JP5977527B2 - Scanning observation device - Google Patents
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- JP5977527B2 JP5977527B2 JP2012017604A JP2012017604A JP5977527B2 JP 5977527 B2 JP5977527 B2 JP 5977527B2 JP 2012017604 A JP2012017604 A JP 2012017604A JP 2012017604 A JP2012017604 A JP 2012017604A JP 5977527 B2 JP5977527 B2 JP 5977527B2
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- 210000001747 pupil Anatomy 0.000 claims description 166
- 230000003287 optical effect Effects 0.000 claims description 130
- 238000001514 detection method Methods 0.000 claims description 55
- 230000004075 alteration Effects 0.000 claims description 28
- 238000005286 illumination Methods 0.000 claims description 13
- 230000005284 excitation Effects 0.000 description 30
- 239000013256 coordination polymer Substances 0.000 description 10
- 238000010586 diagram Methods 0.000 description 10
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- 230000005684 electric field Effects 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- 230000004888 barrier function Effects 0.000 description 2
- 230000004907 flux Effects 0.000 description 2
- 238000013459 approach Methods 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 239000006059 cover glass Substances 0.000 description 1
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Description
本発明は、標本を走査する走査手段を備えた走査型観察装置に関する。 The present invention relates to a scanning observation apparatus provided with scanning means for scanning a specimen.
観察装置の分野においては、標本上の一定の領域から生じた観察光を効率良く検出するための構成として、検出素子の受光面を観察光が入射する対物レンズや集光レンズ（以降、対物レンズと集光レンズのいずれについても集光レンズと記す）の瞳共役面に配置する構成が知られている。このような構成は、例えば、特許文献１で開示されている。 In the field of observation devices, as a configuration for efficiently detecting observation light generated from a certain region on a specimen, an objective lens or a condenser lens (hereinafter referred to as an objective lens) on which the observation light is incident on the light receiving surface of the detection element And the condensing lens are known to be arranged on the pupil conjugate plane of the condensing lens). Such a configuration is disclosed in Patent Document 1, for example.
特許文献１に開示される走査型顕微鏡は、集光レンズの瞳が光検出器の受光面に投影されるように構成されている。標本から集光レンズに入射する観察光の主光線は、その観察光が生じた標本上の位置に関わらず、集光レンズの瞳位置を通過する。このため、受光面が瞳共役面に配置された特許文献１に開示される走査型顕微鏡によれば、走査位置に関わらず標本からの観察光を効率的に検出することが可能である。 The scanning microscope disclosed in Patent Document 1 is configured such that the pupil of the condenser lens is projected onto the light receiving surface of the photodetector. The principal ray of the observation light incident on the condenser lens from the specimen passes through the pupil position of the condenser lens regardless of the position on the specimen where the observation light is generated. For this reason, according to the scanning microscope disclosed in Patent Document 1 in which the light receiving surface is arranged on the pupil conjugate plane, it is possible to efficiently detect the observation light from the specimen regardless of the scanning position.
ところで、検出素子は、外部からの電場や磁場の遮蔽や検出素子の冷却など様々な役割を担うハウジングに収納されていて、通常はハウジングに形成された、光軸と平行な開口の底に配置されている。 By the way, the detection element is housed in a housing that plays various roles such as shielding an electric field and magnetic field from the outside and cooling the detection element, and is usually arranged at the bottom of the opening formed in the housing and parallel to the optical axis. Has been.
集光レンズの光軸からずれた位置から生じた観察光の主光線、即ち、軸外光の主光線は、光軸に対して傾斜した状態で瞳位置に入射することから、軸外光では、ハウジングが障害物となり、ケラレが生じてしまうことがある。 The principal ray of observation light generated from a position deviated from the optical axis of the condenser lens, that is, the principal ray of off-axis light is incident on the pupil position while being inclined with respect to the optical axis. The housing may become an obstacle and vignetting may occur.
このようなハウジングによるケラレを抑制するためには、瞳投影倍率を高くして入射角を浅くする方法が考えられる。しかしながら、瞳投影倍率が高くなると検出素子の受光面に入射する光束が太くなる。このため、投影倍率を高くしすぎると、観察光が受光面からはみ出してしまう事象や、太くなった光束の周辺部分がハウジングに遮られる事象が生じてしまい、かえって効率が低下してしまう。
以上のような実情を踏まえ、本発明では、検出素子を収納するハウジングによる軸外光のケラレを抑制する技術を提供することを課題とする。
In order to suppress such vignetting due to the housing, a method of increasing the pupil projection magnification and reducing the incident angle can be considered. However, when the pupil projection magnification increases, the luminous flux incident on the light receiving surface of the detection element becomes thicker. For this reason, if the projection magnification is too high, the phenomenon that the observation light protrudes from the light receiving surface and the phenomenon that the peripheral part of the thickened light flux is blocked by the housing occur, and the efficiency is lowered.
Based on the above situation, an object of the present invention is to provide a technique for suppressing vignetting of off-axis light by a housing that houses a detection element.
本発明の第１の態様は、光源から射出された照明光で標本を走査する走査手段と、前記照明光が照射された前記標本で発生する観察光を集光する集光レンズと、前記観察光を検出する検出素子と、前記集光レンズの瞳を前記検出素子の前記受光面と略共役にする瞳リレー光学系と、を含み、前記検出素子は、前記瞳リレー光学系から射出された前記観察光の最軸外の主光線と前記瞳リレー光学系の光軸とが前記受光面と前記瞳リレー光学系との間で交差するように、配置され、前記瞳リレー光学系は、前記最軸外の主光線と前記瞳リレー光学系の光軸とが前記受光面と前記瞳リレー光学系との間で交差するような瞳収差を有する走査型観察装置を提供する。 The first aspect of the present invention includes a scanning unit that scans a specimen with illumination light emitted from a light source, a condensing lens that collects observation light generated by the specimen irradiated with the illumination light, and the observation A detection element that detects light, and a pupil relay optical system that conjugates a pupil of the condenser lens with the light receiving surface of the detection element, wherein the detection element is emitted from the pupil relay optical system The principal ray off the outermost axis of the observation light and the optical axis of the pupil relay optical system are arranged so as to intersect between the light receiving surface and the pupil relay optical system, and the pupil relay optical system is Provided is a scanning observation apparatus having a pupil aberration such that an off-axis principal ray and an optical axis of the pupil relay optical system intersect between the light receiving surface and the pupil relay optical system .
