JP2012024061A - Cell dispersion apparatus - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a cell dispersion apparatus for improving cell dispersion efficiency of aggregating cells.SOLUTION: The cell dispersion part 52 (cell dispersion apparatus) disperses the aggregating cells, and includes: a bottom part 62 having a pore part 63 penetrating from an upper surface to a lower surface; a rotor 70 arranged with a predetermined space from the upper surface or lower surface of the bottom part 62, and having a spiral groove 71 at the outer periphery; a motor 80 for rotary-driving the rotor 70; and a pipe 90 arranged outside of the rotor 70. The rotor 70 is rotated by the motor 80 so that the cell aggregated between the bottom part 62 and the rotor 70 is supplied, dispersing the supplied aggregating cells.

Description

  The present invention relates to a cell dispersing apparatus, and more particularly to a cell dispersing apparatus that disperses a plurality of aggregated cells into a single cell.

  When analyzing cells, it is necessary to disperse the aggregated cells. Therefore, a cell dispersion device that disperses aggregated cells is known (see, for example, Patent Document 1).

  Patent Document 1 discloses an apparatus including a cylindrical hollow body with a filter provided at the tip in a solution containing a cell sample. In Patent Document 1, the device is immersed in a solution containing a cell specimen, and the solution containing cells is stirred by rotating a cylindrical hollow body. As a result, a shear force is generated in the cell group to disperse the cells. Next, the air inside the hollow body is sucked in a state where the hollow body is immersed in the solution. A hollow body filter is formed with a plurality of pores (openings) of a size that cannot pass a single cell to be measured but can pass red blood cells or cell pieces having a size smaller than that. For this reason, by suction, a solution containing red blood cells and cell pieces passes through the filter and is sucked into the hollow body, and cells that cannot pass through the filter are adsorbed on the filter surface. Then, after discharging the solution sucked into the hollow body, the filter surface on which the cells are adsorbed is brought into contact with the slide glass, whereby the cells adsorbed on the filter surface are distributed on the slide glass.

Japanese Patent Laid-Open No. 4-232834

  However, in Patent Document 1, since the sample is only stirred by the rotation of the cylindrical hollow body in the sample (solution containing aggregated cells), and shearing force is applied to the aggregated cells by this stirring, There was a problem that cell dispersion efficiency was low.

  The present invention has been made to solve the above-described problems, and one object of the present invention is to provide a cell dispersion apparatus capable of improving the cell dispersion efficiency of aggregated cells. .

Means for Solving the Problems and Effects of the Invention

  In order to achieve the above object, a cell dispersion device according to one aspect of the present invention is a cell dispersion device for dispersing aggregated cells, and includes a first member having a hole penetrating from an upper surface to a lower surface, and a first member. A second member having a spiral groove on the outer periphery thereof, a drive unit that rotationally drives the second member, and a cylinder disposed on the outer side of the second member. And by supplying the aggregated cells between the first member and the second member by rotating the second member by the drive unit, and dispersing the supplied aggregated cells. Yes.

  In the cell dispersion device according to one aspect of the present invention, as described above, the second member is disposed at a predetermined interval from the upper surface or the lower surface of the first member, and the second member having a spiral groove on the outer periphery thereof is provided. By rotating the second member by the driving unit, it is possible to form a flow for moving the sample containing aggregated cells to the first member side by the rotation of the spiral groove of the second member. Thereby, since the supply of the aggregated cells between the first member and the second member can be promoted, it is possible to effectively disperse the aggregated cells between the first member and the second member. it can. Furthermore, since the sample supplied between the first member and the second member is passed through the hole of the first member, the flow of the sample formed by the rotation of the spiral groove is not hindered. Aggregated cells can be supplied one after another between the first member and the second member. As a result, the cell dispersion efficiency of aggregated cells can be improved.

  In the cell dispersion device according to the above aspect, the hole of the first member preferably has a size through which the aggregated cells can pass. If comprised in this way, since the hole part of a 1st member is not obstruct | occluded by agglomerated cells (a hole part is clogged), the sample supplied between the 1st member and the 2nd member is used as the 1st member. It is possible to prevent the flow of the sample formed by the rotation of the spiral groove from being obstructed when passing through the hole. For this reason, since it is possible to effectively supply the aggregated cells between the first member and the second member, it is possible to promote the dispersion of the aggregated cells.

  In the cell dispersion device according to the above aspect, the cylinder preferably has an opening at the tip and side thereof, and one of the opening at the tip and the opening at the side of the cylinder serves as an inlet, and the other is used as the inlet. When the outlet is used, a circulation flow is formed in which cells circulate from the inlet to the inside of the cylinder, the outlet, the outside of the cylinder, and the inlet. If comprised in this way, the flow of the sample formed by rotation of a helical groove | channel can be circulated. Thereby, the supply of the aggregated cells between the first member and the second member can be more effectively performed.

  In this case, preferably, the first member is arranged at the opening at the tip of the cylinder, the opening at the side of the cylinder is used as the inlet, and the hole of the first member arranged at the opening at the tip is the outlet. As a circulation flow through which cells circulate is formed. If comprised in this way, after gathering the aggregated cells to the 1st member side of a front-end | tip by rotation of the helical groove | channel of a 2nd member inside a cylinder, it can be made to flow out from the hole of a 1st member. As a result, the aggregated cells can be reliably supplied between the first member and the second member without hindering the formed circulation flow.

  In the cell dispersion device according to the above aspect, the first member preferably includes a shearing force applying unit for applying a shearing force to the aggregated cells as the second member rotates. If comprised in this way, since a shearing force can be provided to an aggregating cell with rotation of a 2nd member by a shearing force provision part, between a 1st member and a 2nd member by rotation of a 2nd member. Both the supply of the aggregated cells to the cell and the dispersion of the aggregated cells by the shearing force can be performed.

  In this case, preferably, the shearing force application part includes a convex part protruding toward the second member side. If comprised in this way, since the space | interval between a 1st member and a 2nd member becomes small in the area | region between a convex part and a 2nd member, when rotating a 2nd member, this convex part and A shearing force can be applied to the aggregated cells in the region between the second member. Thereby, a shear force can be made to act on an aggregate cell and an aggregate cell can be disperse | distributed easily.

  In the cell dispersion device according to the above aspect, preferably, the hole of the first member has an elongated shape. According to this structure, when the sample flow along the rotation direction is formed by rotating the second member having the spiral groove, the rotation direction extends over a wide range on the long side of the elongated hole. Therefore, the sample can easily pass through the hole of the first member. As a result, the sample containing the dispersed cells can be efficiently discharged from the first member, and aggregated cells can be supplied one after another between the first member and the second member, so the first member and the second member It is possible to promote the supply of aggregated cells to and from.

  In this case, preferably, the hole portion extends in a direction in which the longitudinal direction intersects the rotation direction of the second member. If comprised in this way, since the flow of the rotation direction formed by rotating the 2nd member which has a helical groove | channel and the longitudinal direction of a hole cross | intersect, the flow of a sample is more efficiently carried out to a hole. Can be reached. As a result, the sample containing dispersed cells can be more efficiently discharged from the first member, so that the supply of aggregated cells between the first member and the second member can be further promoted.

  In the cell dispersion device according to the above aspect, the first member preferably has a recess provided on the surface on the second member side, and the hole is disposed in the recess. If comprised in this way, after the sample containing the aggregated cells supplied from the 2nd member side is hold | maintained so that it may accumulate temporarily in the recessed part of a 1st member, a sample flows out from the hole arrange | positioned in a recessed part Can be made. Accordingly, the aggregated cells can be more effectively dispersed between the first member and the second member by temporarily accumulating in the recess, and the sample is allowed to flow out from the hole disposed in the recess. be able to.

