Observation device and observation method

Abstract

The observation device of the present invention is an observation device for observing the state of a cell population composed of multiple cells. The observation device comprises: at least one irradiation unit disposed opposite to the cell population, which irradiates at least one irradiation area set in the cell population with irradiation light having at least one wavelength including the range of 650 nm or more and 850 nm or less and 1060 nm or more and 1090 nm or less; at least one light-receiving unit disposed opposite to the cell population, which receives emitted light emitted from the cell population by irradiating the irradiation area with irradiation light; and a resolution unit communicatively connected to the light-receiving unit, which determines, based on the intensity of the emitted light, at least one of the number of cells and the cell density of the cell population in the irradiation area.

Description

Technical Field

[0001] This invention relates to an observation apparatus and an observation method. This application claims priority to Japanese Application No. 2024-005452, filed on January 17, 2024, and invokes the entire contents of the aforementioned Japanese application. Background Technology

[0002] In recent years, research has been conducted on applying planar or three-dimensional cell populations, referred to as, for example, cell colonies, cell sheets, spheroids, or organoids, to new drug development and regenerative medicine. As a method for non-invasive optical observation of the state of such cell populations, methods for observing the intensity of transmitted light from the cell population using a transmission microscope are known. Other methods for non-invasive optical observation of the state of cell populations are known, for example, the methods disclosed in Patent Documents 1 to 4. Patent Documents 1 and 2 disclose methods for observing the intensity distribution of transmitted or reflected light from the cell population. Patent Documents 3 and 4 disclose methods for observing the internal structure of cell populations using optical coherence tomography (OCT).

[0003] Existing technical documents

[0004] Patent documents

[0005] Patent Document 1: Japanese Patent Application Publication No. 2018-004594;

[0006] Patent Document 2: Japanese Patent Application Publication No. 2020-094925;

[0007] Patent Document 3: International Publication No. 2015 / 004762;

[0008] Patent document 4: International Publication No. 2017 / 216930. Summary of the Invention

[0009] The observation device of the present invention is an observation device for observing the state of a cell population composed of multiple cells. The observation device comprises: at least one irradiation unit disposed opposite to the cell population, which irradiates at least one irradiation area set in the cell population with irradiation light having at least one wavelength including the range of 650 nm or more and 850 nm or less and 1060 nm or more and 1090 nm or less; at least one light-receiving unit disposed opposite to the cell population, which receives emitted light emitted from the cell population by irradiating the irradiation area with irradiation light; and a resolution unit communicatively connected to the light-receiving unit, which determines, based on the intensity of the emitted light, at least one of the number of cells and the cell density of the cell population in the irradiation area. Attached Figure Description

[0010] Figure 1 This is a diagram showing the configuration of the observation device according to the first embodiment.

[0011] Figure 2 It is magnification Figure 1 A diagram of a part of the observation device.

[0012] Figure 3 It is shown Figure 1 A diagram showing the hardware configuration of the parsing unit.

[0013] Figure 4 It is shown Figure 1 The diagram showing the functional structure of the analytical part.

[0014] Figure 5 It is a graph showing the spectrum of changes in light intensity in cells.

[0015] Figure 6 It is a graph showing the spectrum of light intensity changes in the culture medium.

[0016] Figure 7 It shows the use Figure 1 A flowchart of an example of an observation method implemented by an observation device.

[0017] Figure 8 It is shown Figure 1 A diagram showing a modified example of the observation device.

[0018] Figure 9A It is shown Figure 1 A diagram showing a modified example of the observation device.

[0019] Figure 9B It is shown Figure 1 A diagram showing a modified example of the observation device.

[0020] Figure 10 It is shown Figure 1 A diagram showing a modified example of the observation device.

[0021] Figure 11 This is a diagram showing the configuration of the observation device according to the second embodiment.

[0022] Figure 12 It is magnification Figure 11 A diagram of a part of the observation device.

[0023] Figure 13 It is shown Figure 11 The diagram showing the functional structure of the analytical part.

[0024] Figure 14 It is used for explanation Figure 11 A chart showing the method for determining the Rayleigh scattering coefficient and reflection intensity in the analytical part.

[0025] Figure 15 It is a chart used to illustrate the wavelength range suitable for observing the state of cell populations.

[0026] Figure 16 It is shown Figure 11 A diagram showing a modified example of the observation device.

[0027] Figure 17 It is shown Figure 11 A diagram showing a modified example of the observation device.

[0028] Figure 18A It is shown Figure 11 A diagram showing a modified example of the observation device.

[0029] Figure 18B It is shown Figure 11 A diagram showing a modified example of the observation device.

[0030] Figure 19 It is used for explanation Figure 18A and Figure 18B A chart showing the method for determining the Rayleigh scattering coefficient and reflection intensity in the observation device.

[0031] Figure 20 It is shown Figure 11 A diagram showing a modified example of the observation device.

[0032] Figure 21 It is shown Figure 11 A diagram showing a modified example of the observation device. Detailed Implementation

[0033] [The problem this invention aims to solve]

[0034] The aforementioned cell populations are formed, for example, by multiple cells becoming a planar group due to cell proliferation, or by becoming a multi-layered group. Within such cell populations, the number and density of cells can vary depending on their location. Information such as the number and density of cells at each location within the cell population is an important indicator for a detailed evaluation of the cell population's state. When light is irradiated onto a cell population, the intensity of light scattered and reflected within the cell population depends on the number and density of cells within the population. Therefore, by observing the intensity of the emitted light from the cell population, information including the number and density of cells can be obtained. When light is irradiated onto a cell population, in addition to the scattering and reflection of light from the cell population, there is also absorption of light by the cell population and the culture medium. Therefore, the intensity of the emitted light from the cell population reflects a mixture of various phenomena, including light scattering and reflection, and light absorption.

[0035] In the aforementioned method using a transmission microscope, the resulting microscope image reflects a mixture of phenomena such as light scattering, reflection, and absorption. It is difficult to distinguish the effects of scattering and reflection by cell populations, as well as the effects of light absorption by cell populations, from such an observed image (i.e., emitted light). Therefore, this method makes it difficult to obtain detailed information including cell number and cell density. The same applies to the methods disclosed in Patent Documents 1 and 2. Furthermore, while the methods disclosed in Patent Documents 1 and 2 can use the intensity distribution of light to evaluate the quality of cell populations on a population-by-population basis, it is difficult to obtain detailed information such as the number of cells and cell density at specific locations within a cell population.

[0036] In the methods disclosed in Patent Documents 3 and 4 (methods using OCT), observing the state of the cell population is time-consuming because it utilizes light interference to scan the cell population in three dimensions. This method requires observation of the cell population immediately after it has been removed from the culture tank, where the temperature, atmosphere, and other environmental conditions are managed and maintained under conditions suitable for cell culture. Therefore, considering invasiveness, it is difficult to use OCT for quality control or complete inspection of cultured cell populations.

[0037] This invention provides an observation device and method that can observe the state of cell populations in detail and efficiently.

[0038] [Effects of the Invention]

[0039] According to the present invention, an observation apparatus and method are provided that can observe the state of cell populations in detail and efficiently.

[0040] [Description of embodiments of the present invention]

[0041] First, embodiments of the present invention will be described.

[0042] (1) The observation device of the present invention is an observation device for observing the state of a cell population composed of multiple cells. The observation device comprises: at least one irradiation unit disposed opposite to the cell population, which irradiates at least one irradiation area set in the cell population with irradiation light having at least one wavelength including the range of 650 nm or more and 850 nm or less and 1060 nm or more and 1090 nm or less; at least one light receiving unit disposed opposite to the cell population, which receives emitted light emitted from the cell population by irradiating the irradiation area with irradiation light; and a resolution unit communicatively connected to the light receiving unit, which determines, based on the intensity of the emitted light, at least one of the number of cells and the cell density of the cell population in the irradiation area.

[0043] Typically, when light shines on a cell population, phenomena such as light scattering, reflection, and absorption may occur in and around the cell population. These phenomena are reflected in combination in the intensity of light emitted from the cell population according to the illumination. Therefore, by observing the change or relative value of the emitted light intensity relative to the illumination light intensity (hereinafter, "the change or relative value of the emitted light intensity relative to the illumination light intensity" is referred to as "light intensity change"), the intensity of scattering, reflection, and absorption can be observed. Hereinafter, "absorption intensity" and "absorption strength" refer to "light intensity change caused by absorption." "Scattering intensity" and "scattering strength" refer to "light intensity change caused by scattering." "Reflection intensity" and "reflection strength" refer to "light intensity change caused by reflection." Among these phenomena, the light intensity change caused by scattering and reflection depends on at least one of the parameters of the cell number and cell density of the cell population. The light intensity change caused by absorption depends on the chemical reactions in and around the cell population. Therefore, in order to quantitatively determine parameters such as cell number and cell density, the light intensity change caused only by scattering and reflection can be obtained. The inventors have repeatedly studied methods for obtaining changes in light intensity caused solely by scattering and reflection. They have found that irradiating cell populations with light having at least one wavelength in the ranges of 650 nm to 850 nm and 1060 nm to 1090 nm is effective. For example, in a culture medium capable of containing cell populations, the wavelength ranges of 650 nm to 850 nm and 1060 nm to 1090 nm represent a range where the absorption of light by the culture medium is sufficiently small relative to the scattering and reflection of light by the cells; therefore, the effect of the culture medium's absorption of light on the intensity of the emitted light can be ignored. Similarly, in cells, the wavelength ranges of 650 nm to 850 nm and 1060 nm to 1090 nm also represent a range where the absorption of light by the cells is sufficiently small relative to the scattering and reflection of light by the cells; therefore, the effect of the cells' absorption of light on the intensity of the emitted light can be ignored. Therefore, when irradiating a cell population with illumination light having at least one wavelength encompassing the ranges of 650 nm to 850 nm and 1060 nm to 1090 nm, a parameter including at least one of cell number and cell density can be quantitatively determined as an indicator of the cell population's state based solely on the change in light intensity caused only by scattering and reflection. In the aforementioned observation apparatus, illumination light is irradiated onto each irradiated area of ​​the cell population; therefore, a parameter including at least one of cell number and cell density can be determined for each irradiated area based on the intensity of the emitted light from the irradiated area of ​​the cell population. Thus, the state of a specific location within the cell population can be observed. Therefore, according to the aforementioned observation apparatus, the state of the cell population can be observed in detail.According to the above observation device, unlike the method using OCT, the state of the cell population can be observed efficiently in a short time by simply irradiating the cell population with light.

[0044] (2) In the observation apparatus described in (1) above, the irradiation unit may include a laser light source using a semiconductor light-emitting element, which emits light having a wavelength in the range of 650 nm to 850 nm and 1060 nm to 1090 nm as irradiation light. In this case, by using a semiconductor light-emitting element with stable light intensity as the light source, the risk of fluctuations in the intensity of the emitted light caused by factors other than the state of the cell population (e.g., instability in the intensity of the irradiation light) can be reduced. As a result, observation of the state of the cell population based on the intensity of the emitted light can be performed more reliably.

[0045] (3) In the observation apparatus described in (1) or (2) above, the light-receiving part can be positioned on the opposite side of the irradiation part, separated from the cell group, and can receive a portion of the irradiation light emitted from the irradiation part and passing through the cell group as outgoing light. In this case, the position of the light-receiving part relative to the irradiation part can be easily adjusted.

