WO2022153736A1 - 粒子分析装置、粒子分析方法及び光学測定装置 - Google Patents

粒子分析装置、粒子分析方法及び光学測定装置 Download PDF

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Publication number
WO2022153736A1
WO2022153736A1 PCT/JP2021/045300 JP2021045300W WO2022153736A1 WO 2022153736 A1 WO2022153736 A1 WO 2022153736A1 JP 2021045300 W JP2021045300 W JP 2021045300W WO 2022153736 A1 WO2022153736 A1 WO 2022153736A1
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WIPO (PCT)
Prior art keywords
lens
lenses
excitation light
light
particle analyzer
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Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/JP2021/045300
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English (en)
French (fr)
Japanese (ja)
Inventor
丈司 畠山
聡史 長江
幹修 野中
謙次 石田
直道 菊地
隆史 加藤
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Sony Group Corp
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Sony Group Corp
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Filing date
Publication date
Application filed by Sony Group Corp filed Critical Sony Group Corp
Priority to US18/271,408 priority Critical patent/US12379304B2/en
Priority to JP2022575139A priority patent/JP7800447B2/ja
Priority to CN202180089609.9A priority patent/CN116670493A/zh
Publication of WO2022153736A1 publication Critical patent/WO2022153736A1/ja
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Optical investigation techniques, e.g. flow cytometry
    • G01N15/1434—Optical arrangements
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/02—Investigating particle size or size distribution
    • G01N15/0205—Investigating particle size or size distribution by optical means
    • G01N15/0227—Investigating particle size or size distribution by optical means using imaging; using holography
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Optical investigation techniques, e.g. flow cytometry
    • G01N15/1456—Optical investigation techniques, e.g. flow cytometry without spatial resolution of the texture or inner structure of the particle, e.g. processing of pulse signals
    • G01N15/1459—Optical investigation techniques, e.g. flow cytometry without spatial resolution of the texture or inner structure of the particle, e.g. processing of pulse signals the analysis being performed on a sample stream
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/01—Arrangements or apparatus for facilitating the optical investigation
    • G01N21/03—Cuvette constructions
    • G01N21/05—Flow-through cuvettes
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/02—Objectives
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Optical investigation techniques, e.g. flow cytometry
    • G01N15/149—Optical investigation techniques, e.g. flow cytometry specially adapted for sorting particles, e.g. by their size or optical properties
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N2015/1006—Investigating individual particles for cytology
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Optical investigation techniques, e.g. flow cytometry
    • G01N15/1434—Optical arrangements
    • G01N2015/144—Imaging characterised by its optical setup
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Optical investigation techniques, e.g. flow cytometry
    • G01N2015/1493—Particle size

Definitions

  • the present disclosure relates to a particle analyzer, a particle analysis method, and an optical measuring apparatus.
  • a flow cytometer irradiates particles flowing through a flow path formed in a flow cell or microchip with light, detects fluorescence or scattered light emitted from each particle, and executes analysis or the like. It is a device for performing optical measurement using cytometry.
  • Some flow cytometers have a function to separate and collect only particles having specific characteristics based on the analysis results, and a device that specifically targets cells is called a "cell sorter". ..
  • a droplet charging method for charging and separating droplets containing particles is mainly adopted (see, for example, Patent Document 1).
  • a droplet charging type device a fluid discharged from a flow cell, a microchip, etc. is atomized, a positive (+) or negative (-) charge is given to the droplet, and the traveling direction is determined by a deflection plate or the like. By changing it, it will be collected in the specified container.
  • An objective lens used in a general optical measuring device for fluorescence observation or the like is a lens structure formed by combining a plurality of lenses, and an adhesive is used for assembling the objective lens.
  • the optical characteristics of the objective lens deteriorate due to burning of the adhesive due to the strong laser light, or burning of the outgas emitted from the adhesive and adhering to the lens surface due to the excitation light. There was a problem that it could end up.
  • the generated fluorescence extends from the ultraviolet region to the near infrared region, so it is necessary to correct the chromatic aberration of the objective lens over a wide band.
  • the particle analyzer of one form according to the present disclosure focuses on a light source that emits excitation light including light having a wavelength of 400 nm or less and the excitation light at a predetermined position in the flow path.
  • a lens structure to be used a detection unit that detects the light emitted from the particles by being excited by the excitation light, and a processing unit that processes the detection data acquired by the detection unit.
  • the lens structure comprises a plurality of lenses arranged along the optical axis of the excitation light and a lens frame for holding the plurality of lenses, and at least one of the plurality of lenses is attached to the lens. The position in the lens frame is determined by abutting on the adjacent lens.
  • One form of the particle analysis method is to use a lens structure to collect excitation light containing light having a wavelength of 400 nm or less at a predetermined position in a flow path, and particles circulating at the predetermined position.
  • the lens structure comprises a plurality of lenses arranged along the optical axis of the excitation light, including detecting the light emitted from the particle by being excited by the excitation light and processing the detection data.
  • a lens frame for holding a plurality of lenses, and at least one of the plurality of lenses is in contact with a lens adjacent to the lens, so that the position in the lens frame is determined.
  • One form of the optical measuring device includes a light source that emits excitation light including light having a wavelength of 400 nm or less, a lens structure that concentrates the excitation light at a predetermined position in the flow path, and a predetermined position.
  • the lens structure includes a plurality of lenses arranged along the optical axis of the excitation light, and a plurality of lenses, comprising a detection unit for detecting the light emitted from the particles by being excited by the excitation light.
