CN113781378A - Absolute value counting method for obtaining quantity of micro cells in cell suspension - Google Patents
Absolute value counting method for obtaining quantity of micro cells in cell suspension Download PDFInfo
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Abstract
The invention provides a method for obtaining the absolute value of the quantity of trace cells in cell suspension, which adopts a pipettor to take a sample to be measured and measure the volume of the sample to be measured, wherein the sample to be measured is the cell suspension; carrying out fluorescent staining on trace cells in the cell suspension and uniformly mixing; adding the dyed cell suspension into a counting plate, and inserting the counting plate into a full-automatic fluorescent cell analyzer; the image processing system carries out image shooting on the stained cell suspension; the image scanning system scans all the acquired images and acquires the total number of cells (the sum of the total number of live cells and the total number of dead cells), the controller can respectively obtain the absolute value of the total number of the trace cells in the cell suspension through the absolute value counting module according to the volume and the total number of the cells, and the defects of counting deviation caused by uneven distribution and small counting volume of the trace cells in the cell suspension are eliminated.
Description
Technical Field
The invention relates to the field of biomedicine, in particular to a method for obtaining the absolute value of the number of micro cells in cell suspension.
Background
How to accurately count the total number of trace or residual cells in a biological sample has not been solved well.
Currently, there are three types of residual cell counting methods that have been applied: cell counting method of artificial microscope and automatic fine image recognitionCell counting method, flow automatic cell counting method, wherein the principle of the manual microscope cell counting method is as follows: researchers use a blood cell counting plate (national standard JJJG 552-; secondly, counting rule of the pressed line cells: counting up but not counting down, counting left but not counting right; ③ counting the aggregated cells: counting the number of the clusters which are more than or equal to 2 to 1, and counting the clustered samples again after the clustered samples need to be processed; fourthly, a counting formula for counting concentration: cell concentration (one/ml) 4 total cells/4X 104X sample dilution times x specimen dilution times; the method has the advantages of 'what you see is what you get', high accuracy and is the gold standard for cell counting at present. However, when the concentration of the sample is too low, the counting deviation is large;
the principle of image recognition automated cytometry is as follows: the method is derived from a manual microscope cell counting method, the microscope is automatically upgraded, one or more small target surfaces are adopted for counting, and then the cell concentration is calculated: the total number of cells (the sum of live cells and dead cells) is further calculated by the method per milliliter, the method combines the advantages of a manual microscope cell counting method, and simultaneously combines the latest photosensitive chip technology, image recognition software technology and automatic control technology. Not only can reduce the labor intensity of operators, but also can keep original data such as original pictures of cells and the like, and can keep a file for backtracking. However, the method also adopts a concentration counting method, and the counting deviation is larger when the concentration of the sample is too low;
flow-through automated cytometry: using the particle counting principle to queue the cells through the detection window to obtain the electric signal change of each cell, and then reducing the concentration, diameter and number of the cells. The absolute value counting of the flow method is to use a large-volume sample to be injected, select the data of a certain section of volume and calculate the total number of cells in the section of volume;
the three methods have large errors in the counting of low-concentration (trace or residual) cell samples. The main reasons are as follows: firstly, the cell suspension is not a solution, but a suspension, and the cells cannot be uniformly distributed in the liquid; second, even in a low concentration (trace or residual) cell sample, the total number of cells in the sample varies greatly even with the same sampling volume, the same person, and the same sampling method, and therefore the total number calculated by the concentration calculation method varies greatly.
Disclosure of Invention
Aiming at the defects in the prior art and aiming at overcoming the limitation of the existing counting method, the invention provides a method for counting the number of cells in a trace cell suspension, which adopts a full-automatic fluorescence cell analyzer to process images and count the total number of cells, the number of live cells and the number of dead cells.