  In this case, preferably, the first member is circular, extends in a direction intersecting with the rotation direction of the second member, and further has a convex portion protruding toward the second member, and the concave portion is formed on the second member. A plurality of protrusions extending in the direction intersecting the rotation direction are provided as boundaries. If comprised in this way, the sample containing the aggregation cell temporarily stored in the recessed part will flow through the inside of the recessed part in the same direction as the rotation direction of the second member, and extend in a direction crossing the rotation direction of the second member. Reach the convex part. Since the distance between the first member (convex part) and the second member becomes small when this flow tries to get over the convex part, it is possible to easily apply a shearing force to the aggregated cells in the vicinity of the convex part. As a result, the aggregated cells can be dispersed more effectively.

  In this case, preferably, the cell is aggregated between the convex portion of the first member and the second member by rotating the second member by the driving unit. If comprised in this way, since the area | region between the convex part and 2nd member to which the space | interval of a 1st member and a 2nd member becomes small can be made easy to make a shear force act on an aggregated cell. In addition, it is possible to effectively disperse the aggregated cells.

  In the configuration in which the first member has a convex portion extending in a direction intersecting with the rotation direction of the second member, preferably, the hole portion of the first member is a first portion by a drive unit in a plurality of concave portions having the convex portion as a boundary. It arrange | positions at the edge part of the rotation direction side of 2 members, respectively. If comprised in this way, the sample containing the aggregation cell temporarily stored in the recessed part will flow through the inside of the recessed part in the same direction as the rotation direction of the 2nd member, and will reach the convex part of a boundary. Since the hole is formed in this boundary part (end of the second member in the rotation direction), the flow of the sample is divided into a flow that tries to get over the convex part and a flow that flows out of the hole. Is done. As a result, the aggregated cells are dispersed between the convex part and the second member by the flow that tries to get over the convex part, and the aggregated cells are successively collected without flowing into the concave part due to the flow flowing out from the hole part. Is supplied. As a result, it is possible to effectively supply aggregated cells by smoothing the flow of the sample while effectively dispersing aggregated cells.

  In the cell dispersing apparatus according to the above aspect, preferably, the predetermined distance between the upper surface or the lower surface of the first member and the second member is such that the aggregated cells cannot pass and the dispersed single cells pass. It is a possible interval. If comprised in this way, between the 1st member and the 2nd member, while giving a shearing force to the aggregation cell and disperse | distributing a single cell, without giving a shearing force to a single cell, the 1st member and the 1st It can pass between two members. Thereby, the dispersion | distribution efficiency of an aggregated cell can be improved, without damaging a single cell.

It is the perspective view which showed the whole structure of the cell analyzer provided with the cell dispersion | distribution part by one Embodiment of this invention. It is the block diagram which showed the structure of the measuring apparatus of the cell analyzer shown in FIG. It is the block diagram which showed the structure of the sample preparation apparatus of the cell analyzer shown in FIG. It is the block diagram which showed the structure of the data processing apparatus of the cell analyzer shown in FIG. It is the schematic diagram which showed the flow cytometer which comprises the main detection part of the measuring apparatus shown in FIG. It is the schematic diagram which showed the optical system of the flow cytometer shown in FIG. It is the schematic diagram which showed the structure of the preparation device part of the sample preparation apparatus shown in FIG. It is the perspective view which showed the whole structure of the cell dispersion | distribution part by one Embodiment of this invention. It is a longitudinal cross-sectional view of the cell dispersion | distribution part by one Embodiment of this invention shown in FIG. It is the perspective view which showed the perforated member of the cell dispersion part by one Embodiment of this invention shown in FIG. It is a top view of the perforated member shown in FIG. It is sectional drawing of the perforated member along the 500-500 line | wire of FIG. FIG. 9 is a side view showing a rotor of the cell dispersion unit according to the embodiment of the present invention shown in FIG. 8. FIG. 14 is a bottom view of the rotor illustrated in FIG. 13. It is a schematic diagram for demonstrating operation | movement of the cell dispersion | distribution part by one Embodiment of this invention shown in FIG. It is a flowchart for demonstrating the analysis operation | movement of the cell analyzer provided with the cell dispersion | distribution part by one Embodiment of this invention. It is a flowchart for demonstrating the analysis operation | movement of the cell analyzer provided with the cell dispersion | distribution part by one Embodiment of this invention. It is a perspective view which shows the bottom part by Example 2 of this invention.

  DESCRIPTION OF THE PREFERRED EMBODIMENTS Embodiments embodying the present invention will be described below with reference to the drawings.

  First, with reference to FIGS. 1-15, the structure of the cell analyzer 1 provided with the cell dispersion | distribution part 52 (refer FIG. 8) by one Embodiment of this invention is demonstrated. In the present embodiment, an example in which the present invention is applied to a cell dispersion unit of a cell analyzer will be described.

  In the cell analyzer 1, a measurement sample containing cells collected from a patient is caused to flow through the flow cell, and the measurement sample flowing through the flow cell is irradiated with laser light. Then, by detecting light (forward scattered light, side fluorescence, etc.) from the measurement sample and analyzing the optical signal, it is determined whether or not the cell contains cancer cells. More specifically, the cell analyzer 1 uses cervical epithelial cells as analysis targets, and is used for screening cervical cancer.

  As shown in FIG. 1, the cell analyzer 1 includes a measuring device 2 that performs optical measurement with a laser beam on a measurement sample, and a pretreatment such as washing and staining of a biological sample collected from a subject. And a sample preparation device 3 for producing a measurement sample to be supplied to the measurement device 2 and a data processing device 4 for analyzing a measurement result of the measurement device 2 and the like.

  As shown in FIG. 2, the measurement apparatus 2 includes a main detection unit 21, a signal processing unit 22, a measurement control unit 23, and an I / O interface 24. The main detection unit 21 has a function of detecting the measurement target cell and the number and size of the nucleus from the measurement sample. In this embodiment, the flow cytometer 10 shown in FIGS. 5 and 6 is employed for the main detection unit 21.

  The signal processing unit 22 includes a signal processing circuit that performs necessary signal processing on the output signal from the main detection unit 21. The measurement control unit 23 includes a microprocessor 25 and a storage unit 26. The storage unit 26 includes a ROM for storing a control program such as the main detection unit 21 and data, and a RAM.

  The microprocessor 25 of the measurement control unit 23 is connected to the data processing device 4 and a microprocessor 36 (see FIG. 3) of the preparation control unit 33 (see FIG. 3) described later via the I / O interface 24. . Thereby, various data can be transmitted and received between the data processing device 4 and the microprocessor 36 of the preparation control unit 33.

  As shown in FIG. 3, the sample preparation device 3 automatically performs component adjustment on the biological sample by the sub-detection unit 31, the signal processing unit 32, the preparation control unit 33, the I / O interface 34, and the biological sample. And a preparation device unit 35.

  The sub-detecting unit 31 has a function of detecting the number of cells to be measured included in the biological sample. In the present embodiment, a flow cytometer 10 that is substantially the same as that shown in FIGS. The signal processing unit 32 includes a signal processing circuit that performs necessary signal processing on the output signal from the sub-detection unit 31. The preparation control unit 33 includes a microprocessor 36, a storage unit 37, a sensor driver 38, and a drive unit driver 39. The storage unit 37 includes a ROM that stores a control program for controlling the sub-detection unit 31, the preparation device unit 35, and the like, a RAM, and the like.

  The preparation device unit 35 includes a sample setting unit 51, a cell dispersion unit 52 as a cell dispersion device, a sample pipette unit 53, a sample quantification unit 54, a reagent quantification unit 55, and a discrimination / substitution unit 56. ing.

  The specimen setting unit 51 is for setting a plurality of biological containers 6 (see FIG. 7) and product containers 7 (see FIG. 7) that contain biological samples. Further, the cell dispersion unit 52 has a function of forcibly dispersing aggregated cells contained in the biological sample in the biological container 6 to form a single cell.