  • a lens frame for holding the lens is provided, and at least one of the plurality of lenses is in contact with a lens adjacent to the lens, so that the position in the lens frame is determined.
  • FIG. 1 It is a figure which shows the example of the schematic structure of the optical measuring apparatus which concerns on embodiment, and the particle analyzer which includes an optical measuring apparatus. It is a figure which shows the example of the schematic structure of the microchip. It is a figure which shows the example of the schematic structure of the lens structure of 1st Embodiment and 1st Example. It is a figure which shows the example of the schematic structure of the lens structure of 1st Embodiment and 1st Example. It is a figure which shows the example of the lens design of 1st Embodiment and 1st Embodiment. It is a figure which shows the example of the schematic structure of the lens structure of 1st Embodiment and 2nd Embodiment.
  • Embodiment 1.1 Example of the first design condition of the lens structure 1.2.
  • Lens structure of the first embodiment and the second embodiment 1.9.
  • the lens structure of the first embodiment and the fourth embodiment 1.1 1.
  • Modification example 3. Example of effect
  • the optical measuring device exemplified in the embodiment is an open cell sorter.
  • the disclosed techniques are also applicable to closed cell sorters.
  • the microchip method will be described as an example as a method of supplying particles (fine particles) to an observation point (hereinafter, also referred to as a “spot”) on the flow path.
  • various methods such as a droplet method, a cuvette method, and a flow cell method may be adopted.
  • the disclosed techniques include various optical measurements for measuring particles passing through a spot set on the flow path, such as an analyzer-type flow cytometer and a microscope for acquiring an image of particles on the flow path. Applicable to devices.
  • FIG. 1 is a diagram showing an example of a schematic configuration of an optical measuring device according to an embodiment and a particle analyzer including the optical measuring device.
  • the particle analyzer 100 includes an optical measuring device 110 and a processing control unit 120.
  • the particle analyzer 100 includes a light source 1, a mirror group 2, a mirror 3, an objective lens 4, a microchip 5, an imaging lens 6, a detection unit 7, and a detection unit 8.
  • the Z axis corresponding to the direction of the objective lens 4 is shown.
  • the Z-axis positive direction corresponds to the direction from the microchip 5 toward the mirror 3 (the traveling direction of the scattered light BSC or the like described later).
  • the negative Z-axis direction corresponds to the direction from the mirror 3 toward the microchip 5 (the traveling direction of the excitation light EL1 or the like described later).
  • the optical measuring device 110 measures a test object located at a predetermined position in the flow path in the microchip 5.
  • the particles 51 located at the spots 51s are measured as the test object.
  • the optical measuring device 110 irradiates the particles 51 with excitation light and detects the light emitted from the particles 51.
  • the excitation light irradiation will be described with reference to FIG. 1 (A), and the light detection will be described with reference to FIG. 1 (B).
  • the light source 1 emits excitation light, which is light for exciting the particles 51.
  • the excitation light may be coherent light such as a laser or a beam.
  • the excitation light may be pulsed light.
  • the light source 1 emits excitation light including light having a wavelength in the ultraviolet region.
  • the light in the ultraviolet region referred to here may be light having a wavelength of 400 nm or less, and more specifically, light having a wavelength of 350 nm or less.
  • the light source 1 emits excitation light according to the signal Sig3 from the processing control unit 120 described later. As a result, the excitation light is applied to the particles 51 at the timing when the particles 51 are located at the spots 51s.
  • the light source 1 may emit a plurality of excitation lights.
  • the light source 1 includes light sources 11 to 15.
  • the light source 11 emits excitation light having a wavelength of 349 nm.
  • the light source 12 outputs excitation light having a wavelength of 405 nm.
  • the light source 13 emits excitation light having a wavelength of 488 nm.
  • the light source 14 emits excitation light having a wavelength of 561 nm.
  • the light source 15 emits excitation light having a wavelength of 637 nm.
  • An example of the output level of the excitation light is several mW to several tens of mW.
  • excitation lights emitted by the light sources 11 to 15 are referred to as excitation lights EL1 to excitation light EL5 and are shown in the drawing.
  • excitation light EL1 to the excitation light EL5 may be collectively referred to as “excitation light EL1 or the like”.
  • the mirror group 2 guides the excitation light EL1 or the like from the light source 1 to the central portion 31 of the mirror 3, for example, by aligning (combining) their optical axes.
  • the mirror group 2 includes the mirrors 21 to 25.
  • the mirror 21 reflects the excitation light EL1 from the light source 11 toward the central portion 31 of the mirror 3.
  • the mirror 22 transmits the excitation light EL1 from the mirror 21 and reflects the excitation light EL2 from the light source 12 toward the central portion 31 of the mirror 3.
  • the mirror 23 transmits the excitation light EL1 and the excitation light EL2 from the mirror 22, and reflects the excitation light EL3 from the light source 13 toward the central portion 31 of the mirror 3.
  • the mirror 24 transmits the excitation light EL1 to the excitation light EL3 from the mirror 23, and reflects the excitation light EL4 from the light source 14 toward the central portion 31 of the mirror 3.
  • the mirror 25 transmits the excitation light EL1 to the excitation light EL4 from the mirror 24, and reflects the excitation light EL5 from the light source 15 toward the central portion 31 of the mirror 3.
  • the mirror 21 is, for example, a total reflection mirror.
  • the mirrors 22 to 25 are, for example, dichroic mirrors.
  • the mirror 3 is an area division mirror in which an area is divided into a central portion 31 and a peripheral portion 32.
