Spectroscopic measurement device and spectroscopic measurement method

Abstract

A spectrometry device (1) according to the present embodiment comprises: a light source (11) that generates illumination light; a spectrometer (50) that disperses signal light from a sample illuminated by the illumination light, and performs detection via an optical detector (55); a fiber unit (40) that includes multiple fibers disposed within an optical path from the sample S to the spectrometer, such that on an incident end face (411) of the fiber unit (40), a plurality of fibers (45) are arranged adjacent to each other, while on an exit end face of the fiber unit, a plurality of the fibers (45) are arranged in a multi-line shape at intervals from each other; and a processing unit that, with reference to the positional relationships of the plurality of fibers (45) on the incident end face (411) and on the exit end face (421) of the fiber unit (40), generates a spectral image of the sample S from the detection results of the optical detector (55).

Description

[Technical Field]

[0001] The present invention relates to a spectroscopic measurement device and a spectroscopic measurement method, and more particularly to a spectroscopic measurement device and a spectroscopic measurement method that perform spectroscopic measurement of signal light such as Raman scattered light generated in a sample to generate a spectroscopic image. [Background technology]

[0002] Patent Document 1 and Non-Patent Documents 1 to 3 disclose methods for measuring Raman spectra. Patent Document 1 discloses a multifocal confocal Raman microscope using a fiber bundle. Light from a pinhole array is incident on the entrance end of the fiber bundle. In Non-Patent Document 1, the fiber bundle is placed in front of the spectroscope. Also, in Non-Patent Document 1, the fibers are lined up in a row at the exit end.

[0003] In Non-Patent Document 2, multi-focus is generated by modulating illumination light with a liquid crystal spatial light modulator. In Non-Patent Document 3, a multi-focus confocal Raman spectroscopic microscope is disclosed. In Non-Patent Document 3, multi-focus is generated using a microlens array. Furthermore, in Non-Patent Document 3, multi-fiber is used. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] Japanese Patent Application Laid-Open No. 2012-237637 [Non-patent literature]

[0005] [Non-Patent Document 1] “High-resolution confocal Raman microscopy using pixel reassignment” Roider, et al., Optics Letters Vol. 41, Issue 16, pp. 3825-3828 (2016) [Non-patent document 2] “Tissue diagnosis using power-sharing multifocal Raman micro-spectroscopy and auto-fluorescence imaging” Sinjab, et al., Biomed. Opt. Express 7, 2993 (2016). [Non-patent document 3] “Rapid and accurate peripheral nerve imaging by multipoint Raman spectroscopy” Kumamoto, et al., Sci. Rep. 7, 845 (2017). [Non-patent document 4] Internet search: https: / / www.tem-inc.co.jp / products / detail-29.php [Retrieved December 8, 2021] Summary of the Invention

[0006] When performing spectroscopic measurements of weak signal light such as Raman scattered light, the measurement time is long. In particular, when performing imaging, multiple spatial coordinate positions must be measured sequentially, which takes a long time. However, shortening the measurement time per coordinate results in the influence of noise. Therefore, there is a demand for measuring appropriate spectroscopic images in a short time.

[0007] The present disclosure has been made in consideration of the above points, and has an object to provide a spectroscopic measurement device and a spectroscopic measurement method that can generate an appropriate spectroscopic image in a short measurement time.

[0008] The spectroscopic measurement device of this embodiment includes a light source that generates illumination light, a spectrometer that disperses signal light from a sample illuminated with the illumination light and detects it with a two-dimensional array photodetector, a fiber unit having a plurality of fibers arranged in the optical path from the sample to the spectrometer, wherein the plurality of fibers are arranged adjacent to each other at the incident end face of the fiber unit and the plurality of fibers are arranged in a spaced-apart multi-line pattern at the exit end face, and a processing unit that generates a spectral image of the sample from the detection results of the two-dimensional array photodetector by referring to the arrangement relationship of the plurality of fibers at the incident end face and the exit end face of the fiber unit.

[0009] The above spectroscopic measurement device may further include a spatial light modulator that modulates illumination light from the light source so as to selectively illuminate multiple points on the sample, and signal light from the multiple points may be incident on any one of the fibers of the fiber unit.

[0010] The above spectroscopic measurement device may further include a camera that captures an optical image of the sample, and a plurality of points on the sample extracted based on the optical image may be selectively illuminated.

[0011] In the above spectroscopic measurement device, the plurality of fibers may be arranged in a hexagonal close-packed array at the incident end face of the fiber unit.

[0012] In the above spectroscopic measurement device, the incident end face of the fiber unit may be arranged at a position conjugate with the sample.

[0013] In the above spectroscopic measurement device, the signal lights emitted from the different fibers may be detected so as not to overlap on the light receiving surface of the two-dimensional array photodetector.

[0014] The spectroscopic measurement method according to this embodiment includes the steps of illuminating a sample with illumination light from a light source, inputting signal light from the sample into a fiber unit from an entrance end face where a plurality of fibers are arranged adjacent to each other, outputting the signal light from an exit end face of the fiber unit where the plurality of fibers are arranged in a spaced multi-line configuration, spectrally splitting the signal light output from the exit end face and detecting it with a two-dimensional array photodetector, and generating a spectral image of the sample from the detection results of the two-dimensional array photodetector by referring to the arrangement relationship of the plurality of fibers at the entrance end face and the exit end face of the fiber unit.

[0015] In the above spectroscopic measurement method, a spatial light modulator may modulate the illumination light from the light source so as to selectively illuminate multiple points on the sample, and signal light from the multiple points may be incident on any one of the fibers of the fiber unit.