本発明の第２の態様は、光源から射出された照明光で標本を走査する走査手段と、前記照明光が照射された前記標本で発生する観察光を集光する集光レンズと、前記観察光を検出する検出素子と、前記集光レンズの瞳を前記検出素子の前記受光面と略共役にする瞳リレー光学系と、を含み、前記検出素子は、前記瞳リレー光学系から射出された前記観察光の最軸外の主光線と前記瞳リレー光学系の光軸とが前記受光面と前記瞳リレー光学系との間で交差するように、且つ、前記集光レンズの瞳共役位置が前記受光面と前記瞳リレー光学系との間に位置するように、配置されて、前記瞳リレー光学系は、前記最軸外の主光線と前記瞳リレー光学系の光軸とが前記受光面と前記瞳リレー光学系との間で交差するような瞳収差を有する走査型観察装置を提供する。 According to a second aspect of the present invention, there is provided a scanning unit that scans a specimen with illumination light emitted from a light source, a condensing lens that collects observation light generated from the specimen irradiated with the illumination light, and the observation A detection element that detects light, and a pupil relay optical system that conjugates a pupil of the condenser lens with the light receiving surface of the detection element , wherein the detection element is emitted from the pupil relay optical system The principal ray off the most axis of the observation light and the optical axis of the pupil relay optical system intersect between the light receiving surface and the pupil relay optical system, and the pupil conjugate position of the condenser lens is The pupil relay optical system is disposed so as to be located between the light receiving surface and the pupil relay optical system, and the pupil relay optical system has the principal axis off the most axis and the optical axis of the pupil relay optical system as the light receiving surface. Scanning observation apparatus having pupil aberration such that it intersects with the pupil relay optical system To provide.
本発明の第３の態様は、第１の態様または第２の態様に記載の走査型観察装置において、前記走査手段は、照明光路上で、且つ、前記集光レンズの瞳共役位置に配置される走査型観察装置を提供する。 A third aspect of the present invention, in the scanning observation apparatus according to the first aspect or the second state like, said scanning means, in the illumination optical path, and, positioned at the pupil conjugate position of the converging lens A scanning observation apparatus is provided.
本発明の第４の態様は、第１の態様乃至第３の態様のいずれか１つに記載の走査型観察装置において、前記走査型観察装置は、非線形光学顕微鏡である走査型観察装置を提供する。
According to a fourth aspect of the present invention, there is provided the scanning observation device according to any one of the first to third aspects, wherein the scanning observation device is a nonlinear optical microscope. To do.
本発明によれば、検出素子を収納するハウジングによる軸外光のケラレを抑制する技術を提供することができる。 ADVANTAGE OF THE INVENTION According to this invention, the technique which suppresses the vignetting of the off-axis light by the housing which accommodates a detection element can be provided.
図１は、本実施例に係る２光子励起顕微鏡の構成を例示した図である。図１に例示される２光子励起顕微鏡１は、標本Ｓから生じる観察光である蛍光を検出することにより標本Ｓを観察する、スキャナ３を備えた走査観察装置である。 FIG. 1 is a diagram illustrating the configuration of a two-photon excitation microscope according to the present embodiment. A two-photon excitation microscope 1 illustrated in FIG. 1 is a scanning observation apparatus including a scanner 3 that observes a specimen S by detecting fluorescence that is observation light generated from the specimen S.
２光子励起顕微鏡１は、標本Ｓを励起するためのレーザ光を射出するレーザ２と、レーザ２から射出されたレーザ光で標本Ｓを走査する走査手段であるスキャナ３と、瞳リレー光学系４と、レーザ光を反射させて標本Ｓから生じた蛍光を透過させるダイクロイックミラー５と、対物レンズ６と、瞳リレー光学系７と、蛍光を検出する検出素子８ａを備えた検出器８と、を含んでいる。 The two-photon excitation microscope 1 includes a laser 2 that emits laser light for exciting the specimen S, a scanner 3 that is a scanning unit that scans the specimen S with laser light emitted from the laser 2, and a pupil relay optical system 4 A dichroic mirror 5 that reflects laser light and transmits fluorescence generated from the specimen S, an objective lens 6, a pupil relay optical system 7, and a detector 8 that includes a detection element 8a that detects fluorescence. Contains.
スキャナ３は、例えば、ガルバノミラー、ポリゴンミラー、音響光学偏向素子などであり、ダイクロイックミラー５とレーザ２との間の照明光路上で、且つ、対物レンズ６の瞳共役位置に配置されている。瞳リレー光学系４は、レンズ４ａとレンズ４ｂからなり、対物レンズ６の瞳ＰＰがスキャナ３と略共役になるように構成されている。 The scanner 3 is, for example, a galvanometer mirror, a polygon mirror, an acoustooptic deflection element, and the like, and is disposed on the illumination optical path between the dichroic mirror 5 and the laser 2 and at the pupil conjugate position of the objective lens 6. The pupil relay optical system 4 includes a lens 4a and a lens 4b, and is configured so that the pupil PP of the objective lens 6 is substantially conjugate with the scanner 3.
ダイクロイックミラー５は、照明光路と検出光路を分岐させる光路分岐手段である。対物レンズ６は、レーザ光を標本Ｓに照射するとともに、レーザ光が照射された標本Ｓで生じる蛍光を集光する集光レンズである。瞳リレー光学系７は、レンズ７ａとレンズ７ｂからなり、対物レンズ６の瞳ＰＰが検出素子８ａの受光面８ｂと略共役になるように構成されている。 The dichroic mirror 5 is an optical path branching unit that branches the illumination optical path and the detection optical path. The objective lens 6 is a condenser lens that irradiates the sample S with the laser light and collects the fluorescence generated in the sample S irradiated with the laser light. The pupil relay optical system 7 includes a lens 7a and a lens 7b, and is configured such that the pupil PP of the objective lens 6 is substantially conjugate with the light receiving surface 8b of the detection element 8a.
検出器８は、検出素子８ａに加えて、検出素子８ａを収納するハウジング８ｃを含んでいる。外部からの電場や磁場の遮蔽、検出素子８ａの冷却などの為に設けられるハウジング８ｃには、検出素子８ａへ入射する蛍光が通る光軸と平行な開口が形成されている。
なお、検出器８は、例えば冷却式光電子増倍管（フォトマル）であり、検出素子８ａの材質としては、ＧａＡｓＰなどが採用される。
The detector 8 includes a housing 8c that houses the detection element 8a in addition to the detection element 8a. An opening parallel to the optical axis through which the fluorescence incident on the detection element 8a passes is formed in the housing 8c provided for shielding an electric field and a magnetic field from the outside, cooling the detection element 8a, and the like.
The detector 8 is, for example, a cooled photomultiplier tube (photomultiplier), and GaAsP or the like is adopted as the material of the detection element 8a.