The embodiment of the invention provides
The method for obtaining the absolute value of the number of the micro cells in the cell suspension is characterized in that the counting method adopts a full-automatic fluorescent cell analyzer, the analyzer comprises a counting plate for containing the micro cell suspension, a container for storing fluorescent dye, an image scanning system, an image processing system and a controller for counting the number of the cells, and the method comprises the following specific operation steps:
the method comprises the following steps: withdrawing the cell suspension with a pipettor and measuring the volume of the cell suspension;
step two: adding a fluorescent dye into the cell suspension obtained in the step one to dye and uniformly mixing;
step three: adding the cell suspension dyed in the step two into the counting plate, and inserting the counting plate into the full-automatic fluorescent cell analyzer;
step four: the image processing system carries out image shooting on the cell suspension dyed in the third step, all images of the cell suspension on the counting plate are obtained according to the number of the images taken set by the image processing system, and the display areas of all the obtained images completely cover the occupied area of the sample to be detected on the counting plate;
step five: the image scanning system scans all the images acquired in the fourth step, and is used for acquiring the total number of cells, the total number of live cells and the total number of dead cells of the trace amount of cells in the cell suspension in all the images in the fourth step, wherein the total number of the cells is the sum of the total number of the live cells and the total number of the dead cells;
step six: the controller calculates the absolute value of the total number of the cells of the trace cells in the cell suspension through an absolute value counting module in the controller according to the total number of the cells obtained in the step five;
preferably, before the step one, a cell diluent is adopted to dilute the cell mixture to be counted and uniformly mix the cell mixture to obtain the cell suspension;
preferably, the diluted cell suspension and the fluorescent dye are uniformly mixed, the uniformly mixed liquid of the cell suspension and the fluorescent dye is added into the counting plate, and the volume of the diluted cell suspension, the fluorescent dye and the mixed liquid is the same;
preferably, the cell mixture may be selected from anticoagulated whole blood, filtered white blood or other cell culture medium containing a trace amount of cells;
preferably, a full-image large-image splicing function module is arranged in the image processing system and used for splicing all cell images of the cell suspension collected by the image processing system into a full image of the cell suspension, and a display area of the full image completely covers an occupied area of the sample to be detected on the counting plate;
preferably, the first and second liquid crystal materials are,
the absolute value calculation module comprises an absolute value concentration calculation module which calculates the absolute value concentration of the cell suspension through a mathematical calculation formula, wherein,
the mathematical formula is Q ═ n/V, where Q represents the absolute concentration of cells, n represents the total number of cells in all the images, and V represents the volume of the cell suspension;
if Q is the absolute value concentration of the total number of cells, n is the total number of cells in all the images;
if Q is the absolute value concentration of the total number of the living cells, n is the total number of the living cells in all the images;
if Q is the absolute value concentration of the total number of dead cells, n is the total number of dead cells in all the images;
the total cell number absolute value concentration is the sum of the total live cell number absolute value concentration and the total dead cell number absolute value concentration;
preferably, the first and second liquid crystal materials are,
the cell suspension is formed by diluting a cell mixed solution to be counted by a cell diluent or directly formed by the cell mixed solution to be counted, the cell suspension and the fluorescent dye are mixed and diluted and then are added into the counting plate, and counting is started after standing for a preset time, wherein the mathematical calculation formula is Q (n) k f/V, wherein Q represents the number of cells, n represents the total number of the cells in all images, k represents the dilution multiple of the cell mixed solution to be counted by the cell diluent, f represents the dilution multiple of the diluted cell suspension by the fluorescent dye, and V represents the volume of the cell mixed solution to be counted before being released by the cell diluent;
if Q is the absolute value concentration of the total number of cells, n is the total number of cells in all the images;
if Q is the absolute value concentration of the total number of the living cells, n is the total number of the living cells in all the images;
if Q is the absolute value concentration of the total number of dead cells, n is the total number of dead cells in all the images;
the total cell number absolute value concentration is the sum of the total live cell number absolute value concentration and the total dead cell number absolute value concentration;
preferably, the absolute value counting formula is that the absolute value of the total number of cells is equal to the sum of the total number of dead cells and the total number of live cells.
Has the advantages that:
1. the invention adopts a full-automatic fluorescent cell analyzer to carry out image acquisition and image scanning and processing on trace cell suspension after fluorescent staining in a counting plate and automatically calculate the total number of cells (the sum of the number of live cells and dead cells) by a controller, counts the total number of cells in the volume of the cell suspension to be measured, calculates the absolute value number of the total number of cells by using an absolute value counting module, and can also calculate the absolute value concentration of the total number of cells in the unit volume of the cell suspension to be measured by using an absolute value concentration counting module, thereby eliminating the counting deviation, namely the cell concentration deviation, caused by uneven distribution and small counting volume of trace or residual cells in the cell suspension.
2. The invention adopts a full-automatic fluorescent cell analyzer, which also comprises a microscope with visible light and a fluorescent microscopic light path, the dead and alive states of cells after non-staining and trypan blue staining can be counted by using the visible light microscopic light path, the dead and alive states of cells after fluorescent dye staining can be counted by using the fluorescent microscopic light path, and the counting function of the manual microscope is reserved.
