JPH07270302A - Imaging flow sight meter - Google Patents

Abstract

PURPOSE:To achieve a photographing of a transmission light image of particle with a better contrast by a phase difference system. CONSTITUTION:This apparatus is provided with a light transmitting flow cell 1 through which a sample liquid 2 containing particles is made to flow, a laser light source 3 which irradiates a photographing area of the flow cell 1 with light, a video camera 7 to photograph a transmission light image of the particle existing in the photographing area of the flow cell 1, a ring slit 6 for a phase difference provided in an optical path to the flow cell 1 from the laser light source 3 and an objective lens 8 for a phase difference provided in the optical path to the video camera 7 from the flow cell 1.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention is used for observing microorganisms, analyzing blood cells, or analyzing the dynamics of tumor tissue cells, and uses a sample solution containing microorganisms or blood or urine as a flow cell. The present invention relates to an imaging flow cytometer that causes a video camera to image particles such as microorganisms or blood cells contained in blood or cells contained in urine by flowing in a transparent tube.

[0002]

2. Description of the Related Art A flow cytometer (also referred to as "FCM") is known as an apparatus for inspecting, for example, the above-mentioned microorganisms or particle components such as blood cells and cells in a liquid. Imaging flow cytometer ("IFC
(Also referred to as “M”) is an image of a particle, and morphological information of the particle is obtained from the image. Like a flow cytometer, microorganisms and cells are introduced into a thin glass tube called a flow cell, The transmitted light image and the fluorescence image of the microorganisms and cells are captured to obtain their morphological information.

[0003]

Biological samples are exceptionally colored. Therefore, when the transmitted light image of a cell is to be obtained by IFCM, for example, when a cell without color is irradiated with light, the contrast becomes weak and a good image cannot be obtained. Therefore, it is usually necessary to stain the cell. . However, there are cases in which proper dyeing cannot be performed.

This is because, for example, there is no dye that can stain the cells to be observed, or it is desired to observe the cells in a living state without staining the cells, and staining for capturing a fluorescent image. (Fluorescence staining) is also desired to be performed at the same time, but since it is generally difficult to double-stain the transmitted light image and the fluorescent dye, it is not possible to perform the dyeing suitable for capturing the transmitted light image when the fluorescent dye is applied. , And so on.

For these reasons, when proper dyeing cannot be performed, the transmitted light image has a very low contrast, and an image that can be observed by a particle image analyzer cannot be obtained.

As an IFCM, there is a device capable of simultaneously obtaining a fluorescence image and a transmitted light image of the same cell. However, in this device, since the fluorescent dyeing is prioritized, the dyeing for the transmitted light image is carried out, but it is not necessarily the optimum one for the transmitted light image. Had a problem that the contrast was extremely weak, granules, etc. were hardly recognizable, and even the shape of the cell nucleus was difficult to recognize.

By the way, in the field of microscopes, there is a phase contrast microscope capable of observing an object by a phase contrast method. When this phase contrast microscope is used, if there is a difference in the refractive index between the observation target and the surrounding substance, the observation target can be observed with good contrast without dyeing. However, it took time and effort to observe many cells and microorganisms one by one while observing them under a microscope.

The present invention has been made in view of the above points, and by imaging a particle by a phase difference method,
Provided is an imaging flow cytometer capable of capturing a transmitted light image of particles with good contrast even when appropriate dyeing for a transmitted light image cannot be performed.

[0009]

According to the present invention, a light-transmissive flow cell in which a sample solution containing particles flows, and a light irradiating means for irradiating the imaging area of the flow cell with light. A transmitted light image pickup means for picking up a transmitted light image of particles existing in the image pickup area of the flow cell, a ring slit for phase difference provided in the optical path from the light irradiation means to the flow cell, and the flow cell There is provided an imaging flow cytometer including an objective lens for phase difference provided in the optical path from the image pickup means to the transmitted light image pickup means.

The most characteristic point of the present invention is that the transmitted light image of the particles contained in the sample liquid is imaged by the phase difference method. The phase difference method is a method capable of capturing an image of an object with good contrast by utilizing light diffraction, and a method used in a known phase difference microscope can be applied.

