example, in the embodiment shown inFIG. 5, the dip width is only 0.1 nm and the resonant wavelength is shifted as much as 0.86 nm when the adlayer is 1 nm thick and with a refractive index of 1.46. This spectrum shift can be easily measured since the dip width is extremely narrow. Also, an even narrower dip width, and thus more sensitive detection, can be observed by increasing the incident angle and/or the number of alternating layers of the PC structure. It is noted that although the defect member14is absorptive, both immobilized reactants on the operative surface15and mobile reactants that form the adlayer need not be labeled. Thus, it is possible to obtain a label-free measurement for the sample of interest.

In addition to directly measuring the spectrum shift with a spectrometer, a differential reflectance measurement is employed at a fixed wavelength.FIG. 6shows that the reflectance changes as much as 85.24% at the wavelength of a HeNe laser (632.8 nm), when a 0.1-nm thin adlayer with refractive index of 1.46 is formed on the sensor surface. Since a detection accuracy of 10−4˜10−5can be obtained in a typical differential reflectance measurement, an extremely high detection sensitivity of molecular binding on the sensor surface can be achieved. The detectable adlayer thickness can be as small as 10−5˜10−6nm. This is a significant improvement compared with the sensitivity of an SPR based instrument.

Besides higher detection sensitivity, the detection system using the PC sensor10has many other advantages over the SPR based detection, such as larger detection range, better baseline stability and flexible detection configurations. These advantages are described below.

Using the PC sensor10, the adlayer is in the actual optical path of the input light signal, which increases the optical path length of the defect member14. The reflectance dip of the photonic defect state shifts within the photonic bandgap of the PC structure12when the adlayer thickness and/or refractive index changes. The detection range is therefore restricted by the bandgap, which is around 700 nm for the PC structure12with three pairs of alternating dielectric layers and an incident angle of 68°. The detection range can be even wider by increasing the incident angle. In contrast, the detection range of an SPR instrument is limited to the range of the evanescent field, which is typically around 200 nm. Thus, the PC sensor10can be used to detect thicker adlayers or bigger particles compared to SPR measurements.

Also, for the detection range on the small side, a critical factor is the detection baseline stability. A small change in refractive index of the solution on the sensor surface may cause a resonant wavelength variation. There may be several causes for changes in the refractive index, such as the temperature. The evanescent field in SPR has a large portion (relative to the thin adlayer) in the bulk solution. The change in the refractive index of the solution contributes to the shift of SPR wavelength in the same way as the adlayer. A small change in the solution property results in nonnegligible fluctuation in the detection baseline. For example, when the refractive index of solution changes only by 0.0002, the SPR baseline drift is as large as 0.62 nm, which is equivalent to the change when a 0.25-nm adlayer with refractive index of 1.46 is attached to the defect member14. On the other hand, the adlayer interacts with a propagating field in the PC sensor10and increases the actual optical path, while the contribution to the change in resonant wavelength from the solution is due to the Goos Hänchen phase shift, which is very small. Thus, the resonant wavelength is less affected by the solution. For the same change in the solution, the resonant wavelength of the PC sensor10shifts just 0.14 nm.FIG. 7shows the comparison of baseline shift caused by very small fluctuations in the refractive index of bulk solution between SPR and PC-TIR detection.

In the SPR based measurements, the refractive index and thickness of the adlayer cannot be independently determined; one has to use the value of one of the parameters given by other measurements in order to calculate the value of the other parameter. In contrast, the refractive index and thickness of the adlayer can be independently determined using the PC sensor10. The resonant wavelength shift of the defect state varies with the angle or polarization of the incident light, and this dependence is different for different refractive index of the adlayer. Thus, one obtains both the refractive index and adlayer thickness by carrying out the measurement with two different incident angles or two different polarizations.

Referring now toFIG. 8, one embodiment of a detection system20employing a PC sensor10is shown. In this embodiment, the detection system20is a bioassay system. The detection system20is modular because it includes a plurality of light sources22a,22band a plurality of sensing devices24a,24b. In the embodiment shown, the detection system20can be used to detect spectral shift of the output signal and/or differential reflectance of the output signal.

