Optical heterodyne surface plasmon resonance sensor using a dual optical frequency comb light source, wavelength-stabilized orthogonal linearly polarized dual-frequency laser, or wavelength-stabilized left-right circularly polarized dual-frequency laser as the light source.

Abstract

Conventional surface plasmon resonance sensors measure values ​​based on the change in reflected light intensity. However, light intensity measurement is susceptible to the instability of the light source itself and the influence of ambient light, making accurate quantitative measurement of low concentrations of substances difficult in many cases. [Solution] By applying the optical heterodyne method to the measurement of a surface plasmon resonance sensor, and measuring the phase shift between P and S polarized waves rather than the change in reflected light intensity from the sensor, the effects of light source fluctuations and ambient light can be avoided. Furthermore, since the change in phase shift with respect to the change in incident angle near the resonance angle is much steeper and larger than the change in reflected light intensity, extremely high-sensitivity and high-precision measurements become possible.

Description

Technical Field

[0001] The present invention relates to antigen-antibody reaction and high-sensitivity measuring instruments for trace chemical substances, and is a technology for an apparatus that measures with higher precision and sensitivity than conventional apparatuses.

Background Art

[0002] FIG. 1 is a diagram for explaining the principle of a surface plasmon resonance sensor that is a premise for explaining the prior art. A gold thin film 2 with a thickness of about 50 nm is formed on the bottom surface of a prism 1. When laser light 3 is incident on this bottom surface at an incident angle greater than the critical angle, total reflection occurs. Assuming that the laser light is p-polarized light, an interaction occurs between the evanescent wave 4 due to the P-polarized wave generated on the gold thin film side during total reflection and the surface plasmon wave 5 formed by free electrons in the gold thin film. In the interaction region, a surface plasmon polariton in which the evanescent wave and the surface plasmon are combined is generated (note that the S-polarized wave has nothing to do with the interaction with the surface plasmon). When laser light is incident at an angle where the dispersion curve of the evanescent wave coincides with the dispersion curve of the surface plasmon, the interaction resonantly strengthens, the energy of the p-polarized wave of the incident light is strongly absorbed by the surface plasmon, and the reflected laser light 6 is greatly attenuated. The incident angle at which the reflected light is greatly attenuated is called the resonance angle. It is known that the wave number of the surface plasmon is determined by the dielectric constant of the gold thin film and the dielectric constant of the substance attached thereto.

[0003] As shown in Figure 2, if the surface of the gold thin film is modified, for example, by the antibody in the antigen-antibody reaction, i.e., ligand 7, then the wavenumber of the surface plasmon will differ from the dielectric constant of the gold thin film alone, so the dispersion curve will also change, and the resonance angle will shift. Figure 3 shows the angle of incidence of light versus the intensity of p-polarized reflected light. The resonance angle for the case of the metal thin film alone is shown as θ0, and the resonance angle when modified with ligand 7 is shown as θ1. Here, when the antigen of the antigen-antibody reaction, i.e., the analyte 9 (target molecule), solution is delivered to the bottom of the prism by a liquid-phase pump from the flow channel system 8 shown in Figure 2, the dielectric constant of the gold thin film changes further due to the antigen-antibody reaction, and therefore the resonance angle shifts from θ1 to θ2. The antigen-antibody reaction can be quantified from the amount of this angle shift. This is the principle of a conventional surface plasmon resonance sensor. Its features are: ▲1▼If the ligand is determined, the analyte can be measured from the sample solution without labeling (label-free), eliminating the need for solution pretreatment, ▲2▼1 ng / cm 2 The ability to detect trace amounts of something.

[0004] The actual measurement quantification is performed by fixing the incident angle of the p-polarized wave and measuring the change in reflected light intensity ΔR when the resonance angle shifts, as shown in Figure 4. [Overview of the Initiative] [Problems that the invention aims to solve]

[0005] Devices based on the above principles are now widely used, but all of them rely on the change in intensity of reflected light due to surface plasmon resonance. For example, [Non-Patent Literature 1]. However, light intensity measurement is susceptible to the instability of the light source itself and the influence of ambient light, which limits the accurate quantitative measurement of low-concentration analytes. [Means for solving the problem]