As the flow cell, a light-transmissive tube made of, for example, glass or plastic capable of imaging particles from the outside of the flow cell is applied. The flow cell may have various shapes such as a circular cross section (round sheath), a rectangle, a rectangle, and an ellipse.
In order to clearly image the particles, it is necessary to stabilize the focal position with respect to the particles and to image the particles one by one without overlapping. Therefore, from such a point, if the particles are separated, and if the particles are flat, the particles can be oriented so as to face the imaging direction, and the particles can flow. It is suitable to use the flat sheath flow cell of. As such a flat sheath flow cell, a known flow cell capable of forming a flat sample liquid flow having a width of about 50 to 300 μm and a thickness of about 1/10 thereof can be used. For example, those described in JP-A-3-105235 can be applied.

As the sample solution introduced into the flow cell, a sample solution obtained by arbitrarily diluting blood, urine, powder or the like is mainly applied. Depending on the application, the sample may be seawater,
It may be lake water or river water. The particles may be of any size as long as they can be introduced into the flow cell, the material,
The specific gravity, shape, etc. are not particularly limited. Mainly microbes,
Blood cells contained in blood, cells contained in urine, powder and the like are applied as particles.

As the light irradiating means, various light sources capable of irradiating the particles with light having an intensity capable of obtaining a transmitted light image can be used. From the viewpoint of capturing the transmitted light image of 1), it is suitable to use a laser light irradiation device that can irradiate laser light that easily causes diffraction.

As the light source, it is preferable to use a pulsed light emitting type light source in order to image the particles without blurring.
However, it is also possible to use a continuous light emission type light source, but in that case, such as an image intensifier (photomultiplier tube) having a gate function (optical shutter function) in front of the transmitted light image pickup means. An optical amplifier is placed and configured to trigger the gate.

The transmitted light image pickup means may be any one capable of picking up a transmitted light image of a particle by the light applied to the particle. For example, a commercially available video camera can be used.

The phase difference ring slit may have various shapes as used in a known phase difference microscope. The ring slit for phase difference and the objective lens for phase difference must be related to each other, and as the objective lens for phase difference, a substance that shifts the phase (called a phase film) is used. What is applied on the back focal plane of the objective lens in the same shape as the projected image of the ring slit for phase difference is used. In this case, a phase film may be applied to a portion other than the projected image of the phase difference ring slit.

With such a structure, the transmitted light image of the particles can be captured with good contrast without dyeing. Further, by exchanging the ring slit for phase difference, a transmitted light image or a dark field image can be easily captured.

In the imaging flow cytometer of the present invention, in order to image the particles efficiently, a particle passage monitoring system is added, and it is confirmed that the particles have entered the imaging area and then the particles are imaged. Is suitable.

As this particle passage monitoring system, there is a particle passage monitoring system comprising a continuous light emitting light source for detecting particles and a detector for detecting passage of particles through the imaging area of the flow cell by light from the continuous light emitting light source. Applied.

As described above, by adding the particle passage monitoring system, dust and small particles are excluded, only desired particles are selected, and blank shot (images other than particles are shot). It is possible to efficiently capture the transmitted light image of the particles.

When the light irradiation means is a pulse emission type light source, when the particle passage monitoring system is added, a pulse emission type light source is added to the particle passage monitoring system when the passage of particles is detected by the detector. It is preferable to apply the one further provided with a signal processing means for emitting light.

When the light irradiating means is a continuous emission type light source, when the particle passage monitoring system is added,
As described above, the optical amplifier having the gate function is arranged in front of the transmitted light image pickup means. Further, it is preferable to apply a particle passage monitoring system further provided with signal processing means for triggering the gate of the optical amplifier when passage of particles is detected by the detector.

In the imaging flow cytometer of the present invention, a fluorescent image pickup system can be added.
In that case, from the excitation light source that irradiates the imaging region of the flow cell with light for fluorescence excitation, and the fluorescence image capturing means that captures the fluorescence image of the particles existing in the imaging region of the flow cell by the fluorescence excited from the particles. In this configuration, a fluorescent image pickup system is added.

When this fluorescent image pickup system is added, a pulsed light source is used as the light irradiation means. When a pulsed light source is used and a particle passage monitoring system is added as described above, a gate function is provided for opening and closing the optical path entering the fluorescence image pickup means when the fluorescence image pickup system is added. The optical amplifier is arranged in front of the fluorescence image pickup means. The signal processing means of the particle passage monitoring system is configured so that, when the passage of particles is detected by the detector, the gate of the optical amplifier is triggered after a predetermined time to capture the fluorescence image of the particles.

As described above, when the fluorescence image pickup system is added, since it is possible to simultaneously pick up the fluorescence image together with the transmitted light image for one particle, new information (fluorescence amount, fluorescence emission point locality) can be obtained. Can be obtained).