  The specimen pipette unit 53 takes out a biological sample in which cells are dispersed from the biological container 6 (see FIG. 7) and introduces the biological sample into the fluid circuit of the preparation device unit 35, or supplies the prepared product to the product container 7 ( It has a function of returning to the product container 7 or returning to the product container 7 (see FIG. 7). The specimen quantification unit 54 has a function of quantifying a biological sample supplied to the fluid circuit.

  The reagent quantification unit 55 has a function of quantifying a reagent such as a staining solution added to the biological sample. The discrimination / substitution unit 56 has a function of mixing and substituting the biological sample and the diluent, and discriminating the measurement target cell from other cells (red blood cells, white blood cells, etc.) and bacteria. The configuration of a fluid circuit that connects each part of the preparation device unit 35 (sample setting unit 51, cell dispersion unit 52, sample pipette unit 53, sample quantification unit 54, reagent quantification unit 55, and discrimination / substitution unit 56) (FIG. 7). Will be described later.

  The microprocessor 36 of the preparation control unit 33 is connected to the microprocessor 25 (see FIG. 2) of the measurement control unit 23 (see FIG. 2) via the I / O interface 34. Thereby, various data can be transmitted / received to / from the microprocessor 25 of the measurement control unit 23.

  In addition, the microprocessor 36 of the preparation control unit 33 is connected to each unit of the preparation device unit 35 (sample setting unit 51, cell dispersion unit 52, sample pipette unit 53, sample quantification unit 54) via the sensor driver 38 or the drive unit driver 39. The reagent quantification unit 55 and the discrimination / substitution unit 56) are connected to sensors and a drive motor constituting the drive unit. Thereby, the microprocessor 36 executes the control program based on the detection signal from the sensor and controls the operation of the drive unit.

  As shown in FIG. 4, the data processing device 4 in the present embodiment includes a personal computer such as a notebook PC (may be a desktop type), and mainly includes a processing body 41, a display unit 42, and an input unit 43. It is configured.

  The processing body 41 includes a CPU 41a, a ROM 41b, a RAM 41c, a hard disk 41d, a reading device 41e, an image output interface 41f, and an input / output interface 41g, and these units are communicably connected via an internal bus. Has been.

  In addition to various programs such as an operating system and application programs, the hard disk 41d transmits operation commands to the measurement control unit 23 (see FIG. 2) and the preparation control unit 33 (see FIG. 3), and the measurement apparatus 2 (see FIG. 1). The operation program 44 is installed for receiving and analyzing the measurement results performed in step 1 and displaying the processed analysis results. The operation program 44 operates on the operating system.

  An input unit 43 including a keyboard and a mouse is connected to the input / output interface 41g. The input / output interface 41g is also connected to the I / O interface 24 (see FIG. 2) of the measuring apparatus 2 (see FIG. 2). Thereby, it is possible to transmit and receive data between the measuring device 2 and the data processing device 4.

  Next, the flow cytometer 10 constituting the main detection unit 21 of the measuring device 2 will be described. As shown in FIG. 5, the lens system 11 of the flow cytometer 10 has a function of condensing laser light from a semiconductor laser 12 as a light source onto a measurement sample flowing through the flow cell 13. The condensing lens 14 has a function of condensing forward scattered light of the cells in the measurement sample onto a scattered light detector composed of the photodiode 15.

  Specifically, as shown in FIG. 6, the lens system 11 includes, in order from the semiconductor laser 12 side (left side in FIG. 6), a collimator lens 11a, a cylinder lens system (plano-convex cylinder lens 11b + biconcave cylinder lens 11c), and It is composed of a condenser lens system (condenser lens 11d + condenser lens 11e).

  As shown in FIG. 5, the lateral condenser lens 16 has a function of concentrating the side scattered light and the side fluorescence of the measurement target cell or the nucleus in the cell on the dichroic mirror 17. The dichroic mirror 17 is configured to reflect side scattered light to the photomultiplier 18 and transmit side fluorescence to the photomultiplier 19. These lights reflect the characteristics of cells and nuclei in the measurement sample.

  The photodiode 15 and the photomultipliers 18 and 19 convert the received optical signal into an electrical signal, respectively, and forward scattered light signal (FSC), side scattered light signal (SSC), and side fluorescent signal ( SFL) is output. These output signals are amplified by a preamplifier (not shown) and sent to the signal processing unit 22 (see FIG. 2) of the measuring apparatus 2. Each signal FSC, SSC, SFL processed by the signal processing unit 22 of the measuring device 2 is transmitted from the I / O interface 24 to the data processing device 4 by the microprocessor 25.

  The CPU 41a of the data processing device 4 creates a scattergram for analyzing cells and nuclei from each signal FSC, SSC, SFL by executing the operation program 44, and based on this scattergram, It is determined whether or not the cell is abnormal, specifically, whether or not it is a cancerous cell.

  In addition, since the sub-detecting unit 31 of the sample preparation device 3 employs the flow cytometer 10 having substantially the same configuration as the main detecting unit 21 as described above, detailed description thereof is omitted. Since the sub-detection unit 31 preliminarily measures the concentration of the measurement target cell before the main measurement by the measuring device 2, the sub-detection unit 31 outputs a signal for counting the number of cells. It is sufficient if it is possible (if it is possible to acquire a forward scattered light signal (FSC)).

  Next, the preparation device unit 35 will be described in detail. As shown in FIG. 7, the sample setting unit 51 includes a circular rotary table 51a and a drive unit 51b that rotationally drives the rotary table 51a. At the outer peripheral edge of the turntable 51a, there is provided a holding part capable of setting a biological container 6 for storing a biological sample and a product container (microtube) 7 for storing a product after the biological sample is prepared. ing.

  The cell dispersion unit 52 includes a perforated member 60, a rotor 70, and a motor 80 that rotationally drives the rotor 70. A pipe 90 is provided outside the rotor 70, and a perforated member 60 is attached to the opening at the tip of the pipe 90. By inserting these into the biological container 6 and rotating the rotor 70, the aggregated cells contained in the biological sample in the biological container 6 are dispersed into single cells. Details including a specific configuration of the cell dispersion unit 52 will be described later.

  The sample pipette unit 53 sucks the biological sample in the biological container 6 or the measurement sample in the product container 7 and supplies the sample to the sample quantitative unit 54 and the preparation device unit 35. A second pipette 53b for returning the product after the predetermined treatment to the product container 7;

  The second pipette 53 b is connected to the container 56 a of the discrimination / replacement unit 56 by a pipe line, and the liquid from which the measurement target cell is discriminated by the discrimination / replacement unit 56 can be returned to the product container 7. The sample quantification unit 54 includes a quantification cylinder 54a and a drive unit 54b that moves the quantification piston inserted through the quantification cylinder 54a up and down.

  The biological sample sucked from the biological container 6 by the first pipette 53a is introduced into the metering cylinder 54a through the valve V1. The introduced biological sample is sent to the discrimination / substitution unit 56 through the valves V1, V7, and V4 by the movement of the quantitative piston by the drive unit 54b. The quantitative cylinder 54a of the sample quantitative unit 54 is also connected to a diluent unit 57 for preparing a diluent for the biological sample via a pipe.

  The reagent quantification unit 55 includes a pair of quantification cylinders 55a and 55b, and a drive unit 55c that vertically moves a quantification piston inserted into the quantification cylinders 55a and 55b. The reagent in the reagent container is supplied to each quantitative cylinder 55a, 55b. The supplied reagent is quantified by a predetermined amount by the movement of the quantification piston by the drive unit 55c, and the quantified reagent is sent to the second pipette 53b through the valves V2 and V3.

  Therefore, a plurality of types of predetermined amounts of reagents determined by the reagent quantitative unit 55 can be mixed with the discriminated sample returned to the product container 7 of the sample setting unit 51. The reagent weighed with the quantitative cylinder 55a and added to the biological sample is a dye solution for PI staining, and the reagent weighed with the quantitative cylinder 55b and added to the biological sample is an RNase for performing RNA treatment on the cells. It is.

