JP2010127807A - Method for improving power of test for formed element - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for improving power of a test for a formed element in a device to automate the whole of operation or a part thereof associated with formed element analysis (urinary sediment analysis or the like) in a liquid to be inspected, specially urine, without using a flow cell having a pollution risk by a carry-over and a stain with precision and accuracy, and a method therefor. <P>SOLUTION: A method for analysis of a formed component includes the steps of stirring the liquid to be inspected in a sample container, dispensing the liquid to be inspected stirred, dispensing the liquid in a gap of a covered translucent plate integrated translucent plate, imaging the liquid on the translucent plate on an imaging stage, enlarging the sample image of the formed component in the liquid, automatically focusing the sample image of the formed component, capturing the image, and processing the image taken to discriminate into various components. In the method for analysis of a formed component, the method for improving power of a test for a formed element in a liquid to be inspected, in which centrifugal operation of a covered translucent plate integrated translucent plate is performed while the liquid to be inspected is dispensed in a gap of the covered translucent plate integrated translucent plate and a formed element is concentrated. <P>COPYRIGHT: (C)2010,JPO&INPIT

Description

The present invention relates to an apparatus and method for analyzing a formed component in a test liquid, and in particular, for automatically analyzing formed components such as blood cells contained in urine and blood using an image analysis technique. The present invention relates to an apparatus and a method.

Conventionally, analysis of a solid component in a test liquid, for example, analysis of urinary sediment components, [1] centrifuge a urine sample, [2] remove the supernatant using an aspirator or pipette, or by decantation [3] Apply a certain amount of the remaining residue component to the glass slide and use the cover glass as a specimen. [4] Set on a microscope and form components (blood cells, epithelial cells, columnars, microorganisms. Class, crystal salt, etc.). Currently, all of this work is performed manually, which is a heavy burden on the inspection engineer, and because the work is manual, there are large variations, and of course there are individual differences in judgment, and accuracy The problem was also pointed out.

In recent years, as a device that automates these inspection operations, after preparing a coating solution of a specimen without mixing a sample solution, the staining solution is mixed and then suspended, and then flowed through a flow cell, physicostatistical methods, optical methods, etc. There is a method of automatic analysis by the flow cytometer method of analyzing by (Patent Documents 1 to 3).
JP-A-4-337460 JP-A-5-296915 JP-A-5-322885

However, in the method of automatic analysis by the above flow cytometer method, the formed components are always flowing, so it is very difficult to focus on each, and the focus is fixed at a fixed position. The flow cell has a certain thickness. From these things, when the above-mentioned flow cytometer method can accurately measure only the formed component that has flowed to the focused position, and further, when the formed component is located a little away from the position May be blurred as much as the focus shifts, and an image measurement result different from that at the time of focusing may be obtained, resulting in an erroneous determination result.

In this method, since the flow cell is fixed and continuously used, there is a carry-over for the previous sample, and there is a problem in accuracy and precision. In addition, in order to increase the analysis accuracy, when staining a test solution with a staining solution, there is naturally contamination of the flow cell with the staining solution, and the components of the staining solution accumulate, so noise increases over time and the measurement accuracy increases. There is a risk of decline. Furthermore, in a physical statistical measurement method (such as measurement of the scattering intensity of a test component using a laser beam), separation occurs from image observation data obtained by microscopic observation, and the microscopic image must be observed again for the separated specimen. There was a problem.

As another means, there is a method in which a test liquid containing a formed component is placed on a light-transmitting plate without flowing, and this is imaged (Patent Document 4). This method eliminates the shortcomings of flow cytometry, automates all or part of the work involved in the analysis of specimen liquids, especially urine sediments (such as urine sediment analysis). It is possible to provide an apparatus and a method with high precision and accuracy without using a flow cell having a risk of contamination.
Japanese Patent No. 3924870

  However, although not as much as the flow cell, the translucent plate has a certain thickness. For these reasons, components that are difficult to settle, such as red blood cells and bacteria, float, etc., and if the formed component is located slightly away from the bottom surface, which is the imaging position, the component will be blurred by the amount of defocus. Cannot be recognized and the detection power may be reduced. Thereby, it may be measured less than the actual number of formed portions. In order to prevent this, there are only methods such as multiplying the actual measurement value by a fixed correction value or providing a stationary time for allowing the measurement to settle until the start of measurement, which is an error factor.

  The present invention automates all or part of the work involved in the analysis of liquids (especially urine sediment analysis) in test liquids, especially urine, and uses a flow cell that has a risk of contamination with carryover or staining liquid. It is an object of the present invention to provide a method for improving the detectability of a formed component in an apparatus and method with high precision and accuracy.

  As a result of intensive studies, the present inventors performed a centrifugal operation of the coated light transmitting plate-integrated light transmitting plate in a state where the test liquid was dispensed into the gap portion of the coated light transmitting plate integrated light transmitting plate, It has been discovered that the power of detection can be improved by concentrating the formation, and the above object has been achieved.

