JPH06213797A - Method and system for analyzing particle image in flow system - Google Patents

Abstract

PURPOSE:To provide method and system for analyzing particle image in flow system in which the number of particles in sample liquid and the particle concentration can be determined easily over a wide particle concentration range and the distribution thereof can be determined accurately and precisely for various types of particle. CONSTITUTION:A sample liquid is fed to a flow cell 100 and the passage of particles is detected using laser luminous flux 10. A pulse lamp 1 is lighted based on the detection of particle and the static image of particle is imaged by a TV camera 8 in the interlace system thus performing image processing. A number of particle calculating section 40 in a particle analyzing means 102 solves simple first order formulas using the total number of particles Ng subjected to image processing, the number of images ng subjected to image processing, and measurement condition data including the imaging conditions of interlace system thus determining the total number of particles Nm in the sample liquid. Furthermore, particle concentration and the like are calculated using Nm thus classifying and analyzing a plurality of types of particle.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a flow type particle image analyzer and a flow type particle image analyzing method for picking up an image of particles suspended in a flowing sample liquid and analyzing the particles, and in particular to blood. The present invention relates to a flow-type particle image analysis method and a flow-type particle image analysis device suitable for analyzing cells and particles in blood and urine.

[0002]

2. Description of the Related Art Conventionally, cells and particles in a sample liquid such as blood and urine are classified and analyzed by preparing a sample on a slide glass and observing it with a microscope. When the sample liquid is urine, the concentration of particles contained is low,
The sample solution was previously centrifuged and then observed. Then, in order to automate these observations and inspections, the sample solution is smeared on a slide glass and set on a microscope, and then the microscope stage is automatically scanned to obtain a static image of the particle at the position where the particle exists. An apparatus has been developed for imaging particles and performing classification and analysis of particles in a sample liquid by image processing technology using feature extraction and pattern recognition techniques.

However, in the above-mentioned apparatus, it takes time to prepare a sample, and it is necessary to find particles while mechanically moving the microscope stage and move the particles to an appropriate image capturing area in the microscope visual field. Therefore, there are problems that it takes a long time for classification and analysis and that the mechanical mechanism becomes complicated.

On the other hand, a method using a flow cytometer in which the test particles are suspended in the sample liquid and continuously flowed into the flow cell without performing the above-described smear preparation to optically analyze the particles. It has been known. The method using a flow cytometer is for observing fluorescence intensity and scattered light intensity from each particle in a sample liquid, and has a processing capacity of several thousand per second. However, it is difficult to obtain information that reflects the morphological characteristics of particles, and conventional classification of particles based on morphological characteristics cannot be performed.

Further, as an attempt to capture still images of particles in a continuously flowing sample liquid and classify and analyze the particles from the still images of the individual particles, Japanese Patent Publication No. 57-500.
Techniques such as those described in Japanese Patent Laid-Open No. 995 and Japanese Patent Laid-Open No. 63-94156 are known.

In Japanese Patent Publication No. 57-500995, a sample solution is caused to flow through a channel having a special shape having a wide imaging area, and a pulsed light source (hereinafter referred to as a flash lamp) is caused to emit light so that particles in the sample solution are discharged. A method of capturing a still image and performing classification and analysis of particles using the still image is shown. Further, the flash lamp periodically emits light in synchronization with the operation of the CCD camera, and a magnified image of sample particles is projected on the CCD TV camera by using a microscope. The flash lamp emits light for a short time, and the CCD TV camera can capture 30 still images per second, so that still images can be obtained even when particles are continuously flowing.

In Japanese Patent Laid-Open No. 63-94156, an optical system for particle detection, which is different from the optical system for capturing a still image, is provided at a position upstream of the image capturing area. In addition, the sample liquid is used to reduce the influence of the depth of focus.
It is made to flow in a flow path formed of glass or the like called a flow cell so as to have a flat cross-sectional shape in the direction perpendicular to the optical axis of the still image pickup optical system. Then, the passage of the particles is detected by detecting the scattered light of the laser light by the particles in advance in the particle detection optical system, and the image of the particles is captured by the CCD camera when the particles just reach the image capturing area. The flash lamp is turned on after a predetermined delay time from the time of detection. In this method, the still image of the particle is efficiently captured because the still image of the particle is captured at a timing only when the passage of the particle is detected by the particle detection optical system without periodically emitting light from the flash lamp. It is possible to take an image, and even in the case of classification or analysis of particles in a sample liquid having a low concentration, a meaningless image without particles is not taken.

By the way, a still image of continuously flowing particles as described above is picked up by a CCD camera and image processing is performed to classify a plurality of kinds of particles contained in a sample liquid or the number thereof. There are the following problems in counting the number. That is, from the time when particles are detected by the particle detection system, a particle image is picked up by a TV camera such as a CCD camera, converted into a charge signal proportional to a grayscale image of particles, and taken out (transferred) as a video signal for a finite time. Takes. Image transfer work continues during this time,
During that time, the flash lamp is not turned on so as not to disturb the imaged image, and until this transfer work is completed, imaging cannot be performed even if the next particle newly passes, and that information is discarded. There must be. Thus, there is a fixed time during which an image cannot be captured (hereinafter referred to as dead time), and particles that have passed during this dead time are not captured and image processing cannot be performed. With a sample solution with a low concentration, the above-mentioned phenomenon is unlikely to occur because the particles arriving next to the image acquisition area have a low probability of arriving during this dead time, but with a sample solution with a high particle concentration, most of the measurement time is However, it is difficult to accurately know the number of plural kinds of particles contained in the sample liquid from the number of particles subjected to image processing.

To solve such a problem, Japanese Patent Laid-Open No. 4-72
In the apparatus described in Japanese Patent No. 544, a counter for counting the number of passing particles is provided in the particle detection optical system, and the count value of the counter and the existence ratio of each type of particles obtained by image processing are By this, the number of plural kinds of particles in the sample liquid can be obtained. Further, the concentration of a plurality of types of particles in the sample liquid can be known from the number of particles of each type.

An example in which a means for detecting passage of particles is additionally provided in a normal flow cytometer or particle analysis apparatus that does not capture a static image of particles is disclosed in Japanese Patent Laid-Open No. 60-260830. Kaisho 63-231244
Japanese Patent Laid-Open No. 1-245131 and Japanese Patent Laid-Open No. 1-245131 are known.

[0011]

As described above, in the techniques disclosed in Japanese Patent Publication No. 57-500995 and Japanese Patent Laid-Open No. 63-94156, due to the presence of dead time, the particle concentration is low to high. There is a problem in that it is difficult to accurately and accurately obtain a plurality of types of particle numbers and particle concentrations over a wide range.

To solve this problem, Japanese Unexamined Patent Publication No. 4-72544
According to the device described in the publication, the number of particles of each type in the sample liquid can be known over a wide range of particle concentration. In this case, it is assumed that the type-specific distribution of the number of particles to be detected by the particle detection counter matches the type-specific distribution of the number of particles image-processed from the captured still image. However, since the measurement methods of both are different, the distributions by type often do not match.For example, in a particle detection counter, unnecessary signals such as noise signals or stains and dust whose size is not constant are detected and counted by malfunction. However, according to the image processing, such unnecessary particles are regarded as non-measurement targets. Therefore, this device has a problem that the number of particles of each type cannot be accurately determined based on the counting result of the counter.

