JPH10185911A - Device for analyzing cell and method therefor - Google Patents

Abstract

PROBLEM TO BE SOLVED: To automatically determinate detection and measurement, improve precision and reliability, and enable an early detection of cancer by taking the image of a cell, collating measured data with the pattern information of a plurality of previously recognized cells, and classifying them. SOLUTION: Specimen of a plurality of cells extracted from humans or animals are disposed on a stage part 3, and imaged and taken in by the image taking-in part 4 of the CCD camera or the like of a microscope 5. An image processing part 9 displays data of cytoplasm and nucleus on a TV monitor, cytoplasm and nucleus are classified, image-converted, and prescribed necessary items such as the number, area and color of nucleus and small nucleus are measured from the image. Pattern recognition is previously performed, the pattern information of the plurality of the cells input in an information part 11 and the measured data are collated, and it is classified by an analysis part 12 whether they coincide with pattern information or they are similar with it.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a cell analyzing apparatus and method, and more particularly, to a method for automatically and accurately detecting chromosomal abnormalities such as micronuclei by automatic quantification analysis to detect abnormal cells such as cancer cells. Not only analysis of cells as a guide for discovery, but also cells that can obtain high-precision analysis results by quantifying (quantifying) DNA abnormalities in cells, damage frequency and damage to single cells The present invention relates to an analyzer and a method thereof.

[0002]

2. Description of the Related Art For example, cancer, which is said to be the leading cause of sickness and death in Japan, is affected by environmental factors such as food, tobacco, pesticides, pharmaceuticals, alcohol, and radiation damage, and the chromosomes of normal cells are mutated. This is a phenomenon that causes abnormalities and becomes cancerous. Frequently used for early detection of this chromosomal abnormality is the frequency measurement of micronuclei (micronuclei). Conventionally, the conventional method and the cytochalasin method have been used. Was used.

[0003] In the above-mentioned micronucleus, chromosomes cannot move to both poles due to chromosome fragments or delay in cell division during cell division. The frequency of this micronucleus allows for the determination of chromosomal abnormalities.

[0004] As a method for measuring micronuclei, there is, for example, the above-mentioned conventional method, which is performed by visually examining cells extracted from the human body or the like for which the presence of nuclei can be confirmed in the cytoplasm. This is a method of counting the number of micronuclei generated by using a microscope or the like.

[0005]

However, in the measurement of micronuclei by the above-mentioned conventional method, when preparing a cell specimen for observation, cytoplasm often adheres to each other, and these cells are visually separated. Since it is difficult to perform the measurement, the measurement is usually performed only on cells having a sharp outline.

[0006] The same applies to the cytochalasin method.
It is a visual measurement, and therefore, with the conventional method, accurate measurement cannot be performed in any of the former and latter cases, and its quantification is very difficult,
Anyway, clinically requires a lot of more accurate data, which means that DNA abnormalities in cells,
The same applies to the case where the frequency of damage is determined or the damage of a single cell is analyzed. In any case, some solution has been desired.

The present invention has been made in view of the above-mentioned conventional problems, and automatically quantifies the detection / measurement of abnormal cells having chromosomal abnormalities such as micronuclei, which serve as a guide for cancer detection and the like. Thus, the accuracy and reliability are improved, and early detection of cancer is enabled. Further, not only analysis of cells as a guide when detecting chromosomal abnormalities such as the micronucleus, but also DNA abnormalities and damage in cells It is an object of the present invention to provide a cell analysis apparatus and a method for obtaining a highly accurate analysis result by quantifying (quantifying) the frequency of damage and damage to a single cell.

[0008]

Means for Solving the Problems The first technical means taken by the present invention to solve the above-mentioned problems is a cell analysis device, which has previously been subjected to pattern recognition of a plurality of cells. An information unit 11, a stage unit 3 for arranging the extracted cell specimen 2, and an image capturing unit 4 for photographing the specimen
An image processing unit 9 that converts the captured image into a measurement image; performs a predetermined measurement from the processed image; and compares the measured data with the pattern information of the information unit 11 to perform classification. The image processing unit 9 extracts the cytoplasm 6 and the nucleus 7 and measures the number and area of the nucleus 7 and the micronucleus 10 in the image processing unit 9. It becomes.