In this case, an image pickup signal of the transmitted light image of the particle is received from the transmitted light image pickup means, and an image pickup signal of the fluorescent image of the particle is received from the fluorescent image pickup means, and the transmitted light image and the fluorescent image of the imaged particle are received. By providing a configuration in which the image processing means for displaying in a pair on a single screen is further provided, the transmitted light image and the fluorescence image can be displayed in a pair on a single screen. .

[0027]

DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will be described below in detail with reference to the embodiments shown in the drawings. The present invention is not limited to this.

FIG. 1 is an explanatory diagram showing the construction of an embodiment of an imaging flow cytometer according to the present invention. This example shows the most basic configuration. This imaging flow cytometer forms a flat sample flow (also called sample flow) by introducing a sample liquid (also called sample liquid) diluted with blood or urine into a transparent and flat tube called a flow cell. By irradiating the sample flow with pulsed light such as strobe light or pulsed laser light, particles such as blood cells contained in blood or cells contained in urine (sometimes referred to as test particles) are video cameras. It is a device for imaging.

In this figure, 1 is a flow cell made of a transparent and flat tube such as glass or plastic. When the sample liquid 2 is introduced into the flow cell 1, at the same time, the sheath liquid 9 is supplied so as to cover the periphery of the sample liquid 2, and the sample liquid 2 and the sheath liquid 9 become a laminar flow. What is called a "sheath flow" flows in the flow cell 1. In FIG. 1, the sample liquid 2 and the sheath liquid 9 flow in the direction perpendicular to the paper surface. The width of the sample flow in the flow cell 1 in the longitudinal direction (the direction indicated by the arrow A in the figure) is 50 to 300 μm, and the width in the lateral direction (the direction indicated by the arrow B in the figure), that is, the thickness in the imaging direction is about 1 thereof. / 10 to 5 to 30 μm.

Reference numeral 3 is a light source for illuminating the imaging area of the sample liquid 2 flowing in the flow cell 1 with illumination light. The light source 3 for illumination is, for example, a flash lamp or a pulsed light source such as a laser light emitting device that emits pulsed laser light. The light source 3 is caused to periodically emit light to perform imaging. Reference numeral 4 is a condenser lens for condensing light, and 5 is a collimator lens for collimating the light. In the figure, the arrow indicates the traveling direction of light.

Reference numeral 6 denotes a phase difference ring slit provided between the condenser lens 4 and the collimator lens 5, and the phase difference ring slit 6 is provided at the front focal position of the condenser lens 4. Phase difference ring slit 6
An example of the shape of is shown in FIG. The phase difference ring slit 6 may be a pinhole.

Reference numeral 7 denotes a transmitted light image pickup section composed of a video camera, and picks up a transmitted light image of particles existing in the image pickup area of the flow cell 1 by the illumination light emitted from the light source 3.
The transmitted light image pickup unit 7 has a light receiving surface formed of, for example, a CCD for forming a transmitted light image of particles. The transmitted light image captured by the transmitted light image capturing unit 7 is processed by an image processing device (not shown).

Reference numeral 8 denotes a phase difference objective lens provided between the flow cell 1 and the transmitted light image pickup section 7. The phase difference objective lens 8 is coated with a substance that shifts the phase (referred to as a phase film). That is, on the back focal plane of the phase difference objective lens 8, a phase film is applied in the same shape as the image projected by the phase difference ring slit 6. This phase film may be applied to a portion other than the projected image of the phase difference ring slit 6. As the phase film, for example, a known material used in a phase contrast microscope such as a metal film or a dielectric film can be used.

In such a structure, the illumination light from the light source 3 is applied to the phase difference ring slit 6 via the condenser lens 4. The light beam passing through the phase difference ring slit 6 passes through the condenser lens 5 and the flow cell 1
The imaging area of is illuminated with a parallel light beam (that is, Koehler illumination).

The light that passes through the phase difference ring slit 6 and illuminates particles as an object is divided into light that is not diffracted by the particles and light that is diffracted by the particles. The light not diffracted by the particles once forms an image on the phase film of the objective lens. On the other hand, the light diffracted by the particles passes through a portion of the objective lens other than the phase film and forms an image on the light receiving surface of the transmitted light image pickup unit 7.

