JPH03131811A - Confocal scanning type transmission microscope - Google Patents

Abstract

PURPOSE:To eliminate the need for the scanning mechanism of a photodetector and to simplify the structure by scanning a sample in two dimensions and detecting light which is transmitted through the sample while the photodetector is fixed. CONSTITUTION:Luminous flux which is reflected by a corner cube prism 23 is deflected by an optical deflecting means 14 in exactly the same way with illumination light 11 and the light passed through this optical deflecting means 14 travels along the same optical path with illumination light 11 before deflection. Luminous flux 11' branching from the illumination optical path is made incident on an invariably constant position without reference to the deflection of the illumination light 11, so the fixed photodetector 27 detects the luminous flux. Thus, the majority of the optical system which converges the luminous flux transmitted through the sample 20 and guides it to the photodetector 27 is common to a sent light optical system and only an optical branching means such as a half-mirror 13 and a condenser lens 25 need to be provided separately. Consequently, the constitution of the optical system is simplified.

Description

DETAILED DESCRIPTION OF THE INVENTION (Field of Industrial Application) The present invention relates to a confocal scanning microscope, and more particularly to a transmission type confocal scanning microscope that detects light transmitted through a sample.

(Prior Art) Conventionally, illumination light is converged into a minute light spot, and this light spot is scanned two-dimensionally on a sample, and at that time, the light that has passed through the sample or the light that has been reflected there is detected by a photodetector. Optical scanning microscopes are known that detect electrical signals that carry an enlarged image of a sample.

In particular, it is configured to generate illumination light from a light source and image it into a light spot on the sample, and then re-image the light flux from the sample into a point image, which is then detected by a photodetector. A confocal scanning microscope does not require a pinhole on the sample surface, making it easy to implement.

This confocal scanning microscope basically includes a light source that emits illumination light, a sample stage on which a sample is placed, a light transmission optical system that images this illumination light as a minute light spot on the sample, and the above-mentioned. It consists of a light receiving optical system that condenses the light beam from the sample and forms it into a point image, a photodetector that detects this point image, and a scanning mechanism that scans the light spot two-dimensionally on the sample. It is something that

There are two types of confocal scanning microscopes: a reflective type that detects light reflected by a sample, and a transmission type that detects light that has passed through the sample. Incidentally, Japanese Patent Laid-Open Nos. 82-209510 and 1982-308414 disclose an example of this transmission type confocal scanning microscope.

(Problems to be Solved by the Invention) In conventional confocal scanning microscopes, the above-mentioned scanning mechanism includes: ■ a mechanism that moves the sample stage two-dimensionally, or ■ a mechanism that moves the illumination light beam two-dimensionally using an optical deflector. A deflection mechanism was used.

However, when the mechanism (2) was adopted, there was a problem that the sample would fly away when high-speed scanning was performed.

On the other hand, although the mechanism (2) allows sufficiently high-speed scanning, another problem arises when this mechanism is applied to the above-mentioned transmission type confocal scanning microscope.

That is, in that case, since the illumination light is deflected, the imaging position of the light beam transmitted through the sample also changes accordingly, so it is necessary to scan the photodetector in synchronization with the deflection of the illumination light. Providing such a scanning mechanism for the photodetector makes the structure of the confocal scanning microscope extremely complicated.

In view of these circumstances, for example, the above-mentioned Japanese Patent Application Laid-Open No. 62-209
As shown in Publication No. 510, the back surface of the vibrating mirror that deflects the illumination light is also a reflective surface, and the light beam that has passed through the sample is guided to this back surface, and is reflected and deflected so as to cancel out the deflection of the illumination light. There have also been proposals to eliminate the need for photodetector scanning.

However, in such a mechanism, a large number of mirrors and the like are required to guide the light beam that has passed through the sample to the back surface of the vibrating mirror, making the optical system extremely complicated and making its assembly and adjustment extremely difficult.

The present invention has been made in view of the above-mentioned circumstances, and an object of the present invention is to provide a transmission type confocal scanning microscope that does not require scanning a photodetector and has a simple optical system configuration. This is the purpose.

(Means for Solving the Problems) A transmission type confocal scanning microscope according to the present invention includes the above-mentioned sample stage, light source, light transmission optical system, photodetector, and two light points. In a confocal scanning microscope equipped with a dimensional scanning mechanism, the two-dimensional scanning mechanism uses a light deflecting means that deflects the illumination light, and is arranged at a position where the light beam transmitted through the sample is incident, so that the light beam is a corner cube prism that reflects the light along the same path as its incident path, and a light branching device such as a half mirror that branches the reflected light beam from the optical path of the illumination light at a position closer to the light source than the light deflection means. and a condensing lens for condensing the branched light beam to form a point image, and a photodetector was arranged to detect the point image formed by the condensing lens. It is characterized by this.

(Function) In the above configuration, the light beam reflected by the corner cube prism is deflected by the light deflection means in exactly the same way as the illumination light, so after passing through this light deflection means, it is the same as the illumination light before deflection. They will proceed along the same optical path. Therefore, the light beam branched from this illumination light optical path always enters a fixed position regardless of the polarization of the illumination light, so that the light beam can be detected by a fixed photodetector.

Most of the light receiving optical system that condenses the light beam that has passed through the sample and guides it to the photodetector is shared with the light transmitting optical system, and only the light branching means such as the above-mentioned half mirror and the condensing lens are separately provided. The configuration of the optical system becomes extremely simple.

(Example) Hereinafter, the present invention will be described in detail based on an example shown in the drawings.

