JP3460923B2 - Surface plasmon sensor - Google Patents

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a surface plasmon sensor for quantitatively analyzing a substance in a sample by utilizing the generation of surface plasmons, and more particularly to a coupler means for a light beam and a metal film as a sensor. The present invention relates to a surface plasmon sensor configured separately.

[0002]

2. Description of the Related Art In a metal, free electrons oscillate collectively to generate compression waves called plasma waves. And, the quantized compression wave generated on the metal surface is
It is called surface plasmon.

Conventionally, various surface plasmon sensors have been proposed for quantitatively analyzing substances in a sample by utilizing the phenomenon that the surface plasmons are excited by light waves. And as one of the best known of them,
An example is one using a system called the Kretschmann arrangement (see, for example, JP-A-6-167443).

A surface plasmon sensor using the above system is basically a prism, a metal film formed on one surface of the prism and brought into contact with a sample, a light source for generating a light beam, and the light beam for the prism. Through the optical system so that various incident angles can be obtained with respect to the interface between the prism and the metal film, and the intensity of the light beam totally reflected at the interface can be detected at various incident angles. And a detecting means.

In order to obtain various incident angles as described above, a so-called goniometer (see, for example, JP-A-6-50882) for rotating the irradiation system of the light beam is used, or various light beams are used. An optical system is used in which a relatively thick light beam is incident so as to be focused at the interface so that a component incident at an angle is included. In the former case, a light beam whose reflection angle changes with the deflection of the light beam can be detected by a small photodetector that moves synchronously with the deflection of the light beam, or by an area sensor extending along the direction of change of the reflection angle. Can be detected. On the other hand, in the latter case, each light beam reflected at various reflection angles can be detected by an area sensor extending in a direction in which all the light beams can be received.

When a light beam is incident on a metal film at an incident angle θ which is greater than the total reflection angle, a "bleeding wave" called an evanescent wave is generated in the metal film on the reflecting surface. This evanescent wave has an electric field distribution in the sample in contact with the metal film, and surface plasmons are generated at the interface between the metal film and the sample. When the wave number vector of the evanescent wave generated when the p-polarized light beam is incident on the metal film is equal to the wave number vector of the surface plasmon described above and the wave number matching is established, both are in a resonance state and the energy of the light becomes the surface plasmon. The plasmon is excited by the transition. At this time, the intensity of the light totally reflected due to the transfer of light energy is significantly reduced.

Therefore, in the surface plasmon sensor, the light beam incident on the metal film at various incident angles θ is reflected by measuring the intensity of the light beam totally reflected by the metal film. An incident angle θ _sp (total reflection elimination angle) when the phenomenon of a remarkable decrease in intensity occurs is obtained. This total reflection elimination angle θ _sp and the wave vector K _{1 of} incident light are obtained.
Therefore, the resonance wave number K _sp is derived from the relationship of K _sp = K ₁ sin θ _sp . When the wave number K _{sp of the} surface plasmon is known, the dielectric constant of the sample can be obtained. That is, where the angular frequency of the surface plasmon is ω, the speed of light in vacuum is c, and the permittivities of the metal and the sample are ε _m and ε _s , respectively, the following relationships are established.

[0008]

[Equation 1]

If the permittivity ε _s of the sample is known, the concentration of the specific substance in the sample can be known based on a predetermined calibration curve, etc., so that the total reflection elimination angle θ _{sp at which the} reflected light intensity decreases in the end.
By knowing, the specific substance in the sample can be quantitatively analyzed.

[0010]

In the surface plasmon sensor described above, an example in which a metal film as a sensor is directly formed on one surface of a prism which is a coupler of a light beam has been described. For the above reasons, a metal film is formed on one surface of a transparent substrate such as glass separately from the coupler (hereinafter, the substrate on which the metal film is formed is also referred to as a sensor), and the other surface of the substrate is closely attached to the coupler. In many cases, it is configured to allow. Conventionally, in the case of such a separate type configuration, in order to remove the influence of reflection and multiple reflection due to the air layer at the joint between the sensor and the coupler, the refractive index matching oil is applied to the joint surface and Are closely attached.