[0046] (4) In the observation apparatus described in any of (1) to (3) above, the irradiation unit may include one or more light sources that emit irradiation light, and may irradiate each of the multiple irradiation areas set in the cell group. The light receiving unit may include one or more light sensors, and the emitted light is incident on the light sensors, which may receive the multiple irradiation lights passing through the cell group as multiple emitted lights. In this way, when the irradiation light is simultaneously irradiated to each irradiation area of ​​the cell group, compared with the case where the irradiation light is sequentially irradiated to each irradiation area of ​​the cell group, the state of each irradiation area of ​​the cell group can be observed efficiently in a short time.

[0047] (5) In the observation apparatus described in any one of (1) to (4) above, the irradiation unit can irradiate the irradiation area of ​​the cell population with a first irradiation light and a second irradiation light, wherein the first irradiation light has a first wavelength that includes a range of 650 nm or more and 850 nm or less and 1060 nm or more and 1090 nm or less, and the second irradiation light has a second wavelength that includes a range of 650 nm or more and 850 nm or less and 1060 nm or more and 1090 nm or less and is different from the first wavelength. The light receiving unit can receive the first emitted light and the second emitted light that are emitted from the cell population by irradiation with the first irradiation light and the second irradiation light and have wavelengths that are different from each other. The analysis unit can determine the number of cells and the cell density in the irradiation area based on the intensity of the first emitted light and the intensity of the second emitted light, respectively. Among the phenomena of light scattering, reflection and absorption generated in the cell population, light scattering and reflection can be roughly divided into Rayleigh scattering caused by the molecular structure inside the cells constituting the cell population and reflection caused by the density fluctuation inside the cell population. The change in light intensity caused by Rayleigh scattering depends on the number of cells in the cell population. The change in light intensity caused by reflection depends on the cell density of the cell population. The change in light intensity caused by Rayleigh scattering is wavelength-dependent. The change in light intensity caused by reflection is not wavelength-dependent. Therefore, when observing the state of a cell population using light of multiple wavelengths, the difference in their wavelength dependence can be used to distinguish between the change in light intensity caused by wavelength-dependent Rayleigh scattering and the change in light intensity caused by non-wavelength-dependent reflection. In other words, it is possible to differentiate between cell number and cell density and to quantitatively determine the change in light intensity. This allows for a more detailed observation of the state of the cell population.

[0048] (6) In the observation apparatus described in (5) above, the irradiation unit may include: a first light source that emits a first irradiation light; a second light source that emits a second irradiation light; and a switching unit that switches the optical path of the first irradiation light and the optical path of the second irradiation light so that the first irradiation light and the second irradiation light selectively irradiate the irradiated area of ​​the cell population. The light-receiving unit may include a light sensor that selectively receives the first emitted light and the second emitted light emitted from the cell population. In this case, a single light sensor can be used to distinguish and detect the intensity of the first emitted light and the intensity of the second emitted light. As a result, it is not necessary to prepare separate light sensors for detecting the intensity of the first emitted light and light sensors for detecting the intensity of the second emitted light, thus avoiding the complexity of the configuration of the light-receiving unit.

[0049] (7) In the observation apparatus described in (5) above, the irradiation unit may include: a first light source that emits a first irradiation light; a second light source that emits a second irradiation light; and a beam combining unit that combines the first irradiation light and the second irradiation light to generate a beam combining irradiation light, which irradiates the irradiated area of ​​the cell population. The light receiving unit may include: a beam splitting unit that splits the beam combining light emitted from the cell population by the irradiation of the beam combining irradiation light according to wavelength; a first light sensor that detects the intensity of the first emitted light split from the beam combining irradiation light; and a second light sensor that detects the intensity of the second emitted light split from the beam combining irradiation light. In this case, the intensity of the first emitted light and the intensity of the second emitted light can be detected simultaneously using the beam combining irradiation light and the beam combining irradiation light, thus enabling efficient observation of the state of the cell population based on the intensity of the first emitted light and the intensity of the second emitted light in a shorter time.

[0050] (8) In the observation apparatus described in (5) above, the illumination unit may include: a light source that emits light including a first illumination light and a second illumination light; and a filter switching unit that includes a first filter that transmits only the first illumination light and a second filter that transmits only the second illumination light, and switches the position of the first filter and the position of the second filter so that the first filter or the second filter is positioned in the optical path of the light between the light source and the cell cluster. The light receiving unit may include a light sensor that selectively receives the first and second emitted light emitted from the cell cluster. In this case, for example, by using an inexpensive lamp light source as the light source, the cost of the observation apparatus can be reduced compared to using an expensive laser light source as the light source.

[0051] (9) The observation method of the present invention is an observation method for observing the state of a cell population composed of multiple cells. The observation method includes: irradiating at least one irradiation area set in the cell population with irradiation light having at least one wavelength having a range of 650 nm or more and 850 nm or less and 1060 nm or more and 1090 nm or less; receiving emitted light emitted from the cell population by irradiating the irradiation area with irradiation light; and determining, based on the intensity of the emitted light, a parameter including at least one of the number of cells and the cell density of the cell population in the irradiation area. According to this observation method, the same effect as the observation device described above can be obtained.

[0052] (10) In the observation method described in (9) above, in the step of irradiating the irradiated area, a first irradiation light and a second irradiation light can be irradiated onto the irradiated area. The first irradiation light has a first wavelength that includes a range of 650 nm or more and 850 nm or less, and 1060 nm or more and 1090 nm or less. The second irradiation light has a second wavelength that includes a range of 650 nm or more and 850 nm or less, and 1060 nm or more and 1090 nm or less, and is different from the first wavelength. In the step of receiving emitted light, a first emitted light and a second emitted light that have different wavelengths from each other, emitted from the cell population by irradiation with the first irradiation light and the second irradiation light, can be received. In the step of determining parameters, the number of cells and the cell density in the irradiated area can be determined based on the intensity of the first emitted light and the intensity of the second emitted light, respectively. In this case, the same effect as described in (5) above can be obtained.

[0053] [Detailed Description of Embodiments of the Invention]

[0054] Hereinafter, specific examples of the observation apparatus and observation method of the present invention will be described with reference to the accompanying drawings. The present invention is not limited to these examples, but is intended to include all modifications within the meaning and scope equivalent to the claims, as shown by the claims.

[0055] [First Implementation Method]

[0056] Figure 1 This diagram illustrates the configuration of the observation device 1 according to the first embodiment. The observation device 1 is a device for observing the state of the cell group 2. The cell group 2 is a block formed by the aggregation of multiple cells, that is, a block formed by the aggregation of multiple cells together. The cell group 2 can also be described as a single-layer planar cell group, a multi-layer three-dimensional cell group. The cell group 2 includes various forms of cells, such as aggregates of multiple cells called colonies, aggregates arranged in two-dimensional sheets, two-dimensional blocks, and three-dimensional blocks, tissues, and organs, which are called spheroids, organoids, assemblies, etc. The cells included in the cell group 2 include cells collected from animals or humans, stem cells collected from animals or humans, stem cells made based on cells collected from animals or humans, or cells differentiated from stem cells. In a cell group 2, multiple cell types may sometimes coexist.

[0057] The "state" of cell population 2 observed by observation device 1 is represented by a parameter that indicates at least one of the "number of cells" and "cell density" of cell population 2. In this embodiment, a parameter called "agglutination degree," which comprehensively represents the "number of cells" and "cell density" of cell population 2, is used as an indicator of the "state" of cell population 2. "Agglutination degree" is defined by "number of cells" × "cell density" (i.e., the value of the product of "number of cells" and "cell density").

[0058] The observation device 1 illuminates each irradiation region R of the cell population 2 with illumination light L1, and calculates the "agglomeration degree" of the cell population 2 for each irradiation region R based on the intensity of the emitted light L2 from the cell population 2. As a result, for each irradiation region R, information including "cell number" and "cell density" is obtained. The "cell number" in the irradiation region R refers to the total number of cells contained in the irradiation region R. The "cell density" in the irradiation region R refers to the number of cells present per unit volume of the irradiation region R. The cell population 2 is formed by, for example, the proliferation of one cell, resulting in multiple cells stacked in multiple layers. In the cell population 2 thus formed, the "cell number" and "cell density" differ at each location (region). Therefore, obtaining information on the "cell number" and "cell density" in each irradiation region R is important for a detailed assessment of the state of the cell population 2.

[0059] like Figure 1 As shown, the observation device 1 includes, for example, an irradiation module 10, a light-receiving module 20, and a resolution unit 30. The irradiation module 10 is positioned opposite the cell population 2. The light-receiving module 20 is positioned, for example, across the cell population 2, on the opposite side from the irradiation module 10. The cell population 2 is contained, for example, in a container 5 along with culture medium 7. The container 5 is cylindrical with an opening 5b at one end. The observation device 1 may also have a stage for holding the container 5.

[0060] Cell cluster 2 is disposed on the bottom surface 5a of container 5, immersed in culture medium 7. Cell cluster 2 may be in contact with or separated from the bottom surface 5a. Container 5 is formed of, for example, a glass material or resin material capable of transmitting the emitted light L2 emitted from cell cluster 2. Hereinafter, in the vertical direction A1 along the normal of the bottom surface 5a, the direction from the bottom surface 5a toward the opening 5b is sometimes referred to as "up," and the direction from the opening 5b toward the bottom surface 5a is sometimes referred to as "down." The direction perpendicular to the vertical direction A1 is designated as the horizontal direction A2. The transmissible material is one that, for example, has a transmittance of 20% or more for light of, for example, 650 nm or more and 850 nm or less, and 1060 nm or more and 1090 nm or less, with a material thickness of 1 mm.

[0061] The illumination module 10 includes, for example, a plurality of illumination sections 11 arranged in a two-dimensional shape when viewed from above. The plurality of illumination sections 11 are arranged, for example, along a left-right direction A2 perpendicular to the vertical direction A1 and a front-back direction perpendicular to both the vertical direction A1 and the left-right direction A2 (relative to...). Figure 1The cells are arranged in a planar pattern (in the direction of depth of the paper). Multiple irradiation areas R are defined within cell group 2. These multiple irradiation areas R are regions defined to distinguish cell group 2 when viewed from above, and are the regions that become the objects of irradiation by the irradiated light L1. When viewed from above, the multiple irradiation areas R are arranged in a two-dimensional pattern. For example, the multiple irradiation areas R are arranged along the left-right direction A2 and the front-back direction (relative to the depth of the paper). Figure 1 The irradiation units 11 are arranged in a planar pattern (in the depth direction of the paper). Multiple irradiation units 11 are respectively disposed above multiple irradiation areas R. That is, the multiple irradiation units 11 are positioned one-to-one with each of the multiple irradiation areas R in the vertical direction A1. The multiple irradiation units 11 irradiate the multiple irradiation areas R with irradiation light L1. Each irradiation unit 11 may, for example, have the same configuration as each other. Alternatively, each irradiation unit 11 may have different configurations.

[0062] The light-receiving module 20 includes, for example, a plurality of light-receiving parts 21 arranged in a two-dimensional shape when viewed from above. The plurality of light-receiving parts 21 are arranged, for example, along the left-right direction A2 and the front-back direction (relative to...). Figure 1 The light-receiving parts 21 are arranged in a planar pattern (in the depth direction of the paper). Multiple light-receiving parts 21 are disposed below multiple irradiation areas R. The multiple light-receiving parts 21 are positioned opposite each of the multiple irradiation parts 11 in the vertical direction A1, one-to-one, across the cell clusters 2. Each light-receiving part 21 receives outgoing light L2 emitted from the cell cluster 2 after irradiation L1 has been applied to the irradiation area R of the cell cluster 2. In this embodiment, the outgoing light L2 is the transmitted light that passes through the cell cluster 2 in the irradiation light L1. Each light-receiving part 21 outputs an electrical signal S corresponding to the intensity of the outgoing light L2 to the resolution unit 30. Each light-receiving part 21 may, for example, have the same configuration as each other. Alternatively, each light-receiving part 21 may have different configurations.