検出素子８ａは、検出素子８ａの受光面８ｂの法線と瞳リレー光学系７の光軸とが一致するように、その開口の底に配置されている。また、検出素子８ａは、瞳リレー光学系７から射出された蛍光の最軸外の主光線と瞳リレー光学系７の光軸とが、瞳リレー光学系７と受光面８ｂとの間で交差するように、配置されている。 The detection element 8a is disposed at the bottom of the opening so that the normal line of the light receiving surface 8b of the detection element 8a matches the optical axis of the pupil relay optical system 7. In addition, the detection element 8a is configured such that the principal ray off the most axis of fluorescence emitted from the pupil relay optical system 7 and the optical axis of the pupil relay optical system 7 intersect between the pupil relay optical system 7 and the light receiving surface 8b. To be arranged.
図２は、従来技術に係る走査型観察装置と本実施例に係る走査型観察装置の相違点について説明するための図である。図３は、検出素子の配置と開口効率の関係について例示した図である。なお、図３の縦軸は、瞳リレー光学系から射出された蛍光の瞳位置での軸上の光束の面積に対する軸外の光束の面積の割合である開口効率を示し、図３の横軸は、対物レンズの瞳共役位置Ｐ０からの受光面８ｂのずらし量を示す。なお、ずらし量は、受光面８ｂが瞳共役位置Ｐ０よりも瞳リレー光学系から離れた状態でプラスとなるように定義する。 FIG. 2 is a diagram for explaining the difference between the scanning observation apparatus according to the prior art and the scanning observation apparatus according to the present embodiment. FIG. 3 is a diagram illustrating the relationship between the arrangement of the detection elements and the aperture efficiency. The vertical axis in FIG. 3 shows the aperture efficiency, which is the ratio of the area of the off-axis light beam to the area of the light beam on the axis at the pupil position of the fluorescence emitted from the pupil relay optical system, and the horizontal axis in FIG. Indicates the shift amount of the light receiving surface 8b from the pupil conjugate position P0 of the objective lens. The shift amount is defined so as to be positive when the light receiving surface 8b is further away from the pupil relay optical system than the pupil conjugate position P0.
以下、図２及び図３を参照しながら、走査型観察装置である２光子励起顕微鏡の検出素子８ａの配置とその影響について、さらに詳細に説明する。 Hereinafter, with reference to FIGS. 2 and 3, the arrangement of the detection element 8a of the two-photon excitation microscope, which is a scanning observation apparatus, and the influence thereof will be described in more detail.
図２（ａ）には、従来技術に係る走査型観察装置である２光子励起顕微鏡１００に含まれる検出素子８ａの配置が示されている。図２（ｂ）には、本実施例に係る走査型観察装置である２光子励起顕微鏡１に含まれる検出素子８ａの配置が示されている。２光子励起顕微鏡１００と２光子励起顕微鏡１では、レンズ７ｂと検出器８の間の間隔のみが異なっている。 FIG. 2A shows the arrangement of the detection elements 8a included in the two-photon excitation microscope 100, which is a scanning observation apparatus according to the prior art. FIG. 2B shows the arrangement of the detection elements 8a included in the two-photon excitation microscope 1 that is the scanning observation apparatus according to the present embodiment. In the two-photon excitation microscope 100 and the two-photon excitation microscope 1, only the distance between the lens 7b and the detector 8 is different.
図２（ａ）に例示されるように、従来技術に係る２光子励起顕微鏡１００では、検出素子８ａは、受光面８ｂが瞳共役面ＣＰと一致するように光軸上に配置される。この場合、ハウジング８ｃがなければすべての主光線（例えば、軸上光の主光線Ｌ０、軸外光の主光線Ｌ１、軸外光の主光線Ｌ２）が受光面８ｂの中心に位置する瞳共役位置Ｐ０に入射することになるため、主光線により代表される標本Ｓの各位置から生じた光を、その光束径が検出素子８ａのサイズを超えない限りにおいて検出することが可能である。しかしながら、実際にはハウジング８ｃが存在するため、光軸に対して主光線が傾斜している軸外光では、ハウジング８ｃによるケラレが生じてしまう。
なお、ハウジング８ｃの開口の先端には、通常、防塵、保護のため窓レンズと呼ばれるカバーガラス（平行平板）が配置されている。ここでは線のみの記載としているが、厚みが影響するときは、空気換算長を考慮すればよい。
As illustrated in FIG. 2A, in the two-photon excitation microscope 100 according to the related art, the detection element 8a is disposed on the optical axis so that the light receiving surface 8b coincides with the pupil conjugate plane CP. In this case, if there is no housing 8c, all the principal rays (for example, the principal ray L0 of the on-axis light, the principal ray L1 of the off-axis light, and the principal ray L2 of the off-axis light) are located at the center of the light receiving surface 8b. Since the light enters the position P0, it is possible to detect the light generated from each position of the sample S represented by the principal ray as long as the light beam diameter does not exceed the size of the detection element 8a. However, since the housing 8c actually exists, vignetting by the housing 8c occurs in off-axis light whose principal ray is inclined with respect to the optical axis.
A cover glass (parallel plate) called a window lens is usually disposed at the tip of the opening of the housing 8c for dust prevention and protection. Here, only the lines are described, but when the thickness affects, the air equivalent length may be considered.
具体的には、２光子励起顕微鏡１００では、瞳共役位置Ｐ０からの受光面８ｂのずらし量は０であるので、図３に示されるように、主光線Ｌ０に代表される軸上光では開口効率が１００％であるのに対して、主光線Ｌ１に代表される軸外光では開口効率が５５％程度、主光線Ｌ２に代表されるさらに軸外から生じる軸外光では開口効率が０％となる。
なお、図３の開口効率の導出に当たり、ハウジング８ｃの開口形状が円筒形で、その直径ｂがφ６ｍｍ、開口の深さｃが８．９ｍｍ、受光面８ｂの直径ａがφ５ｍｍ、レンズ７ｂの焦点距離ｆが３５．３ｍｍ、レンズ７ｂの入射面側での最も軸外の光束の主光線の光線高が１１．８ｍｍ、レンズ７ｂの射出面側で軸上光束の光束径がφ３．２ｍｍとして計算した。
Specifically, in the two-photon excitation microscope 100, since the shift amount of the light receiving surface 8b from the pupil conjugate position P0 is 0, as shown in FIG. 3, the axial light represented by the principal ray L0 has an aperture. The efficiency is 100%, whereas the off-axis light represented by the principal ray L1 has an aperture efficiency of about 55%, and the off-axis light represented by the off-axis light represented by the principal ray L2 has an aperture efficiency of 0%. It becomes.
