JP6343203B2 - Optical multiple reflection measuring apparatus and optical multiple reflection measuring method - Google Patents

Optical multiple reflection measuring apparatus and optical multiple reflection measuring method Download PDF

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JP6343203B2
JP6343203B2 JP2014165612A JP2014165612A JP6343203B2 JP 6343203 B2 JP6343203 B2 JP 6343203B2 JP 2014165612 A JP2014165612 A JP 2014165612A JP 2014165612 A JP2014165612 A JP 2014165612A JP 6343203 B2 JP6343203 B2 JP 6343203B2
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宏和 阿部
宏和 阿部
靖典 吉富
靖典 吉富
宗弘 岩倉
宗弘 岩倉
彰秀 辺見
彰秀 辺見
田中　敬二
敬二 田中
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Description

本発明は、高速測定可能な光学多重反射測定装置および光学多重反射測定方法に関する。   The present invention relates to an optical multiple reflection measurement apparatus and an optical multiple reflection measurement method capable of high-speed measurement.

表面プラズモン共鳴（Ｓｕｒｆａｃｅ Ｐｌａｓｍｏｎ Ｒｅｓｏｎａｎｃｅ 以下、「ＳＰＲ」という）を利用したＳＰＲセンサーは、分子間の相互作用をリアルタイムでモニタリングできる点に大きなメリットがあり、医薬生化学領域や、食品・環境分析の分野において幅広く利用されている。   The SPR sensor using surface plasmon resonance (hereinafter referred to as “SPR”) has a great merit in that the interaction between molecules can be monitored in real time, and the field of pharmaceutical biochemistry and food / environment analysis. Widely used in

表面プラズモンは、金属表面を伝播するプラズマ波であり、表面プラズモン共鳴は、この表面プラズモンと光波との共鳴によって生じる現象である。光によって表面プラズモンを生じさせるためには、伝播速度が遅い光であるエバネッセント波が用いられる。エバネッセント波は、光が全反射するときに、反射面の反対側に生じる表面波である。   Surface plasmon is a plasma wave propagating on a metal surface, and surface plasmon resonance is a phenomenon caused by resonance between this surface plasmon and a light wave. In order to generate surface plasmons by light, evanescent waves, which are light having a low propagation speed, are used. The evanescent wave is a surface wave that is generated on the opposite side of the reflecting surface when light is totally reflected.

ＳＰＲセンサーで多く用いられる光学系は、金属薄膜がコーティングされたプリズムを用いており、このプリズム表面に光を照射した際に、金属薄膜を通りぬけてその反対側にしみ出したエバネッセント波が金属表面の表面プラズモンを励起する。表面プラズモン共鳴が起こると光のエネルギーは表面プラズモンに移り、反射光強度は減衰するため、光の入射角に対して反射光強度をプロットすると、反射光のある特定の角度で反射光強度が減衰する。この角度をＳＰＲ角度といい、ＳＰＲ角度は、金属薄膜表面近傍の媒質の屈折率に依存して変動する。計測対象となる分子（アナライト）を認識するための分子（リガンド）をセンサーチップ上に固定化し、これにアナライトを含む試料を注入すると、リガンドとアナライトの結合に伴って、センサーチップ表面の質量が増大する。これにより、センサーチップ表面の屈折率が増大し、ＳＰＲ角度が変化する。このＳＰＲ角度の変化を検出することにより、センサーチップ表面における分子間相互作用をリアルタイムに計測することができる。   The optical system often used in the SPR sensor uses a prism coated with a metal thin film. When light is applied to the surface of the prism, the evanescent wave that has passed through the metal thin film and oozed out to the other side is a metal. Excites surface plasmons on the surface. When surface plasmon resonance occurs, the energy of light shifts to surface plasmon and the reflected light intensity attenuates. Therefore, when the reflected light intensity is plotted against the incident angle of light, the reflected light intensity is attenuated at a specific angle of the reflected light. To do. This angle is referred to as an SPR angle, and the SPR angle varies depending on the refractive index of the medium in the vicinity of the metal thin film surface. When a molecule (ligand) for recognizing the molecule to be measured (analyte) is immobilized on the sensor chip and a sample containing the analyte is injected into this, the surface of the sensor chip is accompanied by the binding between the ligand and the analyte. The mass of increases. As a result, the refractive index of the sensor chip surface increases and the SPR angle changes. By detecting the change in the SPR angle, the intermolecular interaction on the surface of the sensor chip can be measured in real time.