[0009] The second technical means is a cell analysis method. In this method, cells are extracted to prepare a specimen 2, the specimen 2 is placed on the stage section 3, and the image is taken by the image capturing means. Photographing, converting the captured image into an image for measurement, performing a predetermined measurement, collating the measured data with pattern information on a plurality of cells that have been subjected to pattern recognition in advance, and performing cell analysis by classifying the data. It is to apply. At this time, in taking in an image, the cytoplasm 6 and the nucleus 7 are specifically extracted, and in the measurement, predetermined necessary items such as the number, area, and color of the nucleus 7 and the micronucleus 10 are measured. It is characterized by:

[0010] Analyzing cells with the above-mentioned cell analyzer is to replace conventional analog analysis with digital analysis (digital image). Statistical processing can be automatically performed.

Further, the specimen 2 placed on the stage section 3 is photographed by the image capturing means using the above-mentioned cell analyzer,
By converting the captured image into an image for measurement and performing a predetermined measurement, and performing a predetermined arithmetic processing on all of the measured data or only the predetermined data, various management data can be obtained.

Thus, as described above, the digital image captured by the image capturing means is used as a measurement image to obtain, as color information, grains of a predetermined substance captured in the nucleus of the cell, or the nucleus extracted as color information. Automatically measure the number of grains in the area and the average number of grains in a predetermined cytoplasmic area located around the nucleus area or the number of grains in a predetermined number of cytoplasmic areas, and further to the display means via the color information. By specifying the coordinates of a specified part of the displayed cell image via a predetermined pointing device and setting it as a measurement area, it is possible to automatically measure the dimensions of each part of the cell, and thus, DNA abnormality and damage in the cell By quantifying and quantifying the frequency of damage and damage to a single cell, analysis can be performed with high accuracy.

[0013]

BEST MODE FOR CARRYING OUT THE INVENTION

<First Embodiment> One embodiment of the present invention will be described below with reference to the drawings.

As shown in FIG. 1, the cell analyzer according to the present invention has a stage section 3 on which a plurality of specimens 1 of cells 1 extracted from humans and animals are arranged, and a CCD camera or the like. A fluorescence microscope 5 or a phase-contrast microscope provided with an image capturing unit 4 for capturing an image of the specimen 2 and capturing the image, and connected to the image capturing unit 4 of the microscope 5 and captured by the image capturing unit 4 An image processing unit that, among the obtained images, displays data of the cytoplasm 6 and the nucleus 7 on a TV monitor 8 or the like, which is one of display means, and separates the cytoplasm 6 and the nucleus 7 into image data for measurement, respectively. 9 and
Predetermined necessary items such as the number, area, and color of the nuclei 7 and micronuclei 10 are measured from the processed image, and the measured data and the pattern recognition of the cytoplasm 6 previously obtained from a plurality of cells 1 (The information input to the information unit 11 recorded on a medium such as HD, FD, CD, or MO), and classify each pattern information as matching or similar. And an analyzing unit 12.

As shown in FIG. 2, a plurality of cells 1 extracted from humans, animals, etc.
It is placed on top and subjected to an appropriate staining treatment such as acridine range staining or Giemsa staining, and then covered with a cover glass.

The stage section 3 of the microscope 5 is an automatic stage. For example, when the sample 2 is set on the automatic stage as shown in an image capturing flowchart shown in FIG. The stage unit 3 is moved at a constant pitch and pattern according to a movement pattern (for example, the number of screens and division designation in the X and Y directions are input).

After the movement, the image capturing section 4 C
Autofocus function provided in CD camera etc. or C
A predetermined auto-focusing device (not shown) is mounted on a CD camera or the like, and thereafter, when the image is focused, the image is captured by the image processing unit 9, saved as image data, and required (set in advance). This operation can be automatically repeated until the number of screens is saved.

When an image is captured from the CCD camera or the like into the image processing unit 9 as described above, if the CCD camera or the like outputs a video signal as an analog signal, the captured video signal is converted to an A signal. After digital / D (analog / digital) conversion, the digital data may be saved as image data. In the case of a model in which a CCD camera outputs a video signal as a digital signal, the captured image of the digital data is directly used. It may be saved as data.

In the above operation, images are sequentially saved. At this time, for example, as shown in FIG. 2, coordinate data 15 is stored in one specimen 2 and the If the numbering is performed in conformity with the coordinate data so that the stage can be recognized, the subsequent processing becomes easier.

The image data saved as described above is respectively subjected to predetermined measurement while being converted into an image for measurement by the image processing section 9. For example, as shown in FIG. (See (a) in the figure), and then only the cytoplasm 6 is projected (extracted) (see (b) in the figure).
Measure the number (area number).