  The discriminating / replacement unit 56 includes an upper opening-shaped storage container 56a, a filtration cylinder 56b provided in the storage container 56a so as to be vertically movable, and a drive unit 56c that moves the filtration cylinder 56b up and down in the storage container 56a. It has.

  The storage container 56a can hold the biological sample quantified by the sample determination unit 54 via the valves V1, V7, and V4 inside the storage container 56a. The filtration cylinder 56b is formed of a hollow cylinder having a filter 56d at the lower portion that does not allow measurement target cells (epithelial cells) to pass therethrough and allows cells having smaller diameters (red blood cells, white blood cells, etc.) to pass therethrough. In addition, the diluting liquid of the diluting liquid unit 57 is supplied to the filtration cylinder 56b through the valve V5.

  The drive unit 56c has a function of moving the filtration cylinder 56b downward with respect to the storage container 56a containing the mixed liquid of the biological sample and the diluent. As a result, the liquid containing only the measurement target cell remains as a residual liquid below the filter 56d, and the liquid containing other cells and impurities passes through the filter 56d and passes above the filter 56d as the filtrate (inside the filtration cylinder 56b). ). As a result, the liquid containing only the measurement target cells remaining as the residual liquid is in a concentrated state.

  The filtrate filtered by the lowering of the filtration cylinder 56b is discarded to the disposal unit 58 through the valve V6. On the other hand, the remaining liquid in the storage container 56a is sent to the flow cell 13 of the sub-detector 31 through the valve V4, and the number of cells is counted. The liquid that has passed through the flow cell 13 is discarded in the discard unit 58. Further, the remaining liquid in the storage container 56 a is sent to the second pipette 53 b of the sample pipette unit 53 and then returned to the product container 7.

  As shown in FIG. 7, the quantification cylinder 54a of the sample quantification unit 54 is also connected to the main detection unit 21 of the measurement apparatus 2 via valves V1 and V7. The measurement sample in the product container 7 is quantified by the sample quantification unit 54 via the first pipette 53a and supplied to the main detection unit 21 of the measurement device 2 through the valves V1 and V7. Yes.

  Next, a specific configuration of the cell dispersion unit 52 according to the present embodiment will be described. As shown in FIGS. 8 and 9, the cell dispersion unit 52 includes a perforated member 60, a rotor 70, and a motor 80 that rotationally drives the rotor 70. In addition, a pipe 90 having a function of guiding the flow of the biological sample is disposed outside the rotor 70.

  As shown in FIGS. 10 to 12, the perforated member 60 is made of stainless steel in consideration of chemical resistance, strength, and the like, so as to block the cylindrical tube portion 61 and the lower end of the tube portion 61. A bottom portion 62 provided with four holes 63 is integrally formed. As shown in FIG. 15, a screw portion 61 a is formed on the inner peripheral surface of the cylindrical portion 61, and is configured to be screwed and fitted into a screw portion 91 formed on the outer peripheral surface at the lower end of the pipe 90. Thus, the perforated member 60 is attached to the opening at the tip of the pipe 90. As shown in FIGS. 11 and 12, the bottom 62 is formed in a circular shape when seen in a plan view, and has four holes 63 penetrating from the upper surface (inner surface) to the lower surface (outer surface). These pores 63 have a size through which aggregated cells (size of about 100 μm or more and about 500 μm or less) can pass.

  On the upper surface of the bottom portion 62, there are four convex portions 64 that extend in a direction (radial direction) orthogonal to the rotation direction (arrow R direction) of the rotor 70 in a plan view and project to the rotor 70 side (upward). Is formed. That is, the four convex portions 64 are formed in a cross shape extending in the radial direction from the center O of the bottom portion 62. In addition, four liquid retaining portions 65 are formed on the upper surface of the bottom portion 62. The four liquid retaining portions 65 are concave portions having the convex portions 64 as boundaries. That is, the circular bottom 62 has a fan-shaped liquid retaining portion (recessed portion) 65 having a substantially quarter circle divided by four cross-shaped convex portions 64 as viewed in a plan view. As shown in FIG. 12, the thickness of the bottom 62 (convex portion 64) is t1 (about 1 mm), and the liquid retaining portion 65 has a depth of t2 (about 0.5 mm).

  As shown in FIG. 11, the four hole parts 63 of the bottom part 62 are respectively arranged in the four liquid retaining parts 65. The hole 63 has an elongated shape having a width W1 (about 0.5 mm) and a length L1 (about 3.25 mm). The hole 63 is formed to extend in a direction orthogonal to the arrow R direction (rotation direction of the rotor 70). More specifically, the hole 63 is adjacent to the protrusion 64 at the end of the rotor 70 in the rotation direction (arrow R direction) side and in parallel with the protrusion 64 in the fan-shaped liquid retaining part 65. It is arranged to extend. The hole 63 is formed to extend from the radially inner end of the fan-shaped liquid retaining portion 65 to the radially outer end.

  As shown in FIG. 9, the rotor 70 is made of stainless steel in consideration of chemical resistance and strength, like the bottom portion 62, and is arranged inside the pipe 90. As shown in FIGS. 13 and 14, the rotor 70 has a spiral groove 71 formed on the outer periphery. The groove 71 is formed with a lead angle α (about 40 degrees). Further, when the rotor 70 rotates in the direction of arrow R (see FIG. 11), the groove 71 sends the biological sample in the downward direction (in the direction of arrow Z2) where the perforated member 60 is arranged by the rotation of the groove 71. Is provided. As a result, when the rotor 70 is rotated around the axis (in the direction of arrow R) by the motor 80, the biological sample can be supplied toward the lower bottom 62 (providing a downward driving force to the biological sample). It is configured as follows. Further, the lower end surface 72 of the rotor 70 is formed flat.

  In the present embodiment, as shown in FIG. 15, the lower end surface 72 of the rotor 70 and the upper surface of the bottom portion 62 of the perforated member 60 attached to the lower end of the pipe 90 (the upper surface of the convex portion 64) are at a predetermined interval. It is configured to be spaced apart from CL1. The interval CL1 has such a size that aggregated cells (100 μm or more) cannot pass and single cells (average 60 μm) can pass. Therefore, the interval CL1 is set in the range of 30 to 80 μm so as to be approximately equal to the size (average 60 μm) of the cervical epithelial cells (single cells) to be measured in this embodiment. In the present embodiment, the distance CL1 is about 50 μm. In FIG. 15, the interval CL1 is exaggerated for the sake of explanation.

  Here, since the aggregated cells have a size exceeding 100 μm, it is possible to effectively apply shear force or dispersion force to the aggregated cells by setting the interval CL1 to about 50 μm. Thus, in this embodiment, the cell dispersion part 52 disperses the aggregated cells between the convex part 64 of the perforated member 60 and the lower end surface 72 of the rotor 70 by rotating the rotor 70 by the motor 80. It is configured as follows. The reason why the distance CL1 is set to 30 to 80 μm is that cells may be crushed if the distance is less than 30 μm, and the dispersion force applied to the aggregated cells may be reduced if the distance CL1 exceeds 80 μm. .

  As shown in FIG. 9, the upper end 73 of the rotor 70 has one end (upper end, an end in the arrow Z1 direction) connected to the output shaft 80a of the motor 80 at the other end (lower end, in the arrow Z2 direction). Attached to the edge). Thereby, the driving force (rotational force) of the motor 80 is transmitted to the rotor 70. A plate member 75 for preventing liquid splash is attached to a connecting portion between the upper end portion 73 of the rotor 70 and the rotating shaft 74. The plate member 75 prevents the biological sample from adhering to the rotating shaft 74 and entering the cylindrical body 82 that supports the rotating shaft 74.