3, the opening shape of the housing 8c is cylindrical, the diameter b is φ6 mm, the opening depth c is 8.9 mm, the diameter a of the light receiving surface 8b is φ5 mm, and the focal point of the lens 7b. Calculation is made assuming that the distance f is 35.3 mm, the principal ray height of the most off-axis light beam on the entrance surface side of the lens 7b is 11.8 mm, and the light beam diameter of the on-axis light beam on the exit surface side of the lens 7b is φ3.2 mm. did.
一方、図２（ｂ）に例示されるように、本実施例に係る２光子励起顕微鏡１では、検出素子８ａは、対物レンズ６の瞳共役位置Ｐ０が受光面８ｂとレンズ７ｂ（瞳リレー光学系７）との間に位置するように、光軸上に配置される。このため、２光子励起顕微鏡１では、ハウジング８ｃに形成された開口の標本側の端部（以降、開口端と記す）において、軸外光の主光線（主光線Ｌ１、主光線Ｌ２）が、走査型観察装置１００の場合に比べて光軸から近い位置を通過することになる。これにより、ハウジング８ｃでのケラレを抑制することができるため、図３に示されるように、受光面８ｃを瞳共役位置Ｐ０からずらすことで、軸外光の開口効率が改善される。 On the other hand, as illustrated in FIG. 2B, in the two-photon excitation microscope 1 according to the present embodiment, the detection element 8a has the pupil conjugate position P0 of the objective lens 6 such that the light receiving surface 8b and the lens 7b (pupil relay optics). It is arranged on the optical axis so as to be located between the system 7). For this reason, in the two-photon excitation microscope 1, the principal ray (principal ray L1, principal ray L2) of off-axis light is generated at the end of the opening formed in the housing 8c on the specimen side (hereinafter referred to as the opening end). Compared to the case of the scanning observation apparatus 100, it passes through a position closer to the optical axis. As a result, vignetting in the housing 8c can be suppressed, and as shown in FIG. 3, the aperture efficiency of off-axis light is improved by shifting the light receiving surface 8c from the pupil conjugate position P0.
なお、瞳共役位置Ｐ０を通過後の軸外光の主光線（主光線Ｌ１、主光線Ｌ２）は、瞳共役位置Ｐ０から離れるほど光軸から広がっていくことになるため、瞳共役位置Ｐ０を開口端に近づけすぎると、瞳共役位置Ｐ０を通過後に主光線が大きく広がってしまい、開口内でケラレが生じてしまう。図３に示されるように、２光子励起顕微鏡１の場合には、受光面８ｃの瞳共役位置Ｐ０からのずらし量としては、４ｍｍから６ｍｍ程度が望ましく、ずらし量を大きくしすぎると、一旦改善された軸外光の開口効率が再び低下してしまう。このため、検出素子８ａの受光面は、瞳共役位置Ｐ０は大きく離れた位置に配置されるべきでなく、瞳共役位置Ｐ０近傍に配置されることが望ましい。
特に、ハウジングの開口の直径ｂに対する深さｃの比ｃ／ｂが１．３から１．６程度のとき、瞳リレー光学系の中間像の後段のレンズ系の焦点距離ｆに対するずらし量ｚの比ｚ/ｆは０．１から０．１８程度であることが望ましい。
Note that the principal rays (principal ray L1 and principal ray L2) of off-axis light after passing through the pupil conjugate position P0 spread from the optical axis as the distance from the pupil conjugate position P0 increases. If it is too close to the aperture end, the chief ray spreads greatly after passing through the pupil conjugate position P0, and vignetting occurs in the aperture. As shown in FIG. 3, in the case of the two-photon excitation microscope 1, the shift amount from the pupil conjugate position P0 of the light receiving surface 8c is preferably about 4 mm to 6 mm, and once the shift amount is increased, it is improved once. The aperture efficiency of the off-axis light that has been generated is reduced again. For this reason, the light receiving surface of the detection element 8a should not be disposed at a position far away from the pupil conjugate position P0, but is desirably disposed in the vicinity of the pupil conjugate position P0.
In particular, when the ratio c / b of the depth c to the diameter b of the housing opening is about 1.3 to 1.6, the shift amount z with respect to the focal length f of the lens system at the rear stage of the intermediate image of the pupil relay optical system The ratio z / f is preferably about 0.1 to 0.18.
このように、瞳共役位置Ｐ０からの受光面８ｃのずらし量ｚには、ケラレの発生を効果的に抑制することができる最適な範囲が存在する。図４は、図１に例示される典型的な開口形状を備えた検出器の拡大図である。図４に例示される検出器８の場合、ケラレの発生を効果的に抑制することができるずらし量ｚの最適な範囲は、以下の式（１）で表される。
ここで、パラメータａは検出素子８ａの受光面径（受光面８ｂの径）であり、パラメータｂはメカ枠径（ハウジング８ｃの開口の径）であり、パラメータｃはメカ枠長（ハウジング８ｃの開口の深さ）である。また、パラメータｈｕは最軸外光の上側光線幅（最軸外光の主光線と上側マージナル光線との間の間隔）であり、パラメータｈｌは、最軸外光の下側光線幅（最軸外光の主光線と下側マージナル光線との間の間隔）であり、パラメータθは、最軸外光の光線入射角度（最軸外光の主光線が瞳共役位置に入射する際の、主光線と光軸とのなす角）である。なお、パラメータａとパラメータｂは、ａ≦ｂの関係にあり、パラメータｃは、開口内に窓レンズが配置されている場合には、空気換算長で表される。 Here, the parameter a is the light receiving surface diameter of the detection element 8a (the diameter of the light receiving surface 8b), the parameter b is the mechanical frame diameter (the diameter of the opening of the housing 8c), and the parameter c is the mechanical frame length (of the housing 8c). The depth of the opening). The parameter hu is the upper ray width of the most off-axis light (the interval between the principal ray of the most off-axis light and the upper marginal ray), and the parameter hl is the lower ray width of the most off-axis light ( The distance between the principal ray of the most off-axis light and the lower marginal ray), and the parameter θ is the ray incidence angle of the most off-axis light (when the principal ray of the most off-axis light is incident on the pupil conjugate position) , The angle between the principal ray and the optical axis). Note that the parameter a and the parameter b have a relationship of a ≦ b, and the parameter c is represented by an air conversion length when a window lens is disposed in the opening.
