CN111398237A - Cell detection device, application and cell classification detection method - Google Patents

Cell detection device, application and cell classification detection method Download PDF

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Publication number
CN111398237A
CN111398237A CN202010380361.9A CN202010380361A CN111398237A CN 111398237 A CN111398237 A CN 111398237A CN 202010380361 A CN202010380361 A CN 202010380361A CN 111398237 A CN111398237 A CN 111398237A
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cell
light source
image
observation window
cells
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廖常宇
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Sichuan danuodi Technology Co., Ltd
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Chengdu Liangyi Technology Co ltd
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/645—Specially adapted constructive features of fluorimeters
    • G01N21/6456—Spatial resolved fluorescence measurements; Imaging
    • G01N21/6458—Fluorescence microscopy
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/01—Arrangements or apparatus for facilitating the optical investigation
    • G01N21/03—Cuvette constructions
    • G01N21/05—Flow-through cuvettes
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N2021/6417—Spectrofluorimetric devices
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/00029—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor provided with flat sample substrates, e.g. slides
    • G01N2035/00099—Characterised by type of test elements
    • G01N2035/00158—Elements containing microarrays, i.e. "biochip"

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Abstract

The invention provides a cell detection device and an application thereof, and a cell classification detection method, and relates to the technical field of cell detection. The cell detection device based on the microfluidic chip provided by the invention has the advantages of simple structure, convenience in operation, no need of professional operation, strong universality, capability of classifying various cells through images shot by image acquisition equipment, effective improvement on cell classification detection efficiency and reduction in detection cost.

Description

Cell detection device, application and cell classification detection method
Technical Field
The invention relates to the technical field of cell detection, in particular to a cell detection device, application and a cell classification detection method.
Background
At present, a centrifugal method, a membrane filtration method and a flow cytometry method are mainly adopted to classify cells, wherein the centrifugal method classification is a technology for classifying different cells from tissue homogenate or blood by utilizing different sedimentation behaviors of cells with different sizes and densities in a centrifugal field, but the centrifugal method has the problems of high cost, long time consumption, need of preparing a medium solution, strict operation and difficult control; the membrane filtration method is a cell classification technology which uses a semipermeable membrane as a selection barrier layer, uses the selectivity (aperture size) of the membrane and uses the energy difference existing on two sides of the membrane as a driving force to allow certain specific types of cells to permeate and retain other types of cells so as to achieve the separation purpose, but the membrane method has the defects that the preparation of the membrane is too complicated, the requirements of different membranes on experimental environments are different, the service life of the membrane is short, and the universality is poor; the flow cytometry method combines a fluorescein labeled antibody with corresponding cells, excites the cells flowing in a single row by using a laser beam, and automatically analyzes or sorts the cells according to fluorescence carried by the cells, has a complex integral system and high price, needs a professional to operate, and is not suitable for being conveniently used in a laboratory.
In view of the above, the present invention is particularly proposed.
Disclosure of Invention
The invention aims to provide a cell detection device for relieving the technical problems of high operation requirement and poor universality in cell classification by a centrifugal method, a membrane filtration method and a flow cytometry method.
The cell detection device provided by the invention comprises a micro-fluidic chip, a light source and image acquisition equipment, wherein an observation window is arranged on the micro-fluidic chip, the light source is used for irradiating the observation window, and the image acquisition equipment is used for acquiring an image of cells flowing through the observation window under the irradiation of the light source.
Further, the micro-fluidic chip includes sample application district, detection zone and waste liquid district, the sample application district the detection zone with the waste liquid district communicates in proper order, the observation window set up in the detection zone.
Further, the image acquisition device is used for acquiring a fluorescence image and a morphological image when the cells flow through the observation window.
Further, the light source includes at least one of an incandescent light source, an HID light source, a fluorescent light source, and an L ED light source.
Further, the light source includes L ED light source and fluorescence light source, and the image capturing device captures morphological images of the cells as they flow through the observation window when L ED light source illuminates the observation window, and captures fluorescence images of the cells as they flow through the observation window when fluorescence light source illuminates the observation window.
Further, a first lens and a first color filter are arranged between the light source and the observation window.
Further, a second lens and a second color filter are arranged between the image acquisition device and the observation window.