  The motor 80 is attached to the upper member 81 a of the support member 81. The output shaft 80a of the motor 80 protrudes downward through a hole 81b formed in the upper member 81a. The lower member 81c of the support member 81 is also provided with a hole 81d corresponding to the hole 81b of the upper member 81a, and a cylinder 82 is attached so as to extend downward from the hole 81d. Inside the cylindrical body 82, a metal bearing 83 is provided at a position from below the upper end portion and the central portion, and a resin bearing member 84 is provided at the lower end portion. The rotation shaft 74 is rotatably supported by the bearing 83 and the bearing member 84. This resin bearing member 84 more reliably prevents the biological sample from entering the cylindrical body 82 during operation.

  The pipe 90 is made of a stainless steel circular pipe having an inner diameter that can accommodate the rotor 70 therein. The upper end of the pipe 90 is connected to the cylinder 82 in which the rotating shaft 74 is inserted. Further, a perforated member 60 (screw part 61a) is screwed and attached to a screw part 91 (see FIG. 15) provided at the lower end (tip side end part) of the pipe 90. The rotor 70 attached to the rotating shaft 74 is arranged so that the side and the lower side are surrounded by the pipe 90 and the perforated member 60. In addition, the outer periphery where the groove 71 of the rotor 70 is formed and the inner peripheral surface of the pipe 90 are slightly separated by a gap CL2 (about 0.3 mm).

  Two elongated openings 92 (see FIGS. 8 and 15) are formed on the side wall of the pipe 90 at a position slightly above the position where the rotor 70 is disposed. The two openings 92 are formed at positions facing each other with the core of the pipe 90 as the center. As shown in FIG. 15, the opening 92 introduces a biological sample outside the pipe 90 into the pipe 90 with the opening 92 as an inlet, and the rotor 70 moves down the pipe 90 from the inlet (arrow Z2). Direction). The biological sample flows out of the pipe 90 with the hole 63 of the perforated member 60 at the lower end as the outflow port, and flows into the opening 92 (inflow port) again. Thereby, in this embodiment, cells in the biological sample circulate from the inlet (opening 92) to the inside of the pipe 90, the outlet (hole 63), the outside of the pipe 90, and the inlet (opening 92). A circulating flow is formed.

  Next, referring to FIGS. 2 to 5, 7, 8, 10, and 15 to 17, the analysis operation of the cell analyzer 1 having the cell dispersion unit 52 according to the embodiment of the present invention. explain. In FIG. 16, the processing flow performed by the control unit (processing body) 41 (see FIG. 4) of the data processing device 4 is shown in the right column, and the processing flow performed by the measurement control unit 23 (see FIG. 2) of the measuring device 2. Is shown in the left column. Moreover, in FIG. 17, the processing flow which the preparation control part 33 (refer FIG. 3) of the sample preparation apparatus 3 performs is shown in a line. These processing flows are executed simultaneously.

  First, in step S <b> 1 of FIG. 16, a menu screen is displayed on the display unit 42 by the processing body 41 of the data processing device 4. Thereafter, in step S <b> 2, a measurement start instruction according to the menu screen is received by the input unit 43. In step S <b> 3, the measurement main body 41 of the data processing device 4 transmits a measurement start signal to the measurement device 2. In step S4, when the measurement start signal transmitted from the data processing device 4 is received by the measurement control unit 23 of the measurement device 2, the process proceeds to step S5. In step S5, a preparation start signal is transmitted to the sample preparation device 3.

  As shown in FIG. 17, when the preparation start signal transmitted from the measuring device 2 is received by the preparation control unit 33 of the sample preparation device 3 in step S6, the process proceeds to step S7. In step S7, a reagent (staining solution, RNase) used for preparing a measurement sample is sucked into a flow path in the apparatus. In step S <b> 8, the biological sample in the biological container 6 in which the biological sample and the preserving liquid containing alcohol as a main component are dispersed in the cell dispersion unit 52.

  The biological sample is dispersed by the cell dispersion unit 52 according to the present embodiment as follows.

  As shown in FIG. 15, first, when the motor 80 (see FIG. 8) is rotationally driven by the preparation control unit 33 (see FIG. 3), the rotation of the motor 80 causes the rotation shaft 74 and the tip of the rotation shaft 74 to be connected. The rotor 70 rotates in the direction of arrow R around the axis. At this time, the biological sample in the groove 71 is sent toward the bottom part 62 (between the lower end surface 72 and the bottom part 62 of the rotor 70) by the rotation of the groove 71 on the outer periphery of the rotor 70. In this embodiment, the rotation speed of the motor 80 is about 10,000 rpm, and rotation (cell dispersion) is continuously performed for about 60 seconds.

  The biological sample sent to the bottom 62 flows into the liquid retaining part (concave part) 65 (see FIG. 10) divided by the convex part 64 and is temporarily stored (liquid retained) in the liquid retaining part 65. At this time, since the lower end surface 72 of the rotor 70 that rotates in the direction of the arrow R exists above the biological sample that has flowed into the liquid holding unit 65 (in the direction of the arrow Z1), the biological sample is arrowed in the liquid holding unit 65. It is moved in the R direction. The flow of the biological sample in the direction of arrow R in the liquid retaining part 65 is blocked by the wall surface of the convex part 64 (see FIG. 12) that divides the liquid retaining part 65. Thereby, the flow of the biological sample in the direction of arrow R follows the flow of the biological sample that tends to flow downward (outside) from the hole 63 formed at the end on the arrow R direction side, and along the wall surface of the convex portion 64. Then, it is divided into the flow of the biological sample that moves upward and gets over the convex portion 64.

  The flow of the biological sample that flows out downward (outside) from the hole 63 flows out (spouts) out of the bottom 62. The biological sample that flows out to the outside is pushed by the flow flowing out from the hole 63 and is drawn into the pipe 90 through the opening 92 in the peripheral wall of the pipe 90 by the rotation of the rotor 70. As a result, a circulating flow is formed that circulates from the inlet (opening 92) to the inside of the pipe 90, the outlet (hole 63), the outside of the pipe 90, and the inlet (opening 92).

  On the other hand, the flow that tries to get over the convex portion 64 passes between the lower end surface 72 of the rotor 70 and the upper surface of the convex portion 64. Here, the interval between the lower end surface 72 and the bottom 62 of the rotor 70 is the minimum CL1 on the upper surface of the convex portion 64, and this interval CL1 is an interval (about 50 μm) through which the aggregated cells cannot pass. For this reason, a shearing force is applied between the rotating rotor 70 (lower end surface 72) and the fixedly installed convex part 64 to the aggregated cells included in the flow that is about to get over the convex part 64, Aggregated cells are dispersed into single cells. Since the interval CL1 is large enough to allow a single cell to pass through, the single cells (and dispersed single cells) included in the flow that is going to get over the convex portion 64 ride on the convex portion. Get over 64. For this reason, the aggregated cells contained in the flow of the biological sample trying to get over the convex portion 64 branched by the wall surface of the convex portion 64 are dispersed one after another, and by circulating the circulating flow for a predetermined time (about 60 seconds), Aggregated cells in the biological sample are dispersed into single cells.

  When the dispersion process is completed, in step S9 in FIG. 17, the adjustment pipetting unit 53 (see FIG. 7) and the sample quantifying unit 54 (see FIG. 7) are operated by the adjustment control unit 33 of the sample preparation device 3, and the dispersed biological body A predetermined amount of the sample is sucked from the living body container 6 into the flow path in the apparatus. In step S10, the aspirated predetermined amount of biological sample is sent to the container 56a of the discrimination / substitution unit 56, and discrimination / substitution processing for the biological sample is performed by the discrimination / substitution unit 56. In this discrimination / substitution process, the preservative containing alcohol as a main component is replaced with the diluent, and the diluent containing only the measurement target cells and the liquid containing other cells and impurities are discriminated (filtered). . Further, in the process of discrimination (filtration), a dilute solution containing only the cells to be measured is concentrated, and this concentrate is obtained as a residual liquid of the discrimination / substitution process.