式（１）は、下記の式（２）から式（５）で算出されるケラレ量が最小化されることを条件に算出することができる。
ここで、ケラレ量ｕａは受光面側での上側ケラレ量（受光面と同一平面における上側マージナル光線と受光面の上端との間の間隔）であり、ケラレ量ｕｂはメカ枠側での上側ケラレ量（開口端と同一平面における上側マージナル光線と上側開口端との間の間隔）であり、ケラレ量ｌａは受光面側での下側ケラレ量（受光面と同一平面における下側マージナル光線と受光面の下端との間の間隔）であり、ケラレ量ｌｂはメカ枠側での下側ケラレ量（開口端と同一平面における下側マージナル光線と下側開口端との間の間隔）である。なお、ケラレ量ｕａ、ケラレ量ｕｂ、ケラレ量ｌａ、ケラレ量ｌｂは、それぞれ、ケラレが生じる場合にはプラスの値として、ケラレが生じない場合にはマイナスの値として算出される。 Here, the shading amount u a is the upper vignetting of the light receiving surface side (distance between the upper end of the light-receiving surface and the upper marginal ray on the light-receiving surface in the same plane), the shading amount u b is at the mechanical frame side amount upper vignetting a (distance between the upper marginal ray and the upper open end at the open end flush), vignetting amount l a is lower marginal in the lower vignetting amount (light receiving surface flush with the light receiving surface side the spacing) between the lower end of the beam and the light receiving surface, the distance between the eclipse amount l b is lower marginal ray and a lower opening end in the lower vignetting amount (opening end flush at the mechanical frame side ). Incidentally, vignetting amount u a, vignetting amount u b, vignetting amount l a, vignetting amount l b, respectively, as positive value when the shading occurs, is calculated as a negative value when the eclipse does not occur .
以上のように構成された本実施例に係る２光子励起顕微鏡１によれば、対物レンズ６の瞳共役位置Ｐ０が、受光面８ｂ近傍で、且つ、受光面８ｂと瞳リレー光学系７との間に位置するように検出素子８ａが配置されることで、ハウジング８ｃによる軸外光のケラレを抑制することができる。 According to the two-photon excitation microscope 1 according to the present embodiment configured as described above, the pupil conjugate position P0 of the objective lens 6 is in the vicinity of the light receiving surface 8b and between the light receiving surface 8b and the pupil relay optical system 7. By disposing the detection element 8a so as to be positioned between them, vignetting of off-axis light by the housing 8c can be suppressed.
図５は、従来技術に係る走査型観察装置と本実施例に係る走査型観察装置の相違点について説明するための図である。図５（ａ）は、従来技術に係る走査型観察装置である２光子励起顕微鏡１００の瞳リレー光学系から検出器８へ向けて射出される光の様子を示している。図５（ｂ）は、本実施例に係る走査型観察装置である２光子励起顕微鏡１０の瞳リレー光学系から検出器８へ向けて射出される光の様子を示している。 FIG. 5 is a diagram for explaining the difference between the scanning observation apparatus according to the prior art and the scanning observation apparatus according to the present embodiment. FIG. 5A shows the state of light emitted from the pupil relay optical system of the two-photon excitation microscope 100, which is a scanning observation apparatus according to the prior art, toward the detector 8. FIG. FIG. 5B shows the state of light emitted toward the detector 8 from the pupil relay optical system of the two-photon excitation microscope 10 which is the scanning observation apparatus according to the present embodiment.
本実施例に係る２光子励起顕微鏡１０は、瞳リレー光学系を構成するレンズ７ｂの代わりにレンズ１７ｂを含む点、及び、受光面８ｂが瞳共役面ＣＰと一致するように検出素子８ａが配置される点が、図１に例示される２光子励起顕微鏡１と異なっている。なお、受光面８ｂが瞳共役面ＣＰと一致するように検出素子８ａが配置される点については、従来技術に係る２光子励起顕微鏡１００と同じである。 In the two-photon excitation microscope 10 according to the present embodiment, the detection element 8a is arranged so that the lens 17b is included instead of the lens 7b constituting the pupil relay optical system, and the light receiving surface 8b coincides with the pupil conjugate plane CP. This is different from the two-photon excitation microscope 1 illustrated in FIG. The detection element 8a is arranged so that the light receiving surface 8b coincides with the pupil conjugate plane CP, which is the same as the two-photon excitation microscope 100 according to the related art.
図５（ｂ）に例示されるレンズ１７ｂと図示しないレンズ７ａとで構成される瞳リレー光学系は、最軸外の主光線とその瞳リレー光学系の光軸とが受光面８ｂと瞳リレー光学系との間で交差するような瞳収差を有している。さらに、その瞳収差は、最軸外において最も大きな球面収差であり、光軸に近づくにつれて球面収差が小さくなるような収差である。 In the pupil relay optical system including the lens 17b illustrated in FIG. 5B and the lens 7a (not shown), the principal ray outside the most axis and the optical axis of the pupil relay optical system are the light receiving surface 8b and the pupil relay. It has pupil aberration that intersects with the optical system. Further, the pupil aberration is the largest spherical aberration off the most axis, and the spherical aberration becomes smaller as it approaches the optical axis.
このため、瞳収差を有する瞳リレー光学系を備えた２光子励起顕微鏡１０では、主光線が光軸と交わる位置が主光線毎に異なることになり、光軸からより離れた軸外光の主光線ほど瞳共役位置Ｐ０から離れた位置で光軸と交わることになる。なお、瞳位置や瞳共役位置は、一般に、近軸光線によって定義される。 For this reason, in the two-photon excitation microscope 10 having a pupil relay optical system having pupil aberration, the position at which the principal ray intersects the optical axis differs for each principal ray, and the principal light of off-axis light that is further away from the optical axis. The light beam intersects the optical axis at a position away from the pupil conjugate position P0. Note that the pupil position and the pupil conjugate position are generally defined by paraxial rays.
図６は、異なる瞳収差を有する瞳リレー光学系の各々から射出される蛍光の様子を例示した図である。図７は、図６に例示される瞳リレー光学系の各々の球面収差図である。図８は、図６に例示される瞳リレー光学系の各々を用いた場合における、検出素子の配置と開口効率の関係について例示した図である。なお、図８の縦軸は、瞳リレー光学系から射出された蛍光に対する検出素子８ａで検出される蛍光の割合である開口効率を示し、図８の横軸は、対物レンズの瞳共役位置Ｐ０からの受光面８ｂのずらし量を示す。なお、ずらし量は、受光面８ｂが瞳共役位置Ｐ０よりも瞳リレー光学系から離れた状態でプラスとなるように定義する。 FIG. 6 is a diagram illustrating the state of fluorescence emitted from each of the pupil relay optical systems having different pupil aberrations. FIG. 7 is a spherical aberration diagram of each of the pupil relay optical systems illustrated in FIG. FIG. 8 is a diagram illustrating the relationship between the arrangement of the detection elements and the aperture efficiency when each of the pupil relay optical systems illustrated in FIG. 6 is used. The vertical axis in FIG. 8 indicates the aperture efficiency, which is the ratio of the fluorescence detected by the detection element 8a to the fluorescence emitted from the pupil relay optical system, and the horizontal axis in FIG. 8 indicates the pupil conjugate position P0 of the objective lens. The shift amount of the light-receiving surface 8b from is shown. The shift amount is defined so as to be positive when the light receiving surface 8b is further away from the pupil relay optical system than the pupil conjugate position P0.