Next, only the nucleus 7 (including the micronucleus 10) is projected (extracted) (see FIG. 4C), and the characteristic values such as the number of the nuclei 7 and the micronuclei 10 and the existing area are determined. Measure (see (d) in the figure).

Therefore, the extracted cytoplasm 6 (see FIG. 4 (b)) is included as one region (see FIG. 4).
(See (c)) By making the nucleus 7 and micronucleus 10 the measurement results,
The nucleus 7 and the micronucleus 10 in one cell can be measured.

That is, by using the extracted cytoplasm 6 as one measurement area and measuring the nucleus 7 and micronucleus 10 in the measurement area, “cells containing nuclei”, “cells containing micronuclei”, “cells containing micronuclei” This makes it possible to classify the cells into cells containing nuclei and micronuclei, and cells not containing nuclei and micronuclei.

These data are tabulated for each stage, for example, as shown in FIG. 5, and each cell is analyzed in detail.

The characteristic values shown in FIG. 5 are derived from values such as the area and circularity of a nucleus (referred to as a main nucleus).
This is a predetermined numerical value for matching with the pattern information.

The analysis is performed by computer control, for example, in a series of control processes from the image capturing unit 4 to the analyzing unit 12, as shown in an automatic analysis flowchart shown in FIG. 6, and printed out by researchers and doctors. Abnormal cells such as cancer are detected based on the analysis data.

According to the cell analysis method using the cell analysis device according to the first embodiment of the present invention, the micronucleus, which serves as a guide for quickly and accurately finding cells with abnormal chromosomes via digital images, can be used. Since analysis is possible and automatic quantification is possible as described above, there is no variation in data, and very reliable results are produced.

In the above-described embodiment, the analysis was performed using the micronucleus as a guide for the purpose of detecting cancer. However, the cell analysis device and method according to the first embodiment of the present invention are not limited to the micronucleus. In other words, the point is to find a chromosomal abnormality in a certain disease, and any cell can be analyzed as long as it has data on which pattern recognition has been performed in advance.

<Second Embodiment> Further, the above-mentioned cell analysis apparatus is provided with a stage section 3 on which an extracted cell specimen 2 is arranged.
An image capturing unit 4 that captures the sample, an image processing unit 9 that converts the captured image into a measurement image, performs a predetermined measurement from the processed image, and outputs all or measured data. Various management data may be obtained by performing predetermined arithmetic processing on only predetermined data. In this case, a so-called cell analyzer is used to test the effect of a test substance on each organ of a living body, so-called, It can be applied to the UDS test.

[0030] Such a test was carried out to test for DN produced by a carcinogen.
When the A damage is removed and repaired, a substance (thymidine) given in advance from the outside is taken up into the cell and the nucleus, and the radioactivity (grain) of the substance (thymidine) taken up in the cell and the nucleus is obtained. This is a test for measuring the particle size, and the grain to be measured can be extracted by preprocessing by the autoradiography method.

That is, the grains of the specimen 2 to be inspected, which have been preprocessed by the autoradiography method, are identified by color information such as chromaticity, lightness, and saturation through a cell analyzer, and the intracellular and intranuclear nuclei are identified. By measuring the number of grains, it is possible to determine the frequency of DNA abnormality and damage due to the frequency of occurrence of the number of grains.

That is, the sample 2 (not shown) extracted based on the color information in the image processing unit 9 of the cell analyzer (not shown)
Automatic measurement of the number of grains in the nuclear area and the average number of grains in a predetermined cytoplasmic area or a predetermined number of grains in a predetermined number of cytoplasmic areas located around the nuclear area, and the measurement parameters processed by the cell analyzer The number of grains in the nuclear area in the cytoplasm, the average number of grains in a predetermined number of cytoplasmic areas, the number of NET grains obtained by subtracting the number of grains in the cytoplasm from the number of grains in the nucleus, and the size, aggregation, and standard deviation of each grain Process the statistics of each data, such as
Moreover, there is an advantage that extraction, identification and measurement of grains that cannot be identified visually can be quantified through operations such as image brightness inversion and image emphasis, so that extremely high-precision results can be obtained. As described above, the procedure for determining the frequency of DNA abnormality and damage may be performed, for example, in accordance with the automatic analysis flowchart shown in FIG.

<Third Embodiment> Further, a cell analyzer configured to obtain various management data by subjecting all of the data measured as described above or only a predetermined data to a predetermined arithmetic processing is provided. The analysis can be applied to a so-called SCG test.