That is, the present invention has the following contents.
[Claim 1]
(1) a step of stirring the test liquid in the sample container;
(2) Dispensing the stirred test solution;
(3) Dispensing the test liquid into the gap portion of the coated translucent plate integrated translucent plate;
(4) imaging the test liquid on the translucent plate with an imaging stage;
(5) a step of enlarging the sample image of the formed component in the test solution;
(6) a step of automatically focusing the sample image of the formed component;
(7) capturing the image;
(8) In the formation analysis method which has the process of processing the imaged image and discriminating into various components,
(9) A step of concentrating the formed component by centrifuging the coated light transmitting plate-integrated light transmitting plate in a state where the test liquid is dispensed into the gap portion of the coated light transmitting plate integrated light transmitting plate. A method for improving the detectability of a formed component in a test liquid, characterized by comprising:
[Section 2]
(3) In the step of dispensing the test liquid into the gap portion of the coated translucent plate integrated translucent plate,
(10) The method for improving the detectability of the formed component according to item 1, comprising a step of mixing with a staining solution before dispensing.
[Section 3]
Item 2. The method for improving the detectability of a formed component according to Item 1, wherein the formed component is any one selected from the group consisting of red blood cells, white blood cells, epithelial cells, and columnars.
[Claim 4]
In addition to epithelial cells, squamous epithelial cells, transitional epithelial cells, tubular epithelial cells, round epithelial cells, urethral columnar epithelial cells, prostate epithelial cells, seminal vesicle epithelial cells, endometrial epithelial cells, oval round fat pad, intracytoplasmic Item 4. The method for improving the detectability of a formed component according to Item 3, wherein the cells are classified into cells containing one or more selected from the group consisting of inclusion cells and polygonal cells.
[Section 5]
Glass cylinder, epithelial cylinder, granule cylinder, waxy cylinder, fat cylinder, erythrocyte cylinder, leukocyte cylinder, cell cylinder, nitric leukocyte cylinder, hemoglobin cylinder, hemosilylidene cylinder, myoglobin cylinder, amyloid cylinder, protein cylinder, vacuole deformation Item 4. The method for improving the detectability of a formed component according to Item 3, wherein the cells are classified into cells containing one or more selected from the group consisting of a column, a platelet column, a bacterial column, a bilirubin column, and a salt column.
[Claim 6]
(1) means for stirring the test liquid in the sample container;
(2) means for dispensing the stirred test solution;
(3) means for dispensing the test solution into the gap portion of the coated translucent plate integrated translucent plate;
(4) an imaging stage for imaging the test liquid on the translucent plate;
(5) means for enlarging the sample image of the formed component in the test solution;
(6) It has a function to automatically adjust the focus of the sample image of the formed object,
(7) means for capturing the image;
(8) In the formed component analyzer having means for processing the captured image and identifying the various components,
(9) Means for concentrating the formed component by centrifuging the coated light transmitting plate-integrated light transmitting plate in a state where the test liquid is dispensed into the gap portion of the coated light transmitting plate integrated light transmitting plate A formed component analyzer characterized by having.
[Claim 7]
(3) In the step of dispensing the test liquid into the gap portion of the coated translucent plate integrated translucent plate,
(10) The formed component analyzer according to item 6, including means for mixing with a staining solution before dispensing.
[Section 8]
(11) The formed component analyzing apparatus according to item 6, comprising means for processing the captured image and storing the result of identifying the various components.
[Claim 9]
(8) The means for processing the captured image and identifying it as various components has the function of calculating the analysis result from all the discrimination results for the preset visual field and the function of outputting the analysis result from the output device. 7. The formed component analyzer according to item 6.
[Section 10]
(8) The tangible component analyzing apparatus according to item 6, wherein the means for processing the captured image and identifying the various components has a learning recognition function.
[Section 11]
(8) The formed component analyzing apparatus according to item 6, wherein the means for processing the captured image to identify various components learns and identifies the range of the feature amount of the formed component.
[Claim 12]
(8) Item 6 wherein the means for processing the captured image to identify various components has a function of calculating the amount of the formed component based on the optical feature amount attributed to the formed component. The formed component analyzer.
[Claim 13]
(5) The formed component analyzer according to item 6, wherein the means for enlarging the sample image of the formed component in the test solution has two or more magnifications.
[Section 14]
Item 7. The formed component analyzer according to Item 6, wherein the material of the light transmitting plate and the coated light transmitting plate is plastic.
[Section 15]
Item 7. The component analysis apparatus according to Item 6, wherein the component is selected from the group consisting of red blood cells, white blood cells, epithelial cells, and columnars.
[Section 16]
In addition to epithelial cells, squamous epithelial cells, transitional epithelial cells, tubular epithelial cells, round epithelial cells, urethral columnar epithelial cells, prostate epithelial cells, seminal vesicle epithelial cells, endometrial epithelial cells, oval round fat pad, intracytoplasmic Item 16. The tangible component analyzer according to Item 15, which is classified into cells containing one or more selected from the group consisting of inclusion body cells and polygonal cells.
[Section 17]
Glass cylinder, epithelial cylinder, granule cylinder, waxy cylinder, fat cylinder, erythrocyte cylinder, leukocyte cylinder, cell cylinder, nitric leukocyte cylinder, hemoglobin cylinder, hemosilylidene cylinder, myoglobin cylinder, amyloid cylinder, protein cylinder, vacuole deformation Item 16. The tangible component analyzer according to Item 15, which is classified into cells containing one or more selected from the group consisting of a column, a platelet column, a bacterial column, a bilirubin column, and a salt column.
[Section 18]
The staining solution is Sternheimer-Malbin staining method, Sternheimer staining method, Prescott-Brodie staining method, Behre-Mühlberg staining method, Sudan III staining method, Lugol staining method, hemosiderin staining method, Papanicolau staining method, Papanicolau staining method Field staining method, Quaglino-Flemans method, Kaplow method, Sato / Sekiya method, Berlin blue method, Giemsa staining method, Wright staining method, Pappenheim staining method, Congo red staining method, methyl green / pyronine staining method, alcian blue staining method , Shawl staining method, Foilgen staining method, oil red O staining method, Brecker method, Heinz body staining method, neutral red / Janus green super-vivo staining method, brilliant cresyl blue Item 8. The formed component analyzer according to Item 7, which is a reagent containing one or more of the components used in at least one of the staining methods.
[Section 19]
(12) The tangible component analyzer according to item 6, further comprising means for calculating the amount of the tangible component based on the optical feature amount attributed to the tangible component.
[Section 20]
Item 20. The formed component analyzer according to Item 19, wherein the optical feature is the amount of light emitted by the reagent for ATP measurement.
[Claim 21]
Item 7. The formed component analyzing apparatus according to Item 6, wherein at least one material of the light-transmitting plate and the coated light-transmitting plate is glass, plastic, chemically-treated glass, or chemically-treated plastic.
[Item 22]
Item (1) (2) (3) (9) (4) (5) (6) (7) (8) To improve the detectability of formed components.
[Section 23]
(1) (2) (3) (9) (4) (5) (6) (7) The item according to item 6, further comprising means for automatically performing each step in order. Formation analysis device.