Using the white light source22a, the reflectance spectrum is obtained using the PC sensor10. A large detection range (about 1-700 nm) of adlayer thickness can be obtained. When the differential reflectance measurement is used, extremely high detection sensitivity can be achieved and the detectable range of the adlayer thickness is from 1 nm down to 10−5nm.

In order to detect spectral shift, light from a broad band white light source22ais directed through a first lens26and is coupled into a single-mode (SM) fiber28to get a good optical mode, and the output beam from the fiber28is collimated with a second lens30. The light from the white light source22ais then reflected by a first reflecting mirror32and through a linear polarizer34to adjust the light polarization. The light is reflected by a second reflecting mirror36that is mounted on a rotator38. The position of the rotator38is controlled by a computer to adjust the incident angle. The light is then reflected again by a third reflecting mirror40. The light enters the PC sensor10through the prism16, is totally internally reflected at the operative surface15for the wavelength component that satisfies the resonance condition, or is reflected by the PC structure12for the other wavelength components. The total output signal from the PC sensor10is reflected by a fourth reflecting mirror42, passes through a third lens44, and is directed into a spectrometer sensing device24a. Thus, the spectral shift of the output signal can be detected.

To detect differential reflectance with the detection system20, a laser22bis used as the light source. In one embodiment, the laser22bis a HeNe laser22bwith the wavelength of 632.8 nm. Also, the PC sensor10includes a two-part structure represented inFIG. 9. On a first portion50of the operative surface15, molecular binding occurs and an adlayer51is formed thereon. A second portion52of the operative surface15remains unchanged. As shown inFIG. 8, the laser beam passes through a fourth lens54, through a pinhole56, through a fifth lens58, and through a diaphragm60. Then, the laser beam is split into two by a splitter62. One of the split beams is reflected by a fifth reflecting mirror64, through the polarizer34, and is reflected by the second and third reflecting mirrors36,40to thereby illuminate the first portion50of the operative surface15having the adlayer51. The other of the split beams is reflected by a sixth and seventh reflecting mirror68,66, through the polarizer34, and is reflected by the second and third reflecting mirrors36,40to thereby be directed at the second portion52of the operative surface15as a reference. The intensities of the two reflected beams are monitored by a second sensing device24bhaving two identical detectors. The intensity ratio between the two beams is recorded, which sensitively reflects the property of the adlayer51.

Referring now toFIG. 10, another embodiment of a detection system21is illustrated. In this embodiment, the PC sensor10includes a defect member14having an operative surface15that is angled. More specifically, the operative surface15is angled so as to define a plane that intersects a plane defined by an opposite side81of the defect member14.

The position where TIR occurs is observed. As shown, for a fixed incident angle and certain wavelength of incident light, the TIR occurs at a position where the defect member14thickness satisfies the resonance condition. Thus, by monitoring the position of TIR one can use the PC sensor10to detect if there is something (e.g. biomolecules) bound to the defect member14or if there is some signal that changes the thickness of the defect member14.

Two techniques can be used to fabricate the PC sensor10for use in the detection system20shown inFIG. 8. First, electron-beam evaporation can be used. Silica, Titania and pure silicon are used as the target materials. Rutherford backscattering spectroscopy (RBS) is used to calibrate the thickness of deposited layers. The transmission spectrum of PC structure12and defect member14composed of silicon and silica at normal incident direction is measured, which is in agreement with simulation results as shown inFIG. 11A. Second, fifteen pairs of GaAs/AlAs layers are grown on a GaAs substrate18by MBE with high accuracy (˜0.1 nm). InxGa1-xAs (x=0.4) is used as the absorptive layer and its thickness (d=8 nm) is decided by considering both resonant dip depth and growing mismatch with GaAs. Another GaAs layer is added on top of InxGa1-x As to satisfy the resonant condition in the defect member.FIG. 11Bshows the measured and calculated transmission spectrum at normal incident direction. The good agreement between the theoretical calculation and experimental result indicate that the PC sensor10is well fabricated.

In one embodiment, the PC sensor10is made by electron-beam evaporation. The thickness of the three pairs of Silica and Titania alternating layers in the PC structure12is 438 nm and 87 nm, respectively. Its defect member14is composed of 18-nm Silicon and 140-nm Silica.