Further, even if all the passing particles are counted by the counter, the accuracy of the distribution of the number of particles by type depends on the accuracy of the number of particles by the image processing. Therefore, even if all the passing particles are counted by the counter, the counting accuracy and classification accuracy of the number of particles of each type are not improved. After all,
Even if the counter counts all the passing particles, the measurement accuracy is not improved. Needless to say, when only one type of particle is present in the sample liquid, it is not necessary to perform image processing to measure the type-specific distribution of the number of particles, and the configuration itself for capturing a still image is unnecessary.

In order to solve the above-mentioned problems of the prior art, the applicant of the present invention can accurately and highly accurately determine a plurality of types of particles and particle concentrations over a wide range from low to high particle concentrations. An analyzer was first proposed and filed (Japanese Patent Application No. 4-300808; filing date November 11, 1992). In the invention of this prior application, a laser beam for particle detection is radiated at a position close to the upstream side of the image capturing area on the flow cell, and the scattered light of the laser light by the particles in the sample liquid flowing in the flow cell is converted into a microscope objective lens. The passage of particles is detected by condensing and detecting with, and when the particles reach the image capturing area, a flash lamp is caused to emit light, a still image is captured by a CCD camera, and image processing is performed based on the image. Then, the number of particles and the particle concentration of each type in the sample liquid are analyzed as follows. That is, the number of particles in the sample liquid is obtained using the concentration corrector from the total number of particles subjected to image processing,
The particle concentration in the sample solution is calculated from the ratio of the number of particles image-processed based on the count value of the total number of particles detected by the particle detection system, and the detection signal waveform in the particle detection system is discriminated into multiple signal classes. By doing so, the particles are roughly classified, and then the particles are correctly classified by image processing. From this, the total number of particles and what kind of particles are present in what proportion are estimated.

However, it has been found that the above-mentioned prior invention has the following points to be further improved. That is, since the formula for obtaining the number of particles has a non-linear complicated shape, it cannot be calculated analytically, and it is necessary to obtain the total number of particles in the measurement sample liquid from the number of particles directly image-processed. It is necessary to calculate the relationship between the total number of particles necessary for correction and the number of particles that have been imaged and image processed in advance, and create and store the relationship, that is, a conversion table, in the density corrector. However, depending on the sample liquid to be measured, there are those in which the measurement range of the particle concentration is as large as 4 to 5 digits or more, and in such a case, a very large conversion table is required, which requires labor and time. Moreover, the conversion table must be rewritten every time the flow condition of the sample liquid, the presence or absence of the staining liquid, the measurement time, and other measurement conditions change.

As a method of not creating the conversion table as described above, there is a method of using a numerical solution method of an equation such as Newton's method, but this requires repeated calculation until the solution converges.

A first object of the present invention is to provide a flow-type particle image analysis method capable of accurately and accurately determining the number and particle concentration of a plurality of types of particles over a wide range from low to high particle concentrations. A flow type particle image analysis device is provided.

A second object of the present invention is to provide a flow-type particle image analysis method and flow capable of easily obtaining the number and concentration of particles in a sample liquid and accurately obtaining the distribution of each type. An object of the present invention is to provide a type particle image analysis device.

[0019]

In order to achieve the above first and second objects, according to the present invention, a sample liquid in which particles are suspended is flown into a flow cell, and an image capturing area in the flow cell is set to the above. Detect that the particles pass, irradiate the flow cell by emitting pulsed light from a pulse light source based on the detection, capture a still image of the passing particles in the flow cell by the interlace method, the obtained still image. In the flow-type particle image analysis method of performing image processing and analyzing particles in the sample liquid, the total number of particles subjected to image processing is N _g , and the next particle detection is performed from the time when the imaging by the previous particle detection is completed. the average time to time the imaging is terminated <T _read>, an image of the mean value of the immobility time of light emission of the pulsed light is blocked for incorporation as <T _dead>

[0020]

[Equation 9]

According to the above, there is provided a flow type particle image analysis method characterized by obtaining the total number N _m of particles in the sample liquid.

Preferably, the number of image-processed images is n _g , and the time required for all measurements is T _m .

[0023]

[Equation 10]

<T _read > is obtained by the following.

Preferably, the number of image-processed images is n _g , the time required for all measurements is T _m , and the frame time in the interlace method is T _f .

[0026]

[Equation 11]

The average number of fields <n> until particle detection is obtained by the following equation, and the first equation based on the statistical event by the Poisson process is calculated from the above <n> and T _f.

[0028]

[Equation 12]

[0029] By obtaining an average time T ₀ from through one particle to the next particle passes through, the T _0, wherein T
_f and the second expression based on the statistical event by the Poisson process from the above <n>

[0030]

[Equation 13]

The average <t> of the time from the output of the n-th field signal to the emission of the pulsed light was obtained, and the image was actually taken with the volume of the sample liquid flowing through the image capturing area during the T _f . From the <t> and the T _f , where k is the ratio of the portion to the volume of the sample liquid,

[0032]

[Equation 14]

By the above, <T _dead > is obtained.

In order to achieve the above first and second objects, according to the present invention, in the above flow type particle image analysis method, the number of image-processed images is n _g , which is required for all measurements. as was the time T _m, the frame time in the interlace method T _f,

[0035]

[Equation 15]

The average number of fields <n> up to particle detection is obtained by the method, and the ratio of the volume of the sample liquid flowing through the image capturing area to the volume of the sample liquid actually imaged during T _f is represented by k With the total number of processed particles being N _g and the total number of frames in T _m being n _f , from <n>,

[0037]

[Equation 16]

According to the above, there is provided a flow-type particle image analysis method characterized by obtaining the total number N _m of particles in the sample liquid.

[0039] wherein preferably, the substituting T _m / T _f to n _f, obtaining the total particle number N _m of the sample solution using instead T _f of n _f.

Preferably, the total particle concentration and the number of particles contained in the volume per image are determined from the total number of particles, the total volume of the sample solution, and the volume of the sample liquid per image, and The number of each type of particles and the concentration of each type of particles existing in the sample liquid are determined from the existence ratio of each type.

In order to achieve the above first and second objects, according to the present invention, a flow cell to which a sample solution in which particles are suspended is supplied, and an image capturing area in the flow cell is provided with the particles. An image pickup that captures a still image of passing particles based on the pulsed light flux by an interlace method, having a particle detection means for detecting passage and a pulse light source for irradiating the flow cell with a pulsed light flux based on the detection by the particle detection means In the flow-type particle image analysis apparatus, which is provided with a means and a particle analysis means for image-processing the obtained still image to analyze the particles in the sample liquid, the particle analysis means has the number of image-processed images n _g. Image counting section for counting the number of particles N
An image processing particle number counting section for counting _g , a time T _m required for all measurements, a total number of frames n _f in the T _m , and a volume of a sample liquid flowing in an image capturing area during a frame time in the interlace method. And a measurement condition data section that stores a ratio k between the volume of the sample solution and the volume of the sampled image, and the n _g , the N _g , the T _m , the n _f , and the k.
And a particle number calculation circuit for calculating the total number N _m of particles in the sample liquid.