  • the mirror 3 is an optical system that guides the excitation light EL1 or the like from the mirror group 2 incident on the central portion 31 to the objective lens 4, and reflects the excitation light EL1 or the like toward the objective lens 4.
  • the light source 1 more specifically, the mirror group 2
  • the objective lens 4 and the detection unit 7 (more specifically, so that the excitation light EL1 or the like from the mirror group 2 is incident at an incident angle of 45 °). More specifically, it is arranged between the imaging lens 6).
  • the objective lens 4 focuses the excitation light EL1 and the like from the mirror 3 on the spot 51s in the microchip 5. The details of the objective lens 4 will be described later with reference to FIGS. 3 and later.
  • the microchip 5 provides a test object to be measured by the optical measuring device 110.
  • the microchip 5 will be described with reference to FIG.
  • FIG. 2 is a diagram showing an example of a schematic configuration of a microchip.
  • the microchip 5 includes a flow path 53 through which the particles 51 flow.
  • the particle 51 is, in this example, a biological particle contained in the biological sample 52.
  • the flow path 53 is configured such that the biological sample 52 flows, in particular, the particles 51 flow in a substantially row.
  • the biological sample 52 and the flow path structure containing the biological sample 52 may be formed of a material such as plastic or glass.
  • the biological sample 52 is, for example, a cell or a non-cellular biological particle.
  • the cell may be a living cell, and more specific examples thereof include blood cells such as erythrocytes and leukocytes, and germ cells such as sperm and fertilized eggs.
  • the cells may be those directly collected from a sample such as whole blood, or may be cultured cells obtained after culturing.
  • non-cellular biological particles include extracellular vesicles, particularly exosomes and microvesicles.
  • the bioparticles may be labeled with one or more labeling substances (eg, dyes (particularly fluorescent dyes) and fluorescent dye labeled antibodies, etc.). Particles other than biological particles may be analyzed by an optical measuring device, and beads or the like may be analyzed for calibration or the like.
  • the particles 51 are irradiated with the excitation light EL1 or the like focused by the objective lens 4, and the particles 51 are excited. Fluorescence is generated due to the excitation of the particles 51. Further, scattered light is generated due to the excitation light EL1 and the like being scattered by the particles 51.
  • the fluorescence generated in the particle 51 the fluorescence that advances toward the objective lens 4 (toward the positive direction of the Z axis) is referred to as fluorescence FL and is illustrated.
  • the fluorescence advancing in the same direction as the excitation light EL1 is referred to as scattered light BSC (backward scattered light) and is shown in the drawing.
  • scattered light FSC forward scattered light
  • the objective lens 4 guides at least a part of the fluorescent FL and the scattered light BSC from the microchip 5 to the peripheral portion 32 of the mirror 3 while condensing them, for example.
  • the mirror 3 transmits the fluorescent FL and the scattered light BSC from the objective lens 4 incident on the peripheral portion 32 and guides them to the imaging lens 6.
  • the mirror 3 also has a function of reflecting the excitation light EL1 or the like from the mirror group 2 incident on the central portion 31 toward the objective lens 4.
  • the central portion 31 of the mirror 3 reflects light in a wavelength band such as excitation light EL1 and transmits light in another wavelength band with respect to, for example, 45 ° incident.
  • the central portion 31 is, for example, a portion where a dichroic mirror coat is formed.
  • the peripheral portion 32 of the mirror 3 constitutes, for example, a portion of the mirror 3 other than the central portion 31, and transmits fluorescent FL and scattered light BSC.
  • the peripheral portion 32 is, for example, a portion where an antireflection coating is formed.
  • the imaging lens 6 concentrates the fluorescent FL and the scattered light BSC transmitted through the mirror 3 on the detection unit 7. As described above, since the objective lens 4 can have a function of condensing the fluorescent FL and the scattered light BSC, the imaging lens 6 cooperates with the objective lens 4 to condense the fluorescent FL and the scattered light BSC. You can.
  • the detection unit 7 detects the fluorescent FL and the scattered light BSC from the imaging lens 6. For example, the detection unit 7 detects light having a wavelength other than the excitation light EL1 or the like in the wavelength band of 360 nm to 850 nm as a component of the fluorescent FL. As a component of the scattered light BSC, the detection unit 7 detects light having the same wavelength as the wavelength of a part of the excitation light (for example, the excitation light EL3 having a wavelength of 488 nm) such as the excitation light EL1 as the scattered light BSC.
  • the detection unit 7 detects light having a wavelength other than the excitation light EL1 or the like in the wavelength band of 360 nm to 850 nm as a component of the fluorescent FL.
  • the detection unit 7 detects light having the same wavelength as the wavelength of a part of the excitation light (for example, the excitation light EL3 having a wavelength of 488 nm) such as the excitation light EL1 as the scattered light BSC.
  • the detection unit 7 detects, for example, the amount of light (spectrum) for each wavelength.
  • the detection unit 7 may include a spectroscopic optical system that disperses incident light for each wavelength, a detector (two-dimensional image sensor, photodiode, etc.) that detects the amount of light for each wavelength, and the like.
  • the detection result acquired by the detection unit 7 is transmitted to the processing control unit 120 as detection data.
  • the signal of the detection data transmitted from the detection unit 7 to the processing control unit 120 is referred to as a signal Sigma1 and is shown in the figure.
  • the detection unit 8 detects the scattered light FSC from the microchip 5.
  • An example of the configuration of the detection unit 8 may be the same as the configuration of the detection unit 7 regarding the scattered light BSC.
