Total internal reflection microscope

Patent metadata

Field	Value
Publication number	US-10241312-B2
Application number	US-201615142804-A
Country	US
Kind code	B2
Filing date	Apr 29, 2016
Priority date	Oct 30, 2013
Publication date	Mar 26, 2019
Grant date	Mar 26, 2019

Abstract

The total internal reflection microscope has an illumination optical system that relays light from a light source with a relay optical system, forms an image of the light source on the incident pupil plane of the objective lens and irradiates a sample with the illumination light via an objective lens, has an angle adjustment mirror for changing the position of the image of the light source in a direction orthogonal to the optical axis, an optical detector for detecting the intensity of the returning illumination light reflected by the sample and collected by the objective lens, and a controller for determining the operation amount of the angle adjustment mirror, wherein the controller determines the operation amount of the angle adjustment mirror so that the illumination light is totally reflected at the sample based on the change in intensity of the returning light when the angle adjustment mirror is changed.

First claim

The invention claimed is:

1. A total internal reflection microscope comprising an illumination optical system for relaying light from a light source with a relay optical system and forming an image of the light source on the incident pupil of an objective lens or the vicinity thereof and irradiating an observation object with the light through the objective lens, the total internal reflection microscope further comprising:
an incident angle adjustment device for changing the position of the image of the light source in a direction orthogonal to a direction of the optical axis,
an optical detector for detecting intensity of the returning light that is the light reflected by the observation object and collected by the objective lens, and
a controller for determining the operation amount of the incident angle adjustment device, wherein
the controller determines the operation amount of the incident angle adjustment device based on the change in intensity of the returning light detected by the optical detector when the incident angle adjustment device is operated.

2. The total internal reflection microscope according to claim 1 , further comprising
a focus lens for changing the position of the image of the light source in the direction of the optical axis, wherein
the controller determines the operation amount of the incident angle adjustment device and the focus lens based on the change in intensity of the returning light when the incident angle adjustment device and the focus lens are operated.

3. The total internal reflection microscope according to claim 2 , wherein
the controller determines the operation amount of the incident angle adjustment device so the light is totally reflected at the sample based on the change in intensity of the returning light when the incident angle adjustment device is operated, and determines the operation amount of the focus lens so the light becomes parallel light based on the change in intensity of the returning light when the focus lens is operated.

4. The total internal reflection microscope according to claim 2 , wherein
the controller adjusts the position of the light source image in the direction orthogonal to the direction of the optical axis in the incident pupil of the objective lens or the vicinity thereof so that the light is totally reflected at the observation object, and adjusts the position of the light source image in the direction of the optical axis in the incident pupil of the objective lens or the vicinity thereof so that the light becomes parallel light.

5. The total internal reflection microscope according to claim 2 , wherein
the controller detects the collecting state of the image of the light source relative to the incident pupil plane based on the change in intensity of the returning light when the position of the image of the light source is changed in the direction of the optical axis by the focus lens.

6. The total internal reflection microscope according to claim 2 , wherein
the controller determines the operation amount of the focus lens so the light becomes parallel light based on the change in intensity of the returning light when the focus lens is operated.

7. The total internal reflection microscope according to claim 2 , wherein
the controller adjusts the position of the light source image in the direction of the optical axis in the incident pupil of the objective lens or the vicinity thereof so that the light becomes parallel light.

8. The total internal reflection microscope according to claim 1 , wherein
the controller identifies the boundary of total reflection in the observation object and non-total reflection in the observation object based on the change in intensity of the returning light.

9. The total internal reflection microscope according to claim 1 , wherein
the controller calculates the incident angle of the light relative to the observation object based on the operation amount of the incident angle adjustment device or the amount of change in the angle formed between the principal ray of the light and the optical axis of the relay optical system, which changes with the operation of the incident angle adjustment device.

10. The total internal reflection microscope according to claim 1 , wherein
the controller calculates at least one of either the refractive index of the observation object or the evanescent field penetration depth based on the operation amount of the incident angle adjustment device, or the amount of change in the angle formed between the principal ray of the light and the optical axis of the relay optical system that changes with the operation of the incident angle adjustment device.

11. The total internal reflection microscope according to claim 1 , wherein
the controller controls the incident angle adjustment device to obtain the desired evanescent field penetration depth.

12. The total internal reflection microscope according to claim 1 , wherein
the optical detector is disposed in a position conjugate to the incident pupil plane or the vicinity thereof.

13. The total internal reflection microscope according to claim 1 , wherein
the controller determines a position of the image of the light source in the incident pupil plane based on the operation amount of the incident angle adjustment device, or the amount of change in the angle formed between the principal ray of the light and the optical axis of the relay optical system that changes with the operation of the incident angle adjustment device.

14. The total internal reflection microscope according to claim 1 , wherein
the controller determines three or more of the positions of the image of the light source in the incident pupil plane based on the change in intensity of the returning light, and determines the center of a circle corresponding to the boundary of total reflection and non-total reflection from the determined positions.

15. The total internal reflection microscope according to claim 1 , wherein
the incident angle adjustment device is on the optical axis of the relay optical system, is disposed so as to intersect a position conjugate to the field of vision of the objective lens or the vicinity thereof, has a reflective surface for reflecting the light, and changes the incident angle by changing the angle of the reflective surface relative to the optical axis with the point of intersection of the reflective surface with the optical axis as the center.

16. The total internal reflection microscope according to claim 1 , wherein
the incident angle adjustment device changes the incident angle by changing the distance of the light source from the optical axis in the plane orthogonal to the optical axis.

17. The total internal reflection microscope according to claim 1 , wherein
the incident angle adjustment device is on the optical axis of the relay optical system and changes the incident angle by rotating with a position conjugate to the field of vision of the objective lens or the vicinity thereof as the center.

18. The total internal reflection microscope according to claim 1 , wherein
the controller determines the operation amount of the incident angle adjustment device so the light is totally reflected at the observation object based on the change in intensity of the returning light when the incident angle adjustment device is operated.

19. The total internal reflection microscope according to claim 1 , wherein
the controller adjusts the position of the light source image in the direction orthogonal to the direction of the optical axis in the

Assignees

• Nikon Corp

Inventors

• Morita Takayuki
• Suzuki Akitoshi
• Kobayashi Tomoko
• Masuda Keita

Classifications

• G02B27/56

  Optics using evanescent waves, i.e. inhomogeneous waves · CPC title

• G02B21/16

  adapted for ultraviolet illumination {; Fluorescence microscopes (G02B21/0076 takes precedence)} · CPC title

• G01N21/648

  using evanescent coupling or surface plasmon coupling for the excitation of fluorescence · CPC title

• G02B21/365

  Control or image processing arrangements for digital or video microscopes (G02B21/361, G02B21/362 take precedence) · CPC title

• G02B21/06 Primary

  Means for illuminating specimens · CPC title