Further, in order to achieve the above first and second objects, according to the present invention, in the above flow type particle image analyzing apparatus, the particle analyzing means has the number of processed images n _g. A processed image counting unit for counting the number of image-processed particles, an image-processed particle number counting unit for counting the total number of image-processed particles N _g ,
The time T _m required for the entire measurement, the frame time T _f in the interlace method, and the ratio k of the volume of the sample liquid flowing through the image capturing area during the T _f and the volume of the sample liquid of the actually imaged portion are stored. Measurement condition data part, and the _ng , the _Ng , the _Tm , the _Tf , and the k.
And a particle number calculation circuit for calculating the total number N _m of particles in the sample liquid.

Here, preferably, the pulsed light source has a capability of generating a pulsed light beam at a cycle of ½ or less of a frame time in the interlace system.

Preferably, the particle analysis means is
The interlaced image pickup means further includes an image memory for storing the image data transferred from the even field and the image data transferred from the odd field at predetermined addresses.

Further, preferably, the total particle concentration is included in the volume per image, depending on the total number of particles, the total volume of the sample liquid, the volume of the sample liquid per image, and the abundance ratio by type of particles. The apparatus further comprises particle concentration calculation means for obtaining the number of particles, the number of each type of particles existing in the sample liquid, and the concentration of each type of particles.

Further, preferably, the particles in the sample solution are biological cells or blood cell components existing in blood.

Further, preferably, the sample liquid is urine or a urine sediment component existing in urine.

[0048]

In the flow type particle image analysis method of the present invention configured as described above, it is first detected that the test particles have passed through the sample liquid flowing in the flow cell before the image capturing area is reached. When particle passage is detected, pulsed light is emitted from the pulse light source to the flow cell based on this detection, and when the analysis particles in the sample solution have just come to the image acquisition area, a still image of the passing particles in the flow cell is interlaced. The still image obtained by the image capturing method according to the method is subjected to image processing and subsequent analysis.

Then, the total number of particles N _g subjected to the image processing described above.
And an average value <T _read > of the time from the time when the imaging by the previous particle detection in the interlace method ends to the time when the imaging by the next particle detection ends, and the above for capturing the image in the interlace method. From the average value of immovable time <T _dead > at which the emission of pulsed light is blocked,

[0050]

[Equation 17]

The total number of particles N _m in the sample liquid can be determined by That is, if the values of <T _read > and <T _dead > can be independently obtained by the measurement conditions even by an indirect method, N _g and N _m can be calculated by using the linear equation (1). It is possible to easily obtain the unknown total particle number N _m in the sample liquid without creating a conversion table.

The value of <T _read > is calculated from the number of image-processed images n _g and the time T _m required for all measurements.

[0053]

[Equation 18]

Is calculated by Of these, n _g is the number of actually processed images, and T _m is one of the interlace measurement conditions that should be set in advance, so that it can be easily known. Therefore, the value of <T _read > can be easily obtained.

Further, the value of <T _dead > is obtained as follows. That is, first, from the above n _g and T _m , and the frame time T _f in the interlace system,

[0056]

[Formula 19]

The average number of fields <n> until particle detection is obtained by the following. Next, from this <n> and the above T _f ,
First formula based on statistical events by Poisson process

[0058]

[Equation 20]

The average time T ₀ from the passage of one particle to the passage of the next particle is obtained in accordance with. Then this T ₀
And the above T _f and <n>, the second equation based on the statistical event by the Poisson process

[0060]

[Equation 21]

Thus, the average <t> of the time from the output of the nth field signal to the emission of the pulsed light is obtained. Next, from this <t>, the ratio k between the volume of the sample liquid flowing through the image capturing region during the above T _f and the volume of the sample liquid of the actually imaged portion, and above T _f ,

[0062]

[Equation 22]

By the above, <T _dead > is obtained. Among the values used here, n _g and the T _m is capable of easily known, as described above, T _f is easily know since it is one of the measurement condition interlace be preset It is possible, and k can be easily known from the conditions of flowing the sample solution into the flow cell and the measurement conditions of the interlace method. Therefore, the value of <T _dead > can be easily obtained.

From the number of image-processed images n _g , the time T _m required for all measurements, and the frame time in the interlace system from T _f ,

[0065]

[Equation 23]

The average number of fields <n> up to particle detection is obtained by the following. From this <n>, the total number of frames n _{f in} T _m , and the above k and N _g ,

[0067]

[Equation 24]

The total number of particles N _m in the sample solution can be determined by This equation (8) is a simple linear equation with respect to N _g and N _m , and among the values included in this equation, n _g is the number of actually image-processed images, T _m and T _f. Is an interlaced measurement condition that should be set in advance, and k
Is a constant determined by the condition of flowing the sample solution into the flow cell and the measurement condition of the interlace method, and therefore each value can be easily known. Therefore, (8)
By using the formula, it is possible to easily obtain the unknown total particle number N _m in the sample liquid without creating a conversion table.

[0069] Further, by substituting the T _m / T _f to the n _f, be calculated on the total particle number N _m of the sample solution using instead T _f of n _f the same result.

As described above, the total number of particles N _m in the sample liquid can be easily known only by solving a simple linear equation, and it is not necessary to prepare a conversion table. For example, the measurement range of particle concentration is 4 to 5 digits. The correct number of particles can be easily obtained for sample liquids to be measured, and it is possible to accurately and accurately obtain multiple types of particle numbers and particle concentrations over a wide range from low to high particle concentrations. Become. Further, even if the conditions of the flow of the sample solution, the presence or absence of the staining solution, the measurement time and other measurement conditions are changed, it is not necessary to rewrite the conversion table each time, and the value necessary for the calculation is simply changed to N _m. Can be determined.

The total number of particles per unit volume is determined by the total number of particles obtained as described above, the total volume of the sample solution known in advance, and the volume of the sample solution per image (one field of view of the microscope). That is, the total particle concentration and the number of particles contained in the volume per image are obtained. Further, the number of particles of a plurality of types of particles existing in the sample liquid and the concentration of each of the plurality of types of particles are obtained based on the existence ratio of each type of particles.

Further, in the particle analyzing means of the flow type particle image analyzing apparatus of the present invention configured as described above, the number of images _ng image-processed by the processed image counting unit is counted and the image processing particle number counting unit is counted. The total number of particles N _g subjected to the image processing in is counted. In the measurement condition data section, the time T _m required for all measurements, the total number of frames n _f in this T _m , and the above T _f.
A ratio k between the volume of the sample liquid flowing through the image capturing area and the volume of the sample liquid in the actually imaged portion is stored. Then, the above n _g , N _g , T _m , n _f , and k are input to the particle number calculation circuit, and using these, the total number N _{m of} particles in the sample liquid is obtained.