The light that is not diffracted by the particles is the background light of the particles, and there is a phase shift between the light that is diffracted by the particles and the light that is not diffracted (that is, background light). When an object having a different refractive index from the surroundings is observed due to the phase difference effect on the light receiving surface, it becomes possible to observe with contrast. As the principle of this phase difference method, the principle of a conventionally known phase difference microscope can be applied as it is.

In this example, the light source 3 for illumination uses a pulse light emission type light source, but in order to capture the cells flowing in such a pulse light emission type light source 3 as a still image without blurring, The light emission time of the light source 3 depends on the flow velocity of the sample flow. In this example, light emission is performed periodically, and the light emission period depends on the number of image capturing frames per second of the transmitted light image capturing unit 7. For example, when the transmitted light image pickup unit 7 is a video camera, it is usually 30 frames / sec. Therefore, if the particles are exactly in the imaging region of the flow cell 1 when the light source 3 for illumination emits light, a transmitted light image of the particles is captured.

Further, a continuous light emission type light source is used as the light source 3 for illumination, and shutter means is provided between the light source 3 and the transmitted light image pickup section 7, whereby a still image of particles can be picked up. When an image intensifier with a shutter function is used as this shutter means, it is arranged in front of the transmitted light image pickup section 7.

If the phase difference ring slit 6 can be easily removed or replaced with a diaphragm or the like, a normal transmitted light image that is not of the phase difference method can be taken. Further, if the phase difference ring slit 6 is changed to a ring slit that is irradiated with an NA larger than the NA (numerical aperture) of the objective lens 8 (however, it does not include NA smaller than the NA of the objective lens 8), Dark field observation is possible.

FIG. 3 is an explanatory diagram showing an example in which a particle passage monitoring function is added. In addition, in FIG.
The same components as those of the above are given the same reference numerals and the description thereof will be omitted.

In the above-mentioned example using the light source 3 of the pulse emission type, the light source 3 is periodically made to emit light for imaging, but when the sample liquid 2 contains a small number of particles, it is imaged. The number of particles is small and the imaging efficiency is poor. Therefore, in this example, a particle passage monitoring function is added, and after confirming that the particles have entered the imaging region of the flow cell 1, the light source 3 is caused to emit light and a particle image is captured.

In this figure, 11 is a light source for particle passage monitoring. The light source 11 is a continuous light emission type laser light source. 12 and 13 are dichroic mirrors, 14
Is a detector for detecting the passage of particles through the imaging region of the flow cell 1 by the light from the light source 11, and 15 is a detector 14
The signal processing unit causes the light source 3 to emit light when passage of particles is detected by. These light source 11, dichroic mirrors 12 and 13, detector 15, and signal processing unit 15
A particle passage monitoring system is constructed from.

As the light source 11 for monitoring particle passage, a continuous irradiation type laser light source is used as described above. The wavelength of the laser light is preferably not in the visible range if possible, but is not particularly limited. As the detector 14 for monitoring particle passage, a one-dimensional image sensor (line sensor), a photodiode having a slit, or the like can be used.

In this example, the transmitted light of the particles or the forward scattered light is detected, but the side scattered light may be detected using a photomultiplier (photomultiplier tube) or the like.

In such a structure, the light emitted from the light source 11 for monitoring particle passage is reflected by the dichroic mirror 12 and narrowed down, for example, to irradiate the sample stream 2.
Then, it is reflected by the dichroic mirror 13 and detected by the detector 14. When a one-dimensional image sensor is used as the detector 14, the light-irradiated region of the sample flow 2 is monitored by the one-dimensional image sensor.

FIG. 4 shows the relationship between the imaging area and the particle passage monitoring area.
Shown in. In the figure, 21 is a flowing particle (in the figure, the arrow indicates the flowing direction of the sample liquid 2), 22 is an imaging region of the transmitted light image capturing unit 7 in the flow cell 1, and 23 is a light source 11 for particle passage monitoring. An irradiation region, 24 is a particle passage monitoring region of the detector 14, and this particle passage monitoring region is a detection region of the one-dimensional image sensor when the one-dimensional image sensor is used as the detector 14.

The signal output from the detector 14 is passed to the signal processing section 15 for processing. Here, if the particles entering the imaging region 22 are the target particles, the light source 3 for illumination is triggered and the light source 3 emits light. Then, similarly to the above-described example, the transmitted light image by the phase difference method is captured by the transmitted light image capturing unit 7.

The above two examples are for capturing only transmitted light images, and these examples are useful when the desired particles cannot be dyed appropriately for capturing transmitted light images.