However, in such a structure in which matching oil is applied to the joint surfaces to bring them into close contact with each other, there is a problem that it takes time to attach and detach the sensor. Since the replacement of the sensor is relatively frequent, it is desirable to make the replacement of the sensor easy. Further, it is difficult to make the subtle film thickness of the matching oil between the coupler and the sensor constant, and the non-uniformity of the film thickness caused when the sensor is replaced has a bad influence on the detection accuracy.

The present invention has been made in view of the above circumstances, and an object thereof is to provide a surface plasmon sensor which realizes simplification of sensor mounting and stabilization of the positional relationship between the sensor and the coupler. is there.

[0013]

The surface plasmon sensor of the present invention comprises a transparent substrate having a predetermined refractive index, and
A sensor unit including a metal film disposed on one surface side of the transparent substrate and another surface side of the transparent substrate opposite to the one surface have a refractive index substantially the same as the predetermined refractive index. Coupling means arranged with a matching liquid sandwiched therebetween, which transmits a light beam input from an input portion formed in a part thereof and makes the light beam incident on an interface between the transparent substrate and the metal film. And a coupler means for transmitting the light beam totally reflected by the output part and outputting it from the output part formed in the other part, where the part through which the light beam passes has a refractive index substantially the same as the predetermined refractive index. In the surface plasmon sensor that inputs the light beam from the input unit and detects the intensity of the light beam that is totally reflected at the interface and output from the output unit, the sensor unit includes the transparent substrate and the transparent substrate. A sensor support means the distance between Ppura means to support always be constant, the matching liquid in the gap is characterized in that it is filled.

The sensor support means may be fixed to a part of the coupler means, for example.

Further, regarding the filling of the matching liquid in the space, the matching liquid supply means for supplying the matching liquid in the space and the matching liquid intrude to a position higher than the other surface of the transparent substrate. Possible, provided with a space portion communicating with the interval, by the matching liquid supply means, may be filled with the matching liquid in the interval, or, of the coupler means,
On the side facing the transparent substrate, a liquid reservoir for accumulating the matching liquid is formed, a waterproof wall is provided on the transparent substrate so as to surround the metal film, and the sensor unit is provided on the transparent substrate for the liquid reservoir. The other surface may be supported in a state of being submerged in the matching liquid, and the space may be filled with the matching liquid.

As the input part and the output part of the coupler means, a prism may be provided in the coupler means and the input part and the output part may be formed in the prism, or the input part and the output part may be formed. The part may be formed of a diffraction grating. Alternatively, the coupler means has a convex portion on a side opposite to the side of the coupler means facing the transparent substrate, and one side surface of the convex portion and the other side surface facing the one side surface are transparent plates. The inside of the convex portion may be filled with the refractive index matching liquid, and the one side surface and the other side surface may serve as the input portion and the output portion, respectively.

Here, the term "coupler means" is a general term for means for coupling the light beam incident on the interface with the surface plasmon resonance generation condition.

The transparent substrate of the sensor unit is
Having a predetermined refractive index, consisting of a main transparent substrate and a holding transparent substrate that are in close contact with each other, the metal film is arranged on the main transparent substrate, the holding transparent substrate via the refractive index matching liquid It may be arranged to face the coupler means. Further, in this case, it is desirable that the main transparent substrate and the holding transparent substrate are brought into close contact with each other via a matching liquid having a refractive index substantially the same as the refractive index.

A binding reaction film is provided on the metal film,
A specific substance that reacts with the binding reaction membrane may be detected. Here, the “binding reaction film” and the “specific substance that reacts with the binding reaction film” are, for example, an antigen (antibody) and an antibody (antigen) that cause an antigen-antibody reaction.