The other purpose of the present invention is to provide the application of the cell detection device based on the microfluidic chip in cell classification detection.
The invention also aims to provide a cell classification detection method, which comprises the following steps:
adding cells into the microfluidic chip, collecting images of the cells flowing through the observation window under the irradiation of the light source by using the image collecting equipment, analyzing the cell images collected by the image collecting equipment, and classifying the cells;
preferably, the fluorescently stained cells are added to a microfluidic chip.
Further, an image acquisition device is used for respectively acquiring a fluorescence image and a morphological image when the cells flow through the observation window, and the cells are classified according to the fluorescence intensity of the cells in the fluorescence image and the cell size in the morphological image.
The cell detection device provided by the invention has the advantages of simple structure, convenience in operation, no need of professional operation, strong universality, capability of classifying various cells through images shot by image acquisition equipment, effective improvement on cell classification detection efficiency and reduction in detection cost.
The cell classification detection method provided by the invention has the advantages that the cell detection device based on the microfluidic chip is used for detection, the universality is high, the classification detection can be carried out on various cells, the operation is convenient, the detection efficiency is improved, and the detection cost is reduced.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.
FIG. 1 is a schematic structural view of a cell detection device provided in example 1 of the present invention;
fig. 2 is a schematic structural diagram of the microfluidic chip in fig. 1.
Icon: 100-a microfluidic chip; 101-a viewing window; 102-a sample addition zone; 103-detection zone; 104-a waste liquid zone; 200-a light source; 201-a first lens; 202-a first color filter; 300-an image acquisition device; 301-a second lens; 302-second color filter.
Detailed Description
The technical solutions of the present invention will be described clearly and completely with reference to the following embodiments, and it should be understood that the described embodiments are some, but not all, embodiments of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
The existing cells are generally classified and detected by a flow cytometer, and the specific operation comprises the following steps: after the cell to be detected is dyed by the fluorescent dye, the cell to be detected is wrapped by the sheath fluid under certain pressure and flows into the chamber, the cells are arranged into a single row, the cells are sprayed out from a nozzle of the flow chamber to form cell fluid flow, and the cell fluid flow is intersected with the incident laser beam. Cells are excited to generate fluorescence by being placed in a position where they intersect with an incident laser beam and a cell flow and with the incident laser beam, the cells are excited to generate fluorescence, forward angle scattered light and side angle scattered light generated from the cells are collected separately by an optical system, the cells are classified by performing arithmetic processing on the forward angle scattered light and the side angle scattered light, and the cells are analyzed comprehensively.
However, the whole system of the flow cytometer is complex, expensive, needs professional personnel to operate, and is not suitable for being used conveniently in a laboratory.
The microfluidic chip technology is a new technology for processing or manipulating micro-fluid in a micron-sized pipeline, can integrate basic operations of sample preparation, reaction, separation, detection and the like in biological, chemical and medical analysis processes on a micron-sized chip, and has the advantages of high integration, high efficiency, low consumption, low cost and the like. The microfluidic chip technology is widely applied to the biomedical fields of in-vitro diagnosis, nucleic acid detection, drug screening and the like, and reports of using the microfluidic chip for cell classification detection are not yet seen at present.
According to a first aspect of the present invention, the present invention provides a cell detection apparatus, including a microfluidic chip, a light source and an image acquisition device, wherein the microfluidic chip is provided with an observation window, the light source is used for illuminating the observation window, and the image acquisition device is used for acquiring an image of a cell flowing through the observation window under the illumination of the light source.
The cell detection device provided by the invention has the advantages of simple structure, convenience in operation, no need of professional operation, strong universality, capability of classifying various cells through images shot by image acquisition equipment, effective improvement on cell classification detection efficiency and reduction in detection cost.
In a preferred embodiment of the present invention, the microfluidic chip comprises a sample addition region, a detection region and a waste liquid region, wherein the sample addition region, the detection region and the waste liquid region are communicated with each other, and the detection region is provided with an observation window.
In a preferred embodiment of the present invention, the sample adding region is used for adding the cells to be tested, and can be configured into a circular or elliptical cavity structure, and can also be configured into other shapes.
In a preferred embodiment of the present invention, the detection zone is disposed between the sample addition zone and the waste liquid zone in the form of a microchannel, the diameter of the microchannel being 10 μm to 4 mm.