[0073] In addition, stored in the measurement condition data unit frame time T _f in interlaced mode in place of the n _f, which enter into the particle number calculating circuit, the total number of particles using the T _f instead of n _f The same result can be obtained by obtaining N _m .
At this time, the fact that n _f is represented by T _m / T _f as described above is used.

Further, since the present invention adopts the interlace system, when the pulsed light flux is generated once, the image transfer for these two fields is completed so as not to disturb the image for two fields. Until then, it is necessary to prevent the generation of the next pulsed light flux. Therefore 1
The minimum time from the generation of one pulse luminous flux to the generation of the next pulse luminous flux is one field time, that is, 1/2 of the frame time.
Becomes Since the pulsed light source of the present invention has the ability to generate a pulsed light flux at a cycle of 1/2 or less of the frame time which is the minimum time, it is necessary to capture an image of one particle for each frame. Even at this time, it is possible to generate a pulsed light flux once per frame and deal with it.

By storing the image data from the even field and the image data from the odd field in the interlace system at a predetermined address of the image memory in advance, which field is used for image capture in relation to the particle detection timing. Even after the start, the subsequent image processing is performed correspondingly, and the image data in each field is not replaced and the image is not disturbed.

Further, in the particle concentration calculation means, the total number of particles obtained as described above and the total volume of the sample liquid,
The total particle concentration, the number of particles contained in the volume per image, the number of particles of a plurality of kinds of particles existing in the sample liquid, and the volume of the sample liquid per image and the existence ratio of each type of particles, and Each particle concentration of a plurality of types of particles is obtained.

[0077]

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS An embodiment of a flow type particle image analysis method and a flow type particle image analysis apparatus according to the present invention will be described with reference to FIGS. First, the configuration of the flow-type particle image analysis apparatus of this embodiment will be described with reference to the overall configuration diagram shown in FIG. As shown in FIG. 1, the flow-type particle image analysis apparatus of this embodiment includes a flow cell 100 to which a sample liquid in which particles are suspended is supplied, an image capturing means 101, a particle analysis means 102, and a particle detection means 103. Prepare

The image pickup means 101 has a function as a microscope and is a flash lamp 1 which is a pulse light source, and a flash lamp driving circuit 1 for causing the flash lamp 1 to emit light.
a, a field lens 2 for making the pulsed light flux from the flash lamp 1 parallel, a microscope condenser lens 3 for focusing the parallel pulsed light flux 10 from the field lens 2 on a sample liquid flow 110 in the flow cell 100, a sample liquid in the flow cell 100 Image data that is an electrical signal that captures the image of the imaging position 6 projected through the microscope objective lens 5 and the projection lens 7 that collects the pulsed light flux irradiated on the stream 110 and forms the image at the imaging position 6 by the interlace method. It has a TV camera 8 for converting it into a signal, a field stop 11 for limiting the width of the pulsed light beam 10 and an aperture stop 12.
As the TV camera 8, a CCD camera or the like having a small afterimage is generally used.

The particle analysis means 102 converts the image data signal transferred from the TV camera 8 into a digital signal A
An image memory 25 that stores data based on signals from the D converter 24 and the AD converter 24 at a predetermined address, an image processing control circuit 26 that controls writing and reading of data in the image memory 25, and an image memory 25 A feature extraction circuit 27 and an identification circuit 28 that perform image processing based on the signal to analyze the number of particles, classification of particles, and the like, a particle number calculation unit 40 that calculates the number of particles in the sample liquid, imaging conditions of the TV camera 8, and the like. Setting conditions for the flow of sample liquid in the flow cell 100, control of the image processing control circuit 26, identification circuit 2
It has a central control unit 29 for storing the image processing result from 8, the data exchange with the particle number calculation unit 40, the display on the display unit 50, and the like.

The particle detecting means 103 includes a semiconductor laser 15 which is a detection light source for emitting a laser beam as a detection beam, a collimator lens 16 for converting the laser beam from the semiconductor laser 15 into a parallel laser beam 14, and a laser from the collimator lens 16. A cylindrical lens 17 that focuses the light flux in only one direction, a reflecting mirror 18 that reflects the light flux from the cylindrical lens 17, and a laser light flux from a reflecting mirror 19 that is provided between the microscope condenser lens 3 and the flow cell 101, on the sample liquid flow 110. A micro-reflecting mirror 19 which guides the light to the adjacent position on the upstream side of the image capturing area, a microscope objective lens 5 which collects the scattered light of the laser light flux by particles, and a beam splitter which reflects the scattered light collected by the microscope objective lens 5. 20, the scattered light from the beam splitter 20 through the diaphragm 21 Having a flash lamp lighting control circuit 23 for controlling the flash lamp driving circuit 1a based on the received electrical signal from the photodetector 22, the photodetector 22 outputs an electrical signal based on the intensity. The microscope objective lens 5 is also used as the image pickup means 101.

Further, the sheath liquid is supplied to the flow cell 100 together with the sample liquid to form a flow in which the sample liquid is wrapped in the sheath liquid. Then, the sample liquid flow 110 becomes a stable steady flow (sheath flow) having a flat cross-sectional shape in the direction perpendicular to the optical axis (microscope optical axis) 9 of the image pickup means 101, and the center of the flow cell 100 is directed downward in the plane of the drawing. Sent to you. The flow velocity of the sample liquid flow 110 is controlled according to the conditions set by the central control unit 29.

Next, the configuration of the particle number calculation unit 40 will be described with reference to FIG. The particle number calculation unit 40 includes a processed image counting unit 41, a measurement condition data unit 42, an image processing particle number counting unit 43, and a particle number calculation circuit 44, and the image processed image from the image processing control circuit 26. The number of particles is counted, and the information of the number of particles subjected to 28 image processing by the feature extraction circuit 27 and the identification circuit is input from the central control unit 29 to be counted, and further the measurement condition data including the interlaced imaging conditions from the central control unit 29. Is input, the calculation of the total number of particles existing in the sample liquid during the measurement time is executed using these values, and the calculation result is sent again to the central control unit 29.

Returning to FIG. 1, the basic operation of the flow type particle image analysis device having the above configuration will be described. The semiconductor laser 15 constantly oscillates continuously, and always observes that particles in the sample pass through the detection region. The laser light flux from the semiconductor laser 15 is converted into a parallel laser light flux 14 by the collimator lens 16 and is focused by the cylindrical lens 17 in only one direction. This laser beam is reflected by the reflecting mirror 18 and the minute reflecting mirror 19 and is irradiated onto the sample liquid flow 110 in the flow cell 4. This irradiation position is a particle detection position where the laser beam is focused by the cylindrical lens 17, and the sample liquid flow 110
It is a position close to the upstream side of the upper image capturing area.