In this test, a specimen 2 (not shown) embedded in the middle layer of agar formed into three layers is subjected to electrophoresis, and then stained with acridine orange or the like. If the DNA is not damaged, the shape is close to a circle, and if the DNA is damaged, it has a tailed shape like a comet. Therefore, it is possible to measure predetermined locations such as the size of the nucleus and the maximum length of the whole cell. By doing so, the degree of damage can be determined.

That is, the image processing unit 9 of the cell analyzer (not shown) extracts the chromaticity, lightness, saturation, etc. of each part of the cell as color information, and, based on the color information, a DNA nucleus part, a DNA damaged part, and the like. Performing a process of identifying a background portion other than DNA, and specifying a coordinate of a predetermined portion of a cell image displayed on a display means such as a TV monitor 8 through color information through a pointing device such as a mouse through color information and setting it as a measurement area. so,
The dimensions of each part of the cell are automatically measured. The cell analyzer obtains a set value of image enhancement (density conversion) based on the color information of the cell, and then classifies the color information into a predetermined stage. DN based on color information such as chromaticity, lightness, and saturation
A The nucleus, the DNA damaged part and the background part are distinguished, and the measurement parameters include the total length of the cell, the diameter of the nucleus, the ratio of the total length to the diameter of the nucleus, the area of the damaged part, the size of the damaged part, the moving distance of the nucleus, By measuring not only the ratio of the damaged part luminance to the total luminance, but also luminance conversion and coloring, etc. Therefore, there is an advantage that extremely high precision results can be obtained. As described above, the procedure for determining the degree of DNA damage may be performed, for example, according to the automatic analysis flowchart shown in FIG.

Further, the cell sample 2 extracted as described above
, An image capturing unit 4 for capturing the sample, an image processing unit 9 for converting the captured image into a measurement image, and performing a predetermined measurement from the processed image. A cell analysis device that can obtain various management data by performing predetermined arithmetic processing on all of the data or only predetermined data is provided with pattern data recognition of a plurality of cells that are pre-ranked based on predetermined conditions. The information section 11 is provided with various management data and the information obtained by performing predetermined arithmetic processing on all or only predetermined data measured from the specimen 2 of each cell in the cell analyzer. When an analysis unit 12 (not shown) is provided for collating the cell pattern data with the cell pattern data in the unit 11 and ranking the sample 2 of each cell, the sample 2 is sequentially set in the cell analyzer. Serial there is an advantage that it is possible to accurately sort organize the data of a large amount of the sample 2 and each target present 2 simply repeating the process steps.

[0037]

Industrial Applicability The present invention provides a conventional analog analysis for automatically quantifying (patterning) the detection / measurement of abnormal cells having chromosomal abnormalities such as micronuclei, which is a standard for finding cancer and the like. By replacing with digital analysis from, it is possible to quickly and accurately analyze cells as a guide when detecting chromosomal abnormalities such as micronuclei, improve precision and reliability, and detect cancer early. Not only is it possible to quantify and quantify not only the analysis of cells as a guide for finding chromosomal abnormalities such as micronuclei, but also the DNA abnormalities in cells, the frequency of damage, and damage to single cells As a result, a special effect that a causal relationship or a prediction of a measurement result can be obtained by statistical processing of a high-precision analysis result has been brought.

[Brief description of the drawings]

FIG. 1 is a schematic diagram of a main part showing one embodiment of a cell analysis device of the present invention.

FIG. 2 is a schematic diagram showing one embodiment of a specimen according to the present invention.

FIG. 3 is an image capture flowchart according to the present invention.

FIG. 4 is a schematic diagram showing an embodiment of image processing according to the present invention.

FIG. 5 is a schematic diagram showing an embodiment of measurement data according to the present invention.

FIG. 6 is an automatic analysis flowchart according to the present invention.

FIG. 7 is an automatic analysis flowchart in the second embodiment of the cell analysis device of the present invention.

FIG. 8 is an automatic analysis flowchart in the third embodiment of the cell analysis device of the present invention.