[0063] Figure 2 This is a diagram showing a portion of the magnified observation device 1. (As shown...) Figure 2As shown, the irradiation unit 11 includes, for example, a light source 13 and a lens 15. The light source 13 is positioned above the irradiation area R of the cell group 2. That is, the light source 13 is positioned opposite the irradiation area R of the cell group 2 in the vertical direction A1. The light source 13 emits irradiation light L1 into the irradiation area R. The optical axis of the irradiation light L1 emitted from the light source 13 is along the vertical direction A1. In one example, the optical axis of the irradiation light L1 is, for example, perpendicular to the bottom surface 5a and parallel to the vertical direction A1. The light source 13 is a laser light source using, for example, a laser diode (LD). The irradiation light L1 has a wavelength encompassing the range of 650 nm to 850 nm and 1060 nm to 1090 nm. The range of 650 nm to 850 nm and 1060 nm to 1090 nm refers to the range encompassing both the range of 650 nm to 850 nm and the range of 1060 nm to 1090 nm. Therefore, it can be said that the irradiating light L1 has a wavelength that includes at least one of the ranges of 650 nm and 850 nm and 1060 nm and 1090 nm.

[0064] Light source 13 is not limited to a laser light source using a laser diode. For example, light source 13 may be a light source using semiconductor light-emitting elements such as laser diodes, light-emitting diodes (LEDs), and superluminescent diodes (SLDs), a laser light source using elements other than semiconductor light-emitting elements, or a lamp light source. For example, in the case of using a broadband light source such as a lamp light source, the illumination light L1 may have one or more wavelengths different from the wavelength range of 650 nm to 850 nm and 1060 nm to 1090 nm. In this case, the one or more wavelengths different from the wavelength may be wavelengths included in the range of 650 nm to 850 nm and 1060 nm to 1090 nm, or wavelengths outside the range of 650 nm to 850 nm and 1060 nm to 1090 nm.

[0065] Lens 15 is positioned in the optical path of the illumination light L1 between the light source 13 and the cell cluster 2. Lens 15 can be, for example, a focusing lens that concentrates the illumination light L1 from the light source 13. Lens 15 can also be a collimating lens that collimates the illumination light L1. The illumination light L1 emitted from the light source 13 passes through lens 15 and illuminates the illumination region R of the cell cluster 2. A portion of the illumination light L1 passes through the cell cluster 2 and exits as the outgoing light L2.

[0066] The light-receiving part 21 includes, for example, a light sensor 23 and a lens 25. The light sensor 23 is disposed below the illumination area R of the cell group 2. That is, the light sensor 23 is disposed on the side opposite to the light source 13 in the vertical direction, across the illumination area R of the cell group 2. Therefore, the light-receiving part 21 is opposite to the light source 13 in the vertical direction A1, across the cell group 2. The light sensor 23 is disposed on the optical path of the emitted light L2 emitted from the cell group 2, that is, on the extension line of the optical axis of the illumination light L1. The optical axis of the emitted light L2 is, for example, perpendicular to the bottom surface 5a on which the cell group 2 is placed, and parallel to the vertical direction A1. The size of the field of view of the light sensor 23 is preferably the same as or smaller than the size of the cells constituting the cell group 2. For example, the diameter of the field of view of the light sensor 23 can be 20 μm or less.

[0067] Lens 25 is positioned in the optical path of the emitted light L2 between the cell cluster 2 and the photosensor 23. Lens 25 can be, for example, an objective lens. Lens 25 can also be a collimating lens. The emitted light L2 from the cell cluster 2 passes through lens 25 and enters the photosensor 23. The photosensor 23 receives the emitted light L2 and outputs an electrical signal S corresponding to the intensity of the emitted light L2. When the illumination light L1 contains multiple wavelengths, a filter that transmits light of a specific wavelength can be positioned at any position between the light source 13 and the photosensor 23.

[0068] like Figure 1 As shown, the analysis unit 30 is communicatively connected to the plurality of light-receiving units 21 of the light-receiving module 20. The analysis unit 30 can also be communicatively connected to the plurality of irradiation units 11 of the irradiation module 10. The analysis unit 30 receives electrical signals S from each light sensor 23. Based on the received electrical signals S, the analysis unit 30 calculates the "aggregation degree" representing the "cell number" and "cell density" of the cell population 2.

[0069] Figure 3 This diagram illustrates the hardware configuration of the parsing unit 30. The parsing unit 30 is physically one or more computers. Figure 3 As shown, the parsing unit 30 physically includes one or more processors 301, main storage device 302, auxiliary storage device 303, input device 304, output device 305, and communication device 306, etc. The parsing unit 30 is composed of one or more physical computers, which are composed of hardware having these physical entities and software such as programs.

[0070] Figure 4 This diagram illustrates the functional configuration of the parsing unit 30. The parsing unit 30, as a functional component, includes, for example, an acquisition unit 31, a storage unit 33, and a conversion unit 35. Each functional component of the parsing unit 30 is implemented by executing a program on the hardware of the aforementioned physical computer.

[0071] The acquisition unit 31 acquires the intensity of the emitted light L2 represented by the electrical signal S from the light receiving module 20 for each irradiated area R of the cell group 2. The acquisition unit 31 provides the light intensity information D1, representing the intensity of the emitted light L2 in each irradiated area R of the cell group 2, to the conversion unit 35.

[0072] Storage unit 33 stores conversion information D2 for determining the "agglomeration degree" of cell population 2 based on the intensity of emitted light L2. Conversion information D2 includes, for example, a standard curve G1 representing the relationship between the change in light intensity obtained from light intensity information D1 and the "agglomeration degree" of cell population 2. Standard curve G1 is a graph plotting the relationship between the change in light intensity and the "agglomeration degree" of cell population 2. The change in light intensity is a relative ratio of the intensity of emitted light L2 to the intensity of irradiated light L1, and is a value after standardizing the intensity of emitted light L2. The change in light intensity is defined by dividing the intensity of emitted light L2 by the intensity of irradiated light L1 (-10Log(intensity of emitted light L2 / intensity of irradiated light L1)).

[0073] like Figure 4 As shown in the standard curve G1, the change in light intensity decreases as the "agglomeration" of cell population 2 decreases and increases as the "agglomeration" increases. Therefore, if the magnitude of the change in light intensity is known, the "agglomeration" can be estimated. By preparing a sample of cell population 2 with a known "agglomeration" (or a sample with a baseline "agglomeration") and irradiating the sample with illumination light L1 using observation device 1, a standard curve G1 representing the relationship between the change in light intensity and the "agglomeration" of the sample can be pre-constructed.

[0074] The storage unit 33 pre-stores a pre-prepared standard curve G1 and provides conversion information D2, including the standard curve G1, to the conversion unit 35. The conversion information D2 may replace the standard curve G1 or, together with the standard curve G1, include a conversion formula (conversion formula) representing the relationship between the change in light intensity and the "agglutination" of the cell population 2. In addition to the standard curve G1, the conversion information D2 may also include the information required to convert the change in light intensity to the "agglutination" of the cell population 2. For example, the conversion information D2 may include the intensity of the irradiation light L1 required for deriving the change in light intensity.

[0075] The conversion unit 35 uses light intensity information D1 and conversion information D2 to calculate the "agglomeration degree" of cell population 2 for each irradiated area R. First, the conversion unit 35 calculates the change in light intensity as a ratio of the intensity of emitted light L2 to the intensity of irradiated light L1. Next, the conversion unit 35 converts the calculated change in light intensity into the "agglomeration degree" of cell population 2 by referring to the standard curve G1. As described above, the "agglomeration degree" of cell population 2 includes parameters of "cell number" and "cell density" of cell population 2, and becomes an indicator of the state of cell population 2. Therefore, by calculating the "agglomeration degree" for each irradiated area R of cell population 2, the state of each irradiated area R of cell population 2 can be quantitatively evaluated. The analysis unit 30 can output the observation results containing the "agglomeration degree" of cell population 2 calculated by the conversion unit 35 using an output device 305 such as a monitor or printer.

[0076] The following is for reference Figure 5 and Figure 6 Explain the reason for using the irradiation light L1, which has a wavelength range of 650 nm to 850 nm and 1060 nm to 1090 nm, to irradiate cell population 2.

[0077] Figure 5 This is a graph showing the spectrum of light intensity changes in cell population 2. Figure 5 In the diagram, the vertical axis represents the change in light intensity of cell group 2, and the horizontal axis represents the wavelength [nm]. Figure 5 The vertical axis can also represent the scattering intensity, reflection intensity, and absorption intensity of cell group 2. Figure 6 This is a graph showing the spectrum of light intensity changes in culture medium 7. Figure 6 In the diagram, the vertical axis represents the change in light intensity in culture medium 7, and the horizontal axis represents the wavelength [nm]. Since no scattering or reflection occurs in culture medium 7, therefore... Figure 6 The vertical axis can also represent the absorption intensity of culture medium 7.

[0078] When light is irradiated onto cell population 2 in culture medium 7, both cell population 2 and culture medium 7 absorb the light, resulting in scattering and reflection of light from cell population 2. The intensity of the light emitted from cell population 2 (transmitted or reflected light) after irradiation reflects a mixture of phenomena such as light scattering and reflection, and light absorption. The intensity of the scattered and reflected light, resulting from these phenomena, depends on parameters such as the "cell number" and "cell density" of cell population 2. Therefore, if the intensity of light arising solely from scattering and reflection can be obtained, the "aggregation" of cell population 2, which includes the "cell number" and "cell density," can be quantitatively determined.

[0079] like Figure 5As shown in Figure G2, in the wavelength range above 1300 nm, there are many peaks indicating abrupt changes in the light intensity of cell group 2. These peaks are caused by the absorption of light by cell group 2. Therefore, in the wavelength range with many peaks, the influence of light absorption on the intensity of light emitted from cell group 2 becomes greater, making it difficult to obtain the light intensity changes caused solely by scattering and reflection. It can be seen that in the wavelength range R1 below 1300 nm, there are no peaks, and the light intensity changes extremely slowly. In this wavelength range R1, the influence of light absorption on the intensity of light emitted from cell group 2 is sufficiently small to be negligible compared to the influence caused by scattering and reflection; therefore, the light intensity changes caused solely by scattering and reflection can be easily obtained.

[0080] like Figure 6 As shown in Figure G3, it can be seen that in the wavelength range R1 below 1300nm, specifically in the wavelength ranges above 650nm and below 850nm, and above 1060nm and below 1090nm, the change in light intensity caused solely by the absorption of light by the culture medium 7 is relative to... Figure 5 The reason shown is that the changes in light intensity due to scattering and reflection by the cells are sufficiently small. Therefore, if the cell population 2 in culture medium 7 is irradiated with light in the wavelength range of 650 nm to 850 nm and 1060 nm to 1090 nm, the effect of light absorption by culture medium 7 can be ignored. In other words, the light absorption by culture medium 7 can be avoided from being reflected in the intensity of light emitted from cell population 2. Culture medium 7 can contain water, amino acids, vitamins, salts, and glucose. Indicators such as phenol red can be added to culture medium 7. If the effect of light absorption by such culture medium 7 can be ignored, the changes in light intensity reflecting only the scattering and reflection by cell population 2 can be obtained. Ultraviolet light with wavelengths shorter than 400 nm may damage cell population 2 and is therefore unsuitable as light for irradiating cell population 2.