[0006] As mentioned above, all conventional devices measure the intensity change of reflected light due to surface plasmon resonance (hereinafter abbreviated as SPR). The intensity change of reflected light is based on the resonant absorption of incident light at the surface plasmon. The presence of resonant absorption simultaneously means that anomalous dispersion occurs in that region. Figure 5 shows an example of theoretical analysis results for reflectance and p-wave phase change near the SPR resonance angle. The p-wave phase change curve shows a much steeper change compared to the absorption curve. The p-wave phase change can be interpreted as a change in refractive index relative to the P-wave and corresponds to the dispersion curve. By utilizing this steep change, which can be called an anomalous dispersion characteristic, even higher sensitivity and accuracy can be expected compared to conventional methods. If this phase change can be measured by the optical heterodyne method, the instability of the light source intensity and the influence of ambient light become extremely small, and as mentioned above, it shows a steeper change than the change in reflectance, making extremely high-sensitivity and high-precision measurements possible.

[0007] The means of achieving this include using (1) a wavelength-stabilized He-Ne transverse Zeeman laser, (2) a wavelength-stabilized He-Ne longitudinal Zeeman laser, (3) a He-Ne laser wavelength-stabilized by the two-longitudinal mode method, or (4) a dual-comb laser. All of these laser light sources can utilize one polarization as a P-wave and the other as a linearly polarized S-wave orthogonal to it. The P-wave, or both P-wave and S-wave, are incident on the SPR sensor, and the beat signal, which is the difference frequency between these two light waves, is observed. Furthermore, the beat signal from the laser light source itself, which does not pass through the sensor, is also monitored, and the phase difference between these two types of beat signals is measured by phase-sensitive detection. The resulting signal will correspond to the anomalous dispersion curve in Figure 5. [Effects of the Invention]

[0008] With the above configuration, it becomes possible to achieve previously unattainable high sensitivity and high precision with SPR sensors. [Prior art documents]

[0009] [Non-Patent Document 1] Hiroyuki Iwasaka; NEW GLASS Vol.24 No.1 (2009) 35. [Example 1]

[0010] (1) Figure 6 shows an example using a wavelength-stabilized He-Ne transverse Zeeman laser 10. Although this laser output is a single longitudinal mode, the Zeeman effect results in orthogonal linear polarization with slightly different frequencies (approximately several hundred kHz to 2 MHz). One linear polarization is used as the P-polarized wave 12, and the other as the S-polarized wave 13. By inserting a linear polarizer 11 at a 45-degree angle to the reflected light from the sensor, a beat signal between the P-wave and S-wave is obtained. A beat signal is also obtained from the laser light source itself. By detecting the phase difference between these two types of beat signals using a phase difference meter 14 or other so-called phase-sensitive detector, measurement information can be obtained. [Example 2]

[0011] (2) Figure 7 shows an example using a wavelength-stabilized He-Ne longitudinal Zeeman laser 15. Although this laser output is a single longitudinal mode, the Zeeman effect results in clockwise and counterclockwise circularly polarized light with slightly different frequencies. This output light is passed through a quarter-wave plate 16 to orthogonal linear polarization to generate p-polarized and s-polarized waves, which are input to a surface plasmon resonance sensor. The light reflected from the sensor is passed through a 45° linear polarizer 11 to extract the beat signal between the P and S polarized waves. Separately from this pair of light waves, left and right circularly polarized light is extracted via an unpolarized light divider 17 placed immediately after the laser light source exit, then orthogonal linear polarization is performed using the quarter-wave plate 16, and the beat signal is detected by the 45° linear polarizer 11. The phase difference between these two beat signals becomes the SPR measurement signal, as shown in Figure 6, and measurement information can be obtained with high sensitivity and accuracy. In this case, the laser light source is smaller than that of a transverse Zeeman laser, which can contribute to the miniaturization of the device. [Example 3]

[0012] (3) Figure 8 shows an example using a wavelength-stabilized He-Ne2 longitudinal mode laser 18. It is known that adjacent longitudinal modes of this laser are affected by the in-plane anisotropy of the laser mirror and are orthogonal to each other in their linear polarization. One of the two sets of S-waves and P-waves, which are divided into two by an unpolarized light divider 17 placed immediately after the output port, is reflected by the SPR sensor and its beat frequency is detected. The other set does not pass through the SPR sensor. When the phase difference between these two beat frequencies is detected by a double-balanced mixer (DBM) 19 or the like, the output becomes an SPR measurement signal, as in the cases of Figures 6 and 7, and measurement information can be obtained with high sensitivity and accuracy. In this case, the beat frequency is the longitudinal mode interval frequency, so the frequency difference is usually on the order of several hundred MHz to 2 GHz, and high-frequency circuit elements such as a DBM (double-balanced mixer) 19 are required for phase-sensitive detection. [Example 4]