FIG. 12Ashows the light intensity of a broad band white light source reflected from the PC sensor10(as indicated by the curve60) and a reference substrate (as indicated by the curve62) used in a total internal geometry.FIG. 12Bshows the experimental and simulated PC-TIR reflectance spectrum. In the calculation, the thickness of different layers are taken from the measured values, and the refractive indices of the substrate18(e.g., BK7), Silica and titania are 1.516, 1.458 and 2.225, respectively, at the wavelength of 600 nm. The refractive index and extinction coefficient of silicon are 3.94 and 0.0183 at 600 nm, respectively. The solvent used on the operative surface15is de-ionized (DI) water with a refractive index of 1.333. The incident angle is 64.5°. Both the simulation and experimental results show a primary resonance dip at 580 nm with a narrow width of 8 nm. The dip width is much narrower than that of a typical SPR resonance spectrum width (40 nm).

Accordingly, the resonant wavelength shifts to longer wavelength with decreasing the incident angle as shown inFIG. 13A. In addition, the dependence of the resonance wavelength on solvents is measured as shown inFIG. 13B. The resonant wavelength shift is only 150 nm per RIU for the solvent with a refractive index close to water. This small shift is preferred in order to have a system with high stability because a small temperature fluctuation may cause the change in solvent refractive index. The refractive index of water is known to decrease by ˜8×10−5RIU per degree K near room temperature. Using the solvent refractive index dependence of the resonance wavelength shift 150 nm/RIU, the temperature stability of the PC-TIR system is −0.012 nm/K, which is remarkably improved comparing with that of an SPR system (i.e., −0.25 nm/K). The dependence of the resonant wavelength shift on the adlayer properties is also shown inFIG. 13C. As shown, the resonant wavelength sensitively shifts to longer wavelength with increasing the adlayer thickness.

In addition, the resonance dip is measured as a function of the incident angle for a fixed wavelength of 580 nm as shown inFIG. 14. The full width at half maximum is only 0.4°, which is about five times smaller than that in an SPR system. The experimental result is again in a good agreement with a simulation.

The PC sensor10also has an extremely high sensitivity in the differential reflectance measurement. Detection of a thin monolayer of small molecules can be carried out. For instance, the PC sensor10is first exposed to 5% APTES solutions for 20 minutes, then rinsed with de-ionized (DI) water and methanol and dried with nitrogen. After the silanization, the PC sensor10is exposed to a 2.5% glutaraldehyde solution in 20 mM HEPES buffer (pH=7.4) for 30 minutes, then rinsed with DI water and dried with nitrogen. A thin layer of Glutaraldehyde molecules are obtained due to its reaction with the amino groups on the silanized surface. The total thickness of the APTES/glutaraldehyde layer is 1.5 nm.

A resonance dip shift by 1.24 nm is observed in the reflectance spectrum measurement as shown inFIG. 15, when the APTES/glutaraldehyde adlayer is formed. In order to achieve higher sensitivity, differential reflectance measurements with a HeNe laser are used.

The intensity stability of the HeNe laser is measured to be about 2% as shown inFIGS. 16A and 16B. A clear improvement of the detection accuracy is obtained by measuring the intensity ratio between these two beams. The fluctuation of the ratio is less than 1.6*10−4as shown inFIG. 16C.

Furthermore, a clear ratio change from 0.8232 to 0.7141 is observed (FIG. 17) when the thin APTES/glutaraldehyde adlayer is formed on the PC sensor10.

The detection sensitivity of the sensor is expressed as,

Since the system has detection accuracy up to 1.6×10−4, and the ratio change is 0.1091 for a 1.5 nm adlayer, the detection sensitivity of the PC sensor10is 0.002 nm, which is significantly improved compared with the sensitivity of an SPR based instrument.

The theoretical detection limit is calculated using our simulation results. For a PC sensor10with three alternating layers in the PC structure12, the resonance dip width is calculated to be 2 nm. A detection limit of 1.5×10−5nm can be predicted for a PC sensor10, which is in contrast to 3×10−3nm for an SPR system. A comparison between the PC sensor10and SPR systems is shown inFIG. 18.

