JP5151171B2 - Microscope image processing system and microscope image processing method - Google Patents

Description

  The present invention relates to a microscope image processing system and a microscope image processing method.

  In the fields of biology and medicine, a spectrum laser microscope is widely used in which a specimen is irradiated with laser light and fluorescence emitted from the specimen is decomposed into a spectrum and observed. Fluorescence emitted from a specimen is emitted as light having a specific spectral component depending on various conditions such as the type of fluorescent reagent and the wavelength to be excited. Therefore, by observing the spectral component of the fluorescence emitted from the specimen, the presence, state, reaction, etc. of the substance in the specimen can be confirmed.

  The result of observing spectral components is displayed in combination with a two-dimensional image of a sample (for example, Patent Document 1).

  Further, the spectral components are processed into band pass data classified into a predetermined band by post-processing and displayed in combination with, for example, a two-dimensional image of a sample. Data processed by post-processing such as averaging spectral components in this way is referred to as post-processed data in this specification.

JP 58-200209 A

  When creating post-processed data, only the results of post-processing such as averaging the spectral components are used. Here, the post-processing processed data is not linked to the original spectral component data. Therefore, the user of the microscope (hereinafter referred to as a user) cannot refer to the spectral data of a specific region characterized by the post-processing processed data on the post-processing processed data screen.

  There is a need for a microscope image processing system and a microscope image processing method capable of creating spectral data of a specific region characterized by post-processing processed data on the post-processing processed data screen.

In the microscope image processing system according to the first aspect of the present invention , when the observation light is irradiated with the scanning light from the light source , the spectral data of the observation sample detected for each irradiation portion of the sample and the spectrum data are detected. Spectral data collection means for collecting position information of an irradiation site in the sample, spectrum data storage means for storing the spectrum data and the position information, processing the spectrum data, creating post-processing processed data, the post-processing processing data screen creation means for creating a treatment processing data screen, and post-processed data storing means for storing the post processed data, the corresponding spectral data of the area identified in the post processed data screen Read from the spectrum data storage means based on the position information or create a specific area spectrum read Characterized in that it comprises the out-creating means.
The microscope image processing system according to the second aspect of the present invention is the spectral data of the observation sample detected for each pixel position on the microscope image surface corresponding to the irradiation site of the sample when the observation sample is irradiated with the scanning light from the light source. Spectrum data collecting means for collecting position information of pixel positions on a microscope image plane corresponding to an irradiation site in the sample from which the spectrum data is detected, spectrum data storing means for storing the spectrum data and the position information Processing the spectrum data, creating post-processing processing data, post-processing processing data screen creation means for creating a post-processing processing data screen, post-processing processing data storage means for storing the post-processing processing data, spectrum corresponding to the spectral data of the specified region in the post processed data screen on the basis of the position information Or read from over data storage unit, or a specific region spectrum read-generating means for generating, characterized in that it comprises a.

In the microscope image processing method according to the third aspect of the present invention , when the observation sample is irradiated with the scanning light from the light source , the spectrum data of the observation sample detected for each irradiated portion of the sample and the spectrum data are detected. A step of collecting position information of an irradiation site in the sample; a step of storing the spectrum data and the position information; and processing the spectrum data to generate post-processing processing data and generating a post-processing processing data screen And a step of reading or creating corresponding spectrum data of the region specified on the post-processing processed data screen based on the position information.

  According to the present invention, since the spectrum data of the detected observation sample and the position information in the sample where the spectrum data is detected are stored, the spectrum data of the area specified in the post-processing processed data screen is stored. , And can be created based on the position information. Therefore, the user can refer to the spectrum data related to the information displayed on the post-processing processed data screen, and can perform an effective analysis.

  According to the present invention, it is possible to obtain a microscope image processing system and a microscope image processing method capable of creating spectrum data of a specific region characterized by post-processing processing data on the post-processing processing data screen.

  FIG. 1 is a diagram showing the configuration of a spectral laser microscope according to an embodiment of the present invention.

  In FIG. 1, the spectrum laser microscope according to the present embodiment includes a laser microscope main body 300, a spectrum detector 200, and a laser microscope controller 100.

  The laser microscope main body 300 includes a laser light source 301 that emits excitation light that irradiates a specimen placed on a stage 309, a collimator lens 303 that collimates the laser light from the laser light source 301, and reflects the laser light toward the specimen. And a dichroic mirror 305 that transmits fluorescence from the specimen, and an XY scanner 311 and an objective lens 307 disposed between the dichroic mirror 305 and the stage 309.

  The spectral detector 200 as spectral data detection means includes a spectroscopic element 201, a multichannel photodetector 203, and an optical signal sampling circuit 205. The optical system of the laser microscope main body 300 and the optical system of the spectrum detector 200 are connected by an optical system (not shown).

  FIG. 2 is a diagram illustrating a spectral component (luminance value) that is an output of the multichannel photodetector 203. The multi-channel photodetector 203 has n channels. Therefore, n output values are obtained corresponding to each part of the sample irradiated with the laser.