Typically, but not by way of limitation, the diameter of the microchannels may be, for example, 10 μm, 12 μm, 15 μm, 18 μm, 20 μm, 30 μm, 50 μm, 80 μm, 1mm, 1.2mm, 1.5mm, 1.8mm, 2mm, 2.5mm, 3mm, 3.5mm or 4 mm.
In a preferred embodiment of the present invention, a filtering region is disposed between the sample application region and the detection region to remove impurities from the cells to be detected, thereby improving the accuracy of the detection result.
In a preferred embodiment of the present invention, the filter area is made of filter paper, and the filter paper is selected from different models according to different analytes.
In a further preferred embodiment of the present invention, the filtration zone may be composed of micro-columns arranged in an array, the diameter of the micro-columns is 1 μm-2mm, and the distance between adjacent micro-columns is 10 μm-5 mm.
Typically, but not limited to, the diameter of the microcolumn is, for example, 1 μm, 2 μm, 8 μm, 10 μm, 12 μm, 15 μm, 18 μm, 20 μm, 30 μm, 50 μm, 80 μm, 1mm, 1.2mm, 1.5mm, 1.8mm or 2 mm.
Typically, but not by way of limitation, the distance between adjacent microcolumns is 10 μm, 12 μm, 15 μm, 18 μm, 20 μm, 30 μm, 50 μm, 80 μm, 1mm, 1.5mm, 2mm, 2.5mm, 3mm, 3.5mm, 4mm, 4.5mm, or 5 mm.
In a preferred embodiment of the present invention, the waste liquid region is used for storing the cells to be tested after the test, and the shape thereof is not limited.
In a preferred embodiment of the present invention, a viewing window is provided at the detection region to facilitate the image collecting apparatus to collect an image of the cell flowing through the detection region through the viewing window to classify the cell according to the image of the cell.
In a preferred embodiment of the invention, the image acquisition device is used for acquiring fluorescence images and morphology images of cells flowing through the observation window.
In the above preferred embodiment of the present invention, the morphological image refers to an image in which the morphology (e.g., size, shape, texture, transparency, etc.) of the cell can be known, such as an image captured by a digital camera.
The fluorescence image of a certain cell flowing through the observation window is collected, the fluorescence intensity of the cell flowing through the observation window can be obtained through calculation, the morphological information such as the size of the cell can be obtained through collecting the morphological image of the cell flowing through the observation window, the fluorescence intensity and the size of the cell flowing through the observation window are comprehensively analyzed, and the cell is classified.
In a preferred embodiment of the invention, the light source comprises at least one of an incandescent light source, a HID light source, a fluorescent light source and an L ED light source.
The term "at least one" as used herein means that the light source may be an incandescent light source, an HID light source, a fluorescent light source, or an L ED light source, or two or more of the above light sources may be used in combination.
The HID light source is referred to as a high pressure gas discharge light source and the L ED light source is referred to as a light emitting diode light source.
In a preferred embodiment of the present invention, when the image capturing device captures an image of a cell flowing through the observation window, both the fluorescence image and the morphological image may be captured under the irradiation of the same light source, or the fluorescence image and the morphological image may be captured under the irradiation of different light sources.
In a preferred embodiment of the invention, the light source comprises an L ED light source and a fluorescent light source, the image capture device capturing morphological images of the cells as they flow through the viewing window when the L ED light source illuminates the viewing window, and the image capture device capturing fluorescent images of the cells as they flow through the viewing window when the fluorescent light source illuminates the viewing window.
When L ED light source shines the observation window, the morphological image that adopts image acquisition equipment to shoot to obtain is more clear, can obtain morphological information such as more accurate cell size through the morphological image, and when adopting fluorescence light source to shine the observation window, under fluorescence light source's excitation, the fluorescence intensity of cell is stronger, and the fluorescence image that obtains is more clear to effectively improve the accuracy of cell classification.
In a preferred embodiment of the present invention, in the cell detection device of the microfluidic chip, a first lens and a first color filter are disposed between the light source and the observation window.
The first lens and the first color filter are arranged between the light source and the observation window, so that the anti-interference capability of the light source is further improved, the divergence angle is reduced, and the illumination effect is improved.
In a preferred embodiment of the invention, a second lens and a second color filter are arranged between the image acquisition device and the viewing window.