  • the detection result of the detection unit 8 is transmitted to the processing control unit 120 as detection data.
  • the signal of the detection data transmitted from the detection unit 8 to the processing control unit 120 is referred to as a signal Sigma 2 and is shown in the figure.
  • the processing control unit 120 has a function as a processing unit that processes the detection data (signal Sig1) from the detection unit 7.
  • An example of the treatment is the analysis treatment of the particles 51.
  • the processing control unit 120 analyzes the component, size, and the like of the particles 51 by evaluating the spectrum of the fluorescent FL. The spectrum image, evaluation result, etc. may be presented to the user.
  • detecting the scattered light BSC it is desirable that the light having a small scattering angle is shielded by a mask or the like, and the light having a large scattering angle is transmitted and detected.
  • the processing control unit 120 also has a function as a control unit that controls the light source 1 based on the detection data (signal Sigma 2) from the detection unit 8. Specifically, the processing control unit 120 identifies the timing at which the next particle 51 passes through the spot 51s of the microchip 5 based on the detection data from the detection unit 8, and at that timing, the excitation light is applied to the next particle 51.
  • the light source 1 is controlled so that EL1 and the like are irradiated.
  • a signal (control signal) for controlling the light source 1 is generated as a signal Sigma 3 and transmitted from the processing control unit 120 to the light source 1. As described above, the light source 1 emits excitation light EL or the like according to the signal Sig3.
  • the objective lens 4 includes a plurality of lenses, but when an adhesive is used for assembling the objective lens 4, the outgas emitted from the adhesive and the adhesive by the laser beam is burnt and the outgas adhering to the lens surface is burnt, and the optical characteristics May deteriorate.
  • the wavelength of the excitation light becomes shorter, that is, when the excitation light having a wavelength in the ultraviolet region of 400 nm or less such as the excitation light EL1 is used, the above-mentioned problems become apparent.
  • Fluorescent FL also passes through the objective lens 4. Since the wavelength range of the fluorescent FL is also wide, for example, from the ultraviolet region to the near infrared region, it is necessary to correct the chromatic aberration of the objective lens 4 in a wide band. Many optical glasses used for lenses have poor transmittance characteristics in the ultraviolet region. In order to correct wideband chromatic aberration, it is necessary to design using a plurality of types of optical glass, but there is a problem that sufficient transmittance characteristics cannot be secured due to absorption of the glass itself.
  • the objective lens 4 is a lens structure designed to address at least a part of the above-mentioned problems.
  • the lens structure includes a plurality of lenses arranged (arranged) along the optical axis (along the Z-axis direction) of the excitation light EL1 and the like, and a lens frame for holding the plurality of lenses. Some examples of lens structure design conditions will be described.
  • the lens structure is designed to satisfy one or more design conditions.
  • the lens frame may hold a plurality of lenses without using an adhesive in the lens frame. At least one of the plurality of lenses may be positioned in the lens frame by abutting on a lens adjacent to the lens. Adjacent lenses form a junction division group.
  • a plurality of joint division groups may be configured, in which case the relative positions between the joint division surfaces are fixed without the use of an adhesive.
  • the inside of the lens frame has an adhesive-less structure, and it is possible to prevent the adhesive from burning and the outgas adhering to the lens surface released from the adhesive from burning. For example, it is possible to suppress deterioration of the optical characteristics of the lenses forming the joint division group.
  • the junction division group By forming the junction division group, it is possible to satisfactorily correct axial chromatic aberration while suppressing an increase in size and cost of the optical system. This effect becomes apparent as the number of junction division groups increases. It also increases the possibility of reducing costs by reducing the number of parts and simplifying the assembly process.
  • the lens structure may have a positive lens on the most object side (Z-axis negative direction side).
  • the glass material of the positive lens may satisfy the following (1) and (2).
  • ⁇ d is the Abbe number on the d line.
  • the transmittance in the ultraviolet region and correct chromatic aberration in a wide band For example, when nd is less than the lower limit of (1) above, the curvature of the positive lens becomes tight, the performance deteriorates, and manufacturing becomes difficult. When nd exceeds the upper limit of (1) above, the glass transmittance with respect to ultraviolet rays deteriorates, the intensity of the laser beam (excitation light EL1 or the like) applied to the particles 51 decreases, and problems such as autofluorescence occur. When ⁇ d is less than the lower limit of (2) above, the glass transmittance with respect to ultraviolet rays is deteriorated, the intensity of the laser beam applied to the particles 51 is lowered, and problems such as autofluorescence occur.
  • the lens structure may have at least three or more positive lenses. These positive lenses may be different from the positive lens on the most subject side (Z-axis negative direction side) under the above-mentioned second design condition.
  • the glass material of three or more positive lenses may satisfy the following (3) to (5). (3) nd ⁇ 1.52 (4) 79 ⁇ d (5) PgF ⁇ 0.54 PgF is a partial dispersion ratio of g-line and F-line.
  • the lens structure may have at least two or more negative lenses.
  • the glass material of these negative lenses may satisfy the following (6) and (7). (6) nd ⁇ 1.78 (7) 29 ⁇ d
  • a plurality of lenses may have a retrofocus configuration (reverse telephoto configuration).
  • the plurality of lenses are arranged in order from the side where the excitation light EL1 or the like is incident to the side where the excitation light EL1 or the like is emitted (toward the negative direction of the Z axis), and are a group of front lenses having a negative refractive power or substantially non-power.
  • a rear lens group having a positive refractive power may include having a negative refractive index or a positive refractive index that does not significantly affect the diffusion and light collection of light, design errors, and the like. ..