  When the discrimination / substitution process is completed, in step S11, the preparation control unit 33 of the sample preparation device 3 sends a concentrated solution (diluted solution that is mostly epithelial cells) to the flow cell 13 (see FIG. 5) of the sub-detection unit 31. To be liquidated. In step S12, the sub-detection unit 31 is used to perform pre-measurement of the concentrated solution (detection of the number of cells contained in the concentrated solution) by flow cytometry. As a result of the pre-measurement, concentration information reflecting the concentration of the measurement target cell (epithelial cell) contained in the biological sample is obtained before the main measurement performed by the measurement device 2 for cancer determination.

  Next, in step S13, the concentrate is discharged out of the fluid circuit by the preparation controller 33 of the sample preparation device 3. In step S14, the concentration of the biological sample is calculated based on the obtained concentration information. In step S15, the sample suction amount of the biological sample for preparing the measurement sample used for the main measurement is determined based on the calculated concentration. That is, based on the concentration of the biological sample used for the pre-measurement (number of cells per unit volume) and the number of significant cells necessary for cancer cell detection in this measurement, A collection fee (liquid amount) of the biological sample necessary for performing the measurement is calculated.

  In step S <b> 16, the preparation control unit 33 of the sample preparation device 3 sucks the biological sample from the biological container 6 by the determined sample suction amount. In step S17, the discrimination / substitution process (step S10) is performed again on the biological sample. As a result, a concentrated solution in which the biological sample of the determined sample suction amount is replaced and discriminated is obtained.

  Next, in step S18, the concentrate obtained in step S17 is supplied to the product container (microtube) 7 by the preparation control unit 33 of the sample preparation device 3. Further, in step S19, the staining solution and RNase stored in the apparatus are supplied to the product container 7 from the reagent quantitative unit 55 (see FIG. 7). In step S20, a DNA sample and RNA treatment are performed in the product container 7 to produce a measurement sample used for the main measurement.

  When the measurement sample is produced, the obtained measurement sample is sent to the main detection unit 21 of the measurement apparatus 2 by the preparation control unit 33 of the sample preparation apparatus 3 in step S21. Note that the preparation control unit 33 of the sample preparation device 3 always determines whether or not the shutdown signal (see point C in FIG. 17) is received from the measurement device 2 in step S22, and has received the shutdown signal. If not, the process returns to step S6 to determine whether a preparation start signal has been received. If the shutdown signal is received, the process proceeds to step S23, where the shutdown process is executed and the sample preparation process is terminated.

  On the other hand, as shown in FIG. 16, the measurement control unit 23 of the measurement device 2 always determines whether or not the measurement sample is supplied from the sample preparation device 3 in step S24 after transmitting the preparation start signal. Yes. Therefore, when the measurement sample is fed from the sample preparation device 3 in step S21 (see FIG. 17) (see point B in FIG. 17), the process proceeds to step S25. In step S <b> 25, the measurement sample is sent to the flow cell 13 of the main detection unit 21 by the measurement control unit 23 of the measurement apparatus 2, and the main measurement is performed on the cells in the measurement sample. After the actual measurement, the obtained measurement data is transmitted to the data processing device 4 in step S26.

  On the other hand, after transmitting the measurement start signal, the processing body 41 of the data processing device 4 always determines whether or not measurement data has been received from the measurement device 2 in step S27. When the measurement data is received from the measurement device 2, the process proceeds to step S28. In step S28, the processing body 41 of the data processing device 4 analyzes the cells and nuclei using the measurement data, and determines whether or not the cells in the measurement sample are cancerous.

  Next, in step S <b> 29, the analysis result is displayed on the display unit 42 by the processing body 41 of the data processing device 4. In step S30, it is determined whether there is a shutdown instruction by a user input. If there is a shutdown instruction, the process proceeds to step S31, and a shutdown signal is transmitted to the measuring apparatus 2 by the processing body 41 of the data processing apparatus 4.

  The measurement control unit 23 of the measuring device 2 always determines whether or not the shutdown signal from the data processing device 4 has been received in step S32. If the shutdown signal has not been received, the process returns to step S4 for determining whether or not the measurement start signal has been received. If the shutdown signal is received, the shutdown signal is transferred to the sample preparation device 3 in step S33. Thereafter, the shutdown is executed in step S34, and the measurement process ends.

  In the present embodiment, as described above, the rotor 70 that is disposed at a predetermined distance CL1 from the upper surface of the bottom 62 and has a spiral groove 71 on the outer periphery thereof is provided, and the rotor 70 is rotated by the motor 80. Thus, by rotating the spiral groove 71 of the rotor 70, it is possible to form a flow that moves the biological sample containing aggregated cells to the bottom 62 side. Thereby, since the supply of the aggregated cells between the bottom 62 and the rotor 70 can be promoted, the cell dispersion of the aggregated cells can be effectively performed between the bottom 62 and the rotor 70. Furthermore, since the biological sample supplied between the bottom 62 and the rotor 70 is passed through the hole 63 of the bottom 62, the flow of the biological sample formed by the rotation of the spiral groove 71 is not hindered. The aggregated cells can be supplied one after another between the bottom 62 and the rotor 70. As a result, the cell dispersion efficiency of aggregated cells can be improved.

  In the present embodiment, as described above, the hole 63 of the bottom 62 (the perforated member 60) is formed to a size that allows the aggregated cells to pass therethrough. According to this configuration, the hole 63 of the bottom 62 is not blocked by the agglutinated cells (the hole 63 is clogged), so that the biological sample supplied between the bottom 62 and the rotor 70 can be removed from the bottom 62. When passing through the hole 63, the flow of the sample formed by the rotation of the spiral groove 71 can be prevented from being hindered. For this reason, since the supply of the aggregated cells between the bottom 62 and the rotor 70 can be performed effectively, the dispersion of the aggregated cells can be promoted.

  Further, in the present embodiment, as described above, the cells pass from the opening 92 (inlet) to the inside of the pipe 90 and the hole 63 of the perforated member 60 (bottom 62) provided in the opening at the tip of the pipe 90. (Circular outlet), the outside of the pipe 90, and a circulating flow that circulates to the opening 92 (inlet) is formed. If comprised in this way, the flow of the biological sample formed by rotation of the helical groove | channel 71 can be circulated. Thereby, it is possible to more effectively supply the aggregated cells between the bottom 62 and the rotor 70.

  Further, in the present embodiment, as described above, the bottom 62 of the perforated member 60 is disposed at the opening at the tip of the pipe 90, and the opening 92 at the side of the pipe 90 is used as the inlet and is disposed at the opening at the tip. The circulated flow in which cells circulate is formed using the hole 63 of the bottom 62 formed as an outlet. With this configuration, the aggregated cells can be collected on the bottom 62 side of the tip by the rotation of the spiral groove 71 of the rotor 70 inside the pipe 90 and then flow out from the hole 63 of the bottom 62. As a result, the aggregated cells can be reliably supplied between the bottom 62 and the rotor 70 without hindering the circulation flow that is formed.

  In the present embodiment, as described above, the bottom portion 62 of the perforated member 60 is provided with the convex portion 64 for applying a shearing force to the aggregated cells as the rotor 70 rotates. With this configuration, since the convex portion 64 can apply a shearing force to the aggregated cells as the rotor 70 rotates, the rotation of the rotor 70 causes the aggregated cells to flow between the bottom 62 and the rotor 70. Both supply and dispersion of aggregated cells by shear force can be performed. Moreover, since the space | interval between the bottom part 62 and the rotor 70 becomes small in the area | region between the convex part 64 and the rotor 70, when rotating the rotor 70, the area | region between this convex part 64 and the rotor 70 is obtained. Thus, a shearing force can be applied to the aggregated cells. Thereby, a shear force can be made to act on an aggregate cell and an aggregate cell can be disperse | distributed easily.