Through set gradually second lens and second color filter between image acquisition equipment and observation window to further reduce the interference, improve the degree of accuracy that detects.
According to a second aspect of the present invention, the present invention provides the use of the microfluidic chip based cell detection device provided in the first aspect in cell classification detection.
The cell detection device based on the microfluidic chip according to the present invention can detect not only cell types but also biological types such as sperm, chromosome, and bacteria.
When the cell detection device based on the microfluidic chip is used for cell classification detection, the cell detection device is high in universality and convenient and fast to detect, the cell classification detection efficiency can be effectively improved, and the detection cost is reduced.
According to a third aspect of the present invention, there is provided a cell sorting method comprising the steps of:
the cell is added into the microfluidic chip, the image acquisition equipment acquires an image of the cell flowing through the observation window under the irradiation of the light source, the cell image acquired by the image acquisition equipment is analyzed, and the cell is classified.
The cell classification detection method provided by the invention has the advantages that the cell detection device based on the microfluidic chip is used for detection, the universality is high, the classification detection can be carried out on various cells, the operation is convenient, the detection efficiency is improved, and the detection cost is reduced.
In a preferred embodiment of the invention, the cells are fluorescently stained cells.
The cells are subjected to fluorescent staining and then added into the microfluidic chip, so that the clear fluorescent image and morphological image of the cells flowing through the observation window can be shot through the observation window under the irradiation of a light source by the image acquisition equipment.
In one embodiment of the invention, the cells are fluorescently stained according to the following steps:
(1) 100 μ L cell suspension (cell density about 1 × 10)6Individual cells);
(2) adding a corresponding amount of fluorescein isothiocyanate or phycoerythrin labeled specific fluorescent direct-labeling monoclonal antibody;
(3) reacting at room temperature in a dark place for a certain time (the time is carried out according to the requirements of a reagent specification), and reacting at room temperature for 15-30 min;
(4) adding 500 mu L phosphate buffer to re-suspend into single cell suspension, completing cell staining, and directly adding into micro-fluidic chip for detection.
In a preferred embodiment of the present invention, in the cell classification detection, the fluorescence image and the morphological image of the cell flowing through the observation window are respectively collected by using an image collecting device, and the cell is classified according to the fluorescence intensity of the cell in the fluorescence image and the cell size in the morphological image.
In one embodiment of the present invention, the cell sorting assay is performed as follows:
(1) adding the cells subjected to fluorescent staining into a microfluidic chip, enabling the cells subjected to fluorescent staining to flow to a detection region from a sample adding region, shooting a fluorescent picture L by an image acquisition device under fluorescent irradiation when the cells flow through an observation window on the detection region, then quickly switching to a L ED light source to irradiate the observation window, and shooting a morphological image F when the cells subjected to fluorescent staining flow through the observation window by the image acquisition device;
(2) by analyzing the fluorescence image L and the morphological image F together, the fluorescence intensity of a particular cell in the fluorescence image L can be obtained as sf1 from the intensity corresponding to that cell, and the size of the cell can be obtained as fsc from the morphological image F corresponding to that cell.
(3) The fluorescence intensity is used as an abscissa, the cell size is used as an ordinate, a two-dimensional scattergram is established, a plurality of detection cells are marked at different positions of the two-dimensional scattergram according to the fluorescence intensity and the cell size of the detection cells, and the positions of different types of cells in the two-dimensional scattergram are different due to the difference of the fluorescence intensity and the cell size of the different types of cells, so that the cells can be classified according to the position of a specific detection cell in the two-dimensional scattergram.
In a preferred embodiment of the present invention, before performing the classification detection of the unknown cell, the fluorescence intensity and the cell size of different types of known cells are measured, a two-dimensional scattergram library of a plurality of types of known cells is created with the fluorescence intensity as an abscissa and the cell size as an ordinate, and then, when the type of the unknown cell is detected, the type of the unknown cell can be clearly determined by the unknown in the two-dimensional scattergram of the unknown cell.
The technical solution provided by the present invention is further described with reference to the following examples.