[0016] In the above spectroscopic measurement method, an optical image of the sample may be captured by a camera, and a plurality of points on the sample extracted based on the optical image may be selectively illuminated.

[0017] In the above spectroscopic measurement method, the plurality of fibers may be arranged in a hexagonal close-packed array at the incident end face of the fiber unit.

[0018] In the above spectroscopic measurement method, an incident end face of the fiber unit may be arranged at a position conjugate with the sample.

[0019] In the above spectroscopic measurement method, the signal lights emitted from the different fibers may be detected so as not to overlap on the light receiving surface of the two-dimensional array photodetector.

[0020] According to the present invention, it is possible to provide a spectroscopic measurement device and a spectroscopic measurement method that can generate an appropriate spectroscopic image in a short measurement time. [Brief explanation of the drawings]

[0021] [Figure 1] FIG. 1 is a schematic diagram showing a spectroscopic measurement device according to a first embodiment. [Figure 2] 1A and 1B are diagrams showing fiber arrangements at the input end and output end of a fiber unit. [Figure 3] FIG. 2 is a diagram showing an image of the light receiving surface of a photodetector. [Figure 4] FIG. 2 is a diagram for explaining an ROI (Region Of Interest) extraction process. [Figure 5] FIG. 10 is a diagram for explaining spectroscopic measurement of signal light from an ROI. [Figure 6] 10A and 10B are diagrams showing images of the input end and output end of the fiber unit 40. FIG. [Figure 7] FIG. 1 is a diagram showing the results of spectroscopic measurement in Example 1. [Figure 8] FIG. 10 is a diagram showing the results of spectroscopic measurement in Example 2. [Figure 9] FIG. 10 is a diagram showing the results of spectroscopic measurement in Example 3. [Figure 10] FIG. 10 is a diagram showing the results of spectroscopic measurement in Example 4. DETAILED DESCRIPTION OF THE INVENTION

[0022] Below, embodiments to which the present invention can be applied are described. The following description is for describing embodiments of the present invention, and the present invention is not limited to the following embodiments. For clarity of explanation, the following description has been omitted and simplified as appropriate. Furthermore, a person skilled in the art would be able to easily modify, add, or convert each element of the following embodiments within the scope of the present invention. Note that elements with the same reference numerals in each drawing indicate similar elements, and descriptions thereof will be omitted as appropriate.

[0023] Embodiment 1 A spectroscopic measurement device and a measurement method thereof according to a first embodiment will be described with reference to Fig. 1. Fig. 1 is a schematic diagram showing an optical system of the spectroscopic measurement device 1. The spectroscopic measurement device 1 is a spectroscopic microscope that captures a spectroscopic image of a sample S. More specifically, the spectroscopic measurement device 1 measures spectral data of Raman scattered light from the sample S.

[0024] The spectroscopic measurement device 1 includes a spectroscopic illumination optical system 10, an observation illumination light source 18, an observation optical system 20, a spectroscopic measurement optical system 30, a fiber unit 40, a spectroscope 50, and a processing device 60. First, the spectroscopic illumination optical system 10 will be described. The spectroscopic illumination optical system 10 is an optical system for guiding illumination light L1 to a sample S. The spectroscopic illumination optical system 10 includes a light source 11, a spatial light modulator 12, a dichroic mirror 13, a dichroic mirror 14, and a lens 15.

[0025] The light source 11 is a laser light source that generates illumination light for spectroscopic measurement. The illumination light for spectroscopic measurement is simply referred to as illumination light L1. The light source 11 is a DPSS (Diode Pumped Solid State) laser that emits CW (Continuous Wave) light with a wavelength of 660 nm. Of course, the type and laser wavelength of the light source 11 are not particularly limited. In spectroscopic measurement, the illumination light L1 serves as excitation light that excites the sample S. Therefore, the light source 11 generates monochromatic illumination light L1.

[0026] Illumination light L1 from the light source 11 is incident on the spatial light modulator 12. The spatial light modulator 12 spatially modulates the illumination light L1 based on a control signal from the processing device 60. The spatial light modulator 12 controls the spatial distribution of the illumination light L1 on the sample S. This allows the area on the sample S onto which the illumination light L1 is incident to have a desired shape and size. For example, the spatial light modulator 12 modulates the illumination light L1 so as to illuminate only the ROI of the sample.

[0027] The spatial light modulator 12 is a liquid crystal device such as LCOS (Liquid Crystal on Silicon). The spatial light modulator 12 is a liquid crystal panel having pixels arranged in an array. The phase of reflected light can be modulated by controlling the voltage applied to each pixel. Of course, the spatial light modulator 12 is not limited to a reflective liquid crystal device such as LCOS, but may also be a transmissive liquid crystal device. Furthermore, the spatial light modulator 12 is not limited to a liquid crystal device, and a DMD or the like may also be used.

[0028] The illumination light L1 from the spatial light modulator 12 is incident on the dichroic mirror 13. The dichroic mirror 13 splits the light according to wavelength. The dichroic mirror 13 reflects the wavelength of the illumination light L1. Therefore, the dichroic mirror 13 reflects the illumination light L1 toward the dichroic mirror 14.

[0029] The dichroic mirror 14 splits the light according to its wavelength. The dichroic mirror 14 transmits the wavelength of the illumination light L1. Therefore, the illumination light L1 that has transmitted through the dichroic mirror 14 is incident on the lens 15. The lens 15 is an objective lens that focuses the illumination light L1 onto the sample S. This illuminates the sample S with the illumination light L1. Furthermore, the illumination light L1 is modulated by the spatial light modulator 12. Therefore, the illumination light L1 can illuminate a desired area on the sample S. The sample S is placed on a stage or the like (not shown). The stage may be a movable stage in order to change the illumination position of the sample S.