  • the outer shape of the lens constituting the front lens group may be substantially increased in the negative direction of the Z axis.
  • the substantially larger outer shape may include that the outer shape of the lens is the same as the outer shape of the adjacent lens.
  • the outer shape of the lens constituting the rear lens group may be substantially reduced in the negative direction of the Z axis.
  • Substantially smaller outer shape may include that the outer shape of the lens is the same as the outer shape of the adjacent lens.
  • the plurality of lenses may satisfy the following (8) and (9). (8) ⁇ 0.24 ⁇ ft / f1 ⁇ 0.08 (9) 0.35 ⁇ ft / f2 ⁇ 0.7 ft is the focal length of the entire plurality of lenses.
  • f1 is the focal length of the front lens group.
  • f2 is the focal length of the rear lens group.
  • the focal length of the objective lens can be shortened as compared with the case where, for example, a telephoto configuration is provided.
  • ft / f1 is lower than the lower limit of the above (8), the negative power becomes too strong and the performance deteriorates.
  • ft / f1 exceeds the upper limit of (8) above or ft / f2 falls below the lower limit of (9) above, the focal length cannot be shortened.
  • ft / f2 exceeds the upper limit of (9) above, the positive power becomes too strong and the performance deteriorates.
  • a plurality of lenses may have a telephoto configuration.
  • the outer shape of the plurality of lenses becomes substantially smaller from the side where the excitation light EL1 or the like is incident to the side where the excitation light EL1 or the like is emitted (Z-axis negative direction).
  • the number of lenses can be reduced and the cost can be reduced as compared with the retrofocus configuration (reverse telephoto configuration).
  • the cost of mechanical parts can be reduced accordingly.
  • first design conditions to sixth design conditions are merely examples, and various other design conditions appearing in the embodiments and examples of the lens structure described later may exist. Based on the above, some embodiments and examples of the lens structure will be described. There are two embodiments described, the first embodiment and the second embodiment. These embodiments differ in that the first embodiment has a retrofocus configuration (corresponding to the fifth design condition), whereas the second embodiment has a telephoto configuration (corresponding to the sixth design condition). do.
  • FIG. 3 is a diagram showing an example of a schematic configuration of the lens structure of the first embodiment and the first embodiment.
  • FIG. 3 shows an example of a cross-sectional structure when the lens structure is cut along a surface including an optical axis such as the excitation light EL1.
  • the illustrated lens structure 41 includes a plurality of lenses 411, a lens frame 412, a fixing member 413, and an aperture diaphragm 414.
  • the focal length is 5 mm
  • the number of lenses is 8
  • NA the numerical aperture NA
  • the objective field of view ⁇ is 0.6 mm.
  • the plurality of lenses 411 are arranged along the optical axis (Z-axis direction) of the excitation light EL1 and the like.
  • the plurality of lenses 411 include lenses G11 to lenses G18 arranged in order in the negative direction of the Z axis.
  • the lenses G11 to G13 form a front lens group 411F arranged on the Z-axis positive direction side.
  • the lenses G14 to G18 form a rear lens group 411R arranged on the negative side of the Z axis.
  • the front lens group 411F and the rear lens group 411R are configured to give a retrofocus configuration.
  • the front lens group 411F has a negative refractive index or is substantially non-power when viewed along the negative direction of the Z axis.
  • the rear lens group 411R has a positive refractive index.
  • the outer shape of the lenses G11 to G13 constituting the front lens group 411F may be substantially increased in the negative direction of the Z axis.
  • the outer shape of the lenses G14 to G18 constituting the rear lens group 411R may be substantially reduced in the negative direction of the Z axis.
  • Some adjacent lenses are in contact with each other to form a joint division group (marginal contact (Butsuke)).
  • the lens G12 and the lens G13 form a junction division group.
  • the lens G15 and the lens G16 form a junction division group.
  • the lens frame 412 is a frame body that holds a plurality of lenses 411.
  • the lens frame 412 includes a front frame body 412F and a rear frame body 412R.
  • the front frame body 412F is a frame body that constitutes a portion of the lens frame 412 on the positive direction side of the Z axis.
  • the rear frame body 412R is a frame body that constitutes a portion of the lens frame 412 on the negative direction side of the Z axis.
  • the lens frame 412 may have a lens barrel shape.
  • the front frame body 412F is a front lens barrel that constitutes a front side portion of the lens barrel shape.
  • the rear frame body 412R is a rear lens barrel that constitutes a rear portion of the lens barrel shape.
  • Examples of materials for the lens frame 412 are metals such as aluminum and brass, alloys, and the like. However, the material is not limited to these materials, and various materials may be adopted in consideration of price, ease of processing, durability, and the like.
  • the lens frame 412 has a configuration suitable for accommodating (fitting) and positioning the lens. Some characteristic parts of the lens frame 412 are illustrated with reference numerals.
  • the front frame body 412F includes an opening 412Fa, an opening 412Fb, a protrusion 412Fc, and a protrusion 412Fd.
  • the opening 412F opens the end on the Z-axis positive direction of the front frame 412F.
  • the opening 412Fb opens the end on the negative side of the Z-axis of the front frame 412F.
  • the protrusions 412Fc and the protrusions 412Fd project toward the inside of the front frame 412F between the lens G11 and the lens G12.
  • the lens G11 comes into contact with the protrusion 412Fc, and the lens G12 comes into contact with the protrusion 412Fd.
  • the rear frame body 412R includes an opening 412Ra, an opening 412Rb, a protrusion 412Rc, and a protrusion 412Rd.