Example 1
Fig. 1 is a cell detection apparatus provided in embodiment 1 of the present invention, and as shown in fig. 1, the cell detection apparatus provided in this embodiment based on a microfluidic chip 100 includes a microfluidic chip 100, a light source 200, and an image acquisition device 300, where the microfluidic chip 100 is provided with an observation window 101, the light source 200 is used to irradiate the observation window 101, and the image acquisition device 300 is used to acquire an image when a cell under irradiation of the light source 200 flows through the observation window 101.
The cell detection device that this embodiment provided is simple structure not only, convenient operation need not the professional operation, and the universality is strong moreover, can classify the multiple cell through the image pair that image acquisition equipment 300 shot, has effectively improved the categorised detection efficiency of cell, has reduced the detection cost.
Fig. 2 is a schematic structural diagram of a microfluidic chip 100 in the cell detection apparatus provided in this embodiment, and as shown in fig. 2, in a preferred embodiment of this embodiment, the microfluidic chip 100 includes a sample addition region 102, a detection region 103, and a waste liquid region 104, the sample addition region 102, the detection region 103, and the waste liquid region 104 are sequentially communicated, and an observation window 101 is disposed in the detection region 103.
In a preferred embodiment of this embodiment, as shown in FIG. 2, the detection zone 103 is in the form of a microchannel having a diameter of 10 μm to 4 mm.
Typically, but not by way of limitation, the diameter of the microchannels may be, for example, 10 μm, 12 μm, 15 μm, 18 μm, 20 μm, 30 μm, 50 μm, 80 μm, 1mm, 1.2mm, 1.5mm, 1.8mm, 2mm, 2.5mm, 3mm, 3.5mm or 4 mm.
In a preferred embodiment of this embodiment, a filtering region can be further disposed between the sample-adding region 102 and the detection region 103 to remove impurities in the cell to be detected, so as to improve the accuracy of the detection result.
In a preferred embodiment of this embodiment, the filtering area is made of filter paper, and the filter paper is selected from different models according to different analytes.
In a preferred embodiment of the present embodiment, the image capturing apparatus 300 can capture both a fluorescence image when the cell flows through the observation window 101 and a morphological image when the cell flows through the observation window 101.
The morphological image refers to an image in which the morphology of the cell (e.g., information on size, shape, texture, transparency, etc.) can be clearly known, such as an image captured by a digital camera.
In a preferred embodiment of this embodiment, the light source 200 is selected from at least one of an incandescent light source 200, an HID light source 200, a fluorescent light source 200, and an L ED light source 200.
In the present embodiment, when the image capturing apparatus 300 captures an image of a cell flowing through the observation window 101, both a fluorescence image and a morphological image may be captured under the irradiation of the same light source 200, or a fluorescence image and a morphological image may be captured under the irradiation of different light sources 200.
In a preferred embodiment of the present embodiment, a first lens 201 and a first color filter 202 are disposed between the light source 200 and the observation window 101.
By disposing the first lens 201 and the first color filter 202 between the light source 200 and the observation window 101, the interference rejection of the light source 200 is further improved, the divergence angle is reduced, and the illumination effect is improved.
In a preferred embodiment of this embodiment, a second lens 301 and a second color filter 302 are arranged between the image capture device 300 and the viewing window 101.
By sequentially arranging the second lens 301 and the second color filter 302 between the image capture device 300 and the observation window 101, interference is further reduced, and the detection accuracy is improved.
In a preferred embodiment of this embodiment, the light source 200 comprises a first light source and a second light source, wherein the first light source is an L ED light source and the second light source is a fluorescent light source.
When the observation window 101 is irradiated by the L ED light source, the morphological image of the cell flowing through the observation window 101 is shot by the image acquisition equipment 300, and when the observation window 101 is irradiated by the fluorescent light source, the fluorescent image of the cell flowing through the observation window 101 is shot by the image acquisition equipment 300. when the observation window 101 is irradiated by the L ED light source, the morphological image shot by the image acquisition equipment 300 is clearer, more accurate morphological information such as cell size can be obtained through the morphological image, when the observation window 101 is irradiated by the fluorescent light source, the fluorescent intensity of the cell is stronger under the excitation of the fluorescent light source, the obtained fluorescent image is clearer, and therefore, the accuracy of cell classification is effectively improved.