[0030] Next, the observation illumination light source 18 and the observation optical system 20 will be described. The observation illumination light source 18 is, for example, a lamp light source, and generates white observation illumination light L4. The observation illumination light L4 from the observation illumination light source 18 illuminates the sample S. Of course, the observation illumination light source 18 is not limited to a white lamp light source. A light source that is not monochromatic can also be used as the observation illumination light source 18.

[0031] The observation optical system 20 is an optical system for guiding observation light L2 from the sample S to the camera 23. The observation light L2 is light from an area illuminated with observation illumination light L4. For example, the observation light L2 is scattered light scattered by the sample S, reflected light reflected by the sample S, or fluorescence generated by the sample S. The observation optical system 20 includes a lens 15, a dichroic mirror 14, a dichroic mirror 13, a filter 21, and a lens 22.

[0032] Observation light L2 from the sample S is refracted by lens 15 and enters dichroic mirror 14. Observation light L2 transmitted through dichroic mirror 14 enters dichroic mirror 13. Observation light L2 transmitted through dichroic mirror 13 enters filter 21.

[0033] The filter 21 is an optical filter that transmits a portion of the light from the sample S. Specifically, the filter 21 is a wavelength filter that transmits or blocks light depending on the wavelength. This allows the camera 23, which will be described later, to detect only the observation light L2 of a desired wavelength. In other words, the light of the wavelength that transmits through the dichroic mirror 14, the dichroic mirror 13, and the filter 21 becomes the observation light L2.

[0034] The observation light L2 transmitted through the filter 21 is incident on the lens 22. The lens 22 is an imaging lens, and forms an image of the sample S on the light receiving surface of the camera 23. The camera 23 CCD The camera 23 is a two-dimensional photodetector such as a Charge Coupled Device (CMOS) camera or a Complementary Metal Oxide Semiconductor (CMOS) image sensor.

[0035] The processing device 60 is an information processing device such as a personal computer. The camera 23 outputs imaging data of the optical image of the sample S to the processing device 60. The processing device 60 stores the imaging data of the optical image in a memory or the like. That is, the processing device 60 stores brightness data corresponding to the amount of light received by each pixel of the camera 23. The processing device 60 has a display or the like for displaying the optical image. A user can extract an ROI of the sample S by checking the optical image captured by the camera 23. For example, the user can select an ROI on the display screen of the processing device 60. This identifies the coordinates in the optical image.

[0036] The dichroic mirror 14 may be disposed so as to be insertable into or removable from the optical path. The dichroic mirror 14 may be removed from the optical path when observing the optical image of the sample S. In other words, the dichroic mirror 14 may be inserted into the optical path when performing spectroscopic measurement.

[0037] Next, we will explain the spectroscopic measurement optical system 30. The spectroscopic measurement optical system 30 is an optical system from the sample S to the photodetector 55 of the spectrometer 50. In other words, the spectroscopic measurement optical system 30 guides the signal light L3 generated in the sample S to the photodetector 55. The spectroscopic measurement optical system 30 includes a lens 15, a dichroic mirror 14, a filter 31, a lens 32, a fiber unit 40, and a spectrometer 50.

[0038] The signal light L3 generated in the sample S is incident on the lens 15. The signal light L3 refracted by the lens 15 is incident on the dichroic mirror 14. The dichroic mirror 14 transmits light of the laser wavelength. The dichroic mirror 14 reflects the signal light L3, which has a wavelength different from that of the illumination light L1, in the direction of the filter 31.

[0039] The signal light L3 reflected by the dichroic mirror 14 is incident on the filter 31. The filter 31 is an optical filter that transmits a portion of the light from the sample S. Specifically, the filter 31 is a wavelength filter that transmits or blocks light depending on the wavelength. Specifically, the filter 31 blocks light of the laser wavelength of the light source 11 and transmits light of a predetermined wavelength band. This allows the spectrometer 50, which will be described later, to perform spectroscopic measurement of the signal light L3 having a wavelength different from the laser wavelength.

[0040] The signal light L3 transmitted through the filter 31 is incident on the lens 32. The lens 32 is an imaging lens, and forms an image of the sample S on the incident end of the fiber unit 40. In other words, the incident end face of the fiber unit 40 is positioned conjugate with the sample S. The fiber unit 40 has multiple fibers, and guides the incident signal light L3 to the spectrometer 50. The fiber unit 40 includes an incident side holder 41, an exit side holder 42, and a connection portion 43.

[0041] The fiber unit 40 is a bundle fiber in which multiple fibers are bundled together. The fiber unit 40 is arranged in the optical path from the sample S to the spectrometer 50. The fiber unit 40 is equipped with an incident side holder 41, an exit side holder 42, and a connection part 43, and the arrangement of the fibers is different at the incident end face and the exit end face of the fiber unit 40.

[0042] The incident side holder 41 is, for example, a cylindrical holder and houses a plurality of fibers therein. The incident side holder 41 fixes the plurality of fibers on the incident end face side of the fiber unit 40. Therefore, on the incident end side of the fiber unit 40, the plurality of fibers are arranged adjacent to each other. Specifically, the plurality of fibers are arranged in a closely packed array.

[0043] The output side holder 42 is, for example, a cylindrical holder and houses multiple fibers therein. The output side holder 42 fixes the multiple fibers on the output end face side of the fiber unit 40. On the output end side of the fiber unit 40, the multiple fibers are arranged in a multi-line configuration with spaces between them. This enables spectroscopic measurement using a multifocal optical system.