図２に、従来の表面プラズモン共鳴の測定系の一例を示す。図２において、光源１から照射された光は、分析対象試料３が搭載されたプリズム２によって反射され、受光素子４に受光されるが、入射光の入射角に応じて反射光の受光角度も機械的に合わせる必要があり、この操作を行うためには、ゴニオメータ等の複雑な機構が必要となる。そのため、装置が複雑で大型化するという問題点がある。また、入射角を繰り返し変化させるために、光源１と受光素子４をそれぞれ図２に示すように往復運動させると、光源１と受光素子４は加速、定速、減速を繰り返すことになるため、高速化が困難であり、連続で観測する場合、高速測定を行うことができなかった。そのため、高分子材料等の濡れ性変化等のように、ミリ秒単位やそれ以下で性状が変化するものを正確に評価することができなかった。   FIG. 2 shows an example of a conventional surface plasmon resonance measurement system. In FIG. 2, the light emitted from the light source 1 is reflected by the prism 2 on which the sample 3 to be analyzed is mounted and is received by the light receiving element 4, but the light receiving angle of the reflected light also depends on the incident angle of the incident light. In order to perform this operation, a complicated mechanism such as a goniometer is required. Therefore, there is a problem that the apparatus is complicated and large. Further, when the light source 1 and the light receiving element 4 are reciprocated as shown in FIG. 2 in order to repeatedly change the incident angle, the light source 1 and the light receiving element 4 repeat acceleration, constant speed, and deceleration. High-speed measurement is difficult, and high-speed measurement cannot be performed when continuously observing. For this reason, it has not been possible to accurately evaluate a material whose property changes in units of milliseconds or less, such as a change in wettability of a polymer material or the like.

特許文献１には、可変光照射装置に微小回動装置を連結し、微小回動装置の駆動に伴って可変光照射装置を所要の角度にて往復回動させて、センサーチップに対する光の入射角度を可変する表面プラズモン共鳴角検出装置が記載されている。   In Patent Document 1, a micro-rotation device is connected to a variable light irradiation device, and the variable light irradiation device is reciprocally rotated at a predetermined angle as the micro-rotation device is driven, so that light is incident on a sensor chip. A surface plasmon resonance angle detection device with variable angle is described.

特開平１１−２７１２１５号公報JP-A-11-271215

本発明は、このような問題点を解決するためになされたもので、高速で性状が変化するものを測定対象としても、正確に高速測定を行うことが可能な光学多重反射測定装置と光学多重反射測定方法を提供することを目的とする。   The present invention has been made to solve such a problem, and an optical multiple reflection measuring apparatus and an optical multiplexing capable of accurately performing high-speed measurement even when a subject whose properties change at high speed are to be measured. An object is to provide a reflection measurement method.

以上の課題を解決するために、本発明の光学多重反射測定装置は、センサーチップが配置され分析対象試料を搭載したプリズムに対して光を照射する光源と、前記プリズムによって反射された光を受光する受光素子と、前記プリズムへの入射光の焦点を中心として前記光源を連続的に回転させる機構部とを備え、前記光源の回転に伴って前記プリズムへの入射光の入射角を変化させて、前記プリズムからの反射光の強度を測定することを特徴とする。   In order to solve the above-described problems, the optical multiple reflection measuring apparatus of the present invention receives a light source that irradiates light to a prism on which a sensor chip is arranged and a sample to be analyzed is mounted, and light reflected by the prism. And a mechanism for continuously rotating the light source around the focal point of the incident light on the prism, and changing the incident angle of the incident light on the prism as the light source rotates. The intensity of reflected light from the prism is measured.