  The microscope controller 100 receives the spectral components of each channel from the optical signal sampling circuit 205 and drives the XY scanner 311 according to instructions from the microscope image processing system 110, the apparatus control circuit 120, and the apparatus control circuit 120 that process them as image data. , An XY scanner driving circuit 140, and a high-voltage power supply 130 that supplies a voltage to the multichannel photodetector 203 according to a command from the apparatus control circuit 120. The microscope image processing system 110 includes a display device 115 that displays a microscope image.

  FIG. 3 is a diagram showing a configuration of the microscope image processing system 110 according to the embodiment of the present invention. The microscope image processing system 110 includes a spectrum data collection unit 1010, a spectrum data storage unit 1030, a post-processing processed data screen generation unit 1050, a specific region spectrum generation unit 1070, and a post-processing processed data storage unit 1090. .

  FIG. 4 is a flowchart showing the processing of the microscope image processing system 110 according to an embodiment of the present invention.

In step S4010 of FIG. 4, the spectrum data collection unit 1010 of the microscope image processing system 110 collects spectrum data. More specifically, the spectral data collection unit 1010 receives the spectral components of each channel from the optical signal sampling circuit 205 and also receives the pixel position data from the device control circuit 120. The pixel position corresponds to the scan position of the XY scanner 311. In this specification, a pixel position corresponding to the scan position of the XY scanner 311 is referred to as a pixel position on the original screen. Since the number of channels is n, n pieces of spectral data are obtained for each pixel position on the original screen. As an example, the number of pixels on the screen
512x512
It is a piece.

  In step S4020 of FIG. 4, the spectrum data collection unit 1010 stores the spectrum data in the spectrum data storage unit 1030.

  FIG. 5 is a diagram illustrating a data structure of the spectrum data storage unit 1030. The spectrum data storage unit 1030 stores n pieces of spectrum data for each pixel position on the original screen.

  In step S4030 of FIG. 4, the post-processing processed data screen creation unit 1050 creates a post-processing processed data screen based on the spectrum data stored in the spectrum data collection unit 1010.

  Below, the specific example of the processing method of post-processing process data is demonstrated.

  The band pass data process is a process for collecting n-channel spectral components into a smaller number of bands. As an example, three bands may be combined and displayed on the post-processing processed data screen in three colors, superimposed on a two-dimensional image of the specimen.

  The processing based on the ratio of spectral components (Ratio processing) is processing for obtaining the ratio of two spectral components at a specific wavelength for each pixel. As an example, the magnitude of the ratio may correspond to the color, and may be displayed on the post-processing processed data screen superimposed on the two-dimensional image of the specimen.

  Spectral component correlation processing (Colocalization processing) is processing for obtaining a correlation between two spectral components at a specific wavelength. As an example, a spectral component at one specific wavelength may be represented on the horizontal axis, and a spectral component at the other specific wavelength may be represented on the vertical axis.

The process of decomposing a spectrum component as a factor (Unmixing process) is a process of decomposing a spectrum pattern caused by a plurality of factors into a spectrum pattern due to individual factors. Each factor is, for example, excitation light having a plurality of wavelengths. FIG. 9A is a diagram showing a spectrum pattern Y generated by a plurality of factors, and FIG. 9B is a diagram showing spectrum patterns Ya, Yb and Yc caused by individual factors. A, b, and c satisfying the following expressions are obtained by calculation.

  As an example, each of a, b, and c may be displayed on the post-processing processed data screen so as to overlap the two-dimensional image of the specimen.

  The post-processing processed data screen creation unit 1050 stores, for example, post-processing processed data created by the above processing method in the post-processing processed data storage unit 1090.

  FIG. 6 is a diagram showing a data structure of the post-processing processed data storage unit 1090. The post-processing processing data storage unit 1090 stores post-processing processing data corresponding to the pixel position for each pixel position on the post-processing processing data screen. The post-processing processed data storage unit 1090 further stores a corresponding pixel position of the original screen for each pixel position of the post-processed processed data screen. When the pixel position on the post-processing processed data screen is the same as the pixel position on the original screen, the pixel position on the post-processed data screen may be used as the pixel position on the original screen. Also, the corresponding pixel position is stored at the time of post-processing processing data storage, but it is not always necessary. For example, the pixel position is also stored only at the time of storing spectrum data, and the post-processing processing data is displayed and displayed. For example, the user may specify the coordinates on the image and read the spectrum data.

  In step S4040 of FIG. 4, the specific region spectrum creation unit 1070 creates a spectrum of the specific region specified on the post-processing processed data screen. More specifically, the specific region spectrum creation unit 1070 accesses the data of the pixel position of the post-processing processed data screen corresponding to the specific region specified on the post-processing processed data screen of the post-processing processed data storage unit 1090, The pixel position of the corresponding original screen is obtained. Next, the specific region spectrum creation unit 1070 accesses the spectrum data storage unit 1030 based on the pixel position of the original screen, and obtains spectral data of the specific region. The spectral data of the specific region may be obtained by averaging the spectral data of the corresponding pixels.