Thus, the PC sensor10offers the ability to perform real-time and label-free quantitative detections, allowing for a wide range of biomolecular assays with a higher sensitivity and better stability compared to known SPR-based instruments. In addition, the PC sensor10can independently determine the refractive index and thickness of the adlayer.

Referring now toFIG. 20, a further embodiment of a system70is illustrated that incorporates a PC sensor10aof the types discussed above. The system70can be used for determining whether interaction occurs between different substances. For instance, the system70can be used for performing a bioassay and determining whether biological substances bind together. Also, in some embodiments, the system70can be used to determine how long the substances bind together, whether the substances bind directly or indirectly to each other, and/or other characteristics of the substances as will be discussed in greater detail below.

As shown schematically inFIG. 20, the system70can include a delivery system72. The delivery system72can be of any suitable type, such as a fluidics system (e.g., a micro-fluidics system) that includes one or more channels73capable of directing a fluid flow therethrough.

The system70can also include a PC sensor10a. The PC sensor10aincluded in the system70can be a PC sensor10aof any of the embodiments discussed above. As such, the PC sensor10acan include a photonic crystal structure12and a defect member14that defines an operative surface15. The PC sensor10acan be in operative communication with the delivery system72in any suitable fashion. For instance, the operative surface15can be exposed to or within the channel73of the delivery system72. As such, the operative surface15can receive a target substance78via the delivery system72as will be discussed in greater detail below. The operative surface15can also receive one or more trial substances80a-80evia the delivery system72. For instance, the trial substances80a-80ecan flow through the channel73toward the PC sensor10aafter the target substance78has been delivered to the PC sensor10a. As such, the trial substances80a-80ecan have an opportunity to interact (e.g., bind) with the target substance78on the operative surface15of the PC sensor10a.

It will be appreciated that the target substance78and the trial substances80a-80ecan be of any suitable type, such as molecules of a protein, etc. Also, it will be appreciated that the delivery system72can deliver any suitable amount and concentration of the target and trial substances78,80a-80e.

Moreover, the system70can include a light source75, such as a laser or other suitable light source75. As discussed above, the light source75can input an input light signal91to the PC sensor10a, the light signal can be internally reflected in the PC sensor10a, and a resultant output light signal93can be outputted. It will be appreciated that, in some embodiments, only a small fraction of the input light signal91(e.g., within a relatively small wavelength range) experiences total internal reflection while the remaining portions of the input light signal91is reflected by the PC sensor10a. Thus, as discussed in detail above, characteristics of the output light signal93can relate to and can be dependent on whether any of the trial substances80a-80einteracts with the target substance78at the operative surface15.

Thus, the PC sensor10acan be used to detect whether binding occurs between any of the trial substances80a-80eand the target substance78. For purposes of clarity, the system70will hereafter be described as detecting whether binding occurs between any of the trial substances80a-80eand the target substance78; however, it will be appreciated that the PC sensor10acan be used to detect any other suitable interaction between the target and trial substances78,80a-80e.

Furthermore, the system70can include an identity detector74athat identifies the trial substances80a-80ethat bind with the target substance78. For instance, the identity detector74acan be a commercially available mass spectrometer76. However, it will be appreciated that the identity detector74acould be of any other suitable type, such as a nuclear magnetic resonance (NMR) device, an enzyme-linked immunosorbent assay (ELISA) device, etc. The identity detector74acan be fluidly connected to the delivery system72. For instance, the identity detector74acan be located downstream of the PC sensor10aand can receive a downstream flow from the PC sensor10avia the delivery system72. Accordingly, one or more of the trial substances80a-80e(and, in some cases, the target substance78) can flow from the PC sensor10ato be received by the identity detector74a. As will be discussed, the identity detector74acan analyze this flow from the PC sensor10ato identify and distinguish which of the trial substances80a-80ebinds with the target substance78.

In addition, the system70can include a controller77. The controller77can be of any suitable type, such as a computerized tool, that is in communication with both the PC sensor10aand the identity detector74a. Also, the controller77can include a processor79for processing data received from both the PC sensor10aand the identity detector74a. The controller77can also include a display (such as a computer monitor, printer, etc.) for displaying the processed results. Moreover, the controller77can control flow through the delivery system72as will be discussed in greater detail below.