また、本発明の光学多重反射測定方法は、センサーチップが配置され分析対象試料を搭載したプリズムに対して光を照射する光源を、前記プリズムへの入射光の焦点を中心として連続的に回転して前記プリズムへの入射光の入射角を変化させ、前記プリズムによって反射された光を受光素子で受光して前記プリズムからの反射光の強度を測定することを特徴とする。   In the optical multiple reflection measurement method of the present invention, a light source that irradiates light to a prism on which a sensor chip is arranged and a sample to be analyzed is continuously rotated about a focal point of incident light on the prism. The incident angle of the incident light to the prism is changed, the light reflected by the prism is received by a light receiving element, and the intensity of the reflected light from the prism is measured.

プリズムへの入射光の焦点を中心として光源を回転させることにより、光源を往復運動させるときのように光源を減速させる必要がなく、一定速度で高速に動かすことができる。そのため、分析対象試料を搭載したプリズムに対して、高速で入射角を変化させて光を投入することができる。これにより、分析対象試料の表面の過渡変化をミリ秒オーダーで観測することが可能となる。   By rotating the light source around the focal point of the incident light to the prism, it is not necessary to decelerate the light source as in the case of reciprocating the light source, and can be moved at a high speed at a constant speed. Therefore, it is possible to inject light with changing the incident angle at a high speed to the prism on which the sample to be analyzed is mounted. Thereby, it becomes possible to observe the transient change of the surface of the sample to be analyzed on the order of milliseconds.

本発明の光学多重反射測定装置においては、前記受光素子は、設置位置を変えることなく反射光を受光することが可能な受光面積を有する大面積型フォトダイオードである。
また、本発明の光学多重反射測定方法においては、前記受光素子は、設置位置を変えることなく反射光を受光することが可能な受光面積を有する大面積型フォトダイオードである。 In the optical multiple reflection measuring apparatus of the present invention, the light receiving element is a large area photodiode having a light receiving area capable of receiving reflected light without changing an installation position.
In the optical multiple reflection measurement method of the present invention, the light receiving element is a large-area photodiode having a light receiving area capable of receiving reflected light without changing an installation position.

受光素子として大面積型フォトダイオードを用いることにより、一つの受光素子で設置位置や角度を変えることなく反射光を受光することができ、角度による光強度の補正をすることで、受光側を固定して表面プラズモンの共鳴角を観測することができる。   By using a large-area photodiode as the light receiving element, reflected light can be received by one light receiving element without changing the installation position and angle, and the light receiving side is fixed by correcting the light intensity according to the angle. Thus, the resonance angle of the surface plasmon can be observed.

また、入射光の焦点を中心として光源を回転させるため、プリズムへの入射光の入射角を大きな範囲で変化させることができる。そのため、表面プラズモン共鳴が起こる全反射角よりも小さい入射角での反射光強度も高速で計測することができる。   Further, since the light source is rotated around the focal point of the incident light, the incident angle of the incident light to the prism can be changed within a large range. Therefore, the reflected light intensity at an incident angle smaller than the total reflection angle at which surface plasmon resonance occurs can be measured at high speed.

本発明によると、高速で性状が変化するものを測定対象としても、正確に高速測定を行うことが可能な光学多重反射測定装置と光学多重反射測定方法を実現することができる。   According to the present invention, it is possible to realize an optical multiple reflection measurement apparatus and an optical multiple reflection measurement method capable of accurately performing high-speed measurement even when an object whose properties change at high speed is a measurement target.

本発明の実施形態に係る光学多重反射測定装置を示す図である。It is a figure which shows the optical multiple reflection measuring apparatus which concerns on embodiment of this invention. 従来の表面プラズモン共鳴の測定系の一例を示す図である。It is a figure which shows an example of the measuring system of the conventional surface plasmon resonance.