  In order to specify a specific area on the post-processing data screen, a display such as a rectangle or a circle is provided on the screen so that the user can click or drag the display and place it in a desired size at a desired position. It may be. In the present embodiment, the position information is grasped by the pixel position, but other coordinates may be provided and the position information may be grasped by the coordinates.

  FIG. 7 is a diagram showing a post-processing processed data screen (correlation processing screen) so that the horizontal axis represents the spectral component at one specific wavelength Si and the vertical axis represents the spectral component at the other specific wavelength Sj. This is called the Scatter gram. This is to look at the distribution of data in the portion surrounded by a frame on the secondary image. According to the present invention, it is possible to refer to spectrum data by designating a specific region as shown by a circle in FIG. 7 on the post-processing processed data screen. This is effective, for example, when comparing the spectrum data after the unmixing process shown in FIG. 9B and the spectrum data before the unmixing process.

  FIG. 8 is a diagram illustrating a result of referring to spectrum data.

  In the conventional image processing system, the data for displaying the post-processing processed data screen is not linked to the spectral data for each pixel stored in the spectral data storage unit 1030. Therefore, spectrum data could not be referred from the post-processing processed data screen.

  On the other hand, in the present invention, the data for displaying the post-processing processed data screen is linked with the spectral data for each pixel stored in the spectral data storage unit 1030. Spectral data can be referred to.

  As described above, according to the present invention, since a spectrum of a specific region characterized by post-processing processed data can be created on the post-processing processed data screen, it provides an effective means for the user to perform analysis. can do.

It is a figure which shows the block diagram of the spectrum laser microscope by one Embodiment of this invention. It is a figure which shows the spectrum component (luminance value) which is the output of the multichannel photodetector 203. FIG. It is a figure which shows the structure of the microscope image processing system 110 by one Embodiment of this invention. It is a flowchart which shows the process of the microscope image processing system by one Embodiment of this invention. It is a figure which shows the data structure of a spectrum data storage part. It is a figure which shows the data structure of a post-processing process data storage part. It is a figure which shows the post-processing process data screen so that the spectral component in one specific wavelength Si may be represented by a horizontal axis, and the spectral component in the other specific wavelength Sj may be represented on a vertical axis | shaft. It is a figure which shows the result which referred spectral data. It is a figure which shows the spectrum pattern Y produced | generated by the some factor, and the spectrum patterns Ya, Yb, and Yc by each factor.

Explanation of symbols

1010: Spectrum data collection unit, 1030 ... Spectrum data storage unit, 1050 ... Post-processing processed data screen creation unit, 1070 ... Specific region spectrum creation unit, 1090 ... Specific region spectrum creation unit

Claims (8)

Spectral data that collects the spectral data of the observation sample detected for each irradiated part of the sample when the observation sample is irradiated with the scanning light from the light source, and the position information of the irradiated part in the sample from which the spectral data was detected Collecting means;
Spectrum data storage means for storing the spectrum data and the position information;
Processing the spectrum data, creating post-processing processing data, post-processing processing data screen creation means for creating a post-processing processing data screen,
Post-processing processing data storage means for storing the post-processing processing data;
A specific region spectrum reading / creating unit that reads or creates corresponding spectrum data of the region identified on the post-processing processed data screen from the spectrum data storage unit based on the position information is provided. Microscope image processing system. And spectral data of the observation sample detected for each pixel position of the microscope image plane corresponding to the irradiated portion of the sample when irradiated with scanning light from a light source to an observation sample, irradiation of the sample to the spectral data is detected Spectral data collection means for collecting position information of the pixel position on the microscope image surface corresponding to the site ;
Spectrum data storage means for storing the spectrum data and the position information;
Processing the spectrum data, creating post-processing processing data, post-processing processing data screen creation means for creating a post-processing processing data screen,
Post-processing processing data storage means for storing the post-processing processing data;
A specific region spectrum reading / creating unit that reads or creates corresponding spectrum data of the region identified on the post-processing processed data screen from the spectrum data storage unit based on the position information is provided. Microscope image processing system. The post processed data storing means, a microscope image processing system according to claim 1 or 2, wherein the location information is stored in the post-treatment processing data. The post-processing processed data storage means stores the position information of the corresponding spectral data for each pixel position on the post-processed processed data screen, thereby associating the post-processed processed data with the spectral data information. The microscope image processing system according to claim 1 or 2 , wherein The post processing data, characterized in that it is a band-pass data of spectral components, the microscopic image processing system according to any one of claims 1 to 4. The microscope image processing system according to any one of claims 1 to 4 , wherein the post-processing data is a ratio or correlation between two spectral components at a specific wavelength. The microscope image processing system according to any one of claims 1 to 4 , wherein the post-processing data is obtained by decomposing a spectral component into a plurality of factors. Collecting the spectral data of the observation sample detected for each irradiation region of the sample when the observation sample is irradiated with the scanning light from the light source, and the position information of the irradiation region in the sample where the spectral data is detected; ,
Storing the spectral data and the position information;
Processing the spectral data, creating post-processed processed data, creating a post-processed processed data screen;
Reading or creating spectral data corresponding to a region specified on the post-processing processed data screen based on the position information.

Images