For example, in the examination of urinary sediment, ordinary staining for urinary sediment (Sternheimer-Marvin staining, etc.)
When subjected to, since the glass column is hardly stained, the contrast is weak, it was difficult to observe, by using the imaging flow cytometer of the present invention,
It is possible to obtain a glass column image with good contrast.

Although it is sometimes desired to observe microorganisms such as plankton alive without staining, even in such a case, by using the imaging flow cytometer of the present invention, contrast can be obtained without staining. It can be observed as a good microbial image.

FIG. 5 is an explanatory view showing an example in which a fluorescent image pickup system is added. In this figure, the same components as those in FIG. 3 described above are designated by the same reference numerals, and the description thereof will be omitted.

In this figure, 31 is an excitation light source for exciting fluorescence, 32 is a dichroic mirror, and 35 is
Is an image intensifier, and 36 is a relay lens. The relay lens 36 may be an optical fiber. Reference numeral 37 is a fluorescent image capturing section including a video camera, and 38.
Is an image processing unit.

The above-mentioned two examples were for picking up only the transmitted light image, but in this example, an image pickup system for the fluorescent image is added to obtain both the transmitted light image and the fluorescent image from the same particle. I am trying to get it. Figure 3 shows the particle passage monitoring system.
It is similar to that described in. In this example, both the light source 3 for illumination and the light source 31 for excitation use a pulse emission type laser light source. As the light source 31 for excitation, a light source having a wavelength that can excite the target fluorescent dye is selected. For example, a Xe flash lamp to which a bandpass filter is added, a pulse laser emitting device, or the like is used.

Since fluorescence is generally very weak, it cannot be imaged with a normal video camera alone because its sensitivity is too low. Therefore, in this example, an image intensifier (also referred to as “II”) 35 is used to amplify weak light two-dimensionally. However, when the emitted fluorescence is strong, the image intensifier 35 is unnecessary. The image intensifier 35 has a gate function (optical shutter function).

In such a configuration, when the passage of the target particles is detected by the particle passage monitoring system, the signal processing unit 15 triggers the light source 3 for illumination.
As a result, the illumination light source 3 emits light, and some of the light amount of the transmitted light image is transmitted through the beam splitter 32, and is captured by the transmitted light image capturing unit 7 by the phase difference method.

At this time, some of the remaining amount of the transmitted light image is reflected by the beam splitter 32 and reaches the image intensifier 35. However, since the gate of the image intensifier 35 is off at this time (the optical shutter is closed), the intense transmitted light image is not amplified by the image intensifier 35, and the image intensifier 35 is not amplified. Fire 35 is not damaged.

After some time, the particles are in the flow cell 1
The light source 31 for excitation is triggered when it moves a little inside, and the light source 31 emits light by this. The gate of the image intensifier 35 is turned on a little before that (several μsec), the optical shutter of the image intensifier 35 is opened, and the weak fluorescence is two-dimensionally amplified, and is passed through the relay lens 36. And fluorescent image pickup unit 3
A fluorescent image is captured at 7. When the light source 31 for excitation has finished emitting light, the gate of the image intensifier 35 is turned off and the optical shutter is closed.

The respective images picked up by the transmitted light image pick-up unit 7 and the fluorescent light image pick-up unit 37 are sent as video signals to the image processing device 36, and the particle shape, the number of fluorescent light emitting units are measured, and the fluorescent light emitting units are measured. The size and position are measured, the shape (circularity, etc.) of the fluorescent light-emitting portion is measured, and transmitted light images and fluorescent images of a large number of imaged cells are paired and displayed together on one screen.

FIG. 6 shows an example of a screen obtained when white blood cells are measured. As shown in this figure, a fluorescence image a and a transmitted light image b by the phase difference method are paired,
It can be displayed together on one screen.

In this example, a pulse emission type light source 31 is used as the excitation light source 31, and a normal image intensifier 35 is used as the image intensifier 35. However, the excitation light source is used. Three
A continuous light emission type may be used as 1, and an image intensifier having a high speed gate function may be used as the image intensifier 35.

To obtain a fluorescent image, the particles are usually subjected to fluorescent staining. However, it is generally difficult to simultaneously perform the fluorescent dyeing and the dyeing for obtaining the transmitted light image. Therefore, when the fluorescent dyeing is performed, the dyeing suitable for the transmitted light image cannot be performed. When it is not possible to stain the transmitted light image in this way, for example, in imaging white blood cells,
When using a conventional imaging flow cytometer that is not a phase difference method, the obtained transmitted light image of white blood cells has weak contrast and even the shape of the nucleus was not clear, but even in such a case, the imaging flow site of the present invention was used. By using the meter, the shape of white blood cells and the like can be imaged with good contrast.