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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 introduced below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to these drawings without creative efforts.
FIG. 1 is a counting plate of the present invention;
FIG. 2 is another embodiment of the counting plate of the present invention;
FIG. 3 is an interface diagram of the operation of the controller of the fully automatic fluorescent cell analyzer of the present invention;
FIG. 4 is a diagram of the interface operated after the relevant information is entered in FIG. 3;
FIG. 5 is a pictorial scan mosaic of a fully automated fluorescence cell analyzer of the present invention;
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are some, but not all, embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without any creative effort belong to the protection scope of the present invention, and the fully automatic fluorescent cell analyzer adopted by the present invention comprises an LED light source, a touch display screen (control operation panel), an image processing system (including a camera for capturing fluorescent signals), an image scanning system (including image recognition and image analysis software for capturing images of fluorescent signals by the camera), a universal fluorescent dye (for performing fluorescent dyeing on living cells and/or dead cells), a disposable cell counting plate (for accommodating a trace amount of cell suspension), and a controller (including a data processing unit for calculating the number of cells or the concentration of cells (the number of cells in a unit volume)), since the fully automatic fluorescent cell analyzer adopted by the present invention is a outsourced product, for implementing the counting method of the present invention, the detailed structure and operation principle of the fully automatic fluorescence cell analyzer will not be described herein.
As shown in fig. 1 to 5, the present invention provides a counting method for obtaining the number of cells in a micro cell suspension, the counting method employs a fully automatic fluorescent cell analyzer (JSY-FL-045, model) including a counter plate for accommodating the micro cell suspension, preferably a disposable counter plate (JSP-GM, model) for storing a fluorescent dye, an image scanning system, an image processing system and a controller for counting the number of cells, and the method includes the following steps: the method comprises the following steps: taking the cell suspension by using a pipettor and measuring the volume of the cell suspension; step two: adding a fluorescent dye into the cell suspension obtained in the step one to dye and uniformly mixing; step three: adding the cell suspension dyed in the step two into the counting plate, and inserting the counting plate into the full-automatic fluorescent cell analyzer; step four: the image processing system carries out image shooting on the cell suspension dyed in the third step, and obtains all images of the sample to be detected on the counting plate according to the quantity of the pictures (the quantity of the pictures is loaded in the image analysis software of the image processing system in advance based on the detection of cells of different blood types, the user only needs to select according to the detected blood types, and the image analysis software automatically sets the quantity of the pictures) set by the image processing system, and the display areas of the obtained images completely cover the occupied area of the sample to be detected on the counting plate, namely the images completely cover the sample to be detected; step five: the image scanning system scans and identifies all the images acquired in the fourth step, and is used for acquiring the total number of cells, the total number of live cells and the total number of dead cells in the cell suspension in all the images in the fourth step, wherein the total number of the cells is the sum of the total number of the live cells and the total number of the dead cells; step six: and the controller calculates the absolute value of the total number of the cells of the trace cells in the cell suspension through an absolute value counting module in the controller according to the total number of the cells acquired in the step five. The method comprises the steps of counting the total number of cells in a cell suspension volume to be measured, calculating by using an absolute value counting module to obtain the absolute value number of the total number of the cells, and calculating the absolute value concentration of the total number of the cells in a unit volume of the cell suspension to be measured by using an absolute value concentration counting module, so that the counting deviation caused by uneven distribution and small counting volume of trace or residual cells in the cell suspension, namely the cell concentration deviation, is eliminated, the trace or residual cells can be subjected to absolute value counting aiming at the number of white blood cells of peripheral blood, and can also be subjected to absolute value counting aiming at the number of all trace cells contained in cell mixed liquor (animal cell mixed liquor or plant cell mixed liquor).
Example 1
Before the first step, i.e. before taking the cell suspension by using a pipettor, preparing the cell suspension, diluting and uniformly mixing a cell mixture to be counted by using a cell diluent to obtain the cell suspension, wherein the cell mixture can be selected from anticoagulated whole blood, filtered white blood or other cell culture solutions containing a small amount of cells, the cell diluent can be selected from normal saline and Phosphate Buffered Saline (PBS), the cell mixture in example 1 is anticoagulated whole blood (with low content of leukocytes), the cell diluent is normal saline, the cell suspension is obtained by diluting the anticoagulated whole blood with normal saline, uniformly mixing the diluted cell suspension with a fluorescent dye, adding the mixed solution of the uniformly mixed cell suspension and fluorescent dye into the counting plate, and the volume of the diluted cell suspension, the fluorescent dye and the mixed solution is the same, and after standing for a preset time, starting counting.