[0044] The connecting portion 43 connects the incident side holder 41 and the output side holder 42. The connecting portion 43 is a flexible portion disposed between the incident side holder 41 and the output side holder 42. The configuration of the fiber unit 40 will be described later.

[0045] The output end of the fiber unit 40 is disposed immediately before the spectrometer 50. The signal light L3 propagated through the fiber of the fiber unit 40 is output from the output end of the fiber unit 40 and enters the spectrometer 50. The signal light L3 from the sample S enters the spectrometer 50 via the fiber unit 40.

[0046] The spectrometer 50 includes a lens 51, a grating 52, a lens 54, and a photodetector 55. The signal light L3 from the fiber unit 40 is incident on the lens 51. The lens 51 refracts the signal light L3 to form a parallel beam. The signal light L3 from the lens 51 is incident on the grating 52. The grating 52 is a wavelength dispersion element that disperses the signal light L3 according to its wavelength. The grating 52 has a diffraction angle according to the wavelength. The direction in which the signal light L3 is dispersed by the grating 52 is defined as the X direction.

[0047] Here, the grating 52 is shown as a transmission type diffraction grating, and therefore the signal light L3 reflected by the grating 52 is incident on the lens 54. Note that the wavelength dispersion element is not limited to a transmission type diffraction grating, and may be a reflection type diffraction grating, a prism, or the like.

[0048] The signal light L3 dispersed by the grating 52 enters a lens 54. The lens 54 focuses the signal light L3 on the light receiving surface of a photodetector 55. The photodetector 55 is a two-dimensional array photodetector such as a CCD (Charge Coupled Device) camera or a CMOS (Complementary Metal Oxide Semiconductor) image sensor. The photodetector 55 has, for example, a plurality of pixels arranged along the X and Y directions. The X and Y directions are directions orthogonal to the optical axis of the optical system. In other words, the XY plane is a plane orthogonal to the optical axis.

[0049] When the wavelength dispersion direction is the X direction, the X coordinate of the photodetector 55 corresponds to the wavelength and spatial position of the signal light L3. An image of the output end of the fiber unit 40 is formed on the light receiving surface of the photodetector 55. The photodetector 55 can capture a spectral image of the sample S dispersed by the spectrometer 50.

[0050] Next, the configuration of the fiber unit 40 will be described with reference to Fig. 2. Fig. 2 is a diagram showing a schematic configuration of the fiber unit 40.

[0051] The fiber unit 40 has a plurality of fibers 45. An incident side holder 41 fixes the plurality of fibers 45 at an incident end surface 411 of the fiber unit 40. The plurality of fibers 45 are arranged adjacent to each other at the incident end surface 411 of the fiber unit 40. The incident ends of the fibers 45 are circular, and the plurality of fibers 45 are arranged so that each circle is in contact with the adjacent circle.

[0052] The incident end face 411 is positioned conjugate with the sample S. An image of the sample S is formed on the incident end face 411. In other words, the signal light L3 from one light irradiation spot on the sample is incident on one or more fibers 45. By arranging the fibers 45 close to each other, the gap between the fibers 45 can be reduced.

[0053] This makes it possible to reduce the area of ​​the sample S that cannot be spectroscopically measured. If the signal light L3 from the sample S enters the gaps between the fibers 45, the signal light L3 will not reach the spectrometer 50. By closely arranging the fibers 45, it is possible to reduce the area of ​​the gaps between the fibers 45. Therefore, it is possible to reduce the area of ​​the area that cannot be spectroscopically measured.

[0054] Non-Patent Document 4 (https: / / www.tem-inc.co.jp / products / detail-29.php) discloses a technique for numbering and mapping multiple fibers. Using this technique, it is possible to eliminate or reduce the gaps between fibers. By fusing the fibers, a honeycomb structure is created, eliminating the gaps between the fibers. Adjacent fibers may also be bonded with an adhesive such as epoxy resin.

[0055] More specifically, 1800 fibers 45 are arranged in a two-dimensional close-packed array at the incident end face 411. The multiple fibers 45 are arranged in a hexagonal close-packed array. Therefore, each fiber 45 is arranged so as to be in contact with six surrounding fibers 45. In other words, each of the multiple fibers 45 is in contact with six other fibers 45. Furthermore, the fibers 45 are not arranged in a matrix. In other words, the two arrangement directions at the incident end face 411 are not orthogonal to each other.

[0056] The incident side holder 41 holds the fibers 45 so that the fibers 45 are closely arranged on the incident end face 411 of the fiber unit 40. In other words, the incident side holder 41 fixes the fibers 45 so that the fibers 45 are arranged adjacent to each other. In this way, the spectroscopic measurement of the sample S can be performed more appropriately.

[0057] An image of the exit end face 421 is formed on the light receiving surface of the photodetector 55. A plurality of fibers 45 are arranged in a multi-line configuration at intervals on the exit end face 421. On the exit end face 421, the exit-side holder 42 holds the plurality of fibers 45 so that the plurality of fibers 45 are arranged at intervals from each other.

[0058] On the output end surface 421, a plurality of fibers 45 are arranged along the X direction and the Y direction. The plurality of fibers 45 are arranged in a two-dimensional matrix. The two arrangement directions on the output end surface 421 are not orthogonal. The plurality of fibers 45 are arranged at equal intervals in the X direction. Similarly, the plurality of fibers 45 are arranged at equal intervals in the Y direction. In this way, the output side holder 42 holds the plurality of fibers 45 so that the plurality of fibers 45 are arranged at regular intervals.

[0059] 1800 fibers 45 are arranged in a 15 x 120 matrix. That is, the fibers 45 are arranged in 15 lines, with each line containing 120 fibers 45. The 120 fibers 45 in each line are arranged parallel to the Y direction.