The figure shows a transmission type confocal scanning microscope according to an embodiment of the present invention. Laser light source 1 as shown
Laser beam (illumination light) which is parallel light emitted from 0
The beam diameter of the beam 11 is expanded by a beam expander 12 , passes through a half mirror 13 , and enters an AOD (acousto-optic optical deflector) 14 . The illumination light 11 is this A
The beam is deflected by the OD 14 in a direction substantially perpendicular to the plane of the paper, passes through a relay lens 15 for aberration correction, and enters a vibrating mirror 16.

The vibrating mirror 16 swings in the direction of arrow A in the figure, thereby deflecting the illumination light 11 in a direction substantially perpendicular to the direction of deflection described above.

This deflected illumination light 11 passes through the relay lens 17 and enters the objective lens 18, and the objective lens 18
A minute light spot P on the sample 20 (on the surface or inside)
image is formed. The sample stage 19 on which the sample 2° is placed is moved by a moving mechanism 21 in the direction of arrow Z, that is, in the optical axis direction of the objective lens 18.

A beam of transmitted light 11' that has passed through the sample 20 is converted into parallel light by an objective lens 22, and then enters a corner cube prism 23. The transmitted light 11' is reflected by the corner cube prism 23 so as to follow the same path of incidence thereon, focused again on the sample 2o by the objective lens 22, and converted into parallel light by the objective lens 18. This transmitted light 11' which has been made into parallel light
, relay lens 17, vibrating mirror 1B,! JL/-L
/ lens 15, AOD 14, and the illumination light 11, it travels in the opposite direction along the common optical path and enters the half mirror 13. The transmitted light 11' is converted into illumination light 11 by this half mirror 13.
It is branched from the optical path of , passes through the relay lens 24 , and is focused into a point image Q by the condensing lens 25 .

Of course, in the portion of the half mirror 13 disposed closer to the laser light source 1a than the AOD 14, the illumination light 11 is not deflected, and the transmitted light 11° is not deflected either. Therefore, this point image Q does not move in response to the deflection of the illumination light 11. This point image Q is the pinhole plate 2B
The light is detected by the photodetector 27 via the photodetector 27. For example, a photodiode, a photomultiplier tube, or the like is used as the photodetector 27, and a signal S indicating the brightness of the point image Q is outputted from the photodetector 27.

As mentioned earlier, the illumination light 11 that irradiates the sample 20 is A
Since it is deflected by the OD 14, the light spot P scans the sample 20 in the X direction, and at the same time, since the illumination light 11 is deflected by the vibrating mirror 16, the light spot P scans the sample 20 in the main direction. Sub-scanning is performed in the Y direction, which is approximately perpendicular to the scanning direction.

As the light spot P scans the sample 20 two-dimensionally in the above manner, the photodetector 27 detects two points on the sample 20.
A time-series signal S carrying a dimensional image is output. This signal S is made into a pixel-divided signal by, for example, integrating at every predetermined period.

Further, in this embodiment, the sample stage 19 is moved by the moving mechanism 21 in the Z direction perpendicular to the main and sub-scanning directions X1Y. If the light spot P is two-dimensionally scanned each time the sample 20 is moved a predetermined distance in the Z direction in this manner, only information on the focused plane is detected by the photodetector 27. Therefore, by importing the output S of the photodetector 27 into the frame memory, it becomes possible to obtain a signal that represents an image in focus on all surfaces within the range in which the sample 20 is moved in the Z direction. .

Note that the light deflection means for deflecting the illumination light 11 is not limited to the AOD 14 and the vibrating mirror 16 described above, but may also include other known means such as a galvanometer mirror, a polygon mirror, and even an EOD (electro-optic light deflector). may be used.

Furthermore, although the confocal scanning microscope of the embodiments described above is for obtaining monochrome microscopic images, the present invention is applicable to conventionally known confocal scanning microscopes for obtaining color images. Of course, this is also possible.

(Effects of the Invention) As explained in detail above, the confocal scanning transmission microscope of the present invention is configured to scan two-dimensionally over a sample and detect the light transmitted therethrough, with the photodetector fixed. Since there is no need for a scanning mechanism for the photodetector, the structure can be simplified.

Furthermore, in the confocal scanning transmission microscope of the present invention, most of the light receiving optical system that guides the transmitted light transmitted through the sample to the photodetector is also used as the light transmitting optical system. Compared to conventional confocal scanning microscopes configured to guide transmitted light, the optical system is significantly simplified, which simplifies assembly and adjustment operations, resulting in significant cost reductions.

[Brief explanation of the drawing]

The figure is a schematic side view showing a confocal scanning microscope according to an embodiment of the present invention. lO... Laser light source 11... Illumination light 11°.
...Transmitted light 13...Half mirror 14...
AOD 15.17.24... Relay lens 16... Vibrating mirror 18.22... Objective lens 19... Sample stage 20... Sample 23.
・Corner cube prism

Claims (1)

[Scope of Claims] A sample stage on which a sample is placed, a light source that emits illumination light, a light transmission optical system that images the illumination light as a minute light spot on the sample, and a light transmission optical system that deflects the illumination light. a corner cube prism that is arranged at a position where the light beam transmitted through the sample is incident and reflects the light beam so as to follow the same path as the incident path; a light branching means for branching the reflected light flux from the optical path of the illumination light at a position closer to the light source than the light deflection means; and a condenser for condensing the branched light flux to form a point image. A confocal scanning transmission microscope consists of an optical lens and a photodetector that detects this point image.