[0020]

The surface plasmon sensor of the present invention comprises a sensor unit comprising a transparent substrate and a metal film arranged on one surface side of the transparent substrate and brought into contact with a sample,
Since the sensor support means is provided to support the transparent substrate and the coupler means so that the distance between them is always constant, and the gap between the transparent substrate and the coupler means is filled with the matching liquid, the matching liquid is applied and brought into close contact. The sensor unit can be replaced very easily as compared with the case. Further, it becomes possible to easily make the interval between the sensor unit and the coupler means constant.

[0021]

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, embodiments of the present invention will be described in detail with reference to the drawings. FIG. 1 shows a side surface shape of a surface plasmon sensor which is a first embodiment of the present invention.

As shown in the figure, the basic structure of the surface plasmon sensor is a sensor unit 1 formed with a metal film of gold, silver or the like that is brought into contact with the sample S to be measured, and a sensor that supports this sensor unit 1. A coupler means 10 provided with a support means 15 and arranged through a refractive index matching oil (matching liquid) 5 with respect to the sensor unit 1, generates a light beam, and causes the light beam to enter the coupler means 10. The light source optical means 20 and the light detecting means 30 for measuring the intensity of the light beam emitted from the coupler means 10.

Next, a detailed configuration of each detailed part in the first embodiment of the present invention will be described.

The sensor unit 1 is formed by forming a metal film 3 of gold, silver or the like on a sensor substrate 2 made of glass. When the metal film 3 of gold or the like is formed on the glass substrate 2, chromium is preliminarily arranged on the glass substrate 2 to have a thickness of about 1 nm. This facilitates the formation of the metal film 3,
Moreover, peeling is suppressed. In addition, in the analysis by the surface plasmon sensor, in general, a binding reaction film (antigen (or antibody)) is formed on the metal film 3 and an antigen-antibody reaction that selectively responds to a specific substance is used to The amount of antibody (or antigen) adsorbed physically is measured as a change in the incident angle.

The coupler means 10 comprises a cell 12 made of glass having a recess 11 formed on the side facing the sensor unit 1, and a prism 13 formed on the other surface of the cell 12. The coupler means 10 is provided with a sensor support means (sensor attachment) 15 for supporting the sensor unit 1 in a part thereof, and the sensor attachment 15 ensures that the distance between the sensor substrate 2 and the cell 12 is always constant. The sensor unit 1 is supported by. A refractive index matching oil 5 is filled between the sensor substrate 2 and the coupler means 10. The sensor substrate 2, the coupler means 10 and the refractive index matching oil 5 have substantially the same refractive index.

The light source optical means 20 comprises a light source 21 composed of a semiconductor laser or the like for generating the light beam L, and a collimator lens 22 and a condenser lens 23 for condensing the light beam L and making it incident from one surface of the prism 13. The light beam L emitted from the light source 21 is p-polarized by a polarizer (not shown) and is incident on the prism 13. The focused light beam L is
It includes components that are incident on the interface 4 between the sensor substrate 2 and the metal film 3 at various incident angles θ. The incident angle θ is set to an angle equal to or greater than the critical angle for total reflection, and the light beam L
Are totally reflected at the interface 4.

The light source optical means is composed of a light source for generating a light beam of a small diameter and a goniometer for rotating the light source, and the light source is rotated by the goniometer to make the incident angle of the light beam. You may make it take various angles. Furthermore, instead of using a goniometer, a configuration may be used in which various incident angles are obtained by deflecting a light beam using a galvanometer mirror (Japanese Patent Application No. 109367/8).

Since the reflection angle of the light beam L reflected from the interface 4 changes in accordance with the change of the incident angle, the light detecting means.
As 30, for example, a CCD line sensor in which light receiving elements are arranged in parallel along the direction of change of the reflection angle is used. In addition, a photodiode, a two-divided photodiode described in Japanese Patent Application No. 8-109366, a photodiode array, or the like may be used as the light detecting means.