[0060] Specifically, on the incident end face 411, 1800 fibers 45 are arranged in a circular area with a diameter of about 2 to 3 cm. The diameter of one fiber 45 is about 45 μm. On the sample S, a circular area with a diameter of about 90 μm is detected by the photodetector 55. In other words, the circular area with a diameter of about 90 μm is enlarged and an image is formed on the incident end face 411 of the fiber unit 40. Therefore, the photodetector 55 simultaneously detects the signal light L3 from this circular area.

[0061] At the output end face 421, 1800 fibers 45 are arranged in a rectangular area of ​​about 14 to 15 cm in the X direction and about 6 to 7 cm in the Y direction. Incidentally, at the output end face 421, multiple slits corresponding to the multiple lines may be provided.

[0062] The output-side holder 42 fixes the plurality of fibers 45 so that the plurality of fibers 45 are arranged at intervals on the output end face 421. The density of the plurality of fibers 45 on the input end face 411 is higher than the density of the plurality of fibers 45 on the output end face 421. In other words, the fibers 45 are sparsely arranged on the output end face 421, and the fibers 45 are densely arranged on the input end face 411.

[0063] Specifically, it is preferable that the density in the area where the fibers 45 are arranged is 60% or more. 、 In the region where the fibers 45 are arranged, the fibers 45 are densely arranged so that 60% or more of the area is occupied by the fibers 45. In other words, it is preferable that the area of ​​gaps between the fibers 45 is less than 40%. Furthermore, it is more preferable that the density of the region where the fibers 45 are arranged is 75% or more. In this case, the fibers 45 are densely arranged so that 75% or more of the area of ​​the region where the fibers 45 are arranged is occupied by the fibers 45. It is preferable that the area of ​​gaps between the fibers 45 is less than 25%.

[0064] In this way, the signal light L3 emitted from adjacent fibers 45 is incident on the photodetector 55 without overlapping. In other words, the signal light L3 from different fibers 45 is detected by different pixels of the photodetector 55. In this way, the spectroscopic image can be measured appropriately.

[0065] As described above, the signal light L3 emitted from the fiber unit 40 is split by the spectrometer 50. Here, the splitting direction of the spectrometer 50 is defined as the X direction. The pixel address in the X direction in the photodetector 55 indicates the split wavelength and spatial position. The pixel address in the Y direction indicates the position in the multi-line.

[0066] The light receiving surface of the photodetector 55 will be described with reference to FIG. 3. FIG. 3 is a diagram showing an image on the light receiving surface of the photodetector 55. The pixels of the photodetector 55 are arranged in the X and Y directions. Furthermore, FIG. 3 schematically shows the fibers 45 included in one line. Here, the arrangement direction of the fibers 45 included in one line is parallel to the Y direction. The arrangement direction of multiple lines is parallel to the X direction. Therefore, the arrangement direction of the fibers 45 and the arrangement direction of the pixels are parallel. Furthermore, the spectral direction of the grating 52 is the X direction.

[0067] Therefore, the spectral data of one line of fibers 45 is measured in a band-shaped detection area on the light-receiving surface. In Fig. 3, the detection area where the spectral data of the fibers 45 included in the first line is measured is shown as detection area D1. Similarly, the detection areas where the spectral data of the fibers 45 in the second to fifteenth lines are measured are shown as detection areas D2 to D15, respectively. Because one line includes 120 fibers 45, spectral data for 120 fibers can be measured in one detection area D1.

[0068] The detection regions D1 to D15 are arranged so as not to overlap one another. For example, the detection region D1 and the detection region D2 are offset in the Y direction. Assume that the spectroscope 50 disperses the signal light on the long wavelength side to the +X side and the signal light L3 on the short wavelength side to the -X side. The pixel address corresponding to the longest wavelength in the detection region D1 is located on the -X side of the pixel address corresponding to the shortest wavelength in the detection region D2. Specifically, the filter 31 limits the wavelength range of the signal light L3 so that the detection regions D1 to D15 do not overlap. Furthermore, the spacing in the X direction of the fibers 45 at the output end face 421 is set to a predetermined value or more.

[0069] Furthermore, the fibers 45 are arranged at a distance from each other on the output end surface 421. For example, as shown in FIG. 3, the fibers 45 included in one line are arranged at intervals in the Y direction. Therefore, the signal light L3 from adjacent fibers 45 is incident on pixels at different Y direction addresses. The signal light L3 from each fiber 45 is detected at a different pixel. In other words, the signal light L3 from one fiber 45 does not enter a pixel into which the signal light L3 from another fiber 45 is incident.

[0070] With this design, the signal light L3 from each fiber 45 is detected without overlapping. Therefore, it is possible to measure the Raman spectra of multiple points on the sample S with a single shot (one frame) of the photodetector 55, without scanning the illumination light L1. This makes it possible to detect Raman scattered light from a two-dimensional area on a plane perpendicular to the optical axis.

[0071] Alternatively, the signal light from each fiber 45 may be detected so that it partially overlaps. In this case, the original spectrum can be restored using mathematical techniques such as compressed sensing. In this case, the signal light L3 from one fiber 45 can be wavelength-dispersed to more pixels. This allows the wavelength range that can be spectroscopically measured to be widened, or wavelength resolution to be improved.

[0072] The signal light L3 from each fiber 45 can be spectroscopically measured independently. This makes it possible to simultaneously spectroscopically measure the signal light L3 from 1800 points on the sample S. In other words, a two-dimensional spectral image with 1800 pixels can be captured in a short time. The signal light L3 from any one light irradiation spot (one location) on the sample S is incident on one fiber 45. Then, the spectrum of the signal light L3 emitted from one fiber 45 corresponds to the spectral data of one light irradiation spot (one location) on the sample S.