The specific implementation method is that a certain amount of anticoagulated whole blood is diluted 5 times by physiological saline, 500 mul of anticoagulated whole blood is mixed with 2ml of physiological saline and stirred uniformly, 15 mul of diluted anticoagulated whole blood is taken by a pipettor and is mixed uniformly with 15 mul of fluorescent dye which is taken in advance to obtain a mixed solution of anticoagulated whole blood and fluorescent dye, 15 mul of mixed solution is taken by a pipettor and is added to the counting plate (as shown in figures 1 and 2), after standing for 1min, an image processing system (a camera with a fluorescent signal capture) of a full-automatic fluorescent cell analyzer is used for shooting the mixed solution in the counting plate for obtaining a cell image, preferably, the mixed solution in the counting plate has an area of 20mm × 40mm (which is slightly larger than the surface area of a groove for containing a cell suspension in the counting plate, the groove adopts a rectangular groove), as shown in figures 3 and 4, the method comprises the steps of self-defining the number of shot images for image analysis software in an image processing system (the number of shot images is not larger than the range of the number of shot images limited by image analysis software of a full-automatic fluorescent cell analyzer), ensuring that all positions of mixed liquid in the area of 20mm multiplied by 40mm are shot, preventing omission, automatically scanning and identifying the obtained images of the mixed liquid in the area by an image scanning system, utilizing a full-image large-image splicing function (image splicing software, namely the image analysis software) arranged in the image processing system to automatically splice a plurality of images to form a complete image (the spliced complete image area is larger than the physical size of a counting plate, namely the area of 20mm multiplied by 40mm) as shown in figure 5, fully covering the occupied area of a sample to be detected on the counting plate by a display area of the complete image, and respectively calculating the absolute value of the total number of cells (the sum of the total number of live cells and the total number of dead cells) in the image by an absolute value counting module in a controller Calculating to obtain the absolute value of the total number of cells in the cell suspension, and obtaining the total cell concentration value by inputting the values of k, f and V into the control panel of the full-automatic fluorescence cell analyzer by an absolute value concentration counting module in the controller through a mathematical formula Q-n-k-f/V (wherein Q represents the absolute value concentration of the cells, n represents the total number of cells in all images, k represents the dilution multiple of the anticoagulated whole blood by the physiological saline, f represents the dilution multiple of the diluted cell suspension by the fluorescent dye, and V represents the volume of the anticoagulated whole blood before dilution by the physiological saline).
Example 2
Before the first step, that is, before the cell suspension is taken by using the pipette, the cell suspension is prepared, and the cell suspension is prepared by directly using the cell mixture, so that in example 2, the cell mixture uses filtered white blood (with low white blood cell content), and does not need to be diluted by using a cell diluent, the cell suspension (filtered white blood) and the fluorescent dye are uniformly mixed in equal amount, the mixed solution of the cell suspension and the fluorescent dye is added to the counting plate, and counting is started after standing for a preset time.
The specific implementation mode is that 15 mul of cell suspension and 15 mul of fluorescent dye are mixed and stirred uniformly in equal quantity, 15 mul of mixed liquid is taken by a pipette and dripped to the counting plate (as shown in figure 1 and figure 2), the counting plate is kept still for 1min and starts to count, an image processing system (a camera with fluorescent signal capture) of the full-automatic fluorescence cell analyzer is used for shooting the mixed liquid in the counting plate for obtaining cell images, preferably, the mixed liquid in the area of 20mm x 40mm (which is slightly larger than the surface area of a groove for containing the cell suspension in the counting plate and adopts a rectangular groove) is used for shooting images, as shown in figure 3 and figure 4, the number of the shot images is defined by image analysis software in the image processing system (which does not exceed the range of the number of the shot images defined by the image analysis software of the full-automatic fluorescence cell analyzer), all positions of the mixed liquid in the area of 20mm x 40mm are ensured to be shot, the method comprises the steps that omission is prevented, the image scanning system automatically scans and identifies the obtained images of the mixed liquid in the area region, as shown in fig. 5, a full-image large-image splicing function (image splicing software, namely image analysis software) arranged in the image processing system is utilized to automatically splice a plurality of images to form a complete image (the area of the spliced complete image is larger than the physical size of the counting plate, namely 20mm multiplied by 40mm), the display region of the complete image completely covers the occupied region of a sample to be detected on the counting plate, an absolute value counting module in a controller respectively calculates the absolute value of the total number of cells (the sum of the total number of living cells and the total number of dead cells) in the image to obtain the absolute value of the total number of cells in the cell suspension, an absolute value concentration counting module in the controller calculates the formula Q n k f/V through a mathematical calculation formula, and k f/V is input into a control panel of the full-automatic fluorescence cell analyzer, f and V can obtain the concentration value of total cells, the concentration value of live cells and the concentration value of dead cells (wherein Q represents the absolute value concentration of cells, n represents the total number of cells in all images, k represents the dilution multiple of the filtered white blood component by the physiological saline, f represents the dilution multiple of the diluted cell suspension by the fluorescent dye, and V represents the volume of the anticoagulated whole blood or the filtered white blood component before being diluted by the physiological saline, and since the cell mixed solution selects the filtered white blood component and does not need to be diluted by the physiological saline, the k value and the f value in the mathematical calculation formula of the absolute value concentration calculation module in the controller are both defaulted to 1).