  In the present embodiment, as described above, the hole 63 of the bottom 62 is formed in an elongated shape. With this configuration, when the flow of the biological sample along the rotation direction (arrow R direction) is formed by rotating the rotor 70 having the spiral groove 71, the elongated hole 63 has a long length. Since the flow in the rotation direction easily reaches a wide range on the side, the biological sample can be easily passed through the perforated member 60 (bottom portion 62). As a result, a biological sample containing dispersed cells can be efficiently discharged from the perforated member 60 (bottom 62), and aggregated cells can be supplied one after another between the bottom 62 and the rotor 70. The supply of aggregated cells to and from the rotor 70 can be promoted.

  In the present embodiment, as described above, the hole 63 is formed so that its longitudinal direction extends in a direction intersecting with the rotation direction of the rotor 70 (arrow R direction). If constituted in this way, since the flow in the rotation direction (arrow R direction) formed by rotating the rotor 70 having the spiral groove 71 and the longitudinal direction of the hole 63 intersect, more efficiently, The flow of the biological sample can reach the hole 63. As a result, the biological sample containing dispersed cells can be more efficiently discharged from the bottom portion 62 (the perforated member 60), thereby further promoting the supply of aggregated cells between the bottom portion 62 and the rotor 70. Can do.

  In the present embodiment, as described above, the hole 63 is disposed in the concave liquid retaining portion 65. If comprised in this way, after the biological sample containing the aggregated cell supplied from the rotor 70 side was hold | maintained so that it may accumulate temporarily in the liquid holding part 65 of the bottom part 62, it was arrange | positioned in the liquid holding part 65 A biological sample can flow out from the hole 63. Thereby, the aggregated cells can be more effectively dispersed between the bottom 62 and the rotor 70 by temporarily accumulating in the liquid retaining part 65, and the hole 63 disposed in the liquid retaining part 65. The biological sample can be discharged from

  In the present embodiment, as described above, the circular bottom portion 62 is provided with the four convex portions 64 extending in the direction intersecting the rotation direction of the rotor 70 (the direction of the arrow R), and the four convex portions 64 are defined as boundaries. A liquid part 65 is provided. If comprised in this way, the biological sample containing the aggregation cell temporarily stored in the liquid holding | maintenance part 65 will flow in the rotation direction (arrow R direction) of the rotor 70 in the liquid holding | maintenance part 65, and rotation of the rotor 70 will be carried out. The projection 64 that intersects the direction (arrow R direction) is reached. Since the gap CL1 between the bottom 62 (convex portion 64) and the rotor 70 (lower end surface 72) decreases when this flow tries to get over the convex portion 64, a shearing force is applied to the aggregated cells in the vicinity of the convex portion 64. As a result, the aggregated cells can be dispersed more effectively.

  Further, in the present embodiment, as described above, the rotor 70 is rotated by the motor 80 so that the aggregated cells are dispersed between the convex portion 64 of the bottom portion 62 (the perforated member 60) and the rotor 70. To do. If comprised in this way, it will be easy to make a shearing force act on an aggregation cell using the area | region (space | interval CL1) between the convex part 64 and the rotor 70 with which the space | interval of the bottom part 62 and the rotor 70 becomes small. Therefore, the aggregated cells can be effectively dispersed.

  Further, in the present embodiment, as described above, a hole is formed at the end on the rotation direction (arrow R direction) side of the rotor 70 by the motor 80 in the four liquid retaining portions 65 with the convex portion 64 of the bottom portion 62 as a boundary. 63 are arranged. If comprised in this way, the biological sample containing the aggregation cell temporarily stored in the liquid holding | maintenance part 65 will flow in the same arrow R direction as the rotation direction of the rotor 70 in the liquid holding | maintenance part 65, and the convex part of a boundary 64 is reached. Since the hole 63 is formed in the boundary portion (the end on the arrow R direction side), the flow of the biological sample is divided into a flow that attempts to get over the convex portion 64 and a flow that flows out of the hole 63. Divided. As a result, aggregated cells are dispersed between the convex portion 64 and the rotor 70 by the flow that tries to get over the convex portion 64, and the biological sample continues to accumulate in the liquid retaining portion 65 by the flow that flows out from the hole portion 63. One after another, aggregated cells are supplied. As a result, it is possible to effectively supply the aggregated cells by smoothing the flow of the biological sample while effectively dispersing the aggregated cells.

  In the present embodiment, as described above, the aggregated cells cannot pass and the dispersed single cells can pass through the predetermined interval between the upper surface of the bottom 62 (convex portion 64) and the rotor 70. The interval CL1 is configured. If comprised in this way, between the bottom part 62 (convex part 64) and the rotor 70, while giving a shearing force to an aggregation cell and disperse | distributing it, the bottom part 62 (without giving a shearing force to a single cell) The projection 64) and the rotor 70 can be passed. Thereby, the dispersion | distribution efficiency of an aggregated cell can be improved, without damaging a single cell.

(Example)
Next, with reference to FIG. 10 and FIG. 18, an experiment for verifying the effect of the present invention will be described. As a comparative example, the number of single cells (single epithelial cells) when the aggregated cells are not dispersed by the cell dispersion unit 52 is measured, and the aggregated cells using the cell dispersion unit 52 according to the present embodiment are measured. When the dispersion treatment was performed (Example 1), the number of single cells (single epithelial cells) was measured, and a comparative experiment was performed to compare changes in the number of single cells (single epithelial cells). Further, a single cell (single epithelial cell) in the case where the shape of the perforated member 60 (bottom 62) of the cell dispersion portion 52 is changed (see FIG. 18) to disperse the aggregated cells (Example 2). The number was measured together.

  First, the structure of the perforated member used in Example 2 will be described. As shown in FIG. 18, the bottom portion 162 of the perforated member is formed in a circular shape when viewed from above, and has four holes penetrating from the upper surface (surface in the direction of arrow Z1) to the lower surface (surface in the direction of arrow Z2). A hole 163 is provided. The four holes 163 have an oblong hole shape extending in the radial direction of the bottom 162. The four holes 163 are disposed in the vicinity of the peripheral edge of the bottom 162 and are disposed at a rotation angle interval of approximately 90 degrees from the center of the bottom 162. In addition, in the bottom part 162 by Example 2 shown in FIG. 18, unlike the bottom part 62 of the cell dispersion | distribution part 52 by this embodiment shown in FIG. 10, a convex part and a liquid holding part (concave part) are formed in the bottom part 162. FIG. The bottom 162 has a flat plate shape. Other configurations of the perforated member used in Example 2 are the same as those of the perforated member 60 of the present embodiment. In Example 2, the porous member having the bottom 162 formed thereon is attached to the pipe 90 of the cell dispersion part 52, so that only the bottom 162 is different from the bottom 62 (Example 1) of the cell dispersion part 52 of the present embodiment. Under the conditions, the aggregated cells are dispersed.

  In Example 1 and Example 2, the number of rotations of the motor 80 was 10,000 rpm, and the dispersion process was performed for 60 seconds. The dispersion treatment and the measurement of the number of single epithelial cells were performed three times each in Example 1, Example 2, and Comparative Example (only the number of cells was measured). As a result, in Example 1 using the cell dispersion unit 52 according to the present embodiment, the number of single epithelial cells measured with respect to the number of single epithelial cells when the aggregated cell dispersion process was not performed (Comparative Example) was measured. The number of cells increased to 151% on average. Further, in Example 2 in which the bottom 62 was changed to the bottom 162, the measured number of single epithelial cells was an average with respect to the number of single epithelial cells when the aggregated cell dispersion treatment was not performed (Comparative Example). Increased to 143%.

  From the above, it was confirmed that the aggregated cells were dispersed by the dispersion treatment using the cell dispersion unit 52 according to the present embodiment, and the number of single cells was increased. In addition, comparing Example 1 and Example 2, compared to Example 2 in which only the hole 163 is formed in the bottom 162, the convex part 64 and the liquid retaining part 65 are formed in the bottom 62. It was confirmed that higher distributed processing efficiency can be obtained.