[0073] For example, the fiber unit 40 has 1800 fibers 45. Therefore, the spectroscopic measurement device 1 can detect 1800 Raman spectra. In other words, the spectroscopic measurement device 1 can perform spectroscopic measurement of the signal light L3 from 1800 points on the sample S. Then, the processing device 60 generates a spectral image of the sample S based on the detection signal of the photodetector 55. In other words, the spectral image is generated based on 1800 pieces of spectral data. The generation process of the spectral image by the processing device 60 will be described below.

[0074] As described above, the incident side holder 41 and the output side holder 42 are connected by the connection part 43. The connection part 43 bundles the plurality of fibers 45 so that the fibers 45 can be deformed. Furthermore, the correspondence between the position of each fiber 45 on the incident end face 411 and the position of each fiber 45 on the output end face 421 is known. For example, the position of one fiber 45a on the incident end face 411 corresponds to the position of the fiber 45a on the output end face 421. Similarly, the position of another fiber 45b on the incident end face 411 corresponds to the position of the fiber 45b on the output end face 421.

[0075] In this way, the position of the exit end face 421 of a fiber 45 located at any position on the incident end face 411 is known. The processing device 60 stores information indicating the arrangement of each fiber 45. In other words, the processing device 60 stores the correspondence between the position of the fiber 45 on the incident end face 411 and the position on the exit end face 421. The processing device 60 then generates a spectral image from the detection results of the photodetector 55 by referencing the arrangement of each fiber 45. The processing device 60 rearranges the detection data for each pixel of the photodetector 55 so that it matches the fiber arrangement on the incident end face 411. In this way, the processing device 60 constructs a two-dimensional spectral image by two-dimensional mapping.

[0076] In this way, the fibers 45 are arranged more densely at the incident end face 411 than at the exit end face 421. Therefore, the photodetector 55 can detect the signal light L3 from more points on the sample S. If the position on the sample corresponds to an area where the fibers 45 are closely arranged, the signal light propagates through one of the fibers 45 and is detected by the photodetector 55.

[0077] Furthermore, since the light beams are arranged in a multi-line pattern on the output end surface 421, the spectrometer 50 can simultaneously perform spectroscopic measurement of the signal light L3 from a plurality of fibers 45. In other words, the signal light L3 output from a fiber 45 is detected independently of the signal light L3 output from other fibers 45. This allows the spectrometer 50 to perform spectroscopic measurement of the signal light L3 from a greater number of fibers 45.

[0078] The spectroscopic measurement device 1 can capture two-dimensional spectroscopic images without scanning with laser light. This makes it possible to measure Raman spectroscopic images in a short time. Furthermore, since Raman spectra can be measured in a short time, measurement with low phototoxicity is possible. Labeling with fluorescent substances or the like is not required, making label-free spectroscopic measurement possible. Furthermore, the signal light L3 from each fiber 45 is separated and detected. This allows spectroscopic measurement with a high SN (Signal to Noise) ratio.

[0079] Next, the details of the illumination method using the spectroscopic illumination optical system 10 will be explained with reference to FIG. 4. First, before spectroscopic measurement, an optical image is captured by optical observation using the camera 23. In optical observation, observation illumination light L4 from the observation illumination light source 18 illuminates the entire field of view of the objective lens. Then, the user or the processing device 60 extracts multiple points on the sample as ROIs based on the optical image. For example, the processing device 60 displays the optical image on a monitor. While viewing the optical image on the monitor, the user specifies a region of interest using a mouse or the like. The processing device 60 stores the coordinates of the specified region. This extracts the ROI.

[0080] The observation illumination light L4 for capturing an optical image is light from a light source different from the illumination light L1 used during spectroscopic measurement. In other words, the observation illumination light source 18 and the light source 11 can be switched between when capturing an optical image and when performing spectroscopic measurement.

[0081] The spatial light modulator 12 controls the beam of illumination light L1 from the light source 11 so as to selectively illuminate the ROI on the sample S. The spatial light modulator 12 modulates the illumination light L1 so that the illumination light L1 is incident only on the ROI. In other words, the spatial light modulator 12 modulates the illumination light L1 so that the illumination light L1 is not irradiated on areas other than the ROI. This allows multiple points extracted as ROIs to be illuminated simultaneously.

[0082] The processing device 60 generates a binary image for controlling the spatial light modulator 12. In the binary image, for example, the position of the ROI is set to 1, and the position outside the ROI is set to 0. The processing device 60 outputs the control binary image to the spatial light modulator 12, which is an LCOS device. The spatial light modulator 12 has a plurality of control pixels and diffracts incident light. The spatial light modulator 12 can control the phase of the diffracted light for each pixel.

[0083] By using the spatial light modulator 12, it is possible to form multi-focus on any multiple points on the sample S. In other words, since the illumination light L1 is focused only on the ROI, it is possible to prevent Raman scattering light from occurring outside the ROI. This enables measurements with a high signal-to-noise ratio. Furthermore, since light is not irradiated outside the ROI, phototoxicity to the sample S can be reduced.

[0084] In Figure 4, the connective tissue and nerves of a biological sample are extracted as ROIs. Therefore, the spatial light modulator 12 modulates the illumination light L1 so that only the connective tissue and nerves are selectively illuminated. Signal light L3 from the ROI propagates to the spectrometer 50 via a fiber unit. The spectrometer 50 performs spectroscopic measurement of the signal light L3 from the area illuminated by the illumination light L1. The processing device 60 generates a spectroscopic image based on the spectral measurement results of the spectrometer 50.

