Surface Plasmon Resonance (SPR) relates to optical excitation of surface plasmon waves along an interface between a conductive film and an adjacent dielectric. At resonance, energy from an incident optical signal is coupled to a surface plasmon wave, resulting in a decrease, or dip, in the intensity of an optical signal that is reflected at the conductive film. The optical wavelength at which the dip occurs, referred to as the resonant wavelength, is sensitive to changes in the refractive index of the dielectric that is adjacent to the conductive film. This sensitivity to changes in refractive index enables the dielectric to be used as a sensing medium, for example to detect and identify biological analytes, or for biophysical analysis of biomolecular interactions. There is a need for measurement schemes that increase the accuracy with which changes in refractive index can be detected. In addition, there is a need for measurement schemes that are scalable for use with analytical systems that include arrays of samples for biochemical sensing.
According to the embodiments of the present invention, an incident signal illuminates an SPR sensor over a wavelength range. Intensity of a reflected signal from the SPR sensor is detected with wavelength discrimination imposed on the incident signal or the reflected signal. The wavelength discrimination is imposed at a predesignated tuning rate within the wavelength range. The detected intensity is then sampled at a sampling rate and an intensity profile associated with the SPR sensor is established from the sampling with a wavelength resolution determined by the tuning rate and the sampling rate.
In a typical SPR sensor 10, the conductive film 1 is a gold layer having an appropriate thickness for an incident optical signal, hereafter signal IINC, at a designated angle of incidence φINC and wavelength, to excite a surface plasmon wave, or surface plasmon, along the conductive film 1. Associated with the surface plasmon is an evanescent tail (not shown) that penetrates into the dielectric 2. Energy in the signal IINC that is not coupled into the surface plasmon is reflected at the conductive film 1 to provide a reflected optical signal, hereafter signal Ir.
Coupling between the signal IINC and the surface plasmon results in a decrease, or dip, in the intensity of the signal Ir. The optical wavelength at which the dip occurs, referred to as the resonant wavelength λR, is indicated in
The optical system 20 includes a tunable optical source 22, typically a tunable laser such as an AGILENT TECHNOLOGIES, INC. model 81680B, that can be tuned at a tuning rate γ within a wavelength range λ1λ2. The wavelength range λ1λ2 in this example spans from at least 1492-1640 nanometers. Spectral bandwidth of the signal IINC provided by the tunable optical source 22 in the optical system is typically less than 100 kHz, which is typically narrower than the shifts Δλ in the resonant wavelength λRdetected or measured by the optical system 20. The tunable optical source 22 is alternatively implemented with a tunable optical filter (not shown) cascaded with a white light or other broadband optical source (not shown) to provide a signal IINC that is spectrally narrow and tunable over the wavelength range λ1-λ2. Examples of tunable optical filters suitable for use in this type of tunable optical source 22 are available from MICRON OPTICS, Inc., Atlanta, Ga., USA.
An erbium-doped fiber amplifier (EDFA), or other type of optical amplifier 24, is optionally cascaded with the tunable optical source 22 to increase the power of the signal IINC that illuminates a region, or target T, of the SPR sensor 10. A collimator 26, or other beam-conditioning element, coupled to the tunable optical source 22 directs the signal IINC to the target T. Typically, the signal IINC includes a p-polarized lightwave and an s polarized lightwave that is orthogonal to the p polarized lightwave, where p, s refer to the conventionally defined polarizations p, s. The signal IINC can also be designated to be p polarized by including a polarization controller (not shown) in the signal path between the tunable optical source 22 and the collimator 26. At a designated angle of incidence φINC, the signal IINC couples to the surface plasmon and causes the signal Ir to undergo the dip in intensity at the resonant wavelength λR, shown for example in the intensity profiles of
A detector 28 intercepts the signal IR as the wavelength λ of the tunable optical source 22 is tuned within a wavelength range λ1-λ2 that includes the resonant wavelength λR of the SPR sensor 10. When the resonant wavelength λR occurs outside the wavelength range λ1-λ2, the angle of incidence φINC can be adjusted so that at an adjusted angle of incidence, the resonant wavelength λR falls within the wavelength range λ1-λ2. Adjusting the angle of incidence φINC is typically enabled by mounting the SPR sensor 10 on a rotation stage 25.