According to the tangible component analysis apparatus and the tangible component analysis method of the present invention, the work related to the analysis of the tangible component in the test liquid is simplified, and the burden on the inspection engineer can be reduced. In addition, since the object to be measured is not a fluid, the sample image can be easily focused, and since a translucent plate is used without using a flow cell, a disposable one can be used. There is no possibility of contamination of the previous specimen with carryover or staining solution.
That is, according to the present invention, it is possible to provide an analysis result with high precision and accuracy.

The present invention can be applied to analysis of body fluids such as urine, blood, serum, plasma, semen, semen, prostate fluid, joint fluid, pleural effusion, ascites, secretory fluid, and water quality tests. In particular, it is effectively used for analysis of urine such as raw urine, concentrated urine, and sediment after centrifugation.

In the present invention, the component to be analyzed is not particularly limited as long as it is dispersed or suspended in the test solution. For example, when the test solution is urine, red blood cells (deformed red blood cells, various red blood cells), white blood cells, epithelial cells (squamous epithelial cells, transitional epithelial cells, tubular epithelial cells, circular epithelial cells, urethral columnar epithelial cells, prostate epithelium Cells, seminal vesicle epithelial cells, endometrial epithelial cells, oval fat pad, intracytoplasmic inclusion body cells, polygonal cells, etc.), cylinders (glass cylinder, epithelial cylinder, granule cylinder, waxy cylinder, fat cylinder) , Erythrocyte cast, leukocyte cast, cell cast, hyaline leukocyte cast, hemoglobin cast, hemosilidene cast, myoglobin cast, amyloid cast, protein cast, vacuolar deformed cast, platelet cast, bacterial cast, bilirubin cast, salt cast, etc.), microorganisms (Fungi, bacteria, fat globules, protozoa, trichomonas, sperm, etc.), crystals and salts (uric acid salt, phosphate salt, calcium oxalate, bilirubin, cystine, cholesterol) , 2,8-dihydroxy-adenine crystals, etc.), other (intranuclear inclusions cells, fat granule cells, macrophages, atypical cells), and the like as solid components to be analyzed.

The apparatus and method of the present invention are characterized in that a test liquid is placed on a translucent plate. The light-transmitting plate may be any material as long as it has a light-transmitting property and can place a test solution, and examples thereof include a slide glass. Since the slide glass is single-use (disposable), there is no possibility of carry-over of the previous specimen or contamination by a staining agent, and a highly reliable measurement result can be provided. The material of the translucent plate is not particularly limited as long as it has translucency such as plastic (synthetic resin) and glass. In the case of plastic, chemical treatment for improving hydrophilicity is preferably performed as necessary.

The test liquid placed on the translucent plate may be covered with a coated translucent plate. An example of the coated translucent plate is a cover glass. Examples of the material of the coated translucent plate include the same materials as those described above, but are not particularly limited.
FIG. 1 is a perspective view of a cover glass-integrated slide glass 1 in which a cover glass serving as a coated translucent plate and a slide glass serving as a translucent plate are integrally formed. In FIG. 1A, two opposite sides of the cover glass portion 3 placed on the slide glass portion 2 are sealed with an adhesive 4 or the like, and the remaining two opposite sides are in an open state. In FIG. 1B, three sides of the cover glass portion 3 placed on the slide glass portion 2 are sealed with an adhesive 4 or the like, and the remaining one side is in an open state. When the test solution is dispensed from the opened side, the test solution is injected into the gap between the slide glass portion 2 and the cover glass portion 3 by capillary action. That is, the test liquid is placed on the slide glass portion 2 which is a light transmitting plate.
If the cover glass-integrated slide glass 1 is used in this way, a predetermined amount can be easily and accurately injected, and laborious sample preparation steps for setting the cover glass can be saved.