The sample analysis by the surface plasmon sensor having the above structure will be described below. The sample S to be analyzed is placed in contact with the metal film 3. A light beam L generated by the light source optical means 20 and set to p-polarized light
Is incident on one surface of the prism 13, passes through the prism 13, and is incident on the interface 4. As described above, the condensed and incident light beam L is incident on the interface 4 between the metal film 3 and the sensor substrate 2 at various incident angles θ. The light beam L is totally reflected by the interface 4, passes through the prism 13 again, and is emitted from the other surface of the prism 13. Then, the emitted light beam L is detected by the light detecting means 30.

The photodetection signal output from each light receiving element of the photodetection means 30 includes the intensity I of the totally reflected light beam L at the interface 4
This is shown for each angle of incidence θ on the incident light, and the relationship between the reflected light intensity I and the angle of incidence θ is approximately as shown in FIG.

Here, the light incident at a specific incident angle (total reflection elimination angle) θ _sp excites surface plasmons at the interface between the metal film 3 and the sample S, so that the reflected light intensity I Sharply drops. Therefore, the total reflection canceling angle θ _sp can be known by using the light detection signal S output for each light receiving element of the light detecting means 30, and the specific substance in the sample S based on the value of this total reflection canceling angle θ _sp. Can be quantitatively analyzed.
The reason is as described in detail above.

Next, FIG. 3 shows a second embodiment of the present invention. Detailed description of the same configurations and operating portions as those of the above-described first embodiment will be omitted (the same applies to the following embodiments).

In the second embodiment, pipes 41 and 42 that allow the matching oil 5 to come in and out are arranged in the gap between the sensor unit 1 and the coupler means 10 so as to penetrate the cell 12, and one pipe 41 is , Has an opening at a position higher in the vertical direction than the bottom surface of the substrate 2 to form a space portion into which the matching liquid 5 can penetrate to a position higher than the bottom surface of the substrate 2, and the other pipe 42 is matched. It is inserted into the matching liquid tank 43 for storing the oil 5, and a pump 44 is arranged in the middle thereof, and the matching liquid tank 43 and the pump 44 form a matching liquid supply means.

In the surface plasmon sensor according to this embodiment, the sensor unit 1 is attached to the sensor attachment.
After being supported by 15, the pump 44 is operated to supply the matching oil 5 between the surfaces from the matching liquid tank 43 through the pipe 42. At this time, by supplying the matching oil 5 until the liquid level of the matching oil 5 entering the one pipe 41 is higher than the bottom surface of the substrate 2, the matching oil 5 can be filled between the surfaces without a gap. it can.

A third embodiment of the present invention is shown in FIG. In the sensor unit 1 according to the third embodiment, the metal film 3 is formed on the glass substrate 2, and the waterproof wall 6 is provided so as to surround the metal film 3.

The sensor attachment 15 'provided on the coupler means 10 has a waterproof wall 6 so that the substrate 2 of the sensor unit 1 is immersed in the matching oil 5.
The sensor unit 1 is supported by a part of the above, and the positional relationship of the sensor unit 1 with respect to the coupler means 10 is determined.

In the above embodiment, the case where the coupler means having the prism is used has been described, but the coupler means may not have the prism. For example, as shown as the fourth embodiment in FIG. 5, the diffraction gratings 50 and 51 may be formed in the cell 12 and the light beam L may be input and output through the diffraction gratings 50 and 51. In this case, the light beam L is diffracted by the input diffraction grating 50, is incident on the interface 4 at an angle θ, is totally reflected by the interface 4, is diffracted by the output diffraction grating 51, and is emitted. Then, the emitted light beam L is detected by the light detecting means.
30 to detect.

Further, as shown as a fifth embodiment in FIG. 6, a downward convex portion is formed on the lower surface of the cell 12, and the convex portion 53 is filled with the refractive index matching oil 5,
Form glass windows 54, 55 on the side, and this glass window 54, 55
It is also possible to adopt a configuration in which the light beam L is input and output.
The glass windows 54 and 55 have the same refractive index as that of the sensor substrate 2 and the matching oil 5.
In this case, the portions of the cell 12 other than the glass windows 54 and 55 need not be transparent.