For further clear description of the technical solution of the present invention, the working principle of the counting method of the present invention is described here, as shown in fig. 1-5, anticoagulant whole blood is selected to be diluted 5 times with normal saline or white blood is directly used to configure cell suspension to be measured, a pipettor is used to accurately stain the cell suspension and fluorescent dye according to the requirements of a full-automatic fluorescent cell analyzer, the cell suspension and fluorescent dye are mixed evenly, cell particles are suspended uniformly as much as possible, sampling errors are reduced, the pipettor is used to accurately suck the cell suspension after fluorescent staining and add the cell suspension into a disposable cell counting plate, the disposable cell counting plate can select different specifications according to the actual requirements of users, as shown in fig. 1, the counting plate is provided with two grooves for accommodating cell suspension and can be used for repeatedly counting the same cell suspension for many times, as shown in fig. 2, the counting plate with a groove for containing cell suspension is provided, the counting plate in fig. 1 and the counting plate in fig. 2 have the same volume, and are convenient to be adapted to the full-automatic fluorescent cell analyzer, and the disposable cell counting plate is inserted into the card slot of the full-automatic fluorescent cell analyzer for placing the disposable cell counting plate, and by clicking or touching the "blood cell" button (i.e. the upper right region in fig. 3) of the control interface or the operation interface of the controller of the full-automatic fluorescent cell analyzer, the counting plate enters the absolute value concentration counting operation interface (i.e. the lower right region in fig. 3), as shown in fig. 4, by inputting information such as blood type, blood cell, blood bag bar code, blood bag number, blood volume, sample dilution factor, and figure number (figure number is set to be greater than the physical size of the disposable counting plate by default value) into the interface, clicking a 'confirmation start' button, automatically finishing image shooting and image scanning by the full-automatic fluorescent cell analyzer, counting the absolute value concentration of the total number of cells (the sum of the total number of living cells and the total number of dead cells) in a unit volume in a scanning set area (preferably, 20mm multiplied by 40mm), displaying a counting result, storing an original picture and a marked picture of the cells, backtracking, finishing counting once, only needing to replace a disposable cell counting plate, repeating the operation, finishing counting of a plurality of times of micro cell suspensions, and so on, reducing the error of the counting result and improving the precision of the cell number counting result.
In the description of the present invention, numerous specific details are set forth. It is understood, however, that embodiments of the invention may be practiced without these specific details. In some instances, well-known methods, structures and techniques have not been shown in detail in order not to obscure an understanding of this description. Similarly, it should be appreciated that in the foregoing description of exemplary embodiments of the invention, various features of the invention are sometimes grouped together in a single embodiment, figure, or description thereof for the purpose of streamlining the disclosure and aiding in the understanding of one or more of the various inventive aspects. However, the disclosed method should not be interpreted as reflecting an intention that: that the invention as claimed requires more features than are expressly recited in each claim. Rather, as the following claims reflect, inventive aspects lie in less than all features of a single foregoing disclosed embodiment. Thus, the claims following the detailed description are hereby expressly incorporated into this detailed description, with each claim standing on its own as a separate embodiment of this invention.
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; such modifications and substitutions do not depart from the spirit and scope of the present invention, and they should be construed as being included in the following claims and description.