The imaging stage for imaging the test liquid on the translucent plate is not particularly limited as long as it can mount the translucent plate, but is movable so that the imaging position can be changed. Is preferred. Although the movement may be performed manually, it is preferably mechanically performed using, for example, a servo motor, a stepping motor, a linear motor, or the like.

Examples of the imaging means include a digital camera and a CCD color video camera. Further, it is preferable to add a function (autofocus function) for automatically focusing the formed specimen image to the imaging means. The magnifying means may optically magnify the specimen image before imaging, or may magnify the captured specimen image by digital processing or the like. Specifically, a zoom lens, an objective lens, or the like attached to the camera can be used.

The formed component in the test solution may range from several μm in size such as blood cells and bacteria to several hundred μm in size such as a cylinder. Accordingly, it is preferable that the magnification means has two or more magnifications rather than only one magnification. In this case, it is possible to analyze more accurately from a small formed component to a large formed component. Further, the magnification may be continuously changed. What is necessary is just to determine an enlargement magnification suitably according to a formation part.

The identifying means classifies and identifies formed components in the captured image based on the form and the like. It is preferable to add a function for calculating the analysis result from all the identification results for the field of view set in advance and a function for outputting the analysis result from the output device to the identification means. Note that the preset visual field here refers to the number of visual fields (images) to be captured. The identification means preferably includes a memory for temporarily storing the captured image.

Examples of the identification means include a computer programmed to perform the above-described identification, an identification device configured with a logic circuit, and the like. Of these, the use of a computer as the identification means is preferable because all the operations such as the operation of each process, image processing, storage, calculation, and output can be performed on software.

The identification means preferably has a learning function. By having a learning recognition function, a measurement result with high accuracy and precision is provided. The identification means includes [1] color extraction range specification for separating red, green, and blue into lightness and chromaticity, [2] binary image processing range specification consisting of filling in a hole, writing a line segment, and separating an image, [3] Image features (area, circularity coefficient, equivalent circle diameter, circumference length, absolute maximum length, ferret diameter X / Y ratio, maximum chord length X / Y ratio, minor axis length / major axis length ratio Etc.) can be learned and identified.

In addition, the above image processing by the identification means classifies and analyzes the formed component based on the form of the formed component by software. If the morphology of renal epithelial cells, red blood cells, white blood cells, bacteria, etc. is similar, the analytical ability will inevitably decrease.

Therefore, in the apparatus of the present invention, a means for adding a reagent such as a staining agent for assisting component identification to the test solution is further added to the apparatus of the present invention. By adding a step of adding a reagent such as a staining agent for assisting in component identification to the liquid, it is possible to classify components similar to the above-described form and improve the analysis ability.

The reagent is not particularly limited, but generally known methods include Sternheimer-Malbin staining method (SM staining method), Sternheimer staining method (modified method of S staining method, NS staining method or Sternheimer staining method), Prescott-Brodie. Staining method, Behr-Muhlberg staining method (BM staining method), Sudan III staining method, Lugor staining method, hemosiderin staining method, Papanicolaou staining method, 4-chloro-1-naphthol method, Field staining method, Quaglino-Klemans method , Sato / Sekiya method, Berlin blue method, Giemsa staining method, Wright staining method, Pappenheim staining method, Congo red staining method, Methyl green / pyronine staining method, Alcian blue staining method, Shawl Staining method, Foilgen staining method, oil red O staining method, Brecker method, Heinz body staining method, neutral red / Janus green superbiological staining method, brilliant cresyl blue staining method ("Clinical laboratory atlas 1 urinary sediment", " Clinical Laboratory Technology Complete Book 3 “Blood test”, “Proposal for clinical test method”, “All dyeing methods: MEDICAL TECHNOLOGY separate volume”), etc., which contain one or more components used in at least one method Can be mentioned.
For example, you can select several dyes and additives from multiple types of dyes and additives used in one dyeing method, and combine them to use as a reagent, or use them in different dyeing methods. A combination of two or more types of staining agents may be used as a reagent.

In addition, in order to improve the storage stability and antiseptic performance of the staining agent, the above-mentioned staining agent includes generally known antiseptic (antibacterial) agents (various antibiotics, EDTA salts, boric acid, citric acid, NaN ₃ , Procrine, benzisothiazolone, pyrithione, N-methylisothiazole, etc.) may be added. In addition, various buffer solutions are added to keep the staining solution at an optimum pH, and various salts (EDTA salts, carbonates, oxalates, citrates, NaCl, KCl to maintain the form of the formed components) , CaCl ₂ , AlCl _3, etc.), various sugars (glucose, fructose, galactose, maltose, xylitol, sorbitol), cyclodextrins, and glutaraldehyde may be added. Furthermore, various surfactants and enzymes may be added to remove insoluble substances that hinder measurement.