[0081] Therefore, when irradiating cell population 2 with irradiation light L1 having a wavelength range R2 encompassing wavelengths of 650 nm to 850 nm and 1060 nm to 1090 nm, as described above, since the influence of cell population 2 and culture medium 7 on the absorption of irradiation light L1 can be ignored, the intensity of emitted light L2, which reflects only the scattering and reflection of irradiation light L1 within cell population 2, can be obtained. As a result, based on the obtained intensity of emitted light L2, the "aggregation degree" of cell population 2 can be quantitatively determined. Thus, in this embodiment, based on the light intensity change spectrum of the cells and the light intensity change spectrum of the culture medium 7, the wavelength range R2 of 650 nm to 850 nm and 1060 nm to 1090 nm is selected as the wavelength range where the influence of cell population 2 and culture medium 7 on the absorption of irradiation light L1 can be ignored.

[0082] Reference Figure 7 The procedure for the observation method implemented using the above-described observation device 1 will be explained. Figure 7 This is a flowchart illustrating the steps involved in the observation method.

[0083] First, each irradiation section 11 arranged in a two-dimensional pattern above the cell group 2 irradiates each irradiation region R of the cell group 2 with irradiation light L1 in the wavelength range of 650 nm to 850 nm and 1060 nm to 1090 nm (step S11). A portion of the irradiation light L1 irradiating each irradiation region R of the cell group 2 passes through the cell group 2 and is emitted from the cell group 2 as outgoing light L2.

[0084] Next, each light-receiving unit 21 arranged in a two-dimensional shape below the cell group 2 receives each emitted light L2 emitted from the cell group 2 (step S12). Each light-receiving unit 21 outputs an electrical signal S corresponding to the intensity of each received emitted light L2 to the resolution unit 30.

[0085] Next, the analysis unit 30 calculates the "cell number" and "cell density" of cell population 2, i.e., "aggregation degree," based on the intensity of the emitted light L2 represented by each electrical signal S, as an indicator of the state of cell population 2 (step S13). Specifically, firstly, the conversion unit 35 of the analysis unit 30 calculates the change in light intensity by dividing the intensity of the emitted light L2 by the intensity of the irradiated light L1. Then, the conversion unit 35 uses a reference... Figure 4 The standard curve G1 shown converts the calculated change in light intensity into the "agglomeration degree" of cell population 2. Using the "agglomeration degree" obtained in this way, the state of each irradiated region R of cell population 2 can be quantitatively observed.

[0086] The effects obtained through the first embodiment described above will be explained. As described above, in the light intensity variation spectrum of the culture medium 7, the wavelength range R2 (refer to) is 650 nm or more and 850 nm or less, and 1060 nm or more and 1090 nm or less. Figure 6 The absorption of light by culture medium 7 containing cell population 2 is relative to... Figure 5 The range of light scattering and reflection by the cells shown is sufficiently small. Therefore, when irradiating cell population 2 with irradiation light L1 having wavelengths R2 including wavelengths above 650 nm and below 850 nm and above 1060 nm and below 1090 nm, the effect of light absorption by the culture medium 7 on the intensity of the emitted light L2 can be ignored. In the light intensity variation spectrum of cell population 2, the wavelength range R2 of 650 nm and below 850 nm and above 1060 nm and below 1090 nm is included in the wavelength range R1 where there are no sharp peaks due to light absorption by the cells (see reference). Figure 5Therefore, the effect of light absorption by cell group 2 on the intensity of emitted light L2 is sufficiently small to be negligible compared to the scattering and reflection generated in cell group 2.

[0087] Therefore, when irradiating cell population 2 with illumination light L1 having a wavelength range R2 of at least 650 nm to 850 nm and 1060 nm to 1090 nm, the intensity of the emitted light L2, which is solely due to scattering and reflection, can be obtained. Therefore, based on the intensity of the emitted light L2, the parameter "agglomeration degree," which includes "cell number" and "cell density," can be quantitatively determined as an indicator of the state of cell population 2. In this embodiment, since illumination light L1 is applied to each irradiated region R of cell population 2, the intensity of emitted light L2 can be distinguished for each irradiated region R. That is, the "agglomeration degree" can be quantitatively determined for each irradiated region R of cell population 2. Thus, information including "cell number" and "cell density" can be obtained at specific locations in cell population 2, allowing for detailed observation of the state of cell population 2.

[0088] According to this embodiment, unlike methods using OCT, the state of cell population 2 can be observed efficiently in a short time through a simple operation of irradiating cell population 2 with illumination light L1. As a result, a complete inspection of all cell populations 2 of the observed object can be performed, avoiding the complexity of process management for observing cell population 2.

[0089] As in this embodiment, the irradiation unit 11 may include a light source 13 using a laser diode, which emits light as irradiation light L1 having a wavelength encompassing a range of 650 nm to 850 nm and 1060 nm to 1090 nm. The light source 13 may be a light source using a semiconductor light-emitting element such as a laser diode, a light-emitting diode, or a superluminescent diode. In this case, by using light from a light source using a semiconductor light-emitting element with stable light intensity as irradiation light L1, the risk of fluctuations in the intensity of the emitted light L2 caused by factors other than the state of the cell population 2 (e.g., instability in the intensity of the irradiation light) can be reduced. Therefore, observation of the state of the cell population 2 based on the intensity of the emitted light L2 can be performed more reliably.

[0090] As in this embodiment, the light-receiving part 21 can be positioned on the opposite side of the irradiation part 11, separated from the cell group 2, and can receive a portion of the irradiation light L1 emitted from the irradiation part 11 and passing through the cell group 2 as outgoing light L2. In this case, the position of the light-receiving part 21 relative to the irradiation part 11 can be easily adjusted.

[0091] As in this embodiment, the multiple irradiation units 11 can irradiate each of the multiple irradiation regions R with irradiation light L1. The multiple light-receiving units 21 can receive a portion of the irradiation light L1 emitted from the multiple irradiation units 11 and passing through the cell group 2 as multiple outgoing light L2. When each irradiation region R of the cell group 2 is simultaneously irradiated with irradiation light L1, compared to irradiating each irradiation region R of the cell group 2 sequentially with irradiation light L1, the state of each irradiation region R of the cell group 2 can be observed efficiently in a short time.

[0092] The observation device and observation method of the present invention are not limited to the first embodiment described above, and can have various other modifications.

[0093] <Variation Example 1>

[0094] Figure 8 This is a diagram showing a modified example of the observation device 1. (For example...) Figure 8 As shown in the observation device 1A, the irradiation module 10A may also include only one irradiation unit 11, and the light-receiving module 20A may also include only one light-receiving unit 21. In this case, the irradiation unit 11 is positioned above any irradiated area R of the cell group 2, irradiating the irradiated area R with irradiation light L1. The light-receiving unit 21 is positioned on the opposite side of the irradiation unit 11, separated from the cell group 2. That is, the light-receiving unit 21 is positioned below the irradiated area R irradiated with irradiation light L1, and is positioned opposite the irradiation unit 11 in the vertical direction A1, separated from the cell group 2.

[0095] The irradiation section 11 and the light-receiving section 21 are configured to be able to move along the left-right direction A2, which is perpendicular to the vertical direction A1, and the front-back direction, which is perpendicular to both the vertical direction A1 and the left-right direction A2 (relative to the vertical direction A1 and the left-right direction A2). Figure 1 The irradiation unit 11 moves along the left-right direction A2 and the front-back direction while sequentially irradiating each irradiation area R of the cell group 2 with irradiation light L1. The light-receiving unit 21 also moves along the left-right direction A2 and the front-back direction in coordination with the movement of the irradiation unit 11 while sequentially receiving each emitted light L2 emitted from the cell group 2 by irradiating each irradiation area R with irradiation light L1.

[0096] The irradiation section 11 and the light-receiving section 21 only need to be configured to be movable relative to the container 5 containing the cell group 2. Therefore, it can be configured such that, with the position of the container 5 fixed, the irradiation section 11 and the light-receiving section 21 move along the left-right direction A2 and the front-back direction. Conversely, it can also be configured such that, with the positions of the irradiation section 11 and the light-receiving section 21 each fixed, the container 5 moves along the left-right direction A2 and the front-back direction.

[0097] exist Figure 8In the observation device 1A shown, the "agglomeration degree," which includes the parameters of "cell number" and "cell density," can be quantitatively determined for each irradiated area R of the cell group 2 based on the intensity of each emitted light L2 from the cell group 2. Therefore, the same effect as in the first embodiment can be obtained. In the observation device 1A, each irradiated area R of the cell group 2 can be set more precisely according to the width of the left-right movement A2 of the irradiation unit 11. Therefore, the "agglomeration degree" in more irradiated areas R of the cell group 2 can be determined. In other words, the state of the cell group 2 can be observed in more detail.

[0098] <Variation Example 2>

[0099] Figure 9A and Figure 9B This is a diagram showing a modified example of the observation device 1. Figure 9A The observation device 1B shown and Figure 9B Each of the observation devices 1C shown has, for example, the above-described Figure 8 The illumination module 10A and the light-receiving module 20A are shown. Each of the observation devices 1B and 1C may also have the illumination module 10 and the light-receiving module 20 of the first embodiment. In the observation devices 1B and 1C, the light-receiving module 20A is disposed above the cell group 2 in the same manner as the illumination module 10A. That is, the light-receiving module 20A is disposed in the same region as the area where the illumination module 10A is located in the two regions separated by the cell group 2.

[0100] exist Figure 9A In the observation device 1B shown, the optical axis of the illumination light L1 emitted from the illumination module 10A is tilted relative to the vertical direction A1. A portion of the illumination light L1 that illuminates the illumination region R of the cell group 2 is reflected on the surface or inside the cell group 2. The reflected light from the cell group 2 is emitted as the outgoing light L2 from the cell group 2. The optical axis of the outgoing light L2 is tilted relative to the vertical direction A1.

[0101] The light-receiving module 20A is disposed in the optical path of the emitted light L2. For example, the light-receiving module 20A is disposed adjacent to the irradiation module 10A in the left-right direction A2. The light-receiving module 20A receives the emitted light L2 and detects its intensity. In the observation device 1B, it is also possible to quantitatively determine the parameter "aggregation degree," which includes "cell number" and "cell density," for each irradiated area R of the cell population 2 based on the intensity of each emitted light L2 from the cell population 2. Therefore, the same effect as in the first embodiment can be obtained.

[0102] exist Figure 9BIn the observation device 1C shown, a circulator 41 is positioned above the irradiation area R of the cell population 2. The irradiation module 10A is positioned further above the circulator 41. The light-receiving module 20A is positioned separately from the circulator 41 in the left-right direction A2. The circulator 41 is positioned at the intersection of the optical axis of the irradiation light L1 emitted from the irradiation module 10A and the optical axis of the outgoing light L2 incident on the light-receiving module 20A. The circulator 41 can also be optically coupled to the irradiation module 10A and the light-receiving module 20A using lenses and optical fibers. A collimating lens or objective lens can also be positioned between the circulator 41 and the cell population 2.

[0103] The circulator 41 is an optical component for separating two beams of light traveling in opposite directions, and has three ports P1, P2, and P3 for the light. Irradiation light L1 emitted from the irradiation module 10A is incident on port P1 of the circulator 41 and exits from port P2 to the irradiation area R of the cell group 2. Outgoing light L2 emitted from the cell group 2 to port P2 exits from port P3 to the light receiving module 20A. In the observation device 1C, the parameter "agglomeration degree," which includes "cell number" and "cell density," can also be quantitatively determined for each irradiation area R of the cell group 2 based on the intensity of each outgoing light L2 from the cell group 2, thus achieving the same effect as in the first embodiment.