図６（ａ）に例示されるレンズ１８ｂを含む瞳リレー光学系は、図７（ａ）に示されるように最軸外で２．５ｍｍ程度の瞳収差（球面収差）を有する瞳リレー光学系であり、レンズ１８ｂを含む瞳リレー光学系を用いた場合の開口効率が図８（ａ）に示されている。図６（ｂ）に例示されるレンズ１７ｂを含む瞳リレー光学系は、図７（ｂ）に示されるように最軸外で５ｍｍ程度の瞳収差（球面収差）を有する瞳リレー光学系であり、レンズ１７ｂを含む瞳リレー光学系を用いた場合の開口効率が図８（ｂ）に示されている。図６（ｃ）に例示されるレンズ１９ｂを含む瞳リレー光学系は、図７（ｃ）に示されるように最軸外で８ｍｍ程度の瞳収差（球面収差）を有する瞳リレー光学系であり、レンズ１９ｂを含む瞳リレー光学系を用いた場合の開口効率が図８（ｃ）に示されている。
ここで、図８の開口効率の導出に当たり、ハウジングの開口は図３の場合と同形状とし、図６（ａ）のレンズ１８ｂ、図６（ｂ）のレンズ１７ｂ、図６（ｃ）のレンズ１９ｂの入射側の面の曲率半径Ｒ１と射出側の面の曲率半径Ｒ２の比（Ｒ１：Ｒ２）は、それぞれ２６．９：∞、５０：５４．６、１００：３５．６として計算した。
The pupil relay optical system including the lens 18b illustrated in FIG. 6A has a pupil aberration (spherical aberration) of about 2.5 mm off the most axis as shown in FIG. 7A. FIG. 8A shows the aperture efficiency when the pupil relay optical system including the lens 18b is used. The pupil relay optical system including the lens 17b illustrated in FIG. 6B is a pupil relay optical system having a pupil aberration (spherical aberration) of about 5 mm off the most axis as shown in FIG. 7B. FIG. 8B shows the aperture efficiency when a pupil relay optical system including the lens 17b is used. The pupil relay optical system including the lens 19b illustrated in FIG. 6C is a pupil relay optical system having a pupil aberration (spherical aberration) of about 8 mm off the most axis as shown in FIG. 7C. FIG. 8C shows the aperture efficiency when a pupil relay optical system including the lens 19b is used.
Here, in deriving the aperture efficiency of FIG. 8, the aperture of the housing has the same shape as in FIG. 3, and the lens 18b of FIG. 6A, the lens 17b of FIG. 6B, and the lens of FIG. 6C. The ratio (R1: R2) of the curvature radius R1 of the incident side surface 19b and the curvature radius R2 of the exit side surface of 19b was calculated as 26.9: ∞, 50: 54.6, and 100: 35.6, respectively.
図８（ｂ）に示されるように、レンズ１７ｂを含む瞳リレー光学系を用いた場合には、受光面８ｂを瞳共役面ＣＰと一致させた状態で、ハウジング８ｃによる軸外光のケラレを抑制して、軸外光に対する高い開口効率を実現している。一方で、図８（ａ）及び図８（ｃ）に示されるように、レンズ１８ｂ、レンズ１９ｂを含む瞳リレー光学系を用いた場合には、それぞれ、瞳収差が不足、過剰な状態にあるため、受光面８ｂを瞳共役面ＣＰと一致させた状態では、特に主光線Ｌ２に代表される軸外光のケラレが大きい。 As shown in FIG. 8B, when a pupil relay optical system including the lens 17b is used, vignetting of off-axis light by the housing 8c is performed with the light receiving surface 8b aligned with the pupil conjugate plane CP. Suppressing and realizing high aperture efficiency for off-axis light. On the other hand, as shown in FIGS. 8A and 8C, when the pupil relay optical system including the lens 18b and the lens 19b is used, the pupil aberration is insufficient and excessive, respectively. Therefore, in the state where the light receiving surface 8b coincides with the pupil conjugate plane CP, the vignetting of off-axis light represented by the principal ray L2 is particularly large.
このように、瞳リレー光学系の瞳収差を利用することによっても、瞳共役位置ＣＰを受光面８ｂに対してずらす場合と同様の効果を得ることが可能である。このため、ある範囲内の瞳収差を利用することで、受光面８ｂを瞳共役位置ＣＰに一致させた状態であっても、ハウジング８ｃでのケラレを抑制して、軸外光の開口効率を改善することができる。 As described above, by using the pupil aberration of the pupil relay optical system, it is possible to obtain the same effect as the case where the pupil conjugate position CP is shifted with respect to the light receiving surface 8b. For this reason, by using pupil aberration within a certain range, vignetting in the housing 8c is suppressed and the aperture efficiency of off-axis light is improved even when the light-receiving surface 8b is aligned with the pupil conjugate position CP. Can be improved.
以上のように構成された本実施例に係る２光子励起顕微鏡１０によれば、対物レンズ６の瞳共役位置Ｐ０が受光面８ｂ近傍（受光面８ｂと一致した状態を含む）に配置され、且つ、最軸外の主光線と瞳リレー光学系の光軸が受光面８ｂと瞳リレー光学系との間で交差するような瞳収差を瞳リレー光学系が有することで、実施例１に係る２光子励起顕微鏡１と同様に、ハウジング８ｃによる軸外光のケラレを抑制することができる。 According to the two-photon excitation microscope 10 according to the present embodiment configured as described above, the pupil conjugate position P0 of the objective lens 6 is disposed in the vicinity of the light receiving surface 8b (including a state in which it coincides with the light receiving surface 8b), and The pupil relay optical system has a pupil aberration such that the off-axis principal ray and the optical axis of the pupil relay optical system intersect between the light receiving surface 8b and the pupil relay optical system. Similar to the photon excitation microscope 1, vignetting of off-axis light by the housing 8c can be suppressed.
なお、瞳リレー光学系の瞳収差１ｍｍ分の変動は、受光面８ｂのずらし量１ｍｍ分の変動と同様に作用する。このため、ケラレの発生を効果的に抑制することができる瞳収差の最適な範囲は、実施例１と同様に、式（１）を用いて算出することができる。 Note that the fluctuation of 1 mm of pupil aberration of the pupil relay optical system acts similarly to the fluctuation of 1 mm of the shift amount of the light receiving surface 8b. For this reason, the optimal range of pupil aberration that can effectively suppress the occurrence of vignetting can be calculated using the equation (1), as in the first embodiment.