[0013] (4) Figure 9 shows an example using a dual-comb laser. A dual-comb laser consists of two optical frequency combs, a Signal comb 20 and a Local comb 21. An optical frequency comb can be described as a comb-shaped optical ruler with an absolute frequency scale for the longitudinal mode interval frequency, for example, in a mode-locked laser, by stabilizing the carrier envelope offset frequency and longitudinal mode interval frequency with high precision, from DC to the optical frequency domain. The polarization direction of the output light of the Signal comb 20 is angle-adjusted with a half-wave plate 22 and incident on the sensor section as a P-polarized wave, entering the photodetector 2. On the other hand, the comb tooth frequency of the Local comb 21 is slightly shifted from that of the Signal comb. The output light of the Local comb 21 enters the photodetector without passing through the sensor section. Many beat frequencies between the two combs are generated in the photodetector, but a BPF 23 (band-pass filter) is configured to allow only the nearest beat signal to pass through, and this is used as the input to a double-balanced mixer 19 (DBM). On the other hand, the Signal comb and Local comb, which do not pass through the SPR sensor section, are similarly introduced to the input of the double-balanced mixer 19 (DBM). As a result, the output of the double-balanced mixer 19 (DBM), i.e., the phase interferogram, which is synchronized with the pulses generated by the combs, records detailed phase changes with respect to the SPR resonance signal. [Explanation of symbols]

[0014] 1 prism 2 gold thin film 3. Laser light 4. Evanescent waves 5. Surface plasmon waves 6. Reflected laser light 7 Ligands 8 Flow System 9 Analyte 10 Wavelength-stabilized transverse Zeeman laser 11 45° linear polarizer 12 P polarized wave 13 S-polarized waves 14 Phase difference meter 15 Wavelength-stabilized He-Ne longitudinal Zeeman laser 16 λ / 4 plate 17 Unpolarized optical splitter 18 Wavelength-stabilized He-Ne dual longitudinal mode laser 19 Double balanced mixer (DBM) 20 Signal comb 21 Local comb 22 λ / 2 plate 23 BPF

Brief Description of the Drawings

[0015] [Figure 1] Diagram showing the principle of a surface plasmon resonance sensor. [Figure 2] Principle diagram of surface plasmon resonance due to antigen-antibody reaction on the surface of a gold thin film. [Figure 3] Diagram showing resonance angle shift dependent on antigen-antibody reaction. [Figure 4] State of resonance angle shift when the incident angle of p-polarized light wave is constant. [Figure 5] An example of theoretical analysis results of reflectance and p-wave phase change near the SPR resonance angle. [Figure 6] Configuration example of a surface plasmon resonance sensor using a wavelength-stabilized He-Ne transverse Zeeman laser. [Figure 7] Configuration example of a surface plasmon resonance sensor using a wavelength-stabilized He-Ne longitudinal Zeeman laser. [Figure 8] Configuration example of a surface plasmon resonance sensor using two longitudinally mode-stabilized He-Ne lasers. [Figure 9] Configuration example of a surface plasmon resonance sensor using a dual-comb laser

Claims

1. A surface plasmon resonance sensor device that uses an optical heterodyne method as a measurement method, employing a laser light source in which the output light of a wavelength-stabilized He-Ne laser consists of two orthogonal linearly polarized beams, and the difference frequency between these two linearly polarized beams is between 10 kHz and 2 GHz.

2. A surface plasmon resonance sensor device that uses an optical heterodyne method as a measurement method, with a laser light source whose output light from a wavelength-stabilized He-Ne laser consists of left-handed and right-handed circularly polarized light, and the difference frequency between these two circularly polarized light waves is from 100 kHz to 100 MHz.

3. A surface plasmon resonance sensor device that uses two optical frequency combs as light sources for a sensor, each having a fixed difference in the longitudinal mode interval frequency between the two optical frequency combs between 100 kHz and 1 GHz, and obtains measurement information from the change in intensity or phase of the beat signal obtained from the difference frequency between these optical frequency combs.

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