[0062]

According to the present invention, by using the phase difference method, it is possible to obtain an image with good contrast without staining the particles such as microorganisms, cells and blood cells to be imaged. . Therefore, it is possible to observe the unstainable particles that were difficult with the conventional imaging flow cytometer, and it is also possible to observe the transmitted light image that could hardly be observed with the conventional imaging flow cytometer with the fluorescent image capturing function. It will be possible. Further, by exchanging the ring slit for phase difference, a transmitted light image or a dark field image can be easily captured. Furthermore, by adding a particle passage monitoring system, dust and small particles are excluded, and only the target particles are selected, and the particles are efficiently ejected without blanking (imaging other than the particles). A transmitted light image can be captured.

[Brief description of drawings]

FIG. 1 is an explanatory diagram showing a configuration of an embodiment of an imaging flow cytometer according to the present invention.

FIG. 2 is an explanatory diagram showing an example of the shape of a phase difference ring slit.

FIG. 3 is an explanatory diagram showing an example in which a particle passage monitoring function is added.

FIG. 4 is an explanatory diagram showing a relationship between an imaging region and a particle passage monitoring region.

FIG. 5 is an explanatory diagram showing an example in which a fluorescent image capturing system is added.

FIG. 6 is an explanatory diagram showing an example of a screen obtained when white blood cells are measured.

[Explanation of symbols]

 1 Flow Cell 2 Sample Liquid 3 Light Source for Illumination Light 4 Condenser Lens 5 Collimator Lens 6 Ring Slit for Phase Difference 7 Transmitted Light Image Capture Section 8 Objective Lens for Phase Difference 9 Sheath Liquid 11 Light Source for Particle Passage Monitoring 12, 13, 32 Dichroic mirror 14 Detector 15 Signal processing unit 21 Particle 22 Imaging region 23 Light source irradiation region 24 Particle passage monitoring region 31 Excitation light source 35 Image intensifier 36 Relay lens 37 Fluorescence image imaging unit 38 Image processing unit a Fluorescence image b Transmitted light image

Claims (7)

[Claims] 1. A light-transmissive flow cell in which a sample liquid containing particles flows inside, a light irradiating means for irradiating light to an imaging area of the flow cell, and the light irradiation means to exist in the imaging area of the flow cell. A transmitted light image pickup means for picking up a transmitted light image of particles, a phase difference ring slit provided in the optical path from the light irradiation means to the flow cell, and an optical path provided from the flow cell to the transmitted light image pickup means. Imaging flow cytometer comprising an objective lens for phase difference. 2. A particle passage monitoring system comprising a continuous light emitting light source for detecting particles and a detector for detecting passage of particles through an imaging region of a flow cell by light from the continuous light emitting light source. 1. The imaging flow cytometer according to 1. 3. The imaging flow cytometer according to claim 1, wherein the light irradiating means is a pulsed light emitting type light source. 4. The signal processing means, wherein the light irradiating means comprises a pulse emission type light source, and the particle passage monitoring system emits a pulse emission type light source when passage of particles is detected by a detector. The imaging flow cytometer according to claim 2, further comprising: 5. An excitation light source for irradiating the imaging region of the flow cell with fluorescence excitation light, and a fluorescence image capturing means for capturing a fluorescence image of the particles existing in the imaging region of the flow cell by fluorescence excited by the particles. The imaging flow cytometer according to any one of claims 1 to 4, further comprising: 6. A fluorescence image pickup system further comprising an excitation light source for irradiating the image pickup region of the flow cell with fluorescence excitation light and a fluorescent image pickup means for picking up a fluorescent image of particles existing in the image pickup region of the flow cell. The fluorescent image capturing system has a gate unit that opens and closes an optical path entering the fluorescent image capturing unit, and when the signal processing unit of the particle passage monitoring system detects passage of particles by a detector, a predetermined time The imaging flow cytometer according to claim 4, wherein the rear gate means is opened to capture a fluorescence image of the particles. 7. The transmitted light image and fluorescence of the captured particles are received from the transmitted light image capturing means and the fluorescent light image capturing signal of the particles. 7. The image processing means further comprising image processing means for pairing images and displaying them together on one screen.
The described imaging flow cytometer.