FIG. 24further illustrates various embodiments of the system70. As shown, the system can include a plurality of PC sensors, namely, a target PC sensor10aand a control PC sensor10b. Also, the system70can include a target identity detector74aand a control identity detector74b. The delivery system72can include a common channel86. The delivery system72can further include a first branch88and a second branch90that are each downstream of the common channel86and that each fluidly branch off of the common channel86. The first branch88can fluidly connect the target PC sensor10ato the common channel86and the target PC sensor10ato the target identity detector74a. Also, the second branch90can fluidly connect the control PC sensor10bto the common channel86and the control PC sensor10bto the control identity detector74b. Also, the system70can include a valve92that controls fluid flow through the first and second branches88,90as will be discussed. Furthermore, as shown inFIG. 24, the controller77can be in communication with both the target and control PC sensors10a,10band both the target and control identity detectors74a,74b. (It will be appreciated thatFIGS. 20 and 21illustrate the first branch88, the target PC sensor10a, and the target identity detector74a.)

During use of the system70, the controller77can control the valve92such that the first branch88is open and the second branch90is closed. Then, the controller77can cause the target substance78to flow through the first branch88to the target PC sensor10ato be deposited on the operative surface15of the target PC sensor10a. For instance, the target substance78can be included at a known concentration within an appropriate coating buffer solution. It will be appreciated that target substance78will be deposited on the operative surface15of the target PC sensor10a, but the target substance78will not be deposited the control PC sensor10b.

Subsequently, the controller77can control the valve92such that both the first and second branches88,90are open, and the controller77can cause the trial substances80a-80eto flow through the first and second branches88,90toward each of the target and control PC sensors10a,10b. For instance, the trial substances80a-80ecan be included at a known concentration as an analyte within a running buffer solution.

As shown inFIG. 21, some of the trial substances80a,80b,80cmay bind to the target substance78on the target PC10awhile other trial substances80d,80emay flow freely away from the target substance78. For instance, trial substances80a,80ccan bind directly to the target substance78. Also, trial substance80bcan bind directly to trial substance80ato indirectly bind to the target substance78. It will be appreciated that some of the trial substances80a,80b,80cmay bind temporarily (directly or indirectly) to the target substance78and then subsequently release to flow toward the identity detector74a. As discussed above, the light source75can emit light in order to detect whether binding is occurring at the operative surface15. It will be appreciated that the PC sensor10acan be used to detect whether binding is occurring in real time. Also, the PC sensor10acan be used to detect whether binding is occurring using relatively low concentrations of the target and trial substances78,80a-80e.

The controller77can receive data from the PC sensor10aand can compile the received data into any suitable form, such as the exemplary graph illustrated inFIG. 22. As shown, the controller77can plot the intensity of the output light signal93from the PC sensor10aversus time. As shown in the exemplary embodiment inFIG. 22, the intensity increases substantially at times t1, t2and t3. For instance, this can indicate that trial substance80abinds at time t1, trial substance80bbinds at time t2, and trial substance80cbinds at time t3.

The target identity detector74acan receive the downstream flow from the target PC sensor10and can identify each of the substances contained therein. Also, the control identity detector74bcan receive the flow through the second branch90and can identify each of the substances contained therein. The identity detectors74a,74bcan be used to detect the concentrations or amounts of the trial substances80a-80ereceived over time. The controller77can compare the results of the target and control identity detectors74a,74b.

More specifically, the controller77can receive data from the target and control identity detectors74a,74band can compile the received data into any suitable form, such as one or more graphs of the type shown inFIGS. 23 and 25. As shown inFIG. 23, the controller77can plot the change in concentration of the trial substance80areceived by the target identity detector74aover time as shown by line97a. Also, the controller77can plot the change in concentration of the trial substance80areceived by the control identity detector74bover time as shown by line99a. Then, by subtracting the target data (point T) from the control data (point C) at a specific time tn, the amount of the trial substance80aat time tncan be detected.

Also, as shown inFIG. 25, a similar graph can be generated for trial substance80d. It will be appreciated that a separate graph can be generated for each of the trial substances80a-80e.