[0104] <Variation Example 3>

[0105] Figure 10 This is a diagram showing a modified example of the observation device 1. (For example...) Figure 10 As shown in the observation device 1D, the illumination module 10B may also include only one illumination section 11A, and the light-receiving module 20B may also include only one light-receiving section 21A. Figure 10 In the example shown, the irradiation unit 11A includes a light source 13 and a lens 15. The light source 13 is configured to uniformly irradiate a wide area with irradiation light L1. The light source 13 is positioned above the cell cluster 2 to uniformly irradiate a wide area with irradiation light L1 that extends in a two-dimensional shape when viewed from above. The range of irradiation light L1 irradiated from the light source 13 can also be along the left-right direction A2 and the front-back direction (relative to the left-right direction A2 and the front-back direction A2). Figure 10 The area extends in a planar shape (the depth direction of the paper surface). The light source 13 simultaneously and uniformly illuminates multiple irradiation areas R set on the cell group 2 along the left-right direction A2 and the front-back direction with irradiation light L1.

[0106] Lens 15 is positioned in the optical path of the illumination light L1 between the light source 13 and the cell group 2. Lens 15 is, for example, a collimating lens that collimates the illumination light L1 emitted from the light source 13 over a wide range. Lens 15 extends in a planar shape along the left-right direction A2 and the front-back direction, so that it is opposed to the light source 13 in the vertical direction A1 and to all the illumination areas R set in the cell group 2 in the vertical direction A1. The illumination light L1 emitted from the light source 13 in a diffuse manner is collimated by lens 15 and illuminates each illumination area R of the cell group 2. Then, a portion of each of the plurality of illumination lights L1 passes through the cell group 2 and is emitted as a plurality of outgoing lights L2.

[0107] The light-receiving unit 21A includes a plurality of light sensors 23 and a lens 25. The plurality of light sensors 23 are respectively disposed below a plurality of irradiation regions R of the cell group 2. The light-receiving unit 21A receives the outgoing light L2 emitted from the cell group 2 by irradiating the irradiation region R of the cell group 2 with irradiation light L1.

[0108] Lens 25 is positioned in the optical path of multiple outgoing light beams L2 between the cell cluster 2 and the multiple photosensors 23. Lens 25 is, for example, an objective lens. Lens 25 can also be a collimating lens. Lens 25 extends in a planar shape along the left-right direction A2 and the front-back direction, so that it is opposed to all illumination areas R set in the cell cluster 2 in the vertical direction A1, and also opposed to all photosensors 23 in the vertical direction A1. Lens 25 is opposed to lens 15 in the vertical direction A1 across the cell cluster 2. Each outgoing light beam L2 emitted from the cell cluster 2 passes through lens 25 and enters each photosensor 23. Each photosensor 23 receives the outgoing light beam L2 and outputs an electrical signal S corresponding to the intensity of the outgoing light beam L2.

[0109] exist Figure 10 In the observation device 1D shown, the parameter "aggregation degree," which includes "cell number" and "cell density," can be quantitatively determined for each irradiated region R of the cell group 2 based on the intensity of each emitted light L2 from the cell group 2. Therefore, the same effect as in the first embodiment can be obtained. In the observation device 1D, a single light source 13 can be used to irradiate each irradiated region R with illumination light L1. Therefore, compared with the case where multiple light sources are used to irradiate each irradiated region R with illumination light L1, the number of light sources can be reduced, thus simplifying the device configuration.

[0110] In the observation device 1D, the illumination unit 11A may also include multiple lenses, each individually provided for a plurality of illumination areas R, instead of a single lens 15. The light receiving unit 21A may also include multiple lenses, each individually provided for a plurality of illumination areas R, instead of a single lens 25. In this case, each lens may be positioned opposite each light sensor 23 in the vertical direction A1.

[0111] [Second Implementation]

[0112] The observation device of the second embodiment will be described below. In the following description of the second embodiment, the descriptions that are repeated with those of the first embodiment will be omitted as appropriate, and the differences from the first embodiment will be mainly described.

[0113] Figure 11 This is a diagram showing the configuration of the observation device 101 according to the second embodiment. Figure 12 It is magnification Figure 11 A diagram showing a portion of the observation device 101. The observation device 101 of this embodiment includes an illumination module 110, a light-receiving module 120, and a resolution unit 130. Each of the plurality of light-receiving units 121 included in the light-receiving module 120 has, for example, the same configuration as the light-receiving unit 21 of the first embodiment. That is, as... Figure 12 As shown, the light-receiving part 121 includes a lens 125 having the same configuration as the lens 25 and a light sensor 123 having the same configuration as the light sensor 23.

[0114] The plurality of light-receiving parts 121 are similar to those in the first embodiment, for example along the left-right direction A2 which is perpendicular to the up-down direction A1, and the front-back direction which is perpendicular to both the up-down direction A1 and the left-right direction A2 (relative to the first embodiment). Figure 1 The irradiation modules 110 are arranged in a planar shape along the depth direction of the paper. The plurality of irradiation units 111 of the irradiation module 110 are also arranged in a planar shape, for example, along the left-right direction A2 and the front-back direction, similar to the first embodiment.

[0115] Each of the plurality of irradiation sections 111 in the irradiation module 110 has a different configuration than the irradiation section 11 in the first embodiment. For example... Figure 12 As shown, the irradiation unit 111 includes a first light source 113A, a second light source 113B, a lens 115, and a switching unit 117. The lens 115 has the same configuration as the lens 15 in the first embodiment. The first light source 113A and the second light source 113B are positioned above an irradiation area R of the cell group 2. Each of the first light source 113A and the second light source 113B is a laser light source using, for example, a laser diode (LD). Each of the first light source 113A and the second light source 113B may be a light source using semiconductor light-emitting elements such as laser diodes, light-emitting diodes (LEDs), and superluminescent diodes (SLDs), or it may be a laser light source or a lamp light source using elements other than semiconductor light-emitting elements.

[0116] The first light source 113A emits a first illumination light L1A having a first wavelength encompassing a range of 650 nm to 850 nm and 1060 nm to 1090 nm. The second light source 113B emits a second illumination light L1B having a second wavelength different from the first wavelength. The second wavelength is, for example, a wavelength different from the first wavelength within the range of 650 nm to 850 nm and 1060 nm to 1090 nm. Each of the first illumination light L1A and the second illumination light L1B may also have multiple wavelengths. That is, the first illumination light L1A may also have other wavelengths different from the first wavelength, and the second illumination light L1B may also have other wavelengths different from the second wavelength.

[0117] A switching unit 117 is disposed between the first light source 113A, the second light source 113B, and the lens 115. The switching unit 117 is optically coupled to the first light source 113A, the second light source 113B, and the lens 115 using an optical fiber and a lens. The switching unit 117 switches the optical path of the first illumination light L1A and the second illumination light L1B, so that the first illumination light L1A from the first light source 113A and the second illumination light L1B from the second light source 113B selectively illuminate the illumination region R of the cell population 2. For example, the switching unit 117 sequentially illuminates the illumination region R of the cell population 2 with the first illumination light L1A and the second illumination light L1B. The switching unit 117 can be, for example, an optical switch that changes the direction of light travel by sliding a mirror or prism. The switching unit 117 can also be an optical switch that uses a dielectric with electro-optic effect, such as LiON3 (lithium nitrate) or LiTaO3 (lithium tantalate), to switch the direction of light travel.

[0118] When the first illumination light L1A illuminates cell group 2, a portion of the first illumination light L1A passes through cell group 2 and is emitted from cell group 2 as the first emitted light L2A. The first emitted light L2A has the same wavelength as the first illumination light L1A. When the second illumination light L1B illuminates cell group 2, a portion of the second illumination light L1B passes through cell group 2 and is emitted from cell group 2 as the second emitted light L2B. The second emitted light L2B has the same wavelength as the second illumination light L1B.

[0119] The light sensor 123 selectively receives a first emitted light L2A and a second emitted light L2B emitted from the cell cluster 2. For example, the light sensor 123 receives the first emitted light L2A and the second emitted light L2B in sequence. The light sensor 123 outputs a first electrical signal SA corresponding to the intensity of the first emitted light L2A and a second electrical signal SB corresponding to the intensity of the second emitted light L2B to the resolution unit 130.

[0120] The analysis unit 130 quantitatively determines the "cell number" and "cell density" of the cell population 2 based on the intensity of the first emitted light L2A and the intensity of the second emitted light L2B. Typically, when light is irradiated onto the cell population 2 in the culture medium 7, the cell population 2 and the culture medium 7 absorb the light, resulting in scattering and reflection of light from the cell population 2. Among these phenomena, the scattering and reflection of light can be broadly categorized into Rayleigh scattering caused by the internal molecular structure of the cells constituting the cell population 2 and reflection caused by fluctuations in the internal density of the cell population 2.

[0121] The intensity of Rayleigh scattering produced by illuminating region R of cell population 2 depends on the "cell number" in region R. The intensity of reflection produced in region R depends on the "cell density" in region R. "Rayleigh scattering intensity" refers to "the change in light intensity caused by Rayleigh scattering." Rayleigh scattering intensity depends on the wavelength of light, while reflection intensity does not. Therefore, when observing the state of cell population 2 using light of multiple wavelengths, the wavelength-dependent Rayleigh scattering intensity and the wavelength-independent reflection intensity can be determined separately by utilizing the differences in their wavelength dependencies. In other words, the "cell number" can be quantitatively determined based on Rayleigh scattering intensity, and the "cell density" can be quantitatively determined based on reflection intensity.

[0122] Figure 13 This diagram illustrates the functional configuration of the analysis unit 130. The analysis unit 130, as a functional component, includes, for example, an acquisition unit 131, a storage unit 133, and a conversion unit 135. The acquisition unit 131 acquires the intensity of the first emitted light L2A represented by the first electrical signal SA and the intensity of the second emitted light L2B represented by the second electrical signal SB for each irradiated region R of the cell population 2. The acquisition unit 131 provides the conversion unit 135 with first light intensity information D11 representing the intensity of the first emitted light L2A in each irradiated region R of the cell population 2 and second light intensity information D12 representing the intensity of the second emitted light L2B in each irradiated region R of the cell population 2.

[0123] Storage unit 133 stores first conversion information D21 for determining the "cell number" of cell population 2 based on the intensity of the first emitted light L2A and the intensity of the second emitted light L2B, and second conversion information D22 for determining the "cell density" of cell population 2 based on the intensity of the first emitted light L2A and the intensity of the second emitted light L2B. The first conversion information D21 includes a first standard curve G11 representing the relationship between the Rayleigh scattering coefficient obtained from the first light intensity information D11 and the second light intensity information D12 and the "cell number" of cell population 2. The "Rayleigh scattering coefficient" is a parameter representing the relationship between Rayleigh scattering intensity and wavelength, which can be derived from the wavelength dependence of Rayleigh scattering intensity. There exists a proportional relationship where a larger Rayleigh scattering coefficient corresponds to a larger Rayleigh scattering intensity. Therefore, by determining either the Rayleigh scattering intensity or the Rayleigh scattering coefficient, the "cell number" can be quantitatively determined. In this embodiment, the case of determining the "cell number" using the Rayleigh scattering coefficient is illustrated. The first standard curve G11 is a graph plotting the relationship between the Rayleigh scattering coefficient and the "cell number". The second conversion information D22 includes a second standard curve G12 representing the relationship between the reflected intensity obtained from the first light intensity information D11 and the second light intensity information D12 and the "cell density" of cell population 2. The second standard curve G12 is a graph plotting the relationship between reflected intensity and "cell density". At least one of the first conversion information D21 or the second conversion information D22 may include the intensity of the first irradiation light L1A used to determine the first light intensity change from the first light intensity information D11, and may also include the intensity of the second irradiation light L1B used to determine the second light intensity change from the second light intensity information D12.