  • the opening 412Ra opens the end on the Z-axis positive direction of the rear frame 412R.
  • the opening 412Rb opens the end on the negative side of the rear frame 412R in the negative direction of the Z axis.
  • the protrusions 412Rc and 412Rd project toward the inside of the rear frame 412R between the lens G17 and the lens G18.
  • the lens G17 comes into contact with the protrusion 412Rc
  • the lens G18 comes into contact with the protrusion 412Rd.
  • the lens frame 412 may have an air hole for allowing internal air to escape when a plurality of lenses 411 are fitted into the lens frame 412.
  • the fixing member 413 fixes the positions of the plurality of lenses 411 so that the plurality of lenses 411 are positioned in the lens frame 412.
  • the fixing member 413 includes a screw N11, a screw N13, a screw N14, a ring RG1415, a ring RG1617, and an adhesive AD18.
  • the ring RG1415 is arranged so as to provide a distance between the lens G14 and the lens G15.
  • the ring RG1617 is arranged so as to provide a distance between the lens G16 and the lens G17.
  • Examples of materials such as screw N11 and ring RG1415 are metals such as aluminum and copper, alloys, and the like. However, the material is not limited to these materials, and various materials may be adopted in consideration of price, ease of processing, durability, and the like.
  • the lens G11 is positioned by the screw N11 and the protrusion 412Fc of the front frame body 412F.
  • the lens G11 is fitted into the front frame body 412F from the opening 412F of the front frame body 412F so that the edge portion of the lens G11 abuts on the protrusion 412Fc.
  • the screw N11 presses (urges) the edge of the lens G11 against the protrusion 412Fc in the negative direction of the Z axis to fix it.
  • the lens G12 and the lens G13 are positioned by the screw N13 and the protrusion 412Fd of the front frame body 412F.
  • the lens G12 and the lens G13 are fitted into the front frame body 412F from the opening 412Fb of the front frame body 412F so that the edge portion of the lens G12 abuts on the protrusion 412Fd.
  • the screw N13 fixes the lens G13 and the edge portion of the lens G12 by pressing them against the protrusion 412Fd in the positive direction of the Z axis.
  • the lenses G14 to G17 are positioned by the screw N14, the ring RG1415, the ring RG1617, and the protrusion 412Rc of the rear frame body 412R.
  • the lenses G14 to G17, the ring RG1415, and the ring RG1617 are fitted into the rear frame 412R from the opening 412Ra of the rear frame 412R so that the edge of the lens G17 abuts on the protrusion 412Rc.
  • the screw N14 presses the edge of the lens G14, the ring RG1415, the edge of the lens G15 and the lens G16, the ring RG1617, and the edge of the lens G17 against the protrusion 412Rc in the negative direction of the Z axis to fix the screw N14.
  • the lens G18 is positioned by the adhesive AD18 and the protrusion 412Rd of the rear frame body 412R.
  • the lens G18 is fitted into the rear frame body 412R from the opening 412Rb of the rear frame body 412R so that the edge portion of the lens G18 abuts on the protrusion 412Rd.
  • the adhesive AD18 fixes the lens G18 to the rear frame 412R on the outside of the rear frame 412R. Since the lens G18 is not sealed by the rear frame 412R, the adhesive AD18 may be used. Instead of using the adhesive AD18, for example, the edge portion of the lens G18 may be pressed against the protrusion 412Rd in the negative direction of the Z-axis by a cap having an open central portion to fix the lens G18.
  • the rear frame body 412R is fitted into the front frame body 412F from the opening 412Fb of the front frame body 412F, and a lens structure 41 in which a plurality of lenses 411 are positioned in the lens frame 412 is obtained.
  • the aperture diaphragm 414 is provided between the front lens group 411F and the rear lens group 411R, and adjusts the amount of passing light. For example, the amount of light incident on the rear lens group 411R from the front lens group 411F is suppressed.
  • FIG. 4 is a diagram showing an example of a schematic configuration of the lens structure of the first embodiment and the first embodiment.
  • FIG. 4 schematically shows some paths (optical paths) of light passing through the lens structure.
  • the lens structure 41 concentrates the excitation light EL1 and the like on the spot 51s, and also concentrates the fluorescent FL and the scattered light BSC toward the detection unit 7 (FIG. 1).
  • the intensity of the excitation light EL1 or the like applied to the particles 51 is increased, and the light to be detected by the detection unit 7 is efficiently guided to the detection unit 7.
  • FIG. 5 is a diagram showing an example of lens design of the first embodiment and the first embodiment. Convex indicates positive power and concave indicates negative power. A plurality of lenses 411 are designed so as to satisfy the above-mentioned first design conditions to fifth design conditions.
  • the lens frame 412 holds a plurality of lenses 411 in the lens frame 412 without using an adhesive.
  • the adjacent lens G12 and lens G13 form a joining division group whose position in the lens frame 412 is determined by abutting against each other.
  • Adjacent lenses G15 and lenses G16 also form a junction division group.
  • the lens G18 is a positive lens located closest to the particle 51 and having positive power.
  • the lens G12, the lens G14, the lens G15, and the lens G17 are positive lenses (three or more positive lenses).
  • the lens G11, the lens G13, and the lens G16 are negative lenses (two or more negative lenses).
  • the lenses G11 to G13 are front lens groups 411F having a negative refractive index or substantially non-power.
  • Lenses G14 to G18 are rear lens groups 411R having a positive refractive index.
  • ft / f1 ⁇ 0.092.