以下に、本発明の光学多重反射測定装置と光学多重反射測定方法を、その実施形態に基づいて説明する。
図１に、本発明の実施形態に係る光学多重反射測定装置を示す。
光学多重反射測定装置は、分析対象試料３を搭載したプリズム２に対して光を照射する光源１と、プリズム２によって反射された光を受光する受光素子４を備えている。光源１は機構部によって、プリズム２への入射光の焦点を中心として、連続的に回転する。光源１の回転により、プリズム２への入射光の入射角を変化させて、プリズム２からの反射光の強度を測定する。断面が半円状のプリズム２の平面側上部にはセンサーチップ６が配置されており、センサーチップ６上に分析対象試料３が搭載されている。分析対象試料３上には気体液体試料用管７が配置される。光源１からプリズム２に入射した光は、センサーチップ６が配置され分析対象試料３が搭載されている、プリズム２の平面側上部にて反射して、反射光が受光素子４に受光される。 Hereinafter, an optical multiple reflection measurement apparatus and an optical multiple reflection measurement method of the present invention will be described based on the embodiments.
FIG. 1 shows an optical multiple reflection measuring apparatus according to an embodiment of the present invention.
The optical multiple reflection measuring apparatus includes a light source 1 that irradiates light to a prism 2 on which an analysis target sample 3 is mounted, and a light receiving element 4 that receives light reflected by the prism 2. The light source 1 is continuously rotated around the focal point of the incident light to the prism 2 by the mechanism. The intensity of the reflected light from the prism 2 is measured by changing the incident angle of the incident light on the prism 2 by the rotation of the light source 1. A sensor chip 6 is disposed on the upper surface of the prism 2 having a semicircular cross section, and the sample 3 to be analyzed is mounted on the sensor chip 6. A gas liquid sample tube 7 is disposed on the sample 3 to be analyzed. The light incident on the prism 2 from the light source 1 is reflected by the upper part on the plane side of the prism 2 where the sensor chip 6 is disposed and the sample 3 to be analyzed is mounted, and the reflected light is received by the light receiving element 4.

センサーチップ６は、数十ｎｍの厚さの金属膜をガラス等の光透過体に形成させた板状のものであり、金属膜として通常はＡｕ膜が使用される。センサーチップ６はプリズム２と別部材として形成してもよく、樹脂成型によりプリズム２と一体として形成することもできる。このセンサーチップ６の金属表面側に分析対象試料３の薄膜を設置してその密度を測定する。センサーチップ６はセンサーセルで固定される。   The sensor chip 6 has a plate shape in which a metal film with a thickness of several tens of nm is formed on a light transmitting body such as glass, and an Au film is usually used as the metal film. The sensor chip 6 may be formed as a separate member from the prism 2, or may be formed integrally with the prism 2 by resin molding. A thin film of the sample 3 to be analyzed is placed on the metal surface side of the sensor chip 6 and its density is measured. The sensor chip 6 is fixed by a sensor cell.

光源１として、特定波長のレーザー光源等を用い、その波長特性を有する受光素子４を組み合わせて使用する。受光素子４は所定の角度で固定する。光源１側の光学系の回転は、機構部に備えられたモータにより行い、光源１側の光学系を所定の速度で一方向へ連続回転させ、回転速度を安定させる。   A laser light source having a specific wavelength is used as the light source 1, and a light receiving element 4 having the wavelength characteristic is used in combination. The light receiving element 4 is fixed at a predetermined angle. The optical system on the light source 1 side is rotated by a motor provided in the mechanism unit, and the optical system on the light source 1 side is continuously rotated in one direction at a predetermined speed to stabilize the rotation speed.

受光素子４として、設置位置を変えることなく反射光を受光することが可能な受光面積を有する大面積型フォトダイオードを用いており、一つの受光素子４で設置位置や角度を変えることなく反射光を受光することができ、角度による光強度の補正をすることで、受光側を固定して表面プラズモンの共鳴角を観測することができる。   As the light receiving element 4, a large-area photodiode having a light receiving area capable of receiving reflected light without changing the installation position is used, and reflected light without changing the installation position or angle by one light receiving element 4. By correcting the light intensity according to the angle, the light receiving side can be fixed and the surface plasmon resonance angle can be observed.