[0124] Figure 14 It is a chart used to illustrate the method for determining the Rayleigh scattering coefficient and reflection intensity. Figure 14 Chart G13 shows the relationship between the change in light intensity and wavelength λ in a state with only Rayleigh scattering and reflection and no absorption. Figure 14 In the diagram, the vertical axis represents the change in light intensity, and the horizontal axis represents one-fourth of the wavelength λ. Figure 14 The diagram shows plotted point P11 representing the relationship between the first light intensity change P(A) and the first wavelength λ(A), and plotted point P12 representing the relationship between the second light intensity change P(B) and the second wavelength λ(B). The first light intensity change P(A) is the relative ratio of the intensity of the first emitted light L2A to the intensity of the first irradiated light L1A, defined by -10Log(intensity of the first emitted light L2A / intensity of the first irradiated light L1A). The second light intensity change P(B) is the relative ratio of the intensity of the second emitted light L2B to the intensity of the second irradiated light L1B, defined by -10Log(intensity of the second emitted light L2B / intensity of the second irradiated light L1B).

[0125] exist Figure 14In the graph, the change in light intensity is represented by the sum of Rayleigh scattering intensity and reflected intensity. Wavelength-independent reflected intensity is represented by the intercept of the line connecting plotted points P11 and P12 (i.e., the intensity P(C) shown at the intersection of the line and the vertical axis of the graph). Wavelength-dependent Rayleigh scattering intensity is represented by the product of the slope of the line connecting plotted points P11 and P12 (i.e., the Rayleigh scattering coefficient) and the fourth power of the wavelength λ. Therefore, the Rayleigh scattering coefficient Pr1 can be calculated using the following mathematical formula (1), and the reflected intensity Pr2 can be calculated using the following mathematical formula (2). If the Rayleigh scattering coefficient Pr1 is calculated, the Rayleigh scattering intensity can also be calculated using the above relationship.

[0126] [Mathematical Expression 1]

[0127]

[0128] [Mathematical Expression 2]

[0129]

[0130] like Figure 13 As shown, the storage unit 133 pre-stores first conversion information D21, which includes a first standard curve G11 representing the relationship between the Rayleigh scattering coefficient and the "cell number", and second conversion information D22, which includes a second standard curve G12 representing the relationship between the reflection intensity and the "cell density". By preparing a sample of cell population 2 with known "cell number" and "cell density" (or a sample with "cell number" and "cell density" as a reference), and irradiating the sample with a first irradiation light L1A and a second irradiation light L1B using the observation device 101, it is possible to pre-create the first standard curve G11 representing the relationship between the Rayleigh scattering coefficient and the "cell number" of the sample, and the second standard curve G12 representing the relationship between the reflection intensity and the "cell density".

[0131] The conversion unit 135 uses the first light intensity information D11 and the second light intensity information D12 from the acquisition unit 131, and the first conversion information D21 and the second conversion information D22 from the storage unit 133 to calculate the "number of cells" and "cell density" for each irradiated area R of the cell population 2. Specifically, firstly, the conversion unit 135 calculates the relative ratio of the intensity of the first emitted light L2A to the intensity of the first irradiated light L1A, i.e., the first light intensity change P(A), and calculates the relative ratio of the intensity of the second emitted light L2B to the intensity of the second irradiated light L1B, i.e., the second light intensity change P(B). Next, the conversion unit 135 uses the first light intensity change P(A) and the second light intensity change P(B) to calculate the Rayleigh scattering coefficient Pr1 shown in mathematical formula (1). The conversion unit 135 uses the Rayleigh scattering coefficient Pr1 to calculate the reflection intensity Pr2 shown in mathematical formula (2).

[0132] Then, the conversion unit 135 converts the Rayleigh scattering coefficient Pr1 into "cell number" by referring to the first standard curve G11. The conversion unit 135 converts the reflection intensity Pr2 into "cell density" by referring to the second standard curve G12. Thus, the "cell number" and "cell density" of each irradiated area R of the cell group 2 can be calculated separately.

[0133] Figure 15 This is a chart illustrating the wavelength range suitable for observing the state of cell population 2. Figure 15 Chart G14, represented by a solid line, shows the wavelength dependence of light intensity changes, reflecting the effect of cell and culture medium 7 on light absorption. Figure 15 Chart G15, indicated by dashed lines, shows the wavelength dependence of light intensity variation in the absence of the influence of cells and culture medium 7 on light absorption. Figure 15 As shown in Figure G14, the wavelength ranges R11 and R12 contain peaks where the light intensity changes sharply, and the wavelength ranges R21 and R22 do not contain such peaks. In the wavelength ranges R11 and R12 where such peaks exist, the Rayleigh scattering coefficient may be difficult to determine accurately due to the influence of the peaks. Therefore, to accurately determine the Rayleigh scattering coefficient, it is suitable to use wavelengths contained in the wavelength ranges R21 and R22 where the peaks do not exist.

[0134] In this embodiment, the first wavelength λ(A) of the first irradiation light L1A and the second wavelength λ(B) of the second irradiation light L1B are both contained in the wavelength range R2 (refer to) of 650 nm or more and 850 nm or less, and 1060 nm or more and 1090 nm or less. Figure 6 In ), wavelengths above 1060nm and below 1090nm are... Figure 15 The wavelengths in graph G14 that do not contain peaks are included in wavelength range R21, and wavelengths above 650 nm and below 850 nm are included in wavelength range R22. Therefore, when the first irradiation light L1A and the second irradiation light L1B described above are irradiated onto the irradiation region R of cell population 2, the change in light intensity without the influence of cell population 2 and culture medium 7 on the absorption of the irradiated light can be obtained. Therefore, the Rayleigh scattering coefficient can be accurately determined based on the change in light intensity.

[0135] The effects obtained through the second embodiment described above will be explained. As described above, when light is irradiated onto cell population 2, Rayleigh scattering occurs due to the molecular structure within the cells constituting cell population 2, and reflection occurs due to fluctuations in the density within cell population 2. The intensity of Rayleigh scattering depends on the "cell number." The intensity of reflection depends on the "cell density." The intensity of Rayleigh scattering is wavelength-dependent. The intensity of reflection is not wavelength-dependent. Therefore, when observing the state of cell population 2 using light of multiple wavelengths, the wavelength-dependent Rayleigh scattering intensity and the wavelength-independent reflection intensity can be distinguished and determined by utilizing the differences in their wavelength dependencies. In other words, the "cell number" and "cell density" can be distinguished and quantitatively determined. As a result, the state of cell population 2 can be observed in more detail.

[0136] As in this embodiment, the irradiation unit 111 may include a first light source 113A, a second light source 113B, and a switching unit 117. The light-receiving unit 121 may include a light sensor 123. In this case, a single light sensor 123 can be used to distinguish and detect the intensity of the first emitted light L2A and the intensity of the second emitted light L2B. As a result, it is not necessary to prepare separate light sensors for detecting the intensity of the first emitted light L2A and for detecting the intensity of the second emitted light L2B, thus avoiding the complexity of the configuration of the light-receiving unit 121.

[0137] The observation device and observation method of the present invention are not limited to the second embodiment described above, and can have various other modifications.

[0138] <Variation Example 1>

[0139] Figure 16 This is a diagram showing a modified example of the observation device 101. (As shown...) Figure 16 As shown in the observation device 101A, the irradiation module 110A may also include only one irradiation unit 111, and the light-receiving module 120A may also include only one light-receiving unit 121. In this case, the irradiation unit 111 is positioned above any irradiation area R of the cell group 2, irradiating the irradiation area R with a first irradiation light L1A and a second irradiation light L1B. The light-receiving unit 121 is positioned on the opposite side of the irradiation unit 111, separated from the cell group 2. That is, the light-receiving unit 121 is positioned below the irradiation area R irradiated by the first irradiation light L1A and the second irradiation light L1B, and is positioned opposite the irradiation unit 111 in the vertical direction A1, separated from the cell group 2.

[0140] The irradiation section 111 and the light-receiving section 121 are configured to be able to move along the left-right direction A2, which is perpendicular to the vertical direction A1, and the front-back direction, which is perpendicular to both the vertical direction A1 and the left-right direction A2 (relative to the vertical direction A1 and the left-right direction A2). Figure 16The irradiation unit 111 moves along the left-right direction A2 and the front-back direction while sequentially irradiating each irradiation area R of the cell group 2 with the first irradiation light L1A and the second irradiation light L1B. The light-receiving unit 121 also moves along the left-right direction A2 and the front-back direction in coordination with the movement of the irradiation unit 111, while sequentially receiving the first emitted light L2A and the second emitted light L2B emitted from the cell group 2 by irradiating each irradiation area R with the first irradiation light L1A and the second irradiation light L1B.

[0141] The irradiation section 111 and the light-receiving section 121 only need to be configured to be movable relative to the container 5 containing the cell group 2. Therefore, it can be configured such that, with the position of the container 5 fixed, the irradiation section 111 and the light-receiving section 121 move along the left-right direction A2 and the front-back direction. Conversely, it can also be configured such that, with the positions of the irradiation section 111 and the light-receiving section 121 each fixed, the container 5 moves along the left-right direction A2 and the front-back direction.

[0142] exist Figure 16 In the observation device 101A shown, the "cell count" and "cell density" can also be quantitatively determined for each irradiated area R of the cell population 2 based on the intensity of the first emitted light L2A and the intensity of the second emitted light L2B. Therefore, the same effect as in the second embodiment can be obtained. In the observation device 101A, each irradiated area R of the cell population 2 can be set more precisely according to the width of the left-right movement A2 of the irradiation unit 111. Therefore, the "cell count" and "cell density" in more irradiated areas R of the cell population 2 can be determined. In other words, the state of the cell population 2 can be observed in more detail.

[0143] <Variation Example 2>

[0144] Figure 17 This is a diagram showing a modified example of the observation device 101. (As shown...) Figure 17As shown in the observation device 101B, the illumination unit 111A may include a beam combiner 119 instead of the switching unit 117. The beam combiner 119 is optically coupled to the first light source 113A, the second light source 113B, and the lens 115 using an optical fiber and a lens. The beam combiner 119 is disposed between the first light source 113A, the second light source 113B, and the lens 115. The beam combiner 119 combines the first illumination light L1A from the first light source 113A and the second illumination light L1B from the second light source 113B, and illuminates the illumination region R of the cell group 2 as the combined illumination light L3. The combined illumination light L3 is a combined light including the first illumination light L1A and the second illumination light L1B, and has a first wavelength λ(A) and a second wavelength λ(B). When the combined illumination light L3 illuminates the illumination region R of the cell group 2, a portion of the combined illumination light L3 passes through the cell group 2 and is emitted from the cell group 2 as the combined outgoing light L4. The output beam L4 has a wavelength corresponding to the first wavelength λ(A) and the second wavelength λ(B) (e.g., the same wavelength as the first wavelength λ(A) and the second wavelength λ(B)). The multiplexing section 119 is, for example, a dichroic mirror. The multiplexing section 119 can also be an optical fiber multiplexer such as a WDM coupler.