  • ft / f2 0.565. Therefore, (8) ⁇ 0.24 ⁇ ft / f1 ⁇ 0.08 and (9) 0.35 ⁇ ft / f2 ⁇ 0.7 are satisfied.
  • FIGS. 6 and 7 are diagrams showing an example of a schematic configuration of the lens structures of the first embodiment and the second embodiment.
  • the illustrated lens structure 42 includes a plurality of lenses 421, a lens frame 422, a fixing member 423, and an aperture diaphragm 424.
  • the focal length is 5 mm
  • the number of lenses is 8
  • the numerical aperture NA is 0.85
  • the objective field of view ⁇ is 0.6 mm.
  • the plurality of lenses 421 include lenses G21 to lenses G23 constituting the front lens group 421F and lenses G24 to lenses G28 constituting the rear lens group 421R.
  • the lens G21 and the lens G22 form a junction division group.
  • the lens G25 and the lens G26 also form a joint division group.
  • the lens frame 422 includes a front frame body 422F and a rear frame body 422R.
  • the front frame body 422F includes an opening 422Fa, an opening 422Fb, a protrusion 422Fc, and a protrusion 422Fd.
  • the rear frame body 422R includes an opening 422Ra, an opening 422Rb, a protrusion 422Rc, and a protrusion 422Rc.
  • the fixing member 423 includes a screw N21, a screw N23, a screw N24, a ring RG2425, a ring RG2627, and an adhesive AD28.
  • the ring RG2425 is arranged so as to provide a distance between the lens G24 and the lens G25.
  • the ring RG2627 is arranged so as to provide a distance between the lens G26 and the lens G27.
  • the aperture diaphragm 424 is provided between the front lens group 421F and the rear lens group 421R, and adjusts the amount of passing light.
  • the lens G21 and the lens G22 are positioned by the screw N21 and the protrusion 422Fc of the front frame body 422F.
  • the lens G21 and the lens G22 are fitted into the front frame body 422F from the opening 422F of the front frame body 422F so that the edge portion of the lens G22 abuts on the protrusion 422Fc.
  • the screw N21 presses the edges of the lens G21 and the lens G22 against the protrusion 422Fc in the negative direction of the Z axis to fix the lens G21 and the lens G22.
  • the lens G23 is positioned by the screw N23 and the protrusion 422Fd of the front frame body 422F.
  • the lens G23 is fitted into the front frame body 422F from the opening 422Fb of the front frame body 422F so that the edge portion of the lens G23 abuts on the protrusion 422Fd.
  • the screw N23 fixes the edge of the lens G23 by pressing it against the protrusion 422Fd in the positive direction of the Z axis.
  • the lenses G24 to G27 are positioned by the screw N24, the ring RG2425, the ring RG2627, and the protrusion 422Rc of the rear frame body 422R.
  • the lens G24 to the lens G27, the ring RG2425, and the ring RG2627 are fitted into the rear frame body 422R from the opening 422Ra of the rear frame body 422R so that the edge portion of the lens G27 abuts on the protrusion 422Rc.
  • the screw N24 presses the edge of the lens G24, the ring RG2425, the edge of the lens G25 and the lens G26, the ring RG2627, and the edge of the lens G27 against the protrusion 422Rc in the negative direction of the Z axis to fix the screw N24.
  • the lens G28 is positioned by the adhesive AD28 and the protrusion 422Rd of the rear frame body 422R.
  • the lens G28 is fitted into the rear frame body 422R from the opening 422Rb of the rear frame body 422R so that the edge portion of the lens G28 abuts on the protrusion 422Rd.
  • the adhesive AD28 fixes the lens G28 to the rear frame 422R on the outside of the rear frame 422R.
  • the rear frame body 422R is fitted into the front frame body 422F from the opening 422Fb of the front frame body 422F, and a lens structure 42 in which a plurality of lenses 421 are positioned in the lens frame 422 is obtained.
  • the light As shown in FIG. 7, as the light travels in the negative direction of the Z axis, the light is generally diffused in the front lens group 421F, and the light is generally focused in the rear lens group 421R.
  • FIG. 8 is a diagram showing an example of lens design of the first embodiment and the second embodiment.
  • a plurality of lenses 421 are designed so as to satisfy the above-mentioned first design conditions to fifth design conditions.
  • the lens frame 422 holds a plurality of lenses 421 in the lens frame 422 without using an adhesive.
  • the adjacent lens G21 and lens G22 form a joining division group whose position in the lens frame 422 is determined by abutting against each other.
  • Adjacent lenses G25 and lenses G26 also form a junction division group.
  • the lens G28 is a positive lens located closest to the particle 51 and having positive power.
  • the lens G23, the lens G24, the lens G25, and the lens G27 are positive lenses (three or more positive lenses).
  • the lens G21 and the lens G26 are negative lenses (two or more negative lenses).
  • the lenses G21 to G23 are front lens groups 421F having a negative refractive index or substantially non-power.
  • Lenses G24 to G28 are rear lens groups 421R having a positive refractive index.
  • FIGS. 9 and 10 are diagrams showing an example of a schematic configuration of the lens structure of the first embodiment and the third embodiment.
  • the illustrated lens structure 43 includes a plurality of lenses 431, a lens frame 432, a fixing member 433, and an aperture diaphragm 434.
  • the focal length is 5 mm
  • the number of lenses is 8
  • the numerical aperture NA is 0.85
  • the objective field of view ⁇ is 0.6 mm.
  • the plurality of lenses 431 include a lens G31 and a lens G32 that form a front lens group 431F, and lenses G33 to a lens G38 that form a rear lens group 431R.