Further, FIG. 7 shows a sixth embodiment. In this embodiment, the sensor unit 1 is the main transparent substrate 10
2 and a sensor section 104 composed of the metal film 103 arranged on the main transparent substrate 102, and a sensor holding section 105 for holding the sensor section 104. The lower surface of the sensor holding portion 105 is composed of a holding transparent substrate 106 having a refractive index similar to that of the main transparent substrate 102 of the sensor portion 104.
The measurement is performed in a state where the transparent substrate 102 of the sensor unit 104 is brought into close contact with the substrate 6 via the matching oil. It is assumed that the binding reaction film is formed on the metal film 103 as in the above embodiment. To replace the sensor, the sensor unit 101 may be replaced, or only the sensor unit 104 may be replaced. The sensor unit 104 has a simple structure, is inexpensive, and can be easily replaced. In addition, a free-form substrate that does not use a substrate of uniform size can be used as the sensor unit 104.

In the coupler means 10, similar to the fifth embodiment, a downward convex portion is formed in a part of the cell 112, and a glass window 114, formed on the side surface of the convex portion 113. 11
A structure that allows the light beam to enter and exit from 5 is used.
At the time of measurement, the cell 112 and the optical system casing 141 described later are used.
The matching oil 5 is filled in the concave portion surrounded by and so that the lower surface of the sensor holding portion 105 is immersed in the matching oil 5.

The light source optical means 20 for emitting the light beam L and making it enter the coupler means 10 is provided with a semiconductor laser 121 and a collimator lens 122, and emits the laser light L as parallel light. It also comprises a mirror 124 for reflecting the laser light L and a condenser lens 125. The light beam L emitted from the light source 123
Is p-polarized by a polarizer (not shown) and is incident on the coupler means 10. The focused light beam L
Contains a component that enters the interface 107 between the transparent substrate 102 and the metal film 103 at various incident angles θ. In addition,
The incident angle θ is set to an angle equal to or greater than the total reflection critical angle so that the light beam L is totally reflected at the interface 107.

A condenser lens 131 is also arranged on the side of the detecting means 30 for detecting the light beam reflected by the interface 107 and output from the window 115 of the coupler means 110, so that the light beam is always detected by the detecting means 30. It is configured to be detected.

The above-mentioned coupler means 10 and light source optical means 20
And the detection means 30 are all arranged on the base 140. The coupler means 10 is attached to the optical system casing 141 formed on the base 140 so as to surround the light source optical means 20 and the like.
Is fixed, and the condenser lens 122 of the light source optical means 20 and the condenser lens 131 on the side of the detection means 30 are supported on the lower surface of the coupler means 10.

On the base 140, an optical system casing 141 is further provided.
A casing 142 is formed so as to surround the sensor unit 102, and a sensor attachment 143 that supports the sensor unit 101 is provided on the upper surface of the casing 142. The sensor unit 10 is attached to the sensor attachment 143 fixed to the housing 142.
1 is fixed and supported so as to maintain a constant distance from the coupler means 10.

Further, in addition to the method of obtaining the total reflection elimination angle θ _sp from the reflection intensities from various incident angles as described above, the reflection intensity at a constant incident angle changes according to the value of the total reflection elimination angle θ _sp. By utilizing this, for example, the incident angle of the light beam is set to a constant angle smaller than the total reflection elimination angle θ _sp ,
It is also possible to obtain the total reflection elimination angle θ _sp based on the reflection intensity at this time.

[Brief description of drawings]

FIG. 1 is a side view of a surface plasmon sensor according to a first embodiment of the present invention.

FIG. 2 is a graph showing a schematic relationship between an incident angle of a light beam on a surface plasmon sensor and a light intensity detected by a light detecting means.

FIG. 3 is a side view of a surface plasmon sensor according to a second embodiment of the present invention.

FIG. 4 is a side view of a surface plasmon sensor according to a third embodiment of the present invention.