[0145] The light-receiving unit 121A replaces the light sensor 123 with a beam-splitting unit 129, a first light sensor 123A, and a second light sensor 123B. The first light sensor 123A and the second light sensor 123B are disposed below the cell group 2, adjacent to each other along the left-right direction A2. The beam-splitting unit 129 is disposed between the cell group 2 and the first light sensor 123A and the second light sensor 123B. The beam-splitting unit 129 splits the combined outgoing light L4 according to wavelength. For example, the beam-splitting unit 129 splits the combined outgoing light L4 into a first outgoing light L2A having a first wavelength λ(A) and a second outgoing light L2B having a second wavelength λ(B). The first light sensor 123A receives the first outgoing light L2A split from the combined outgoing light L4 by the beam-splitting unit 129 and outputs a first electrical signal SA representing the intensity of the first outgoing light L2A. The second optical sensor 123B receives the second outgoing light L2B split from the combined outgoing light L4 by the beam splitter 129, and outputs a second electrical signal SB representing the intensity of the second outgoing light L2B. The beam splitter 129 is, for example, a dichroic mirror. The beam splitter 129 can also be a fiber optic beam splitter such as a WDM coupler.

[0146] exist Figure 17In the observation device 101B shown, the "cell number" and "cell density" can also be quantitatively determined for each irradiated area R of the cell population 2 based on the intensity of the first emitted light L2A and the intensity of the second emitted light L2B, thus achieving the same effect as in the second embodiment. According to the observation device 101B, the intensity of the first emitted light L2A and the intensity of the second emitted light L2B can be detected simultaneously using the combined irradiation light L3 and the combined emitted light L4, thus enabling more efficient observation of the state of each irradiated area R of the cell population 2 based on the intensity of the first emitted light L2A and the intensity of the second emitted light L2B in a shorter time.

[0147] <Variation Example 3>

[0148] Figure 18A and Figure 18B This is a diagram showing a modified example of the observation device 101. Figure 18A The observation device 101C shown and Figure 18B Each of the observation devices 101D shown has, for example, Figure 16 The illumination module 110A and the light-receiving module 120A are shown. Each of the observation devices 101C and 101D may also have the illumination module 110 and the light-receiving module 120 of the second embodiment. In the observation devices 101C and 101D, the light-receiving module 120A is disposed above the cell group 2 in the same manner as the illumination module 110A. That is, the light-receiving module 120A is disposed in the same region as the region where the illumination module 110A is located, in one of the two regions separated by the cell group 2.

[0149] exist Figure 18A In the observation device 101C shown, the optical axes of the first illumination light L1A and the second illumination light L1B (hereinafter referred to as "illumination lights L1A and L1B") emitted from the illumination module 110A are inclined relative to the vertical direction A1. A portion of the first illumination light L1A illuminating the illumination region R of the cell group 2 is reflected on the surface or inside the cell group 2. The reflected light reflected from the cell group 2 is emitted from the cell group 2 as the first outgoing light L2A. A portion of the second illumination light L1B illuminating the illumination region R of the cell group 2 is reflected on the surface or inside the cell group 2. The reflected light reflected from the cell group 2 is emitted from the cell group 2 as the second outgoing light L2B. The optical axes of the first outgoing light L2A and the second outgoing light L2B are inclined relative to the vertical direction A1.

[0150] The light-receiving module 120A is disposed in the optical path of the emitted light L2A and L2B. The light-receiving module 120A is disposed, for example, adjacent to the illumination module 110A in the left-right direction A2. The light-receiving module 120A receives the first emitted light L2A and the second emitted light L2B (hereinafter simply referred to as "emitted light L2A and L2B"), and detects the intensity of the emitted light L2A and L2B. The observation device 101C, similar to the observation device 101 of the second embodiment, calculates the Rayleigh scattering coefficient and reflection intensity based on the intensity of the emitted light L2A and L2B.

[0151] Figure 19 This is a chart illustrating the method for determining the Rayleigh scattering coefficient and reflection intensity. Figure 19 In the diagram, the vertical axis represents the change in light intensity, and the horizontal axis represents one-fourth of the wavelength λ. Figure 19 The diagram shows plotted point P21 representing the relationship between the first light intensity change P(A) and the first wavelength λ(A) of the first emitted light L2A, and plotted point P22 representing the relationship between the second light intensity change P(B) and the second wavelength λ(B) of the second emitted light L2B. The first light intensity change P(A) is the relative ratio of the intensity of the first emitted light L2A to the intensity of the first irradiated light L1A, defined by -10Log(intensity of the first emitted light L2A / intensity of the first irradiated light L1A). The second light intensity change P(B) is the relative ratio of the intensity of the second emitted light L2B to the intensity of the second irradiated light L1B, defined by -10Log(intensity of the second emitted light L2B / intensity of the second irradiated light L1B).

[0152] exist Figure 19 In graph G23, the change in light intensity is represented by the sum of Rayleigh scattering intensity and reflection intensity. Wavelength-independent reflection intensity is represented by the intercept of the line connecting plotted points P21 and P22 (i.e., the intensity P(C) shown at the intersection of the line and the vertical axis of the graph). Wavelength-dependent Rayleigh scattering intensity is represented by the product of the slope of the line connecting plotted points P21 and P22 (i.e., the Rayleigh scattering coefficient) and the fourth power of the wavelength λ. Therefore, the Rayleigh scattering coefficient Pr1 can be obtained using the above mathematical formula (1). The reflection intensity Pr3 can be obtained using the following mathematical formula (3). Then, by converting the Rayleigh scattering coefficient Pr1 into "cell number" and the reflection intensity Pr3 into "cell density," each of the "cell number" and "cell density" can be quantitatively determined. Therefore, the same effect as in the second embodiment can be obtained in the observation device 101C.

[0153] [Mathematical Expression 3]

[0154]

[0155] exist Figure 18BIn the observation device 101D shown, a circulator 141 is disposed above the irradiation area R of the cell population 2. An irradiation module 110A is disposed further above the circulator 141. A light-receiving module 120A is disposed at a position separated from the circulator 141 in the left-right direction A2. The circulator 141 is disposed at the intersection of the optical axes of the irradiation light L1A, L1B emitted from the irradiation module 110A and the optical axes of the outgoing light L2A, L2B incident on the light-receiving module 120A.

[0156] Circulator 141 is an optical component for separating two beams of light traveling in opposite directions, and has three ports P1, P2, and P3 for light. Irradiation light L1A and L1B emitted from irradiation module 110A is incident on port P1 of circulator 141 and exits from port P2 to the irradiation area R of cell group 2. Outgoing light L2A and L2B emitted from cell group 2 to port P2 are exited from port P3 to light receiving module 120A. In observation device 101D, the “cell number” and “cell density” of cell group 2 can also be quantitatively determined for each irradiation area R of cell group 2 based on the intensity of outgoing light L2A and L2B from cell group 2, thus achieving the same effect as in the second embodiment.

[0157] <Variation Example 4>

[0158] Figure 20 This is a diagram showing a modified example of the observation device 101. (As shown...) Figure 20 As shown in the observation device 101E, the illumination module 110B may also include only one illumination section 111B, and the light-receiving module 120B may also include only one light-receiving section 121B. Figure 20 In the example shown, the irradiation unit 111B includes a light source 114 and a lens 115. The light source 114 is configured to uniformly irradiate a wide area with irradiation light L1A and L1B. The light source 114 is positioned above the cell group 2 to uniformly irradiate the wide area with irradiation light L1A and L1B that extends in a two-dimensional shape when viewed from above. The range of irradiation light L1A and L1B irradiated from the light source 114 can also be along the left-right direction A2 and the front-back direction (relative to the left-right direction A2 and the front-back direction A2). Figure 20 The area extends in a planar shape (in the depth direction of the paper). The light source unit 114 simultaneously and uniformly irradiates multiple irradiation areas R set on the cell group 2 and arranged along the left-right direction A2 and the front-back direction with irradiation light L1A or irradiation light L1B.

[0159] The light source unit 114 may, for example, include a first light source 113A, a second light source 113B, and a switching unit 117 (see reference). Figure 12 It can also be a configuration that includes a first light source 113A, a second light source 113B, and a wave combiner 119 (see reference). Figure 17Alternatively, the light source unit 114 may also be configured as described later, including the light source 113C, lens 115, and filter switching unit 118 (see below). Figure 21 The light source unit 114 is not limited to these configurations; it can be any configuration that allows for the illumination of the illumination light L1A and L1B to be extended in a two-dimensional manner when viewed from above.

[0160] Lens 115 is disposed in the optical path of the illumination light L1A and L1B between the light source 114 and the cell group 2. Lens 115 is, for example, a collimating lens that collimates the illumination light L1A and L1B emitted from the light source 114 over a wide range. Lens 115 extends in a planar shape along the left-right direction A2 and the front-back direction, so that it is opposed to the light source 114 in the vertical direction A1 and to all the illumination areas R set in the cell group 2 in the vertical direction A1. The illumination light L1A and L1B emitted from the light source 114 in a diffuse manner is collimated by lens 115 and illuminates each illumination area R of the cell group 2. A portion of the illumination light L1A passes through the cell group 2 and is emitted as the outgoing light L2A, and a portion of the illumination light L1B passes through the cell group 2 and is emitted as the outgoing light L2B.

[0161] The light-receiving part 121B includes a plurality of light sensors 123 and a lens 125. The plurality of light sensors 123 are respectively disposed below a plurality of irradiation regions R of the cell group 2. Each light sensor 123 selectively receives outgoing light L2A and L2B emitted from the cell group 2 by irradiating the irradiation regions R of the cell group 2 with irradiation light L1A and L1B.

[0162] Lens 125 is disposed in the optical path of multiple outgoing light beams L2A and L2B between cell cluster 2 and multiple photosensors 123. Lens 125 is, for example, an objective lens. Lens 125 extends in a planar shape along the left-right direction A2 and the front-back direction, so that it is opposed in the vertical direction A1 to all illumination areas R set in cell cluster 2 and to all photosensors 123 in the vertical direction A1. Lens 125 is opposed in the vertical direction A1 to lens 115 across cell cluster 2. Each outgoing light beam L2A and L2B emitted from cell cluster 2 is incident on each photosensor 123 through lens 125. Each photosensor 123 selectively receives the outgoing light beams L2A and L2B and outputs electrical signals SA and SB corresponding to the intensity of the outgoing light beams L2A and L2B.

[0163] exist Figure 20In the observation device 101E shown, the "cell number" and "cell density" can also be quantitatively determined for each irradiated area R of the cell group 2 based on the intensity of each emitted light L2A and L2B from the cell group 2. Therefore, the same effect as in the second embodiment can be obtained. In the observation device 101E, one light source unit 114 can be used to irradiate each irradiated area R with irradiation light L1A and L1B. Therefore, compared with the case where multiple light source units are used to irradiate each irradiated area R with irradiation light L1A and L1B, the number of light source units can be reduced, and the device configuration can be simplified.

[0164] In the observation device 101E, the illumination unit 111B may also include multiple lenses, each individually provided for a plurality of illumination areas R, instead of a single lens 115. The light receiving unit 121B may also include multiple lenses, each individually provided for a plurality of illumination areas R, instead of a single lens 125. In this case, each lens is positioned opposite each light sensor 123 in the vertical direction A1, one-to-one.

[0165] <Variation Example 5>

[0166] Figure 21 This is a diagram showing a modified example of the observation device 101. (As shown...) Figure 21 As shown, the observation device 101F may also have an illumination unit 111C instead of the illumination unit 111. The illumination unit 111C includes a light source 113C, a lens 115, and a filter switching unit 118. The light source 113C is a halogen lamp (light source) that emits illumination light L1 containing multiple wavelengths. At least one wavelength contained in the illumination light L1 is in the range of 650 nm or more and 850 nm or less, and 1060 nm or more and 1090 nm or less. Other wavelengths contained in the illumination light L1 may be in the range of 650 nm or more and 850 nm or less, and 1060 nm or more and 1090 nm or less, or may be outside the range of 650 nm or more and 850 nm or less, and 1060 nm or more and 1090 nm or less.