  The embodiment disclosed this time should be considered as illustrative in all points and not restrictive. The scope of the present invention is shown not by the above description of the embodiments but by the scope of claims for patent, and further includes all modifications within the meaning and scope equivalent to the scope of claims for patent.

  For example, in the above-described embodiment, the example in which the cell dispersion device of the present invention is applied to the cell dispersion unit 52 of the cell analysis device 1 is shown, but the present invention is not limited to this. For example, it is good also as a structure which uses the cell dispersion apparatus of this invention independently. Further, the present invention can be applied to any apparatus other than the cell analysis apparatus as long as the apparatus performs a dispersion treatment of aggregated cells.

  In the above-described embodiment, an example in which cervical epithelial cells are analyzed by a cell analyzer, and aggregated cells obtained by aggregating the epithelial cells are dispersed into single cells by a cell dispersion unit. It is not limited to this. In the present invention, cells other than cervical epithelial cells may be dispersed by the cell dispersion unit.

  Moreover, in the said embodiment, although the example which formed the bottom part and the cylinder part integrally in the perforated member was shown, this invention is not limited to this. In this invention, the structure attached so that a bottom part may be inserted in opening of the front-end | tip of a pipe, without providing a cylinder part may be sufficient.

  Moreover, although the example which attached the perforated member to opening of the front-end | tip of a pipe was shown in the said embodiment, this invention is not limited to this. In this invention, you may comprise so that a perforated member may be arrange | positioned at intervals CL1 without attaching a perforated member to a pipe.

  Moreover, in the said embodiment, although the example which provided four holes which penetrate from the upper surface of a bottom part to a lower surface was shown, this invention is not limited to this. In the present invention, the number of holes may be 3 or less, or 5 or more.

  Moreover, in the said embodiment, although the example which formed the four hole parts in the elongate shape of the width W and the length L was shown, this invention is not limited to this. The hole may be, for example, circular or rectangular. Moreover, it is not necessary that each hole has the same shape, and holes having different shapes may be provided.

  From the results confirmed experimentally, in the case of the elongated hole 63 of the above embodiment, the width W of the hole 63 is preferably in the range of about 0.3 mm to about 1 mm.

  In the above-described embodiment, an example in which the four holes are formed to have a size through which aggregated cells (about 100 μm or more and about 500 μm or less) can pass is shown. In the present invention, the hole may be formed in a size that prevents the aggregated cells from passing therethrough, or a plurality of holes having different sizes may be provided.

  In the above-described embodiment, an example in which the elongated hole portion is formed so that its longitudinal direction extends in a direction orthogonal to the rotation direction of the rotor (arrow R direction) is shown, but the present invention is not limited thereto. Absent. In the present invention, the longitudinal direction of the hole may intersect with the rotation direction of the rotor at a predetermined angle other than a right angle. Moreover, you may form a hole so that it may extend in the direction in alignment with the rotation direction of a rotor.

  Moreover, in the said embodiment, although the example which formed four liquid holding parts which consist of a recessed part in the bottom part was shown, this invention is not limited to this. In the present invention, the number of liquid retaining portions may be three or less, or five or more. In addition, as in the bottom portion 162 used in the second embodiment shown in FIG.

  Moreover, although the example which formed the four convex parts used as the boundary of four liquid holding | maintenance parts in the bottom part was shown in the said embodiment, this invention is not limited to this. In the present invention, the number of convex portions may be three or less, or five or more. At this time, the convex portion does not need to be the boundary of the liquid retaining portion, and for example, a columnar convex portion protruding from the liquid retaining portion (concave portion) may be provided. Further, unlike the bottom portion 162 used in the second embodiment shown in FIG. 18, it is not necessary to provide a convex portion at the bottom portion.

  Moreover, in the said embodiment, although the hole was arrange | positioned in the edge part of the rotation direction (arrow R direction) side of a rotor inside the fan-shaped liquid holding | maintenance part, this invention is limited to this. Absent. In the present invention, the hole portion may be disposed inside the liquid retaining portion at a position other than the end portion on the rotation direction side of the rotor, or the hole portion may be disposed outside the liquid retaining portion.

  Moreover, in the said embodiment, although the example which has arrange | positioned the rotor above the bottom part (arrow Z1 direction) was shown, this invention is not limited to this. In the present invention, a rotor may be disposed below the bottom.

  Moreover, in the said embodiment, although the example which provided the pipe which has an opening in a front-end | tip and a side wall was shown in the outer side of the rotor, this invention is not limited to this. In the present invention, the pipe need not be provided.

  Note that the various dimensions in the above embodiment (intervals CL1, CL2, thickness (t1) of the bottom (convex part), depth t2 of the liquid retaining part, etc.) are merely examples, and the present invention is not limited thereto. . The dimensions of each part may be changed according to the amount of biological sample to be dispersed at a time and the types of cells to be dispersed.

1 Cell analyzer 52 Cell dispersion unit (cell dispersion device)
62, 162 Bottom (first member)
63, 163 hole 64 convex part (shearing force application part)
65 Liquid retention part (recess)
70 rotor (second member)
71 groove 80 motor (drive unit)
90 pipe (cylinder)
CL1 interval

Claims (13)

A cell dispersion device for dispersing aggregated cells,
A first member having a hole penetrating from the upper surface to the lower surface;
A second member disposed at a predetermined interval from the upper surface or the lower surface of the first member and having a spiral groove on the outer periphery thereof;
A drive unit that rotationally drives the second member;
A cylinder disposed outside the second member,
By rotating the second member by the drive unit, the aggregated cells are supplied between the first member and the second member, and the supplied aggregated cells are dispersed. Cell dispersion device.   The cell dispersion device according to claim 1, wherein the hole portion of the first member has a size through which aggregated cells can pass. The cylindrical body has an opening at its tip and side,
When one of the opening at the tip and the opening at the side of the cylinder is an inflow port and the other is an outflow port, the cells pass from the inflow port to the inside of the cylinder, the outflow port, the outside of the cylinder, The cell dispersion device according to claim 1 or 2, wherein a circulating flow that circulates to the inflow port is formed. The first member is disposed in an opening at the tip of the cylindrical body,
An opening of the side part of the cylindrical body is used as an inflow port, and a circulation flow in which cells circulate is formed using a hole part of the first member arranged in the opening of the tip as an outflow port. The cell dispersion apparatus according to claim 3.   The cell dispersion device according to any one of claims 1 to 4, wherein the first member includes a shearing force applying unit that applies a shearing force to the aggregated cells as the second member rotates.   The cell dispersing apparatus according to claim 5, wherein the shearing force applying part includes a convex part protruding toward the second member.   The cell dispersion device according to claim 1, wherein the hole of the first member has an elongated shape.   The cell disperser according to claim 7, wherein the hole portion extends in a direction in which a longitudinal direction thereof intersects a rotation direction of the second member. The first member has a recess provided on the surface on the second member side,
The cell dispersion device according to any one of claims 1 to 8, wherein the hole is disposed in the recess. The first member is circular, and further includes a convex portion that protrudes toward the second member while extending in a direction intersecting with the rotation direction of the second member,
The cell dispersing device according to claim 9, wherein a plurality of the concave portions are provided with the convex portion extending in a direction intersecting with the rotation direction of the second member as a boundary.   The cell according to claim 10, wherein the cell is configured to disperse the aggregated cells between the convex portion of the first member and the second member by rotating the second member by the driving unit. Distributed device.   The hole portion of the first member is respectively disposed at an end portion on the rotation direction side of the second member by the driving portion in the plurality of concave portions having the convex portion as a boundary. The cell disperser described.   The predetermined interval between the upper surface or the lower surface of the first member and the second member is an interval at which aggregated cells cannot pass and a single dispersed cell can pass through. The cell dispersing apparatus according to any one of the above.

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