Claims (8)
1. A method for obtaining absolute values of the number of micro cells in a cell suspension, which is characterized in that the counting method adopts a fully automatic fluorescent cell analyzer, the analyzer comprises a counting plate for containing the micro cell suspension, a container for storing fluorescent dye, an image scanning system, an image processing system and a controller for counting the number of the cells, and the method comprises the following specific operation steps:
the method comprises the following steps: withdrawing the cell suspension with a pipettor and measuring the volume of the cell suspension;
step two: adding a fluorescent dye into the cell suspension obtained in the step one to dye and uniformly mixing;
step three: adding the cell suspension dyed in the step two into the counting plate, and inserting the counting plate into the full-automatic fluorescent cell analyzer;
step four: the image processing system carries out image shooting on the cell suspension dyed in the third step, all images of the cell suspension on the counting plate are obtained according to the number of the images taken set by the image processing system, and the display areas of all the obtained images completely cover the occupied area of the sample to be detected on the counting plate;
step five: the image scanning system scans all the images acquired in the fourth step, and is used for acquiring the total number of cells, the total number of live cells and the total number of dead cells of the trace amount of cells in the cell suspension in all the images in the fourth step, wherein the total number of the cells is the sum of the total number of the live cells and the total number of the dead cells;
step six: and the controller calculates the absolute value of the total number of the cells of the trace cells in the cell suspension through an absolute value counting module in the controller according to the total number of the cells obtained in the step five.
2. The method according to claim 1, wherein before the step one, the cell suspension is obtained by diluting and mixing the cell mixture to be counted with a cell diluent.
3. The method according to claim 2, wherein the diluted cell suspension and the fluorescent dye are uniformly mixed, the uniformly mixed solution of the cell suspension and the fluorescent dye is added to the counting plate, and the volume of the diluted cell suspension is the same as that of the fluorescent dye and the mixed solution.
4. The method according to claim 3, wherein the cell mixture is selected from the group consisting of anticoagulated whole blood, filtered white blood, and other cell culture media containing a trace amount of cells.
5. The method according to claim 1, wherein a full-image map stitching function module is disposed in the image processing system, and is configured to stitch all cell images of the cell suspension acquired by the image processing system into a full image of the cell suspension, and a display area of the full image fully covers an occupied area of the sample to be measured on the counting plate.
6. The method according to claim 1, wherein the absolute value calculating module comprises an absolute value concentration calculating module, and the absolute value concentration calculating module calculates the absolute value concentration of the cell suspension by a mathematical calculation formula, wherein,
the mathematical formula is Q ═ n/V, where Q represents the absolute concentration of cells, n represents the total number of cells in all the images, and V represents the volume of the cell suspension;
if Q is the absolute value concentration of the total number of cells, n is the total number of cells in all the images;
if Q is the absolute value concentration of the total number of the living cells, n is the total number of the living cells in all the images;
if Q is the absolute value concentration of the total number of dead cells, n is the total number of dead cells in all the images;
the total number of cells absolute value concentration is the sum of the total number of living cells absolute value concentration and the total number of dead cells absolute value concentration.
7. The method according to claim 6, wherein the cell suspension is obtained by diluting the cell mixture to be counted with a cell diluent or directly consists of the cell mixture to be counted, the cell suspension is mixed and diluted with the fluorescent dye and then added to the counting plate, and the counting is started after standing for a preset time, wherein the mathematical formula is Q n k f/V, wherein Q represents the number of cells, n represents the total number of cells in all images, k represents the dilution multiple of the cell mixture to be counted with the cell diluent, f represents the dilution multiple of the diluted cell suspension with the fluorescent dye, and V represents the volume of the cell mixture to be counted before being released by the cell diluent;
if Q is the absolute value concentration of the total number of cells, n is the total number of cells in all the images;
if Q is the absolute value concentration of the total number of the living cells, n is the total number of the living cells in all the images;
if Q is the absolute value concentration of the total number of dead cells, n is the total number of dead cells in all the images;
the total number of cells absolute value concentration is the sum of the total number of living cells absolute value concentration and the total number of dead cells absolute value concentration.
8. The method of claim 1, wherein the absolute value of the count is equal to the sum of the dead cells and the live cells.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010431054.9A CN113781378A (en) | 2020-05-20 | 2020-05-20 | Absolute value counting method for obtaining quantity of micro cells in cell suspension |
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| CN116769871A (en) * | 2023-08-15 | 2023-09-19 | 深圳市卫光生物制品股份有限公司 | Method for counting chick embryo cells |
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