When the particles to be measured cross the laser beam, the laser beam is scattered, the scattered light is condensed by the microscope objective lens 5 shared by the image pickup means 101, and is reflected by the beam splitter 20. The observation area on the sample liquid flow 110 is limited by the diaphragm 21, and the light is received by the photodetector 22 and converted into an electric signal based on its intensity.

The electric signal from the photodetector 22 is input to the flash lamp lighting control circuit 23, where it is judged whether or not the electric signal has a predetermined signal level or more. If the electric signal has a predetermined signal level or more, image processing is performed. It is considered that the particles of interest have passed and a detection signal is sent to the flash lamp drive circuit 1a. This detection signal indicates the distance between the particle detection position and the image capturing area and the sample liquid so that the flash lamp 1 emits light to capture an image when the particles have just reached a predetermined position in the image capturing area of the TV camera 8. It is sent to the flash lamp driving circuit 1a after a predetermined delay time determined by the flow velocity. However, this delay time is very short because the distance between the particle detection position and the image capture area is very short, and the accuracy of particle detection and analysis is not affected by the flow velocity and particle concentration of the sample solution. Absent. Further, the flash lamp lighting control circuit 23 sends a lamp light emission ready signal, which will be described later, simultaneously with the above detection signal,
The light emission timing of the flash lamp is controlled based on the timing of the interlaced field signal. Further, the detection signal of the flash lamp lighting control circuit 23 is also sent to the image processing control circuit 26.

The detection signal is the flash lamp drive circuit 1a.
Then, the flash lamp drive circuit 1a causes the flash lamp 1 to emit light. The pulsed light emitted from the flash lamp 1 travels on the optical axis 9 of the microscope, becomes parallel light by the field lens 2, is focused by the condenser lens 3 of the microscope, and flows in the sample solution 11 in the flow cell 100.
0 is illuminated. The field stop 11 and the aperture stop 1
2 limits the width of the pulsed light flux 10.

Sample liquid flow 11 in flow cell 100
The pulsed light flux irradiated to 0 is condensed by the microscope objective lens 5 to form an image at the image forming position 6. The image at the image forming position 6 is projected onto the image pickup surface of the TV camera 8 by the projection lens 7 and converted into an image data signal by the interlace method. This means that a still image of the particles has been captured. The image pickup conditions of the TV camera 8 are preset in the central control unit 29, and the image pickup operation of the TV camera 8 is controlled by this. This interlaced imaging will be described later.

The image data signal output from the TV camera 8 is converted into a digital signal by the AD converter 24, and the data based on this is stored in a predetermined address of the image memory 25 under the control of the image processing control circuit 26. Remembered. The data stored in the image memory 25 is read out under the control of the image processing control circuit 26, is input to the feature extraction circuit 27 and the identification circuit 28, and is subjected to image processing to analyze the number of particles and the classification of particles. The processing is executed, and the result is stored in the central control unit 29. The measurement condition data including the image processing result and the imaging condition of the interlace method, and the number of processed images from the image processing control circuit 26 are input to the particle number calculation unit 40, and the particle number calculation unit 40 calculates the total number of particles. Calculated. The calculation result is input to the central control unit 29, and is displayed on the display unit 50 together with the image processing result from the identification circuit 28.

Next, the interlace type imaging condition in the TV camera 8 and the calculation in the particle number calculating section 40 using the measurement condition data including this imaging condition and the image processing result will be described. As the TV camera 8, a CCD camera having a small afterimage is used as a general two-dimensional image pickup device. As described above, the flash lamp 1 emits light to capture a still image of a particle only when the particle detecting unit 103 detects the particle, but the TV camera 8 operates continuously regardless of the particle detecting unit 103. It is assumed that

FIG. 3 shows the field signal of the TV camera 8,
The relationship between the CCD transfer signal in the even field (e) and the odd field (o), the electric signal from the photodetector 22, the light emission of the flash lamp, the lamp light emission ready signal, and the dead time generated by the image capturing is shown.

In FIG. 3, a field signal having a field time T _f as a cycle is generated from a clock built in the TV camera 8. An image is captured in an even field (e) and an odd field (o), and these two fields are combined to form one frame. In the even field (e) and the odd field (o), image capture (accumulation) and transfer are performed alternately by shifting the frame period, that is, T _f / 2. Therefore, one field time is half the frame time T _f /
It is 2. This frame time is set to 1/30 seconds, for example. In each field, there is an image storage time and a transfer time during a frame time for obtaining one image. The amount of light incident on the image pickup surface is accumulated as electric charge during the image storage time, and this accumulated during the transfer time. Transfer charge. After the transfer, the charge amount becomes 0, and the image storage period starts again. In FIG. 3, CCD transfer signals of even field (e) and odd field (o) are shown, but the transfer time of these is very short.

The CCD transfer signal shown in FIG. 3 shows only the image when the image of the particle is detected and the image of the particle is picked up for simplification, but each CCD is actually shown.
The transfer signal is always output every time a field signal is output.
Further, it is assumed that the process relating to the preceding detection particle has already been completed at the position of 0 of the field signal at the left end in the figure.

The CCD transfer signal of each field is digitized by the A / D converter 24 and temporarily stored in each predetermined address of the image memory 25, and then the image data of the two fields are combined to form one image. Made The timing of image capturing of particles, that is, whether image capturing starts from the even field (e) or the odd field (o) depends on the relationship between the particle detection timing and the field signal at that time. Then, the image data from these fields is stored in a predetermined address of the image memory 25, that is, at an address determined for the even field image and the odd field image, so that the image capturing starts from which field. However, the subsequent image processing is performed for each field, the image data in each field is not replaced, and the image is not disturbed.

After n fields have passed after the detection of the previous particle (after the nth field signal is output), the next particle is detected and the photodetector 22 emits an electric signal.
The flash lamp 1 emits light after a delay time T _d until the particles come to an appropriate position in the image capturing area. At this time, the time from the output of the nth field signal to the light emission of the pulse lamp is represented by t in the figure. The lamp emission ready signal has passed two fields (one frame) from the time when the electric signal from the photodetector 22 for particle detection is output (n
The time until the +2) th field signal is output is turned off, and the emission of the flash lamp 1 is blocked even if the next particle that arrives at this time is detected. Since the light emission of the flash lamp 1 is blocked in this way, the particle image is not disturbed by the next detected particles.
When the (n + 2) th field signal is output, the process for this particle ends, and the count of the field signal for the next detected particle restarts from 0.

The minimum off time of the lamp light emission ready signal is the minimum time required to transfer the image data of two fields, that is, one field time (T _f /
2). The flash lamp 1 has the ability to generate a pulsed light flux at a cycle of one field time or less, which is the minimum time, and for example, even when it is necessary to capture an image of one particle for each frame, one frame Per 1
It is possible to generate the pulsed light flux of the number of times and respond to it.