[0085] The processing device 60 performs decoding processing of the spectral measurement results and the segment images. Therefore, the processing device 60 generates an image based on the segment images of the optical image in the region outside the ROI, and generates an image based on the spectral measurement results in the ROI. This allows a more appropriate spectral image to be generated and displayed. The spectral image contains spectral information in the ROI.

[0086] 5 is a diagram for explaining the fiber unit 40 when the illumination light L1 selectively illuminates only the ROI. In FIG. 5, an incident end face 411 and an exit end face 421 of the fiber unit 40 are shown. Furthermore, the light receiving surface of the photodetector 55 that detects the signal light L3 emitted from the fiber unit 40 is The face As shown

[0087] In FIG. 5, the fiber 45 corresponding to the ROI is shown as fiber 451, and the fiber 45 corresponding to the region other than the ROI is shown as fiber 452. An image of the ROI is formed on the incident end face 411. The position of the fiber 451 on the incident end face 411 corresponds to the ROI. Signal light L3 from the ROI is incident on the fiber 451. The region other than the ROI is not illuminated with the illumination light L1. Therefore, the signal light L3 is not generated in the region other than the ROI. Therefore, the signal light L3 is not incident on the fiber 452.

[0088] When the signal light L3 is emitted from the fiber 451 at the output end face 421, the spectrometer 50 disperses the signal light L3. On the light receiving surface of the photodetector 55, the signal light L3 is dispersed in the X direction. The signal light L3 from each fiber 451 is detected so as not to overlap on the light receiving surface. Therefore, the signal light L3 from each point on the sample S can be appropriately spectroscopically measured. Since light from areas other than the ROI can be reduced, noise light can be suppressed. Therefore, measurement with a high S / N ratio is possible. Therefore, an appropriate spectral image can be generated with a short measurement time.

[0089] 5, the arrangement direction of the multi-lines and the arrangement direction of the pixels are parallel, but they do not have to be parallel.Furthermore, the direction of light distribution and the arrangement direction of the pixels do not have to be parallel.

[0090] Fig. 6 is a diagram showing images of an incident end face 411 and an exit end face 421. At the incident end face 411, a plurality of fibers 45 are arranged closely together. The plurality of fibers 45 are arranged in a closely packed array. At the exit end face 421, the plurality of fibers 45 are arranged in a multi-line array. In Fig. 6, 15 lines M1 to M15 are formed at the exit end face 421.

[0091] The spectroscopic measurement method according to this embodiment includes the steps of illuminating a sample with illumination light from a light source, inputting signal light from the sample into a fiber unit from an entrance end face where a plurality of fibers are arranged adjacent to each other, outputting the signal light from an exit end face of the fiber unit where the plurality of fibers are arranged in a spaced multi-line configuration, spectrally splitting the signal light output from the exit end face and detecting it with a two-dimensional array photodetector, and generating a spectral image of the sample from the detection results of the two-dimensional array photodetector by referring to the arrangement relationship of the plurality of fibers at the entrance end face and the exit end face of the fiber unit.

[0092] Example 1 Example 1 will be described with reference to Fig. 7. Fig. 7 shows an optical image of sample S and the results of spectroscopic measurement of Raman scattered light. The results of spectroscopic measurement indicate the amount of light detected by each pixel of the photodetector 55. In Example 1, the sample S is polystyrene. Here, The spectroscopic illumination optical system 10 selectively illuminates laser spots A and B as ROIs. Then, signal light L3 from laser spots A and B enters the spectrometer 50 via the fiber unit 40. On the light-receiving surface of the photodetector 55, the signal light from laser spot A and laser spot B enter different pixels. Therefore, the respective Raman spectra can be measured by different pixels of the photodetector 55.

[0093] Example 2 Example 2 will be described with reference to FIG. 8. FIG. 8 shows an optical image of sample S and the results of spectroscopic measurement of Raman scattered light. In Example 1, sample S contains polystyrene and calcium carbonate (CaCO3). Spots 1 and 2 in FIG. 8 are ROIs.

[0094] Here, polystyrene and calcium carbonate are each extracted as ROIs. Specifically, spot 1 corresponds to polystyrene, and spot 2 corresponds to calcium carbonate. Signal light L3 from spot 1 and signal light L3 from spot 2 are incident on different pixels on the light-receiving surface. Therefore, the respective Raman spectra can be measured at different pixels of the photodetector 55. The peak wavelength of the Raman spectrum of polystyrene is different from the peak wavelength of the Raman spectrum of calcium carbonate.

[0095] Example 3 FIG. 9 shows an image illustrating ROI extraction and ROI illumination, in which the ROI is selectively illuminated. In Example 3, the sample S is polystyrene. The ROI is extracted from the optical image of the sample S. The ROI is then selectively illuminated. FIG. 9 shows an image of the incident end face when ROI is illuminated and the results of spectroscopic measurement. Signal light from multiple points on the sample S is detected at different pixels.

[0096] In the above embodiment, the signal light has been mainly described as being Raman scattered light, but the signal light may be light other than Raman scattered light. Therefore, the spectroscopic measurement device according to this embodiment may be a spectroscopic measurement device other than a Raman spectrometer. For example, it may be a spectroscopic measurement device that detects fluorescence excited by excitation light, or a spectroscopic measurement device that measures ultraviolet absorption spectra or near-infrared absorption spectra. These spectroscopic measurement devices can also measure spectra with a high signal-to-noise ratio. This is particularly suitable for spectroscopic measurement devices that require high-speed measurement or repeated measurements.