[0167] Lens 115 is optically coupled to light source 113C using an optical fiber bundle or similar fiber. A filter switching unit 118 is disposed in the optical path of the illumination light L1 between lens 115 and cell cluster 2. The filter switching unit 118 maintains a first filter 118A and a second filter 118B, such that either the first filter 118A or the second filter 118B is disposed in the optical path of the illumination light L1. The first filter 118A is a bandpass filter that transmits only the first wavelength λ(A) of the plurality of wavelengths contained in the illumination light L1. The second filter 118B is a bandpass filter that transmits only the second wavelength λ(B) of the plurality of wavelengths contained in the illumination light L1. Figure 21As shown in the upper right part, when the filter switching section 118 is viewed along the vertical direction A1, each of the first filter 118A and the second filter 118B is, for example, circular.

[0168] like Figure 21 As shown, when the filter switching unit 118 configures a first filter 118A in the optical path of the illumination light L1, the first illumination light L1A with a first wavelength λ(A) passing through the first filter 118A illuminates the cell population 2. When the filter switching unit 118 configures a second filter 118B in the optical path of the illumination light L1, the second illumination light L1B with a second wavelength λ(B) passing through the second filter 118B illuminates the cell population 2. The filter switching unit 118 switches the position of the first filter 118A and the position of the second filter 118B so that either the first filter 118A or the second filter 118B is configured in the optical path of the illumination light L1 between the light source 113C and the cell population 2. Thus, the filter switching unit 118 selectively illuminates the cell population 2 with the first illumination light L1A and the second illumination light L1B.

[0169] When the first irradiation light L1A irradiates the cell group 2, a portion of the first irradiation light L1A passes through the cell group 2 and is emitted from the cell group 2 as the first emitted light L2A. When the second irradiation light L1B irradiates the cell group 2, a portion of the second irradiation light L1B passes through the cell group 2 and is emitted from the cell group 2 as the second emitted light L2B. The light receiving unit 121 selectively receives the first emitted light L2A and the second emitted light L2B emitted from the cell group 2. The light receiving unit 121 outputs a first electrical signal SA corresponding to the intensity of the first emitted light L2A and a second electrical signal SB corresponding to the intensity of the second emitted light L2B to the analysis unit 130.

[0170] exist Figure 21 In the observation device 101F shown, the "cell number" and "cell density" can also be quantitatively determined for each irradiated area R of the cell population 2 based on the intensity of the first emitted light L2A and the intensity of the second emitted light L2B, thus achieving the same effect as in the second embodiment. As with the observation device 101F, by using a light source 113C, which is an inexpensive lamp light source, the cost of the observation device 101F can be reduced compared to using an expensive laser light source.

[0171] This invention is not limited to the embodiments and modifications described above, and various other modifications are possible. For example, the embodiments and modifications described above can be combined with each other without contradiction, depending on the desired purpose and effect. In the embodiments and modifications described above, the case of containing a single cell population in a container has been explained, but multiple cell populations can also be contained in the container. The cell population can be suspended in the culture medium. In the embodiments and modifications described above, the case of irradiation module being arranged above the cell population and light-receiving module being arranged below the cell population has been mainly explained, but the irradiation module and light-receiving module can also be interchanged. That is, the irradiation module can be arranged below the cell population, and the light-receiving module can be arranged above the cell population.

[0172] In the various embodiments and modifications described above, the case where the irradiation light irradiating the cell population includes at least one wavelength within the range of 650 nm to 850 nm and 1060 nm to 1090 nm has been explained. However, it is also possible for the irradiation light to have all wavelengths within the range of 650 nm to 850 nm and 1060 nm to 1090 nm. That is, the irradiation light may only have wavelengths within the range of 650 nm to 850 nm and 1060 nm to 1090 nm. In this case, the reduction in measurement accuracy caused by the presence of wavelengths outside the range of 650 nm to 850 nm and 1060 nm to 1090 nm mixed in with the wavelengths of the irradiation light can be avoided. The analysis unit may also process only the information (irradiation light intensity or emitted light intensity) obtained from wavelengths within the range of 650 nm to 850 nm and 1060 nm to 1090 nm.

[0173] Explanation of reference numerals in the attached figures

[0174] 1, 1A, 1B, 1C, 1D, 101, 101A, 101B, 101C, 101D, 101E, 101F: Observation devices;

[0175] 2: Cell population;

[0176] 5: Container;

[0177] 5a: Bottom surface;

[0178] 5b: Open;

[0179] 7: Culture medium;

[0180] 10, 10A, 10B, 110, 110A, 110B: Irradiation modules;

[0181] 11, 11A, 111, 111A, 111B, 111C: Irradiation section;

[0182] 13: Light source (laser light source);

[0183] 15, 25, 115, 125: Lenses;

[0184] 20, 20A, 20B, 120, 120A, 120B: Light receiving modules;

[0185] 21, 21A, 121, 121A, 121B: Light-receiving parts;

[0186] 23, 123: Light sensor;

[0187] 30, 130: Analysis section;

[0188] 31, 131: Acquisition Department;

[0189] 33, 133: Storage section;

[0190] 35, 135: Transformer;

[0191] 41, 141: Circulator;

[0192] 113A: Primary light source;

[0193] 113B: Secondary light source;

[0194] 113C: Light source;

[0195] 114: Light source section;

[0196] 117: Switching unit;

[0197] 118: Filter switching unit;

[0198] 118A: First filter;

[0199] 118B: Second filter;

[0200] 119: Combined wave department;

[0201] 123A: First optical sensor;

[0202] 123B: Second optical sensor;

[0203] 129: Spectrometer;

[0204] 301: Processor;

[0205] 302: Main storage device;

[0206] 303: Auxiliary storage device;

[0207] 304: Input device;

[0208] 305: Output device;

[0209] A1: Up and down direction;

[0210] A2: Left and right directions;

[0211] D1: Light intensity information;

[0212] D2: Transformation information;

[0213] D11: First light intensity information;

[0214] D12: Second light intensity information;

[0215] D21: First conversion information;

[0216] D22: Second conversion information;

[0217] G1: Standard curve;

[0218] G11: First standard curve;

[0219] G12: Second standard curve;

[0220] L1: Illumination light;

[0221] L1A: First illumination light;

[0222] L1B: Second illumination light;

[0223] L2: Outgoing beam;

[0224] L2A: First emitted beam;

[0225] L2B: Second outgoing beam;

[0226] L3: Combined illumination light;

[0227] L4: Combined output beam;

[0228] P1, P2, P3: Ports;

[0229] P(A): First change in light intensity;

[0230] P(B): The second change in light intensity;

[0231] Pr1: Rayleigh scattering coefficient;

[0232] Pr2, Pr3: Reflection intensity;

[0233] R: Irradiated area;

[0234] R1, R2, R11, R21, R22: Wavelength range;

[0235] λ: wavelength;

[0236] λ(A): First wavelength;

[0237] λ(B): Second wavelength;

[0238] S: Electrical signal;

[0239] SA: First electrical signal;

[0240] SB: Second electrical signal.

Claims

1. An observation device for observing the state of a cell population consisting of multiple cells, said observation device having: At least one irradiation unit is disposed at a position opposite to the cell group and irradiates at least one irradiation area set in the cell group with irradiation light having at least one wavelength including the range of 650 nm or more and 850 nm or less and 1060 nm or more and 1090 nm or less. At least one light-receiving portion, disposed opposite to the cell group, receives outgoing light emitted from the cell group by irradiating the irradiated area with the irradiated light; and The analysis unit, which is communicatively connected to the light-receiving unit, determines, based on the intensity of the emitted light, at least one of the cell number and cell density of the cell population in the irradiated area.

2. The observation device according to claim 1, wherein, The irradiation unit includes a light source using a semiconductor light-emitting element, which emits light having a wavelength in the range of 650 nm to 850 nm and 1060 nm to 1090 nm as the irradiation light.

3. The observation device according to claim 1 or 2, wherein, The light-receiving part is positioned on the opposite side of the irradiation part, separated from the cell group, and receives a portion of the irradiation light emitted from the irradiation part and passing through the cell group as the outgoing light.

4. The observation device according to any one of claims 1 to 3, wherein, The irradiation unit includes one or more light sources that emit the irradiation light, irradiating each of the plurality of irradiation regions located in the cell population with the irradiation light. The light-receiving part includes one or more photosensors that receive the incident light from the emitted light, and receives the plurality of illumination lights passing through the cell group as the plurality of emitted lights.

5. The observation device according to any one of claims 1 to 4, wherein, The irradiation unit irradiates the irradiated area of ​​the cell population with a first irradiation light and a second irradiation light. The first irradiation light has a first wavelength that includes a range of 650 nm to 850 nm and 1060 nm to 1090 nm. The second irradiation light has a second wavelength that includes a range of 650 nm to 850 nm and 1060 nm to 1090 nm and is different from the first wavelength. The light-receiving part receives first and second emitted light, which have different wavelengths from each other, emitted from the cell group by the first and second irradiation lights. The analysis unit calculates the number of cells and the cell density in the irradiated area based on the intensity of the first emitted light and the intensity of the second emitted light, respectively.

6. The observation device according to claim 5, wherein, The irradiation unit includes: A first light source, which emits the first illumination light; A second light source, which emits the second illumination light; and The switching unit switches the optical path of the first irradiation light and the optical path of the second irradiation light, so that the first irradiation light and the second irradiation light selectively irradiate the irradiated area of ​​the cell population. The light-receiving part includes a light sensor that selectively receives the first emitted light and the second emitted light emitted from the cell group.

7. The observation device according to claim 5, wherein, The irradiation unit includes: A first light source, which emits the first illumination light; A second light source, which emits the second illumination light; and The beam combiner combines the first and second irradiation lights to generate a combined irradiation light, which then irradiates the irradiated area of ​​the cell population. The light-receiving part includes: The beam splitter separates the combined outgoing light emitted from the cell group by the combined irradiation light according to wavelength. A first optical sensor detects the intensity of the first emitted light split from the combined emitted light; and The second optical sensor detects the intensity of the second emitted light split from the combined emitted light.

8. The observation device according to claim 5, wherein, The irradiation unit includes: A light source that emits light comprising the first illumination light and the second illumination light; and A filter switching unit includes a first filter that transmits only the first irradiation light and a second filter that transmits only the second irradiation light, and switches the positions of the first filter and the second filter so that either the first filter or the second filter is positioned in the optical path of the light between the light source and the cell group. The light-receiving part includes a light sensor that selectively receives the first emitted light and the second emitted light emitted from the cell group.

9. An observation method for observing the state of a cell population consisting of multiple cells, the observation method comprising: The step of irradiating at least one irradiation region set in the cell population with irradiation light having at least one wavelength in the range of 650 nm or more and 850 nm or less and 1060 nm or more and 1090 nm or less; The steps of receiving emitted light from the cell population by irradiating the irradiated area with the irradiated light; and The step of determining, based on the intensity of the emitted light, a parameter including at least one of the number of cells and the cell density of the cell population in the irradiated area.

10. The observation method according to claim 9, wherein, In the step of irradiating the irradiated area The irradiated area is irradiated with a first irradiation light and a second irradiation light. The first irradiation light has a first wavelength that includes a range of 650 nm to 850 nm and 1060 nm to 1090 nm. The second irradiation light has a second wavelength that includes a range of 650 nm to 850 nm and 1060 nm to 1090 nm and is different from the first wavelength. In the step of receiving the emitted light Receives first and second emitted light rays, having different wavelengths from each other, emitted from the cell population after being irradiated by the first and second irradiation lights. In the step of determining the parameters... Based on the intensity of the first emitted light and the intensity of the second emitted light, the number of cells and the cell density in the irradiated area are calculated respectively.

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