Furthermore, when sufficient results cannot be expected by classification analysis according to form, the apparatus of the present invention has a function of calculating the amount of the formed component based on the optical feature amount attributed to the formed component in the identifying means. In the method of the present invention, it is preferable to calculate the amount of the formed component based on the optical feature amount attributed to the formed component.

For example, when measuring the amount of microorganisms such as bacteria in the test solution, a luminescent reagent can be added to the test solution for the purpose of further improving analysis accuracy. However, since the degree of staining of bacteria varies depending on the state (dead bacteria, viable bacteria), the analytical ability is lowered. Therefore, in order to further improve the analytical performance, the amount of bacteria is accurately measured by adding the step of measuring ATP in the test solution by utilizing the production of ATP (adenosine triphosphate) by microorganisms. be able to. ATP can be measured by using an ATP reagent containing luciferase, luciferin, and the like and detecting the amount of luminescence (optical feature amount) corresponding to the amount of ATP (see Chemical Formula 1).

Further, the apparatus of the present invention is preferably provided with means (image storage device) for storing the result of processing the captured image and identifying the various components. Examples of means for storing include auxiliary storage devices such as a magneto-optical disk, a fixed disk, a digital video disk, and a CD-R having a large storage capacity. Examples of the output device used in the device of the present invention include output devices such as a CRT display, a liquid crystal display, and a printer, and the auxiliary storage device.

When a display is used as the output device, depending on the menu selection, in addition to measurement results and image data, time, current device status (measurement status of each sample, measurement end scheduled time of each sample or selected sample or A function that displays the required remaining time, amount of waste liquid in the waste liquid tank, remaining amount of pure water tank, remaining amount of each reagent, remaining amount of detergent, remaining number of slide glass, etc. A function of enlarging the display so that it can be read from the image may be added.

In the apparatus of the present invention, the identification means has a function of classifying the formed component that cannot be classified into an item of “other components”, or its image is called later, and an engineer makes a direct visual judgment, and the judgment result is Functions that can be added or modified to the data may be added.

The apparatus of the present invention may be added with a function that allows remote control of selection of a display menu on the display and operation of the apparatus using a remote control device. Furthermore, in the apparatus of the present invention, when any accident occurs, when the waste liquid tank becomes full, when the remaining of pure water, each reagent, detergent, slide glass, etc. decreases, a screen display, A function of issuing a warning by sound or signal may be added.

The apparatus according to the present invention has a function of giving priority to an emergency sample (a sample requiring urgent analysis) next to a sample being analyzed in order to cope with an urgent analysis, or an emergency sample immediately after pausing the analysis. A function for analyzing the above may be added. In addition, since an urgent analysis needs to be performed quickly, it is preferable to install an emergency analysis button, for example, so that the apparatus can perform an urgent analysis only by operating the button.

The centrifugal operation of the present invention is not particularly limited as long as the light-transmitting plate can be centrifuged, but can be easily performed by a centrifugal apparatus as shown in FIG. The light transmitting plate in which the test solution is dispensed in the gap portion of the coated light transmitting plate-integrated light transmitting plate is automatically or manually applied to the recess 27 having a size that can be accommodated by the light transmitting plate provided in the rotor 25. Then, the centrifugal operation is performed around the rotating shaft 26. Further, if a balance 24 is provided in advance so as to be in contact with the translucent plate, the operation is easy. At this time, the direction of the rotor, the radius of rotation, and the direction of rotation are not limited, but the erection direction of the translucent plate floats on the translucent plate bottom by erection so that the bottom side faces the centrifugal direction (outward). Collect the solid and concentrate. In addition, a plurality of recesses 27 may be arranged so that a plurality of light transmitting plates can be centrifuged at the same time, and the rotation time and rotation speed can be arbitrarily set. Preferably, the rotation time is 0.5 to 5 minutes, The rotation speed is 500G to 3000G.

The method of the present invention can be easily performed by using the above-described present invention in an apparatus. That is,
(1) a step of stirring the test liquid in the sample container;
(2) Dispensing the stirred test solution;
(3) Dispensing the test liquid into the gap portion of the coated translucent plate integrated translucent plate;
(9) a step of concentrating the formed component by centrifuging the coated translucent plate integrated translucent plate in a state where the test liquid is dispensed into the gap portion of the coated translucent plate integrated translucent plate; ,
(4) imaging the test liquid on the translucent plate with an imaging stage;
(5) a step of enlarging the sample image of the formed component in the test solution;
(6) a step of automatically focusing the sample image of the formed component;
(7) capturing the image;
(8) processing the captured image and identifying the various components;
It is a formed-parts analyzer which has a function which carries out each process continuously and automatically in the order of the above-mentioned description. Thus, if the apparatus of this invention is used, the method of this invention can be performed fully automatically.

Hereinafter, embodiments of the apparatus and method of the present invention will be described with reference to the drawings. However, the apparatus and method of the present invention are not limited to these embodiments.

Example 1
FIG. 2 is a diagram showing an example of the component analyzer of the present invention, and shows an analyzer of urine sediment components.
In the example of FIG. 2, a slide glass 5 is used as the translucent plate. As shown in FIG. 1A, the slide glass 5 is formed integrally with the cover glass to form a cover glass-integrated slide glass. The slide glass 5 is placed on an XY table 6 serving as an imaging stage. The XY table 6 incorporates a step motor as a drive source. The XY table 6 is driven in response to a signal from the drive circuit 11, and moves and stops the slide glass 5 freely in the X coordinate axis direction and the Y coordinate axis direction to change the imaging position.