また、実施例１では、瞳共役位置ＣＰを受光面８ｂに対してずらすことにより、実施例２では、瞳リレー光学系が瞳収差を有することにより、ハウジング８ｃでのケラレを抑制する例を示したが、これらを組み合わせて用いてもよい。例えば、受光面８ｂを瞳共役位置ＣＰに対して５ｍｍずらした状態でケラレを最も効果的に抑制することができる場合には、例えば、最軸外の瞳収差が２ｍｍの瞳リレー光学系を用いた上で、受光面８ｂを３ｍｍだけずらすことで、ケラレを抑制してもよい。 In the first embodiment, the pupil conjugate position CP is shifted with respect to the light receiving surface 8b. In the second embodiment, the pupil relay optical system has pupil aberration, thereby suppressing vignetting in the housing 8c. However, these may be used in combination. For example, when vignetting can be most effectively suppressed with the light receiving surface 8b shifted by 5 mm with respect to the pupil conjugate position CP, for example, a pupil relay optical system having an off-axis pupil aberration of 2 mm is used. In addition, vignetting may be suppressed by shifting the light receiving surface 8b by 3 mm.
図９、本実施例に係る２光子励起顕微鏡の構成を例示した図である。図９に例示される２光子励起顕微鏡２０は、標本Ｓから生じる蛍光を観察光として検出することにより標本Ｓを観察する、スキャナ２２を備えた走査観察装置である。 FIG. 9 is a diagram illustrating the configuration of a two-photon excitation microscope according to the present embodiment. A two-photon excitation microscope 20 illustrated in FIG. 9 is a scanning observation apparatus including a scanner 22 that observes the specimen S by detecting fluorescence generated from the specimen S as observation light.
２光子励起顕微鏡２０は、標本Ｓを励起するためのレーザ光を射出するレーザ２１と、レーザ２１から射出されたレーザ光で標本Ｓを走査する走査手段であるスキャナ２２と、瞳リレー光学系２３と、レーザ光を標本Ｓに照射するレンズ２４と、標本Ｓを挟んでレンズ２４と向かい合わせに配置されたレンズ２５と、瞳リレー光学系２６と、レーザ光を遮断し蛍光を透過させるバリアフィルタ２７と、蛍光を検出する検出素子２８ａを備えた検出器２８と、を含んでいる。 The two-photon excitation microscope 20 includes a laser 21 that emits laser light for exciting the specimen S, a scanner 22 that is a scanning unit that scans the specimen S with the laser light emitted from the laser 21, and a pupil relay optical system 23. A lens 24 that irradiates the sample S with laser light, a lens 25 that is disposed facing the lens 24 across the sample S, a pupil relay optical system 26, and a barrier filter that blocks the laser light and transmits fluorescence. 27 and a detector 28 having a detection element 28a for detecting fluorescence.
スキャナ２２は、例えば、ガルバノミラー、ポリゴンミラー、音響光学偏向素子などであり、標本Ｓとレーザ２１との間の照明光路上で、且つ、レンズ２５の瞳共役位置に配置されている。瞳リレー光学系２３は、レンズ２３ａとレンズ２３ｂからなり、レンズ２５の瞳ＰＰと光学的に共役な位置に形成されるレンズ２４の瞳がスキャナ２２と共役になるように構成されている。 The scanner 22 is, for example, a galvano mirror, a polygon mirror, an acousto-optic deflection element, and the like, and is disposed on the illumination optical path between the sample S and the laser 21 and at the pupil conjugate position of the lens 25. The pupil relay optical system 23 includes a lens 23 a and a lens 23 b, and is configured such that the pupil of the lens 24 formed at a position optically conjugate with the pupil PP of the lens 25 is conjugate with the scanner 22.
レンズ２５は、レーザ光が照射された標本Ｓで生じる蛍光を集光する集光レンズである。瞳リレー光学系２６は、レンズ２６ａとレンズ２６ｂからなり、レンズ２５の瞳ＰＰが検出素子２８ａの受光面２８ｂと略共役になるように構成されている。 The lens 25 is a condensing lens that condenses the fluorescence generated in the sample S irradiated with the laser light. The pupil relay optical system 26 includes a lens 26a and a lens 26b, and is configured such that the pupil PP of the lens 25 is substantially conjugate with the light receiving surface 28b of the detection element 28a.
検出器２８は、検出素子２８ａに加えて、検出素子２８ａを収納するハウジング２８ｃを含んでいる。外部からの電場や磁場の遮蔽、検出素子２８ａの冷却などの為に設けられるハウジング２８ｃには、検出素子２８ａへ入射する蛍光が通る光軸と平行な開口が形成されている。 The detector 28 includes a housing 28c that houses the detection element 28a in addition to the detection element 28a. An opening parallel to the optical axis through which the fluorescence incident on the detection element 28a passes is formed in the housing 28c provided for shielding an electric field and a magnetic field from the outside and cooling the detection element 28a.
検出素子２８ａは、検出素子２８ａの受光面２８ｂの法線と瞳リレー光学系２６の光軸とが一致するように、その開口の底に配置されている。また、検出素子２８ａは、瞳リレー光学系２６から射出された蛍光の最軸外の主光線と瞳リレー光学系２６の光軸とが、瞳リレー光学系２６と受光面２８ｂとの間で交差するように、配置されている。 The detection element 28a is arranged at the bottom of the opening so that the normal line of the light receiving surface 28b of the detection element 28a and the optical axis of the pupil relay optical system 26 coincide. Further, in the detection element 28a, the principal ray off the most axis of fluorescence emitted from the pupil relay optical system 26 and the optical axis of the pupil relay optical system 26 intersect between the pupil relay optical system 26 and the light receiving surface 28b. To be arranged.
より具体的には、実施例１と同様に、検出素子２８ａがレンズ２５の瞳共役位置が受光面２８ｂとレンズ２６ｂとの間に位置するように配置されてもよく、実施例２と同様に、瞳リレー光学系２６が最軸外の主光線とその瞳リレー光学系の光軸とが受光面８ｂと瞳リレー光学系との間で交差するような瞳収差を有していてもよい。 More specifically, similarly to the first embodiment, the detection element 28a may be arranged so that the pupil conjugate position of the lens 25 is located between the light receiving surface 28b and the lens 26b. The pupil relay optical system 26 may have a pupil aberration such that the off-axis principal ray and the optical axis of the pupil relay optical system intersect between the light receiving surface 8b and the pupil relay optical system.