Moreover, the controller77can compare the data from the target PC sensor10ato the data from the identity detectors74a,74bin order to detect which of the trial substances80a-80eare binding to the target substance78and which of the trial substances80a-80eare not. More specifically, the target identity detector74acan determine a time t1when binding is occurring on the target identity detector74aas discussed above and as shown inFIG. 22. Then, the amount of each substance80a-80eapproximately at time t1can be detected (tn≈t1) using the data generated by the target and control identity detectors74a,74b. For instance, if time tninFIGS. 23 and 25is approximately equal to time t1, the amount of trial substance80ais relatively low, and the amount of trial substance80dis relatively high. Thus, it can be reasonably assumed that substance80ais binding with the target substance78while substance80dis not binding with the target substance78. A similar analysis can be performed for each of the target substances80a-80efor time t1. Furthermore, a similar analysis can be performed for each of times t1, t2and t3.

Moreover, it will be appreciated that the system70can be used to determine how long the trial substances80a-80ebind with the target substance78. More specifically, as shown inFIG. 26, the identity detector74acan detect that the received amount of trial substance80bis relatively low at or near time tn, and the amount of trial substance80bis substantially higher at time tn+1. Thus, it can be reasonably assumed that substance80bbinds for a time approximately equal to tn+1minus tn.

Subsequently, a regenerate buffer solution can be delivered through the first and second branches88,90of the delivery system72to clean the operative surface15of the PC sensor10a. As such, the trial substances80a-80eand the target substance78can be removed from the PC sensor10a. The analyses discussed above can be performed in conjunction with the delivery of the regenerate buffer solution.

Furthermore, in some embodiments, a wash buffer solution can be delivered through the first and second branches88,90. The wash buffer can be different than the buffer used when the trial substances80a-80ewere previously delivered. For instance, the wash buffer can have a different concentration of salts. As such, the wash buffer can remove those trial substances80a-80ethat are weakly bound to the target substance78. Thus, by performing the tests described above in combination with a plurality of wash buffer solutions, the strength of binding of the trial substances80a-80ewith the target substance78can be tested.

In addition, a plurality of tests can be performed to determine if the trial substances78,80a-80eare binding directly or indirectly to the target substance78. For instance, the trial substance80aand the trial substance80bcan be tested, and binding of both trial substances80a,80bwill likely be detected. Subsequently, the trial substance80bcan be tested alone, and binding is unlikely to be detected. Thus, it can be reasonable to assume that trial substance80bdirectly binds to the trial substance80aand the trial substance80bindirectly binds to the target substance78(seeFIG. 21).

Accordingly, the system70can be used for accurate, real time analysis of the interaction of the target and trial substances78,80a-80e. Furthermore, the system70can be used for label-free analysis of these interactions. Moreover, the system70can be used to detect interactions at relatively low concentrations of the target and trial substances78,80a-80e.

APPLICATION EXAMPLE 2

Ultrasonic Detector

The PC sensor10can also be employed in an ultrasonic detector. Specifically, the exposed operative surface15enables use of the PC sensor10as an ultrasonic detector. Optical detection of ultrasound by using an etalon has been demonstrated as shown inFIG. 19A. As shown, the incident and reflected optical signals are presented as spatially separate but, in practice, are collinear. Acoustic displacement at the etalon surface changes the cavity length, which in turn changes the intensity of the optical signal reflected from the etalon. However, the closed structure of the etalon limits the detection sensitivity.

On the other hand, by using the PC sensor10, the etalon cavity is opened so that the sensor interface can directly interact with the ultrasonic signals. As such, a material with large elasticity can be used as the defect member14, which is difficult to use in a closed etalon cavity because the two high reflection surfaces are intended to remain parallel to each other and flat. Thus, larger change of the optical response to an ultrasonic signal can be obtained compared with prior art detection using an etalon. In addition, the PC sensor10has advantages in optical generation of ultrasound. Because the optical field is enhanced at the operative surface15of the defect member14of the PC sensor10, the ultrasonic signal is generated and propagates directly from the operative surface15without attenuation by any other additional layers, thus allowing strong ultrasound generations. The PC sensor10therefore makes it possible to integrate a strong optical ultrasound generator with a sensitive optoacoustic receiver.