A CCD camera 10 with an autofocus function is used as the imaging means. An objective lens is attached to the CCD camera 10 as an enlargement means.
The identification means includes an image processing control circuit 12, an image memory 14, a feature extraction circuit 15, an identification calculation circuit 16, a central control unit 17, and a calculation circuit 22. In the figure, the identification means is surrounded by a dotted line. Reference numeral 23 denotes a display used as an output device. Reference numeral 18 denotes an image storage device for storing the results identified for various components. Next, processing performed in the apparatus of the present invention will be described in time series.

First, a required amount of a urine sample as a test solution is dispensed from the sample container into the three reaction tubes 19a, 19b, and 19c with a probe. In order to dispense more uniformly, the urine sample is agitated before dispensing. If a container with a barcode attached in advance is used as the sample container, the barcode is read in advance with a barcode reader. If the presence / absence, type, and specimen number of a sample are read by using a barcode, the subsequent processing data and the sample can be easily identified.
Next, a required amount of the dyeing solution is injected into the reaction tube 19a, and the formed portion is dyed. After about 2 minutes, the reaction solution of the test solution and the staining solution is collected from the reaction tube 19a and injected into the cover glass-integrated slide glass (slide glass 5). The dyeing reaction time and reaction temperature are arbitrarily set.

Next, the dispensed cover glass-integrated slide glass (slide glass 5) is automatically or manually stored in the centrifuge of FIG. 3, and the centrifugal operation is performed around the rotary shaft 3. About the installation direction of a translucent plate, the formation part which has floated on the translucent plate bottom is collected and concentrated by installing so that a bottom side may face a centrifugal direction (outward direction). The rotation time is 0.5 minutes and the rotation speed is 1000G. The cover glass-integrated slide glass (slide glass 5) that has been centrifuged is moved manually or automatically to a position where the next imaging step is performed.

Next, in order to image the test solution, light is irradiated from the lamp 7 serving as a light source. The light (dotted line portion) irradiated from the lamp 7 travels on the optical axis, passes through the condenser lens 8, and is condensed on the test liquid on the slide glass 5. The formed sample image in the test solution is formed at the imaging position by the objective lens 9, and the sample image at this imaging position is projected as an image on the imaging surface of the CCD camera 10 and subjected to photoelectric conversion.

The image processing control circuit 12 controls the drive circuit 11 to control the movement or stop of the XY table 6 and causes the CCD camera 10 to take an image when the XY table 6 stops. An image obtained by the CCD camera 10 is digitized by the A / D converter 13. The image processing control circuit 12 stores the digitized image data in the image memory 14.

After completing the storage in the image memory 14, the image processing control circuit 12 further moves the XY table 6 to change the imaging position (field of view), and performs imaging, A / D conversion, and storage in the memory in the same manner as described above. . The image processing control circuit 12 repeats this series of operations until the number of fields of view set in advance is reached. The movement of the XY table 6 for changing the imaging position may be controlled to be performed after the image analysis for one field of view is completed.

Next, the image processing control circuit 12 inputs the image data stored in the image memory 14 to the feature extraction circuit 15. The feature extraction circuit 15 uses the feature amount of the image (for example, area of formation, circularity coefficient, equivalent circle diameter, circumference length, absolute maximum length, Ferret diameter X / Y ratio, maximum chord length X / Y ratio, short Axis length / major axis length ratio etc.) are extracted as primary parameters. The image processing control circuit 12 inputs these primary parameters and secondary parameters generated by the combination calculation to the identification calculation circuit 16.

The identification operation circuit 16 classifies the formed component using a neural network. The neural network performs learning using a large amount of data in advance based on an expert's judgment and optimizes the coupling coefficient between the neurons. Therefore, the discrimination calculation circuit 16 performs a neural network calculation using the input primary and secondary parameters, and automatically classifies the formed component as a target. Instead of the neural network calculation, the identification calculation circuit 16 may perform automatic classification of the formed component using the statistical learning recognition method.

The central control unit 17 stores the classification result and the image data in the image storage device 18. In this embodiment, a magneto-optical disk having a large storage capacity is used as the image storage device 18.

On the other hand, in order to measure the amount of microorganisms such as bacteria in the urine specimen, trichloroacetic acid is added to the reaction tube 19b, a part of the specimen sample is taken, and an ATP measurement reagent is added to emit light. In the reaction tube 19c, without adding trichloroacetic acid, a part of the sample sample is taken as a blank, and the ATP measurement reagent is added to emit light in the same manner as described above. Luminescence is detected by the luminometer 20. The integrating circuit 21 counts and integrates light emission detected by the luminometer 20 within a predetermined time. Next, the arithmetic circuit 22 converts the information on the number of emitted light (number of photons) integrated by the integration circuit 21 into an ATP amount based on a calculation formula calculated in advance by a standard product of ATP, and calculates the number of microorganisms. The calculation result is input to the central control unit 17.