Here, when the ratio of the volume of the sample solution flowing in the imaging region during the frame time T _f to the volume of the sample in the actually imaged portion is k, the time of T _f / k is the actual imaging of particles. Is the time (effective part).
The time obtained by excluding this time of T _f / k from the time when the lamp light emission ready signal is turned off is the actual immovable time during which the light emission of the flash lamp is blocked for image capture, that is, a shaded line in the figure. The dead time is T _dead .
It should be noted that k is a condition (flow velocity, size of image capturing range, etc.) of flowing the sample liquid in the flow cell and frame time T _f.
It is decided by.

From FIG. 3, the dead time T _dead is expressed by the following equation.

[0098]

[Equation 25]

In the equation (9), the time t is a value that is randomly distributed depending on the particle detection timing. When the particle concentration is low and the particle detection interval is long, the detection timing is 1 field time T _f / 2. It is thought that the average value approaches T _f / 4, and conversely, when the particle concentration is high and the particle detection interval is short, the detection timing is within 1 field time T _f / 2. It is considered to be densely distributed, and its average value approaches 0. Letting the average value of t be <t>, the average value of dead time <T _dead > is expressed by the following equation.

[0100]

[Equation 26]

On the other hand, when the image pickup by the previous particle detection is completed, that is, from the position of 0 of the field signal in the figure, CC
Average value of time until transfer end of D transfer signal <T _read >
Is the average value of n, that is, the average number of fields until particle detection is <n>,

[0102]

[Equation 27]

It is represented by Here, T _m is the time required for all measurements, and _ng is the number of images that have been image-processed, that is, the number of images.

Since the ratio of the number of passing particles counted down due to the dead time T _dead generated by one-time image capturing is <T _dead > / <T _read >, the total number of particles actually image-processed is N. _{If g} and the total number of particles in the measurement sample liquid are N _m , these relationships are as follows.

[0105]

[Equation 28]

From this equation (12), N _m can be obtained as the total number of particles in the measurement sample liquid. However, when a plurality of types of particles are present in one image, N _m is the _sum of these.

<T _read > can be easily obtained by the equation (11). Further, <T _dead > is obtained from the equation (10) if <t> is known. The calculation method will be described below. The Poisson process is commonly assumed in statistical events that count particles passing through a flow cell. If this assumption is satisfied, the average time from one particle passes until the next particle passes through a T _O <n> and <t> is expressed as follows.

[0108]

[Equation 29]

If T _o and <n> are known in the above equation (14), <t> can be obtained. First, according to equation (11), <n
> Look, solving for T _O by substituting this result into equation (13). Then, substituting the solutions of <n> obtained by resolution with recent (11) of the T _O to (14) is <t> obtained. Further, <T _dead > is obtained by substituting the value of <t> into the equation (10).

As described above, <T _read > and <T _dead >
N _m is equal to N _g because the value of
Can be easily known by solving the linear equation for.

Returning to FIG. 2, in the particle number calculation unit 40,
Every time one image processing is performed, a signal is sent from the image processing control circuit 26 to the processed image number counting section 41, and the count value of the processed image number counting section 41 is incremented by one. At the end of the measurement, the processed image number counting unit 41 stores the total count value, that is, the number _ng of images subjected to image processing during the measurement time.

Further, the measurement condition data section 42 receives and stores the measurement condition data including the interlaced imaging conditions, that is, the above-mentioned T _f , T _m and k from the central control section 29.

Further, every time one image processing is performed, the number of particles image-processed in the feature extraction circuit 27 and the identification circuit 28 is sent to the image-processed particle number counting section 43 via the central control section 29, and the image is processed. It is sequentially added to the count value of the processed particle number counting unit 43. Then, at the end of the measurement, the total number of particles N _g subjected to image processing is stored in the image processing particle number counting section 43.

The above-mentioned (10) is applied to the particle number calculation circuit 44.
Expression (14) and its calculation procedure are stored,
The values of _{ng of} the processed image number counting unit 41, T _f , T _m , and k of the measurement condition data unit 42, and the value of N _g of the image processed particle number counting unit 43 are input, and equation (10) is changed to equation (14). It is used to calculate the total number of particles N _m in the sample liquid.

The total number of particles in the sample liquid thus obtained is returned to the central controller 29. Then, if necessary, the total number of particles, the total volume of the measurement sample solution stored in the central control unit 29, and the volume of the sample solution per image (one field of view of the microscope) are used to determine the total volume per unit volume. The number of particles, that is, the total particle concentration and the number of particles contained in the volume per one field of the microscope are calculated. Further, when a plurality of types of particles are present, the number of particles of each type, particle concentration, etc. are calculated using the existence ratio of each type of particles obtained by the identification circuit 28. The results are output to the display unit 50 as described above. These calculations are performed by a particle concentration calculating means (not shown) provided in the central control unit 29.

As described above, according to this embodiment, <T _read
Since the value of <> and the value of <T _dead > can be obtained from the measurement result including the image processing result and the imaging condition of the interlace method, the above-mentioned <T _read can be performed without requiring a complicated procedure for creating a conversion table. The total number of particles N _m in the sample solution can be easily obtained by only solving the linear equations relating to N _g and N _m using the ratio of> and <T _dead >. Therefore, for example, the correct number of particles can be easily obtained even for a sample liquid to be measured having a particle concentration measuring range of 4 to 5 digits or more,
It is possible to accurately and accurately obtain the number of different types of particles and the particle concentration over a wide range of particle concentration. Further, even if the measurement conditions change, it is not necessary to rewrite the conversion table each time, and N _m can be determined only by changing the value required for calculation.

Further, the total particle concentration and the per image are determined by the total number of particles obtained as described above, the total volume of the sample liquid known in advance, and the volume of the sample liquid per one image (one field of view of the microscope). The number of particles contained in the volume of can be calculated. Further, the number of particles of a plurality of types of particles existing in the sample liquid and the concentration of each of the plurality of types of particles can be obtained based on the existence ratio of each type of particles.

In addition, the flash lamp 1 has a minimum field time (T _f
/ 2) Since it has the ability to generate a pulsed light flux with a cycle of the following, even when it is necessary to capture an image of one particle per frame, one pulse per frame can be assured without disturbing the image. A luminous flux can be generated and dealt with.

Further, since the respective image data from the even field (e) and the odd field (o) are stored in a predetermined address of the image memory 25, which field is used for image capture in relation to the timing of particle detection. Even if it starts, the subsequent image processing is performed correspondingly, and the image data in the field is not replaced and the image is not disturbed.

Next, another embodiment of the flow type particle image analysis method and flow type particle image analysis apparatus according to the present invention will be described with reference to FIG. The flow-type particle image analysis device of the present embodiment is the same as the above-described embodiment, except that the measurement condition data used in the particle number calculation unit and the calculation formula and the calculation procedure stored in the particle number calculation circuit are different. is there. As shown in FIG. 4, the particle number calculation unit 40a includes a processed image counting unit 41, a measurement condition data unit 42a, an image processing particle number counting unit 43, and a particle number calculation circuit 44a.