FIG. 5 is a side view of a surface plasmon sensor according to a fourth embodiment of the present invention.

FIG. 6 is a side view of a surface plasmon sensor according to a fifth embodiment of the present invention.

FIG. 7 is a side view of a surface plasmon sensor according to a sixth embodiment of the present invention.

[Explanation of symbols]

1 sensor unit 2 sensor board 3 metal film 4 interfaces 5 Refractive index matching oil 10 coupler means 12 cells 13 prism 15 sensor attachment 20 Light source optical means 30 Light detection means L light beam

─────────────────────────────────────────────────── ─── Continuation of the front page (58) Fields investigated (Int.Cl. ⁷ , DB name) G01N 21/00-21/01 G01N 21/17-21/61 PATOLIS EUROPAT (QUESTEL) WPI / L (QUESTEL)

Claims (9)

(57) [Claims] 1. A sensor unit comprising a transparent substrate having a predetermined refractive index and a metal film disposed on one surface side of the transparent substrate; and another sensor unit opposite to the one surface of the transparent substrate. On the surface side, a coupler means arranged with a matching liquid having a refractive index substantially the same as the predetermined refractive index sandwiched, through which a light beam input from an input portion formed in part is transmitted,
The light beam is made incident on the interface between the transparent substrate and the metal film, and the light beam totally reflected at the interface is transmitted and output from the output part formed in the other part. A coupler means having a refractive index substantially equal to a predetermined refractive index, the light beam is input from the input unit, and the intensity of the light beam totally reflected at the interface and output from the output unit is detected. in the surface plasmon sensor, the sensor unit, a distance between the coupler means and the transparent substrate is always supported so as to be constant, the mosquito
A surface plasmon sensor comprising a sensor support means fixed to a part of the puller means, and the matching liquid being filled in the space. 2. A matching liquid supply means for supplying the matching liquid to the gap, and a space portion for allowing the matching liquid to penetrate to a position higher than the other surface of the transparent substrate and communicating with the gap. , by the matching liquid supply means, the surface plasmon sensor of claim 1 Symbol mounting the matching liquid in the gap is characterized in that it is filled. 3. A liquid reservoir for accumulating the matching liquid is formed on a side of the coupler means facing the transparent substrate, and a waterproof wall is provided on the transparent substrate so as to surround the metal film. unit, the surface plasmon sensor of claim 1 Symbol mounting the other surface of the transparent substrate to the liquid reservoir portion is characterized in that it is supported in a state of immersion in the matching liquid. 4. A becomes provided with the coupler means prism, the input unit and the output unit, the surface plasmon sensor according to any one of claims 1 3, characterized in that it is formed on the prism. Wherein said input and said output of said coupler means, the surface plasmon sensor according to any one of claims 1 3, characterized in that it consists of a diffraction grating. 6. The coupler means has a convex portion on the side opposite to the side of the coupler means facing the transparent substrate, and the convex portion is on one side surface and one side surface of the convex portion. The other opposite side surface is formed of a transparent plate, the inside of the convex portion is filled with the refractive index matching liquid, and the one side surface and the other side surface are the input portion and the output portion, respectively. The surface plasmon sensor according to any one of claims 1 to 3, wherein 7. The transparent substrate of the sensor unit comprises a main transparent substrate and a holding transparent substrate which have the predetermined refractive index and are in close contact with each other, and the metal film is arranged on the main transparent substrate, surface plasmon sensor according to any one of claims 1 to 6, wherein the holding transparent substrate is arranged so as to oppose to said coupler means through said refractive index matching liquid. 8. The surface plasmon sensor according to claim 7, wherein the main transparent substrate and the holding transparent substrate are brought into close contact with each other through a matching liquid having a refractive index substantially the same as the refractive index. 9. provided the binding reaction film on the metal film, the surface plasmon sensor according to any one of claims 1 8, characterized in that in order to detect a specific substance binding reaction with the bonding reaction film .