以上のように構成された本実施例に係る２光子励起顕微鏡２０によっても、実施例１及び実施例２に係る２光子励起顕微鏡と同様に、ハウジング２８ｃによる軸外光のケラレを抑制することができる。 The two-photon excitation microscope 20 according to the present embodiment configured as described above can also suppress vignetting of off-axis light by the housing 28c, similarly to the two-photon excitation microscope according to the first and second embodiments. it can.
実施例１から実施例３では、走査型観察装置として２光子励起顕微鏡を例示したが、本願発明が適用される走査型観察装置は、２光子励起顕微鏡に限られない。軸外からの光を効率的に検出することが求められる走査型観察装置であればよく、例えば、第２高調波（ＳＨＧ）顕微鏡などの蛍光顕微鏡以外の顕微鏡であってもよい。 In the first to third embodiments, the two-photon excitation microscope is illustrated as the scanning observation apparatus. However, the scanning observation apparatus to which the present invention is applied is not limited to the two-photon excitation microscope. Any scanning observation apparatus that is required to efficiently detect off-axis light, for example, a microscope other than a fluorescence microscope such as a second harmonic (SHG) microscope may be used.
なお、非線形光学現象を利用した非線形光学顕微鏡の場合、光は一点からのみ生じるが標本中での散乱などにより広がって標本から射出される。このため、これらの散乱した光を無駄なく取り込む必要がある非線形光学顕微鏡は、本願発明の適用対象として、特に好適である。 In the case of a non-linear optical microscope using a non-linear optical phenomenon, light is generated only from one point, but is spread out due to scattering in the sample and emitted from the sample. For this reason, the nonlinear optical microscope which needs to take in these scattered light without waste is especially suitable as an application object of this invention.
１、１０、２０、１００・・・２光子励起顕微鏡
２、２１・・・レーザ
３、２２・・・スキャナ
４、２３・・・瞳リレー光学系
４ａ、４ｂ、７ａ、７ｂ、１７ｂ、１８ｂ、１９ｂ、２３ａ、２３ｂ、２４、２５、２６ａ、２６ｂ・・・レンズ
５・・・ダイクロイックミラー
６・・・対物レンズ
７、２６・・・瞳リレー光学系
８、２８・・・検出器
８ａ、２８ａ・・・検出素子
８ｂ、２８ｂ・・・受光面
８ｃ、２８ｃ・・・ハウジング
２７・・・バリアフィルタ
Ｐ０・・・瞳共役位置
ＰＰ・・・瞳面
ＣＰ・・・瞳共役面
Ｌ０、Ｌ１、Ｌ２・・・主光線
Ｓ・・・標本
1, 10, 20, 100... 2 photon excitation microscope 2, 21... Laser 3, 22, scanner 4, 23, pupil relay optical system 4 a, 4 b, 7 a, 7 b, 17 b, 18 b, 19b, 23a, 23b, 24, 25, 26a, 26b ... lens 5 ... dichroic mirror 6 ... objective lens 7, 26 ... pupil relay optical system 8, 28 ... detectors 8a, 28a ... detecting elements 8b, 28b ... light receiving surfaces 8c, 28c ... housing 27 ... barrier filter P0 ... pupil conjugate position PP ... pupil plane CP ... pupil conjugate planes L0, L1, L2 ... chief ray S ... specimen
Claims (4)
前記照明光が照射された前記標本で発生する観察光を集光する集光レンズと、
前記観察光を検出する検出素子と、
前記集光レンズの瞳を前記検出素子の前記受光面と略共役にする瞳リレー光学系と、を含み、
前記検出素子は、前記瞳リレー光学系から射出された前記観察光の最軸外の主光線と前記瞳リレー光学系の光軸とが前記受光面と前記瞳リレー光学系との間で交差するように、
配置され、
前記瞳リレー光学系は、前記最軸外の主光線と前記瞳リレー光学系の光軸とが前記受光面と前記瞳リレー光学系との間で交差するような瞳収差を有する
ことを特徴とする走査型観察装置。 Scanning means for scanning the specimen with illumination light emitted from a light source;
A condensing lens that condenses the observation light generated in the specimen irradiated with the illumination light;
A detection element for detecting the observation light;
A pupil relay optical system that makes the pupil of the condenser lens substantially conjugate with the light receiving surface of the detection element;
In the detection element, the principal ray off the most axis of the observation light emitted from the pupil relay optical system and the optical axis of the pupil relay optical system intersect between the light receiving surface and the pupil relay optical system. like,
Arranged ,
The pupil relay optical system has a pupil aberration such that the off-axis principal ray and the optical axis of the pupil relay optical system intersect between the light receiving surface and the pupil relay optical system. A scanning observation apparatus characterized by that.
前記照明光が照射された前記標本で発生する観察光を集光する集光レンズと、
前記観察光を検出する検出素子と、
前記集光レンズの瞳を前記検出素子の前記受光面と略共役にする瞳リレー光学系と、を含み、
前記検出素子は、前記瞳リレー光学系から射出された前記観察光の最軸外の主光線と前記瞳リレー光学系の光軸とが前記受光面と前記瞳リレー光学系との間で交差するように、且つ、前記集光レンズの瞳共役位置が前記受光面と前記瞳リレー光学系との間に位置するように、配置されて、
前記瞳リレー光学系は、前記最軸外の主光線と前記瞳リレー光学系の光軸とが前記受光面と前記瞳リレー光学系との間で交差するような瞳収差を有する
ことを特徴とする走査型観察装置。 Scanning means for scanning the specimen with illumination light emitted from a light source;
A condensing lens that condenses the observation light generated in the specimen irradiated with the illumination light;
A detection element for detecting the observation light;
A pupil relay optical system that makes the pupil of the condenser lens substantially conjugate with the light receiving surface of the detection element;
In the detection element, the principal ray off the most axis of the observation light emitted from the pupil relay optical system and the optical axis of the pupil relay optical system intersect between the light receiving surface and the pupil relay optical system. And the pupil conjugate position of the condenser lens is disposed between the light receiving surface and the pupil relay optical system ,
The pupil relay optical system has a pupil aberration such that the off-axis principal ray and the optical axis of the pupil relay optical system intersect between the light receiving surface and the pupil relay optical system. A scanning observation apparatus characterized by that.
前記走査手段は、照明光路上で、且つ、前記集光レンズの瞳共役位置に配置される
ことを特徴とする走査型観察装置。 The scanning observation apparatus according to claim 1 or 2 ,
The scanning observation apparatus, wherein the scanning unit is arranged on an illumination optical path and at a pupil conjugate position of the condenser lens.
前記走査型観察装置は、非線形光学顕微鏡である
ことを特徴とする走査型観察装置。 The scanning observation apparatus according to any one of claims 1 to 3 ,
The scanning observation apparatus is a nonlinear optical microscope.
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