The detector 28 is typically a photodiode, photosensor or other transducer suitable for converting an intercepted optical signal into a corresponding electrical signal, hereinafter referred to as detected signal IDET. The detected signal IDET is provided to a processing unit 30 that in this example includes an analog to digital converter 32 that acquires samples of the detected signal IDET. This acquisition of the samples is triggered by a trigger signal TRIG provided by the tunable optical source 22, which indicates initiation of the tuning or sweeping of the tunable optical source 22. The rate of the sample acquisitions, or sample rate, is determined by a clock rate fCLOCK established by a clock 34. The acquisitions result in a set S of samples of the detected signal IDET that is stored in a memory 36. Samples Si in the set S represent the detected intensity of the signal Ir at the wavelengths λi of the tunable optical source 22. Each integer sample number i corresponds to a wavelength λi within the wavelength range λ1-λ2. For example, the wavelength λi of the sample number i of the sample Si in the set S of samples is determined by the relationship λi=λ1+(γ/fCLOCK)i.
Although the detector 28 is typically a broadband detector, to accommodate the wavelength range λ1-λ2 of the tunable optical source, the signal Ir intercepted by the detector is spectrally narrow at the wavelengths of the samples Si, so that wavelength resolution of the acquired samples Si in the set S is not compromised by the spectral width of the signal Ir. With the wavelength λ of the tunable optical source 22 being swept or tuned at the tuning rate γ, the wavelength resolution with which the samples Si in the set S are acquired is based on the ratio of the clock rate fCLOCK and the tuning rate γ. Increasing the clock rate fCLOCK relative to the tuning rate γ increases the wavelength resolution, enabling the intensity of the signal Ir to be accurately represented in an intensity profile as a function of wavelength λ. Curve fitting, averaging or applying other signal processing techniques to the acquired set S of samples enables an accurate representation of an intensity profile associated with the SPR sensor 10. These signal processing techniques are readily performed via a computer or other type of processor (not shown) coupled to the memory 36.
The intensity profile enables an accurate determination of the resonant wavelength λR of the SPR sensor 10, which can be used to accurately determine the resonant wavelength λR of the SPR sensor 10, or shifts Δλ in the resonant wavelength λR, such as those shifts Δλinduced by changes Δn in the refractive index nS of the dielectric 2 of the SPR sensor 10. For example, resonant wavelength λR can be determined from derivatives of the intensity profile to find the minimum of the intensity profile that corresponds to the resonant wavelength λR, or from any other suitable technique for identifying the resonant wavelength λR at the dip in the intensity profile. Shifts Δλ in the resonant wavelength λR between two or more intensity profiles can be detected and quantified by determining the difference in resonant wavelengths λR of the two or more intensity profiles. Shifts in the intensity profile can also be associated with a change in one or more attributes of the SPR sensor such as a change in refractive index in a sensing medium of the SPR sensor 10.
The detected shifts Δλ in the resonant wavelength λR detected from the samples of the detected signal IDET can then be mapped to changes Δn in refractive index nS of the dielectric 2 that induce the shifts Δλ in the resonant wavelength λR. In one example, mapping between the shifts Δλ and the changes Δn is established from computer simulation of the SPR sensor 10 using MATLAB or other suitable program or environment that solves the Fresnel reflections at the interface between the conductive film 1 and the dielectric 2. The computer simulation models the sensitivity dλ/dnS of the resonant wavelength λR to refractive index nS. From the sensitivity dλ/dnS, each shift Δλ in resonant wavelength λR can be mapped to a corresponding change Δn in refractive index nS. In another example, multiple targets T having dielectrics 2 with different known refractive indices nS1, nS2 . . . nSx are illuminated sequentially or simultaneously by optical signals IINC1, IINC2 . . . IINC3 at wavelengths λ in the vicinity of the resonant wavelength λR. From detection and sampling of reflected optical signals Ir1, Ir2 . . . Irx by the detector and processing unit of the optical system, resonant wavelengths λR1, λR2 . . . λRX corresponding to each of the refractive indices nS1, nS2 . . . nSx are determined. Curve-fitting of the resonant wavelengths λR1, λR2 . . . λRX to refractive indices nS1, nS2 . . . nSx, interpolation, or other suitable techniques are then used to establish a mapping between shifts Δλ in resonant wavelength λR and changes Δn in refractive index nS.