分析対象試料３を囲むフローセル５を用いることにより、温度、圧力、雰囲気ガス等の環境を制御することが可能である。そのため、これらの環境変化による材料の変化も観測することができ、従来観測できなかった材料特性を調べることができる。ピペットによる手動操作またはポンプ等の送液装置により各種の液体や気体を接触させ、その前後のデータを連続して取得する。   By using the flow cell 5 surrounding the sample 3 to be analyzed, it is possible to control the environment such as temperature, pressure, and atmospheric gas. Therefore, changes in materials due to these environmental changes can also be observed, and material properties that could not be observed in the past can be investigated. Various liquids and gases are brought into contact with each other by manual operation using a pipette or a liquid delivery device such as a pump, and data before and after that are continuously acquired.

測定結果のデータ処理は、測定値とリファレンス値との差分を採るため、光源１側の光学系を通してセンサーチップ６の裏面へ照射した光の反射光と、リファレンス光の差分をとってレーザー光そのものの変動を相殺し、分析対象試料３の変性による応答の変化を捉える。   Since the data processing of the measurement result takes the difference between the measurement value and the reference value, the difference between the reflected light of the light irradiated to the back surface of the sensor chip 6 through the optical system on the light source 1 side and the reference light is taken to obtain the laser light itself. The change in the response due to the denaturation of the sample 3 to be analyzed is captured.

プリズム２に対する入射光の投入角度に応じて反射光も変化するが、大面積フォトダイオードである受光素子４は所定の角度で固定されているため、受光素子４への入射角が大きくなるにつれて受光強度が低減する。このため、受光角度に応じて補正するプログラムを備えることにより、反射角による受光強度の補正を行う。   Although the reflected light also changes according to the incident angle of incident light to the prism 2, the light receiving element 4, which is a large area photodiode, is fixed at a predetermined angle, and therefore receives light as the incident angle to the light receiving element 4 increases. Strength is reduced. For this reason, the correction of the received light intensity by the reflection angle is performed by providing a program for correcting according to the received light angle.

上述した測定方法は、単層膜に対して適用されるばかりでなく、多層膜に対してもスネル法則を利用することにより各膜の密度を知ることができるため、多層膜に対しても適用することができ、単層膜、多層膜の区別なく適用対象を広く設定できる。   The measurement method described above is not only applied to single-layer films, but can also be applied to multilayer films because the density of each film can be determined by using Snell's law for multilayer films. The application object can be set widely without distinction of a single layer film and a multilayer film.

本発明においては、光源１を機構部により回転させることにより、光源１を一定速度で高速に動かすことが可能である。そのため、分析対象試料３を搭載したプリズム２に対して、高速で入射角を変化させて光を投入することができ、分析対象試料３の表面の過渡変化をミリ秒オーダーで観測することが可能となる。このようにして、表面プラズモン共鳴の高速な変化を捉えることができるため、高分子材料等の濡れ性の変化など、ミリ秒単位以下で性状が変化するものの評価に用いることができる。具体的な測定対象として、印刷物（紙、インク、塗料等）、タイヤゴム、化粧品等を挙げることができる。   In the present invention, it is possible to move the light source 1 at a constant speed and at a high speed by rotating the light source 1 by the mechanism. Therefore, it is possible to inject light with changing the incident angle at a high speed to the prism 2 on which the analysis target sample 3 is mounted, and it is possible to observe a transient change in the surface of the analysis target sample 3 in the order of milliseconds. It becomes. In this way, since a high-speed change in surface plasmon resonance can be captured, it can be used to evaluate changes in properties in units of milliseconds or less, such as a change in wettability of a polymer material or the like. Specific examples of the measurement target include printed matter (paper, ink, paint, etc.), tire rubber, cosmetics, and the like.