[0097] The sample S to be measured can be a biological sample or a medical sample. Samples for pathological diagnosis, drug discovery samples, food, cosmetics, advanced materials, devices, etc. can also be used as the sample S. The results of spectroscopic measurement can be used to contribute to quality control. Furthermore, in the field Science It is also possible to use various samples (space, ocean, biology, etc.) as the sample S. By combining a spectroscopic measurement device with an endoscope, it becomes possible to inspect the inside of an object. The endoscope may be a rigid or flexible endoscope, and may be a medical or industrial endoscope.

[0098] Example 4 FIG. 10 shows the measurement results of a biological sample. In this example, an optical image of adipose tissue as a sample and the spectroscopic measurement results are shown. An ROI is extracted from the optical image. In FIG. 10, the ROI is shown as a laser irradiation spot. Then, the ROI is selectively irradiated with laser light, which serves as excitation light.

[0099] Furthermore, part or all of the processing in the processing device 60 described above can be implemented as a computer program. Such a program can be stored in and supplied to a computer using various types of non-transitory computer-readable media. Non-transitory computer-readable media include various types of tangible recording media. Examples of non-transitory computer-readable media include magnetic recording media (e.g., flexible disks, magnetic tapes, hard disk drives), magneto-optical recording media (e.g., magneto-optical disks), CD-ROMs (Read Only Memory), CD-Rs, CD-R / Ws, and semiconductor memories (e.g., mask ROMs, PROMs (Programmable ROMs), EPROMs (Erasable PROMs), flash ROMs, and RAMs (Random Access Memory)). The program may also be supplied to a computer by various types of temporary computer-readable media. Examples of temporary computer-readable media include electrical signals, optical signals, and electromagnetic waves. The temporary computer-readable media can be supplied to a computer via wired communication paths such as electric wires and optical fibers, or via wireless communication paths.

[0100] The invention made by the present inventor has been described above based on the embodiments. specific However, the present invention is not limited to the above-described embodiment, and it goes without saying that various modifications can be made without departing from the spirit of the present invention.

[0101] This application claims priority based on Japanese Patent Application No. 2022-9137, filed on January 25, 2022, the disclosure of which is incorporated herein in its entirety. [Explanation of symbols]

[0102] 1 Spectrometer 10 Spectral illumination optical system 11 Light source 12 Spatial Light Modulator 13 Dichroic mirror 14 Dichroic mirror 15 Lenses 18 Observation illumination light source 20 Observation optical system 21 Filters 22 Lens 23 Camera 30 Spectroscopic measurement optical system 31 filters 32 Lens 40 Fiber Unit 41 Incident side holder 42 Output side holder 45 Fiber 50 spectrometer 51 Lens 52 Grating 54 Lens 55 Photodetector 60 Processing equipment L1 illumination light L2 observation light L3 signal light

Claims

1. a light source that generates illumination light; a spectroscope that separates signal light from the sample illuminated with the illumination light and detects the separated signal light with a two-dimensional array photodetector; a fiber unit having a plurality of fibers arranged in an optical path from the sample to the spectrometer, the plurality of fibers being arranged adjacent to each other at an incident end face of the fiber unit and the plurality of fibers being arranged in a multi-line configuration with gaps at an exit end face; a processing unit that generates a spectral image of the sample from the detection result of the two-dimensional array photodetector by referring to the arrangement relationship of the plurality of fibers at the incident end face and the exit end face of the fiber unit; a spatial light modulator that modulates illumination light from the light source so as to selectively illuminate a plurality of points in a region of interest on the sample; The signal light from the plurality of points is incident on the fibers of the fiber unit, a spectroscopic measurement device, wherein the incident end face of the fiber unit is arranged at a position conjugate with the sample.

2. further comprising a camera for capturing an optical image of the sample; 2. The spectroscopic measurement device according to claim 1, wherein a plurality of points in the region of interest on the sample extracted based on the optical image are selectively illuminated.

3. 3. The spectrometer according to claim 1, wherein the plurality of fibers are arranged in a hexagonal close-packed array at the incident end face of the fiber unit.

4. 3. The spectroscopic measurement device according to claim 1, wherein the signal lights emitted from the different fibers are detected so as not to overlap on the light receiving surface of the two-dimensional array photodetector.

5. illuminating the sample with illumination light from a light source; a step of inputting signal light from the sample into a fiber unit from an incident end face on which a plurality of fibers are arranged adjacent to each other; a step of emitting light from an output end surface of the fiber unit in which the plurality of fibers are arranged in a multi-line configuration with intervals; a step of separating the signal light emitted from the emission end face and detecting the separated signal light with a two-dimensional array photodetector; generating a spectral image of the sample from the detection results of the two-dimensional array photodetector by referring to the arrangement relationship of the plurality of fibers at the incident end face and the exit end face of the fiber unit; a spatial light modulator modulating the illumination light from the light source to selectively illuminate a plurality of points in a region of interest on the sample; The signal light from the plurality of points is incident on the fibers of the fiber unit, an incident end face of the fiber unit is arranged at a position conjugate with the sample; Spectroscopic measurement method.

6. capturing an optical image of the sample with a camera; The spectroscopic measurement method according to claim 5 , wherein a plurality of points in the region of interest on the sample extracted based on the optical image are selectively illuminated.

7. 7. The spectroscopic measurement method according to claim 5, wherein the plurality of fibers are arranged in a hexagonal close-packed array at the incident end face of the fiber unit.

8. 7. The spectroscopic measurement method according to claim 5, wherein the signal lights emitted from the different fibers are detected so as not to overlap on the light receiving surface of the two-dimensional array photodetector.

Images