The mapping between shifts Δλ in resonant wavelength λR and changes Δn in refractive index nS can also be established by matching appropriate wave vectors at the interface between the conductive film 1 and the dielectric 2. This includes equating the wave vector kSPR=w/c ((ε1nS2)(ε1+nS2))1/2 of the surface plasmon to the wave vector kx=n4(2π/λ)sinφINC of the optical signal IINC, where ε1 is the dielectric constant of the conductive film 1, where n4 is the refractive index of the prism 4, and where φINC is the angle of incidence of the optical signal Ic. The change Δn in refractive index nS can be derived from the equation of the wave vectors kSPR, kx, as equation (1), where the imaginary component of the dielectric constant ε1 of the conductive film 1 is set to zero.
The alternatives presented for mapping detected shifts in the resonant wavelength to changes Δn in refractive index nS are exemplary. It is appreciated that any suitable scheme is alternatively used to establish this mapping.
According to an alternative embodiment of the present invention shown in
In one embodiment, the detector 28 intercepts a resulting filtered signal IF from the tunable optical filter 44 as the passband of the tunable optical filter 44 is tuned within a wavelength range λ1-λ2 that includes the resonant wavelength λR of the SPR sensor 10. When the resonant wavelength λR occurs outside the wavelength range λ1-λ2, the angle of incidence φINC of the signal IWINC can be adjusted via the rotational stage 25 so that at an adjusted angle of incidence, the resonant wavelength λR falls within the wavelength range λ1-λ2. In response to the intercepting the filtered signal IF, the detector 28 produces the signal IDET. The detected signal IDET is then provided to the processing unit 30, which acquires the set S of samples. As with the embodiment shown in
Alternative embodiments of the present invention, shown in
The detected signals IDET1-IDETN from the array of detector elements D1-DN are then provided to the processing unit 30, which acquires sets S1-SN of samples that correspond to each of the targets T1-TN. As with the embodiment shown in
According to the embodiment of the present invention shown in
In the examples presented, shifts Δλ in resonant wavelength λR have been mapped to changes Δn in refractive index nS of the dielectric 2. These changes Δn in refractive index nS can then be used to detect and identify biological analytes, or for biophysical analysis of biomolecular interactions. However, according to alternative embodiments of the present invention, the shifts Δλ in the resonant wavelength λR are mapped to the presence or identity of biological analytes, to biophysical analyses of biomolecular interactions, or to any suitable attributes or features of the SPR sensor 10 that induce the shifts Δλ in the resonant wavelength λR.
Conventional SPR sensing techniques provide for detection of small and medium size analytes, with large analytes being difficult to detect. Surface Plasmon Resonance Biosensors, by Homola et al., page 243, reports that the sensitivity of conventional sensor techniques is not adequate for detecting larger analytes, such as bacteria and cells. However, the embodiments of the present invention accommodate longer wavelengths λ within the wavelength range λ1-λ2 over which the signal IINC illuminates the SPR sensor 10. These longer wavelengths provide correspondingly deeper penetration of the evanescent field into the dielectric 2 of the SPR sensor 10, which enables larger analytes to be detected, identified, monitored, or otherwise measured using the optical systems and methods according to the embodiments of the present invention.
According to the embodiments of the present invention, the resonant wavelength associated with the SPR sensor 10 is typically the wavelength at which the dip in the intensity profile occurs, as shown for example in
Step 56 of the measurement method 50 includes sampling the detected intensity at a sampling rate. Step 58 includes establishing an intensity profile associated with the SPR sensor from the sampling of step 56, where the intensity profile has a wavelength resolution determined by the tuning rate Δ and the sampling rate. The measurement method 60 optionally comprises step 59, which includes adjusting the angle of incidence of the incident signal on the SPR sensor 10 when an identified resonant wavelength λR associated with the SPR sensor 10 occurs outside the wavelength range λ1-λ2, so that at an adjusted angle of incidence, the resonant wavelength λR of the SPR sensor 10 falls within the designated wavelength range λ1-λ2.
While an SPR sensor 10 has been included in the embodiments of the present invention, SPR sensors in these embodiments are meant to include resonant mirror transducers, or any other type of transducer providing reflected optical signals Ir having associated intensity profiles dependent on attributes of a sensing medium that are sensed by penetration of an evanescent wave into the sensing medium.
While the embodiments of the present invention have been illustrated in detail, it should be apparent that modifications and adaptations to these embodiments may occur to one skilled in the art without departing from the scope of the present invention as set forth in the following claims.