The central control unit 17 integrates the classification results for each visual field of the captured image for all visual fields, and together with the number of microorganisms input from the arithmetic circuit 22, the boundary value (standard value for formation) input in advance. Is converted into qualitative data (for example,-, ±, +, ++, ++, etc.), and the obtained qualitative data and image data are output to an output device 23 such as a display or a printer.

FIG. 4 is a diagram showing an example of a centrifuge arranged in the formed component analyzer. Hereinafter, FIG. 4 will be described for each step.

[Sample preparation process]
This step is a step of placing the test liquid on the translucent plate. In this step, 0.015 ml is dispensed from the test solution by the dispensing unit 28 and dispensed into the gap portion of the cover glass-integrated slide glass to prepare a sample.

[Centrifuge process]
The prepared specimen was moved to the light-transmitting plate housing recess 27 of the rotor 25 by the slide push-out rod 29, and centrifuged at a rotation speed of 1000 G for a rotation time of 0.5 minutes.

[Sample movement process]
The prepared specimen is set on the XY table 6 (imaging stage) with the slide push-out rod 29. The imaging stage moves the set specimen to the imaging position.

[Imaging process]
This step is a step of enlarging the sample image of the formed component in the test solution and a step of automatically focusing the enlarged sample image of the formed component and capturing the sample image of the formed component. . In this step, first, the specimen image is magnified by the objective lens 9 (magnifying means) attached to the CCD camera 10. Next, the CCD camera set on the imaging stage automatically focuses and takes an enlarged image.

[Image processing and storage process]
In this step, as in the example of FIG. 4, the captured image is stored, the urine components are classified into various components from the image data, and counted for each component. The classification result and coefficient result are transmitted to the control calculation process. The imaging process and the image processing / storing process are repeated until the number of fields of view reaches 100 while changing the imaging position by moving the imaging stage.

[Control calculation process]
In this step, the results obtained from the image processing / storage step are integrated and output to a display, a printer, or the like.

Using the above-mentioned apparatus, a U-SCANNER dedicated quality control reagent (manufactured by Toyobo Co., Ltd.), which was visually adjusted to 200 / μl, was measured.
As a comparative example, the other measurements were performed in the same manner under the condition that the actual measurement value was not multiplied by a certain correction value without a centrifugal operation. The results are shown in Table 1.

Particles that are floating and could not be recognized by the present invention can be measured, and the number of measurements is almost the same as that of visual observation without the need to multiply the actual measurement value by a fixed correction value, and the detection power is improved.

As described above, according to the present invention, it is possible to improve the detection power of the formed component in the sample, and it is possible to observe or analyze reliably, quickly and simply. For this reason, industrial applicability exists in a wide range of fields such as pharmaceuticals, foods, and environmental fields as well as medical fields such as medical and diagnostic devices.

1 is a perspective view of a cover glass-integrated slide glass 1. FIG. It is a figure which shows an example of the formation component analyzer of this invention. It is a figure which shows the example of the centrifuge of this invention. It is a figure which shows an example of the centrifuge which has been arrange | positioned in the tangible component analyzer of this invention.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Cover glass integrated type slide glass 2 Slide glass part 3 Cover glass part 4 Adhesive 5 Slide glass 6 XY table 7 Lamp 8 Condenser lens 9 Objective lens 10 CCD camera 11 Drive circuit 12 Image processing control circuit 13 A / D converter 14 Image memory 15 Feature extraction circuit 16 Identification calculation circuit 17 Central control unit 18 Image storage device 19 Reaction tube 20 Luminometer 21 Accumulation circuit 22 Calculation circuit 23 Output device 24 Balance 25 Rotor 26 Rotating shaft 27 Translucent plate storage recess 28 Dispensing part 29 Slide Extrusion Bar 30 Slide Conveying Unit

Claims (23)