The formula for calculating the total number of particles in the sample liquid stored in the particle number calculating circuit 44a and the calculation procedure will be described below. The above-mentioned T _O , that is, the average time from the passage of one particle to the passage of the next particle is the value obtained by dividing the time T _m required for all measurements by the total number of particles N _m in the sample liquid. To T _m / N _m . Using these facts, by summarizing the equations (10) to (14), the relation between the total number of particles N _m in the measurement sample and the actual image-processed total particles N _g is expressed by the following equation. Become.

[0122]

[Equation 30]

Here, n _f is the total number of frames in the time T _m required for all measurements. In this equation (15), <n>
Is an amount that changes depending on N _m , and this <n> is obtained from the relationship of the above-mentioned equation (11). Any other value n _f
Since k and k are values preset in the imaging condition and the measurement condition, N _m can be easily known by solving a linear equation with respect to N _g . However, the above n _f is a value obtained by dividing the time T _m required for all the measurements by the frame time T _f , so T _m / T _f
Can be replaced with

In FIG. 4, in the measurement condition data section 42a, as the measurement condition data from the central control unit 29, the volume of the sample solution flowing in the imaging region during the frame time T _f and the volume of the sample of the portion actually imaged are recorded. The ratio k and the time T _m required for all measurements, as well as the total number of frames n _{f in} T _m
Is input and stored. The operations in the processed image counting unit 41 and the image processed particle number counting unit 43 are the same as those in FIG. 2, and the image processed image number _ng and the image processed total particle number _Ng are respectively counted and stored.

The above-mentioned (1
The equations 1) and (15) and the calculation procedure thereof are stored, and n _{g of} the processed image number counting unit 41 and the measurement condition data unit 4 are stored.
2 _f n _f , T _m , k, N _{g of the} image processing particle number counting unit 43
Is input, and the total number of particles N _m in the sample liquid is calculated using the equations (11) and (15). However, the above n _f
Can be replaced by T _m / T _f , so that T _f instead of n _f may be input from the central control unit 29 to the particle number calculation circuit 44a via the measurement condition data unit 42a. In this case, n _f is replaced with T _m / T _f in the equation (15) for calculation.

The total number of particles in the sample liquid thus obtained is returned to the central control unit 29, and the central control unit 29
In the particle concentration calculation means (not shown) provided in
Similar to the above-described embodiment, the total particle concentration, the number of particles contained in the volume of one field of view of the microscope, the number of plural types of particles, the particle concentration, etc. are calculated. The results are output to the display unit 50.

As described above, according to this embodiment, the same effect as that of the above-described embodiment can be obtained, the calculation procedure is simplified, and the total number of particles N _m in the sample liquid can be obtained more easily. You can

In the above embodiment, the particle number calculation unit 40
Alternatively, in 40a, the total number of particles N _m in the sample liquid is obtained and returned to the central control unit 29, and the total particle concentration, the number of particles contained in the volume of one field of view of the microscope, and the number of particles and the particle concentration of a plurality of types are obtained. However, the total particle concentration, the number of particles contained in the volume per one field of view of the microscope, and the number and types of particles of a plurality of types may be calculated in the particle number calculation unit 40 or 40a. In this case, the central control unit 29
As the measurement condition data to be input to the measurement condition data section 42, the entire volume of the measurement sample liquid or one image (one field of view of the microscope)
Add the volume of sample solution per
4 or 44a may be calculated using these values and the total number of particles N _m in the sample liquid.

The flow type particle image analysis method and flow type particle image analysis device of the present invention are present in biological samples and cells suspended in a liquid, blood cell components such as red blood cells and white blood cells in blood, or urine. It is effective in classifying and analyzing urinary sediment components. In particular, in counting the number of particles of the urinary sediment component in urine and classifying the particles, the number of particles present in a normal sample and an abnormal sample may differ by several digits or more. Since it is possible to know the total number of particles in the sample liquid from the image processing result by the image pickup means and the particle analysis means, it is possible to correctly obtain the number of particles of the kind to be analyzed even in the above cases where the particle concentrations are greatly different. it can.

In the above embodiments, the case where a still image of particles contained in the sample liquid continuously flowing in the flow cell is obtained and the image processing is performed is described. This method can also be applied to particle image analysis of moving slide specimens. Furthermore, the case where the laser light flux from the semiconductor laser is used as the detection light in the particle detection means and the scattered light of the laser light flux scattered by the particles is used has been described, but the present invention is not limited to this, and fluorescence or transmitted light from the particles is used. You can also do 1
A method of detecting particles by a three-dimensional image sensor or a method of detecting particles by a change in electric resistance due to passage of particles can also be used.

[0131]

According to the present invention, the average value <T _read > of the time from the end of the image pickup by the previous particle detection to the end of the image pickup by the next particle detection and the acquisition of the image Since the average value of immovable time <T _dead > at which the emission of pulsed light is blocked can be obtained from the measurement conditions,
The total number of particles N _m in the sample liquid can be easily obtained only by solving the linear equation without making a conversion table. Therefore, the correct number of particles can be easily obtained even for a sample solution having a large measurement range, and a plurality of types of particles and particle concentrations can be obtained accurately and highly accurately over a wide range of particle concentrations. Further, even if the measurement conditions change, it is not necessary to rewrite the conversion table each time.

The total particle concentration, the total volume of the sample solution, the volume of the sample solution per image (one field of view of the microscope), and the abundance ratio of each type of particle are used to determine the total particle concentration. It is possible to obtain the number of particles contained in the volume per image, the number of plural kinds of particles existing in the sample liquid, and their particle concentrations.

Further, the pulse light source is 1 / of the frame time.
Since it has the ability to generate a pulsed light flux at a cycle of 2 or less, it is possible to generate a pulsed light flux at least once per frame without disturbing the image.

Further, since the image data transferred from the even field and the image data transferred from the odd field are stored in a predetermined address of the image memory, no matter which field the image capture starts in relation to the timing of particle detection, the image will be displayed. Not disturbed.

Therefore, according to the present invention, the number of particles in the sample liquid and the particle concentration can be easily obtained over a wide range of the particle concentration, and their kind-based distributions can be obtained accurately and highly accurately. .

[Brief description of drawings]

FIG. 1 is a diagram showing an embodiment of a flow type particle image processing method and a flow type particle image processing apparatus according to the present invention.

FIG. 2 is a diagram showing a configuration of a particle number calculation unit in a particle analysis unit and an exchange of data around the particle number calculation unit.

FIG. 3 is a time chart showing an interlaced imaging condition, particle detection timing, flash lamp emission timing, and the like in a TV camera.

FIG. 4 is a diagram showing another embodiment of the flow-type particle image processing method and the flow-type particle image processing device according to the present invention, showing the configuration of the particle number calculation unit in the particle analysis means and the exchange of data around it. FIG.