また、プリズム２への入射光の焦点を中心として光源１を回転させるため、プリズム２への入射光の入射角を大きな範囲で変化させることができ、表面プラズモン共鳴が起こる全反射角よりも小さい入射角での反射光強度も高速で計測することができる。そのため、表面プラズモン共鳴に限らず、反射光強度の計測から得られる分析対象試料３の情報を広く採取することが可能な光学多重反射測定装置と光学多重反射測定方法として機能する。   Further, since the light source 1 is rotated around the focal point of the incident light to the prism 2, the incident angle of the incident light to the prism 2 can be changed in a large range, and is smaller than the total reflection angle at which surface plasmon resonance occurs. The reflected light intensity at the incident angle can also be measured at high speed. Therefore, it functions not only as surface plasmon resonance but also as an optical multiple reflection measurement apparatus and an optical multiple reflection measurement method capable of widely collecting information on the sample 3 to be analyzed obtained from the measurement of reflected light intensity.

本発明は、高速で性状が変化するものを測定対象としても、正確に高速測定を行うことが可能な光学多重反射測定装置と光学多重反射測定方法として広く利用することができる。   INDUSTRIAL APPLICABILITY The present invention can be widely used as an optical multiple reflection measurement apparatus and an optical multiple reflection measurement method capable of accurately performing high-speed measurement even when a subject whose properties change at high speed is a measurement target.

１ 光源
２ プリズム
３ 分析対象試料
４ 受光素子
５ フローセル
６ センサーチップ
７ 気体液体試料用管 DESCRIPTION OF SYMBOLS 1 Light source 2 Prism 3 Sample to be analyzed 4 Light receiving element 5 Flow cell 6 Sensor chip 7 Gas liquid sample tube

Claims (4)

センサーチップが配置され分析対象試料を搭載したプリズムに対して光を照射する光源と、前記プリズムによって反射された光を受光する受光素子と、前記プリズムへの入射光の焦点を中心として前記光源を連続的に回転させる機構部とを備え、前記光源の回転に伴って、前記光源を減速させずに一定速度で動かすことにより、前記プリズムへの入射光の入射角を変化させて、前記プリズムからの反射光の強度を測定することを特徴とする光学多重反射測定装置。 A light source that irradiates light to a prism on which a sensor chip is mounted and on which a sample to be analyzed is mounted; a light receiving element that receives light reflected by the prism; and A mechanism unit that continuously rotates, and by rotating the light source at a constant speed without decelerating, the incident angle of the incident light to the prism is changed, An optical multiple reflection measuring apparatus characterized by measuring the intensity of reflected light. 前記受光素子は、設置位置を変えることなく反射光を受光することが可能な受光面積を有する大面積型フォトダイオードであることを特徴とする請求項１記載の光学多重反射測定装置。   2. The optical multiple reflection measuring apparatus according to claim 1, wherein the light receiving element is a large area type photodiode having a light receiving area capable of receiving reflected light without changing an installation position. センサーチップが配置され分析対象試料を搭載したプリズムに対して光を照射する光源を、前記プリズムへの入射光の焦点を中心として連続的に回転して、前記光源を減速させずに一定速度で動かすことにより、前記プリズムへの入射光の入射角を変化させ、前記プリズムによって反射された光を受光素子で受光して前記プリズムからの反射光の強度を測定することを特徴とする光学多重反射測定方法。 A light source that irradiates light to a prism on which a sensor chip is mounted and on which a sample to be analyzed is mounted is continuously rotated around the focal point of incident light on the prism, and the light source is not decelerated at a constant speed. The optical multiple reflection is characterized in that the incident angle of the incident light to the prism is changed by moving, the light reflected by the prism is received by a light receiving element, and the intensity of the reflected light from the prism is measured. Measuring method. 前記受光素子は、設置位置を変えることなく反射光を受光することが可能な受光面積を有する大面積型フォトダイオードであることを特徴とする請求項３記載の光学多重反射測定方法。   4. The optical multiple reflection measurement method according to claim 3, wherein the light receiving element is a large area type photodiode having a light receiving area capable of receiving reflected light without changing an installation position.
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