(1) a step of stirring the test liquid in the sample container;
(2) Dispensing the stirred test solution;
(3) Dispensing the test liquid into the gap portion of the coated translucent plate integrated translucent plate;
(4) imaging the test liquid on the translucent plate with an imaging stage;
(5) a step of enlarging the sample image of the formed component in the test solution;
(6) a step of automatically focusing the sample image of the formed component;
(7) capturing the image;
(8) In the formation analysis method which has the process of processing the imaged image and discriminating into various components,
(9) A step of concentrating the formed component by centrifuging the coated light transmitting plate-integrated light transmitting plate in a state where the test liquid is dispensed into the gap portion of the coated light transmitting plate integrated light transmitting plate. A method for improving the detectability of a formed component in a test liquid, characterized by comprising: (3) In the step of dispensing the test liquid into the gap portion of the coated translucent plate integrated translucent plate,
(10) The method for improving the detectability of the formed component according to claim 1, comprising a step of mixing with a staining solution before dispensing.     The method for improving detectability of a formed component according to claim 1, wherein the formed component is any one selected from the group consisting of red blood cells, white blood cells, epithelial cells, and columnars.     In addition to epithelial cells, squamous epithelial cells, transitional epithelial cells, tubular epithelial cells, round epithelial cells, urethral columnar epithelial cells, prostate epithelial cells, seminal vesicle epithelial cells, endometrial epithelial cells, oval round fat pad, intracytoplasmic The method for improving detectability of a formed component according to claim 3, wherein the method comprises classifying cells into one or more cells selected from the group consisting of inclusion body cells and polygonal cells.     Glass cylinder, epithelial cylinder, granule cylinder, waxy cylinder, fat cylinder, erythrocyte cylinder, leukocyte cylinder, cell cylinder, nitric leukocyte cylinder, hemoglobin cylinder, hemosilylidene cylinder, myoglobin cylinder, amyloid cylinder, protein cylinder, vacuole deformation The method for improving the detectability of a formed component according to claim 3, wherein the cells are classified into cells containing one or more selected from the group consisting of a column, a platelet column, a bacterial column, a bilirubin column and a salt column. (1) means for stirring the test liquid in the sample container;
(2) means for dispensing the stirred test solution;
(3) means for dispensing the test solution into the gap portion of the coated translucent plate integrated translucent plate;
(4) an imaging stage for imaging the test liquid on the translucent plate;
(5) means for enlarging the sample image of the formed component in the test solution;
(6) It has a function to automatically adjust the focus of the sample image of the formed object,
(7) means for capturing the image;
(8) In the formed component analyzer having means for processing the captured image and identifying the various components,
(9) Means for concentrating the formed component by centrifuging the coated light transmitting plate-integrated light transmitting plate in a state where the test liquid is dispensed into the gap portion of the coated light transmitting plate integrated light transmitting plate A formed component analyzer characterized by having. (3) In the step of dispensing the test liquid into the gap portion of the coated translucent plate integrated translucent plate,
(10) The formed-component analyzer according to claim 6, comprising means for mixing with a staining solution before dispensing. (11) The formed component analyzing apparatus according to claim 6, further comprising means for storing a result of processing the captured image and identifying the various components. (8) The means for processing the captured image and identifying it as various components has the function of calculating the analysis result from all the discrimination results for the preset visual field and the function of outputting the analysis result from the output device. The formed-component analyzer according to claim 6. (8) The formed component analyzing apparatus according to claim 6, wherein the means for processing the captured image to identify various components has a learning recognition function. (8) The formed component analyzing apparatus according to claim 6, wherein the means for processing the captured image and identifying the various components learns and identifies the range of the feature amount of the formed component. (8) The means for processing the captured image and identifying the various components has a function of calculating the amount of the formed component based on the optical feature amount attributed to the formed component. Described component analyzer. (5) The formed component analyzer according to claim 6, wherein the means for enlarging the sample image of the formed component in the test solution has two or more magnifications.     The material analysis apparatus according to claim 6, wherein the material of the light transmitting plate and the coated light transmitting plate is plastic.   The formed component analyzer according to claim 6, wherein the formed component is any one selected from the group consisting of red blood cells, leukocytes, epithelial cells, and cylinders.     In addition to epithelial cells, squamous epithelial cells, transitional epithelial cells, tubular epithelial cells, round epithelial cells, urethral columnar epithelial cells, prostate epithelial cells, seminal vesicle epithelial cells, endometrial epithelial cells, oval round fat pad, intracytoplasmic The tangible component analyzer according to claim 15, wherein the apparatus is classified into cells including one or more selected from the group consisting of inclusion body cells and polygonal cells.     Glass cylinder, epithelial cylinder, granule cylinder, waxy cylinder, fat cylinder, erythrocyte cylinder, leukocyte cylinder, cell cylinder, nitric leukocyte cylinder, hemoglobin cylinder, hemosilylidene cylinder, myoglobin cylinder, amyloid cylinder, protein cylinder, vacuole deformation The tangible component analyzer according to claim 15, which is classified into cells containing one or more selected from the group consisting of a column, a platelet column, a bacterial column, a bilirubin column and a salt column.   The staining solution is Sternheimer-Malbin staining method, Sternheimer staining method, Prescott-Brodie staining method, Behre-Mühlberg staining method, Sudan III staining method, Lugol staining method, hemosiderin staining method, Papanicolau staining method, Papanicolau staining method Field staining method, Quaglino-Flemans method, Kaplow method, Sato / Sekiya method, Berlin blue method, Giemsa staining method, Wright staining method, Pappenheim staining method, Congo red staining method, methyl green / pyronine staining method, Alcian blue staining method , Schole staining method, Foilgen staining method, oil red O staining method, Brecker method, Heinz body staining method, neutral red / Janus green super-vivo staining method, brilliant clay Le blue staining material ingredient analyzer according to claim 7, wherein a reagent containing one or more kinds of ingredients which are used in at least one method of. (12) The tangible component analyzer according to claim 6, further comprising means for calculating the amount of the tangible component based on the optical feature amount attributed to the tangible component.     20. The formed component analyzer according to claim 19, wherein the optical characteristic amount is a light emission amount by the reagent for ATP measurement.     The material analysis apparatus according to claim 6, wherein at least one material of the light-transmitting plate and the coated light-transmitting plate is glass, plastic, glass subjected to chemical treatment, or plastic subjected to chemical treatment. (1) (2) (3) (9) (4) (5) (6) (7) (8) The method for improving the detectability of the formed component.   (1) (2) (3) (9) (4) (5) (6) (7) (8) It has a means to carry out each process automatically and automatically in the order. Formed component analyzer.