[Explanation of symbols]

1 Flash Lamp 1a Flash Lamp Driving Circuit 3 Microscope Condenser Lens 5 Microscope Objective Lens 8 TV Camera 14 Laser Beam 15 Semiconductor Laser 17 Cylindrical Lens 19 Micro Reflector 22 Photodetector 23 Flash Lamp Lighting Control Circuit 24 AD Converter 25 Image Memory 26 Image processing control circuit 27 Feature extraction circuit 28 Discrimination circuit 29 Central control unit 40, 40a Particle number calculation unit 41, 41a Processed image counting unit 42 Measurement condition data unit 43 Image processing particle number counting unit 44, 44a Particle number calculation circuit 50 Display Part 100 Flow cell 101 Image capturing means 102 Particle analyzing means 103 Particle detecting means 110 Sample flow N _g Total number of particles image-processed N _m Total number of particles in sample solution n _g Number of image-processed images T _m Required for all measurements Time T _f Inter Frame time in the race system n _f Total number of frames in time required for measurement k T _f Ratio of the volume flowing in the imaging region to the volume of the actually imaged part during T _f

Claims (14)

[Claims] 1. A sample solution in which particles are suspended is flown into a flow cell, it is detected that the particles pass through an image capturing area in the flow cell, and pulse light is emitted from a pulse light source based on the detection. In the flow-type particle image analysis method of irradiating the flow cell, capturing a still image of passing particles in the flow cell by an interlace method, performing image processing on the obtained still image, and analyzing particles in the sample liquid, image processing The total number of particles is N _g , the average value of the time from the time when the imaging by the previous particle detection is finished to the time when the imaging by the next particle detection is finished is <T _read >, for capturing the image. _Let <T _dead > be the average value of the immovable time during which the emission of the pulsed light is blocked, and A flow-type particle image analysis method, characterized in that the total number of particles N _m in the sample liquid is calculated by 2. The number of image-processed images is n _g , and the time required for all measurements is T _m , where The flow-type particle image analysis method according to claim 1, wherein the <T _read > is obtained by 3. The number of image-processed images is n _g , the time required for all the measurements is T _m , and the frame time in the interlace method is T _f , The average number of fields <n> until particle detection is obtained by the following equation, and from the above <n> and T _f , the first equation based on statistical events by the Poisson process The average time T ₀ from the passage of one particle to the passage of the next particle is calculated by the following _equation , and from the T ₀ , the T _f , and the <n>, the second equation based on the statistical event by the Poisson process is obtained. [Equation 5] The average time from out n-th field signal to said pulse light is emitted seek <t>, wherein T _f
Let k be the ratio of the volume of the sample liquid flowing in the image capturing area to the volume of the sample liquid in the portion actually imaged,
From the <t> and the T _f , The flow-type particle image analysis method according to claim 1, wherein the <T _dead > is obtained. 4. A sample solution in which particles are suspended is flown into a flow cell, it is detected that the particles pass through an image capturing area in the flow cell, and pulse light is emitted from a light source based on the detection, and the light is emitted. In a flow type particle image analysis method of irradiating a flow cell, capturing a still image of passing particles in the flow cell by an interlace method, and processing the obtained still image to analyze particles in the sample liquid, image processing is performed. The number of images is n _g , the time required for all measurements is T _m , and the frame time in the interlace method is T _f. The average number of fields <n> up to particle detection was obtained by means of k, and the ratio of the volume of the sample liquid flowing through the image capturing area during T _f to the volume of the sample liquid of the actually imaged portion was subjected to image processing. Letting N _{g be the} total number of particles and n _{f be} the total number of frames in T _m , from <n>, A flow-type particle image analysis method, characterized in that the total number of particles N _m in the sample liquid is calculated by 5. substituting T _m / T _f to the n _f, the flow of claim 4, wherein the using the T _f instead of n _f seek total particle number N _m of the sample liquid Particle image analysis method. 6. The total particle concentration and the number of particles contained in the volume per image are obtained from the total number of particles, the total volume of the sample liquid, and the volume of the sample liquid per image, and further, it is determined according to the type of particles. The number of particles of a plurality of types of particles existing in the sample liquid and the concentration of each particle of a plurality of types of particles are determined from the abundance ratio.
The flow-type particle image analysis method described. 7. A flow cell to which a sample liquid in which particles are suspended is supplied, and a particle detection means for detecting passage of the particles through an image capturing area in the flow cell,
An image capturing unit that has a pulse light source that irradiates the flow cell with a pulsed light flux based on the detection by the particle detection unit and captures a still image of passing particles based on the pulsed light flux by an interlace method, and image processing the obtained still image. In a flow-type particle image analysis device comprising a particle analysis means for analyzing particles in the sample liquid, the particle analysis means includes a processed image counting section for counting the number of image-processed images _ng , and an image processing means. An image processing particle number counting section for counting the total number of particles N _g , a time T _m required for all measurements, a total number of frames n _f in the T _m , and an image capturing area during a frame time in the interlace method. A measurement condition data section that stores a ratio k of the volume of the sample liquid flowing through the sample liquid to the volume of the sample liquid of the actually imaged portion,
A particle number calculation circuit for calculating the total particle number N _m in the sample liquid by inputting the n _g , the N _g , the T _m , the n _f , and the k. apparatus. 8. A sample liquid in which particles are suspended is supplied.
Flow cell and image capturing area in the flow cell
Particle detection means for detecting that the particles pass through,
Based on the detection by the particle detection means,
The pulsed light having a pulsed light source for irradiating the flow cell
Interlaced still images of passing particles based on bundles
Image capturing means that captures more and the obtained still image
Particle analysis means for processing and analyzing particles in the sample liquid
In the flow-type particle image analysis device including:_gCounting
A processed image counting unit for performing the image processing, and the total number N of particles subjected to image processing_gTo
Image processing particle number counting section to count and time required for all measurements
T_m, Frame time in the interlace method
T_f, And the above T_fA sump that flows through the image capture area inside
Volume of the sample solution and the volume of the sample solution of the part actually imaged
A measurement condition data section for storing a ratio k of_g, The above
N_g, Said T _m, Said T_f, And input k and sample
Total number of particles in liquid N_mAnd a particle number calculation circuit for obtaining
A flow type particle image analysis device characterized by the above. 9. The flow type particle image analyzer according to claim 7, wherein the pulsed light source has a capability of generating a pulsed light flux at a cycle of 1/2 or less of a frame time in the interlace method. . 10. The particle analysis means further comprises an image memory for storing image data transferred from an even field and image data transferred from an odd field in the interlaced image pickup means at respective predetermined addresses. The flow type particle image analysis device according to claim 7 or 8, characterized in that. 11. The total number of particles, the total volume of the sample liquid,
The total particle concentration, the number of particles contained in the volume per image, the number of particles of a plurality of kinds of particles existing in the sample liquid, and the volume of the sample liquid per image and the existence ratio of each type of particles, and 9. The flow-type particle image analysis device according to claim 7, further comprising a particle concentration calculation unit that calculates each particle concentration of a plurality of types of particles. 12. The flow-type particle image analyzer according to claim 7, wherein the particles in the sample liquid are biological cells. 13. The particles in the sample liquid are blood cell components present in blood, according to claim 7.
1. The flow type particle image analysis device according to any one of 1. 14. The sample liquid according to claim 7, which is urine or a urinary sediment component present in urine.
1. The flow type particle image analysis device according to any one of 1.