[Explanation of symbols]

 2 specimen 3 stage part 4 image capturing part 6 cytoplasm 7 nucleus 9 image processing part 11 information part 10 micronucleus 12 analysis part

Claims (16)

[Claims] An information section (11) in which a plurality of cell patterns have been recognized in advance, a stage section (3) for arranging an extracted cell specimen (2), and an image capturing section for photographing the specimen.
(4), an image processing unit (9) that converts the captured image into a measurement image, performs a predetermined measurement from the processed image, and measures the measured data and the pattern of the information unit (11). A cell analysis device, comprising: an analysis unit (12) for collating and classifying information. 2. The cell analysis system according to claim 1, wherein the image processing unit extracts specific color information such as chromaticity, lightness, and chroma of the cytoplasm and the nucleus. apparatus. 3. The cell analyzer according to claim 1, wherein the number and area of the nuclei (7) and the micronuclei (10) are measured in the image processing unit (9). 4. A stage section (3) for placing a specimen (2) of an extracted cell, and an image capturing section for photographing the specimen.
(4) an image processing unit (9) for converting the captured image into a measurement image, performing a predetermined measurement from the processed image, and performing a predetermined operation on all of the measured data or only the predetermined data. Characterized in that it is configured to obtain various management data by performing the arithmetic processing of (1). 5. The image processing unit (9) performs a process of extracting grains generated in cells and nuclei as color information such as chromaticity, lightness, and saturation. Item 5. The cell analysis device according to item 4. 6. The image processing section (9), wherein the number of grains in a nuclear area extracted from the color information and the number of grains in a predetermined cytoplasmic area located around the nuclear area or a number of grains in a predetermined number of cytoplasmic areas are determined. The cell analyzer according to claim 5, wherein the average number of grains is automatically measured. 7. The image processing unit (9) extracts chromaticity, lightness, saturation, etc. of each part of the cell as color information, and extracts a DNA core, a DNA damaged part, and a background part other than DNA from the color information. The identification process is performed, and the coordinates of a predetermined portion of the cell image displayed on the display means are displayed via the color information via a predetermined pointing device and set as a measurement area. The cell analyzing apparatus according to claim 4, wherein the cell analyzing apparatus is configured to perform the following. 8. The cell analysis device is provided with an information section (11) in which pattern data of a plurality of cells classified in advance based on predetermined conditions is recognized. Various management data obtained by subjecting all of the data measured from the cell specimen (2) or only predetermined data to predetermined arithmetic processing and cell pattern data in the information section (11) are compared with each other. The cell analyzer according to any one of claims 4 to 7, further comprising an analysis unit (12) for classifying the cell specimen (2). 9. A sample (2) is prepared by extracting cells,
(2) is arranged on the stage section (3), photographed by the image capturing means, the captured image is converted into a measurement image, a predetermined measurement is performed, and the measured data and a plurality of patterns which have been subjected to pattern recognition in advance. A cell analysis method characterized by performing cell analysis by collating and classifying the information with pattern information of cells. 10. The method according to claim 10, wherein the capturing of the image
10. The cell analysis method according to claim 9, wherein specific color information such as chromaticity, lightness, and saturation of (6) and the nucleus (7) is extracted. 11. In the measurement, the nucleus (7) and the micronucleus (1
The cell analysis method according to claim 9 or 10, wherein predetermined required items such as the number, area, and color of (0) are measured. 12. A sample (2) is prepared by extracting cells, the sample (2) is arranged on a stage (3), photographed by an image capturing means, and the captured image is converted into a measurement image. A predetermined measurement, and performing predetermined calculation processing on all of the measured data or only predetermined data to obtain various management data. 13. The image for measurement is obtained by extracting grains of a predetermined substance taken into a nucleus of a cell as color information such as chromaticity, lightness, and saturation. The cell analysis method as described above. 14. An average number of the number of grains in a nuclear area extracted as color information and the number of grains in a predetermined cytoplasmic area located around the nuclear area or the number of grains in a predetermined number of cytoplasmic areas in the measurement. The cell analysis method according to claim 13, wherein is automatically measured. 15. The image for measurement includes chromaticity of each part of a cell,
The brightness, saturation, and the like are obtained by extracting the color information, and a process of identifying a DNA core portion, a DNA damaged portion, and a background portion other than DNA is performed from the color information, and then displayed on the display means via the color information. 13. The cell analysis method according to claim 12, wherein coordinates of a predetermined part of the obtained cell image are designated and set as a measurement area, thereby automatically measuring characteristic values of each part of the cell. 16. A method in which the cell analysis apparatus performs various arithmetic data obtained by performing predetermined arithmetic processing on all of the measured data or only predetermined data, and a plurality of management data that are pre-ranked based on predetermined conditions. The cell analysis method according to any one of claims 12 to 15, wherein the specimen (2) is ranked by comparing with the cell pattern data.