Example 2
The embodiment provides a cell classification detection method, which is used for cell classification detection by using the cell detection device based on the microfluidic chip provided by the embodiment 1 of the invention, and specifically comprises the following steps:
(1) adding the cells subjected to fluorescent staining into a microfluidic chip, enabling the cells subjected to fluorescent staining to flow to a detection region from a sample adding region, shooting a fluorescent picture L by an image acquisition device under fluorescent irradiation when the cells flow through an observation window on the detection region, then rapidly switching to a L ED light source to irradiate the observation window, and shooting a morphological image F of the cells subjected to fluorescent staining when the cells flow through the observation window by the image acquisition device, wherein the cells are stained by a conventional method to obtain the cells subjected to fluorescent staining, and the description is omitted;
(2) by analyzing the fluorescence image L and the morphological image F together, the fluorescence intensity of a particular cell in the fluorescence image L can be obtained as sf1 from the intensity corresponding to that cell, and the size of the cell can be obtained as fsc from the morphological image F corresponding to that cell.
(3) The fluorescence intensity is used as an abscissa, the cell size is used as an ordinate, a two-dimensional scattergram is established, a plurality of detection cells are marked at different positions of the two-dimensional scattergram according to the fluorescence intensity and the cell size of the detection cells, the positions of different types of cells in the two-dimensional scattergram are different due to the difference of the fluorescence intensity and the cell size of the different types of cells, and therefore the cells are classified according to the positions of a specific detection cell in the two-dimensional scattergram.
In a preferred embodiment of this embodiment, before performing the classification detection of the unknown cell, the fluorescence intensity and the cell size of different types of known cells are measured, a two-dimensional scattergram library of a plurality of types of known cells with the fluorescence intensity as the abscissa and the cell size as the ordinate is created, and then when detecting the type of the unknown cell, the type of the unknown cell can be clearly determined by the position of the unknown cell in the two-dimensional scattergram.
Finally, it should be noted that: the above embodiments are only used to illustrate the technical solution of the present invention, and not to limit the same; while the invention has been described in detail and with reference to the foregoing embodiments, it will be understood by those skilled in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some or all of the technical features may be equivalently replaced; and the modifications or the substitutions do not make the essence of the corresponding technical solutions depart from the scope of the technical solutions of the embodiments of the present invention.

Claims (10)

1. The cell detection device is characterized by comprising a micro-fluidic chip, a light source and image acquisition equipment, wherein an observation window is arranged on the micro-fluidic chip, the light source is used for irradiating the observation window, and the image acquisition equipment is used for acquiring an image of cells under the irradiation of the light source when the cells flow through the observation window.
2. The cell detection device according to claim 1, wherein the microfluidic chip comprises a sample application region, a detection region and a waste liquid region, the sample application region, the detection region and the waste liquid region are sequentially communicated, and the observation window is disposed in the detection region.
3. The cell detecting apparatus according to claim 1, wherein the image acquiring device is configured to acquire a fluorescence image and a morphology image of the cell as it passes through the observation window.
4. The cell detection apparatus of claim 1, wherein the light source comprises at least one of an incandescent light source, an HID light source, a fluorescent light source, and an L ED light source.
5. The cell detection apparatus according to claim 1, wherein the light source comprises L ED light source and a fluorescent light source, and the image capturing device captures a morphological image of the cell as it flows through the observation window when the L ED light source illuminates the observation window, and the image capturing device captures a fluorescent image of the cell as it flows through the observation window when the fluorescent light source illuminates the observation window.
6. The cell detecting apparatus according to claim 1, wherein a first lens and a first color filter are provided between the light source and the observation window.
7. The cell detecting apparatus according to claim 1, wherein a second lens and a second color filter are provided between the image pickup device and the observation window.
8. Use of the cell detection device according to any one of claims 1 to 7 in a cell sorting test.
9. A method for detecting a cell classification, comprising the steps of:
adding cells into the microfluidic chip, collecting images of the cells flowing through the observation window under the irradiation of the light source by using the image collecting equipment, analyzing the cell images collected by the image collecting equipment, and classifying the cells;
preferably, the fluorescently stained cells are added to a microfluidic chip.
10. The method for detecting cell classification according to claim 9, characterized by comprising the steps of: and respectively acquiring a fluorescence image and a morphological image of the cell flowing through the observation window by adopting an image acquisition device, and classifying the cell according to the fluorescence intensity of the cell in the fluorescence image and the cell size in the morphological image.
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