Observation apparatus and observation method

Abstract

An observation apparatus and an observation method capable of changing the stimulation position in accordance with the position and the shape of a cell and of observing the cell immediately after stimulation are provided. The observation apparatus 1 includes a light source 11 that emits illumination light used to observe a specimen A to which a fluorescent substance that specifically binds to or is expressed in a stimulus target has been supplied, an illumination optical system 10 that radiates the illumination light emitted from the light source 11 onto the specimen A, a deflecting device 15 that changes an area of the specimen A to be irradiated with the illumination light, a wavelength selecting section that selects the wavelength of illumination light to be radiated onto the specimen A, an observation optical system 20 that collects light from the specimen A, a detector 25 that detects the light collected by the observation optical system 20 , an image processing section 13 that generates an image from the light detected by the detector 25 , and a control section 14 that controls these components.

1 . An observation apparatus comprising:
 at least one light source that emits illumination light used to observe a specimen to which a fluorescent substance that specifically binds to or is expressed in a stimulus target has been supplied;   an illumination optical system that radiates the illumination light emitted from the light source onto the specimen;   a deflecting section that changes an area of the specimen to be irradiated with the illumination light;   a wavelength selecting section that selects the wavelength of the illumination light to be radiated onto the specimen;   an observation optical system that collects light from the specimen;   at least one light detector that detects the light collected by the observation optical system;   an image generation section that generates an image from the light detected by the light detector; and   a control section that controls these components,   wherein the control section performs:   fluorescent-substance extraction processing in which light having a first wavelength used to excite the fluorescent substance that specifically binds to or is expressed in the stimulus target included in the specimen is selected by the wavelength selecting section, and the fluorescent substance is extracted from a fluorescence image generated by the image generation section based on fluorescence produced from the fluorescent substance when the light having the first wavelength is radiated;   stimulation processing in which light having a second wavelength used to stimulate the stimulus target is selected by the wavelength selecting section, and the illumination optical system radiates the light having the second wavelength onto the position of the fluorescent substance extracted in the fluorescent-substance extraction processing, through the deflecting section; and   image generation processing in which light having a third wavelength used to acquire an image of the specimen is selected by the wavelength selecting section, and an image of the specimen that includes the stimulus target is generated by the image generation section.   
     
     
         2 . An observation apparatus according to  claim 1 , comprising:
 a first light source that emits light having the first wavelength and light having the third wavelength; and   a second light source that emits light having the second wavelength,   wherein the light detector detects fluorescence produced from the specimen when the light having the first wavelength and the light having the third wavelength are radiated.   
     
     
         3 . An observation apparatus according to  claim 1 , comprising:
 a first light source that emits light having the third wavelength;   a second light source that emits light having the first wavelength and light having the second wavelength;   a first light detector that detects fluorescence produced from the specimen when the light having the third wavelength is radiated; and   a second light detector that detects fluorescence produced from the specimen when the light having the first wavelength is radiated,   wherein the control section extracts the stimulus target from an image generated by the image generation section based on the fluorescence detected by the second light detector.   
     
     
         4 . An observation apparatus according to  claim 1 , comprising:
 a first light source that emits light having the third wavelength;   a second light source that emits light having the second wavelength;   a third light source that emits light having the first wavelength;   a first light detector that detects fluorescence produced from the specimen when the light having the third wavelength is radiated; and   a second light detector that detects fluorescence produced from the specimen when the light having the first wavelength is radiated,   wherein the control section extracts the fluorescent substance from a fluorescence image generated by the image generation section based on the fluorescence detected by the second light detector.   
     
     
         5 . An observation apparatus according to  claim 1 , wherein the observation optical system has a galvanometer mirror. 
     
     
         6 . An observation apparatus according to  claim 1 , wherein the deflecting section is a galvanometer mirror. 
     
     
         7 . An observation apparatus according to  claim 1 , wherein the deflecting section is a microdevice array on which a plurality of microdevices that reflect or transmit light radiated from the illumination optical system are arrayed. 
     
     
         8 . An observation apparatus according to  claim 1 , comprising a recording section that records the image generated by the image generation section,
 wherein the control section records in the recording section a fluorescence image of the specimen generated by the image generation section when the light having the first wavelength is radiated.   
     
     
         9 . An observation method comprising:
 a fluorescent-substance extraction step of supplying a fluorescent substance that specifically binds to or is expressed in a stimulus target to a specimen to label the stimulus target with the fluorescent substance and of extracting the fluorescent substance by processing a fluorescence image that is acquired by irradiating the specimen with light having a first wavelength used to excite the fluorescent substance;   a stimulation step of radiating light having a second wavelength used to stimulate the stimulus target, onto the position of the fluorescent substance extracted in the fluorescent-substance extraction step; and   an image generation step of radiating light having a third wavelength used to acquire an image of the specimen, onto the specimen to generate an image of the specimen that includes the stimulus target stimulated in the stimulation step.