  • the lens G31 and the lens G32 form a joint division group.
  • the lens G33 and the lens G34 also form a joint division group.
  • the lens G36 and the lens G37 also form a joint division group.
  • the lens frame 432 includes a front frame body 432F and a rear frame body 432R.
  • the front frame body 432F includes an opening 432Fa, an opening 432Fb, and a protrusion 432Fd.
  • the rear frame body 432R includes an opening 432Ra, an opening 432Rb, a protrusion 432Rc, and a protrusion 432Rd.
  • the fixing member 433 includes a screw N32, a screw N33, a ring RG3435, and a ring RG3536.
  • the ring RG3435 is arranged so as to provide a distance between the lens G34 and the lens G35.
  • the ring RG3536 is arranged so as to provide a distance between the lens G35 and the lens G36.
  • the aperture diaphragm 434 is provided between the front lens group 431F and the rear lens group 431R, and adjusts the amount of passing light.
  • the lens G31 and the lens G32 are positioned by the screw N32 and the protrusion 432Fd of the front frame body 432F.
  • the lens G31 and the lens G32 are fitted into the front frame body 432F from the opening 432F of the front frame body 432F so that the edge portion of the lens G31 abuts on the protrusion 432Fd.
  • the screw N32 presses the edges of the lens G32 and the lens G31 against the protrusion 432Fd in the positive direction of the Z axis to fix the lens G32 and the lens G31.
  • the lenses G33 to G37 are positioned by the screw N33, the ring RG3435, the ring RG3536, and the protrusion 432Rc of the rear frame body 432R.
  • the lens G33 to the lens G37, the ring RG3435, and the ring RG3536 are fitted into the rear frame body 432R from the opening 432Ra of the rear frame body 432R so that the edge portion of the lens G37 abuts on the protrusion 432Rc.
  • the screw N33 fixes the edge of the lens G33 and the lens G34, the ring RG3435, the edge of the lens G35, the ring RG3536, and the edge of the lens G36 and the lens G37 by pressing them against the protrusion 432Rc in the negative direction of the Z axis. do.
  • the lens G38 is positioned by the protrusion 432Rd of the rear frame body 432R.
  • the lens G38 is fitted into the rear frame body 432R from the opening 432Rb of the rear frame body 432R so that the edge portion of the lens G38 abuts on the protrusion 432Rd.
  • the lens G38 may be fixed to the rear frame 432R with an adhesive or the like on the outside of the rear frame 432R.
  • the rear frame body 432R is fitted into the front frame body 432F from the opening 432Fb of the front frame body 432F, and a lens structure 43 in which a plurality of lenses 431 are positioned in the lens frame 432 is obtained.
  • the light As shown in FIG. 10, as the light travels in the negative direction of the Z axis, the light is generally diffused in the front lens group 431F, and the light is generally focused in the rear lens group 431R.
  • FIG. 11 is a diagram showing an example of lens design of the first embodiment and the third embodiment.
  • a plurality of lenses 431 are designed so as to satisfy the above-mentioned first design conditions to fifth design conditions.
  • the lens frame 432 holds a plurality of lenses 431 in the lens frame 432 without using an adhesive.
  • the adjacent lens G31 and lens G32 form a joining division group whose position in the lens frame 432 is determined by abutting against each other.
  • Adjacent lenses G33, lenses G34, G36 and lenses G37 also form a junction division group.
  • the lens G38 is a positive lens located closest to the particle 51 and having positive power.
  • the lens G32, the lens G33, the lens G35, and the lens G36 are positive lenses (three or more positive lenses).
  • the lens G31, the lens G34, and the lens G37 are negative lenses (two or more negative lenses).
  • the lens G31 and the lens G32 are front lens groups 431F having a negative refractive index or substantially non-power.
  • Lenses G33 to G38 are rear lens groups 431R having a positive refractive index.
  • ft / f1 ⁇ 0.159.
  • ft / f2 0.483. Therefore, (8) ⁇ 0.24 ⁇ ft / f1 ⁇ 0.08 and (9) 0.35 ⁇ ft / f2 ⁇ 0.7 are satisfied.
  • FIGS. 12 and 13 are diagrams showing an example of a schematic configuration of the lens structure of the first embodiment and the fourth embodiment.
  • the illustrated lens structure 44 includes a plurality of lenses 441, a lens frame 442, a fixing member 443, and an aperture diaphragm 444.
  • the focal length is 5 mm
  • the number of lenses is 9
  • the numerical aperture NA is 0.85
  • the objective field of view ⁇ is 0.6 mm.
  • the plurality of lenses 441 include lenses G41 to G44 constituting the front lens group 441F and lenses G45 to G49 constituting the rear lens group 441R.
  • the lens G41 and the lens G42 form a joint division group.
  • the lens G43 and the lens G44 also form a joint division group.
  • the lens G45 and the lens G46 also form a joint division group.
  • the lens G47 and the lens G48 also form a joint division group.
  • the lens frame 442 includes a front frame body 442F and a rear frame body 442R.
  • the front frame body 442F includes an opening 442Fa, an opening 442Fb, and a protrusion 442Fd.
  • the rear frame body 442R includes an opening 442Ra, an opening 442Rb, a protrusion 442Rc, and a protrusion 442Rd.
  • the fixing member 443 includes a screw N42, a screw N43, a ring RG4445, a ring RG4647, and an adhesive AD49.
  • the ring RG4445 is arranged so as to provide a distance between the lens G44 and the lens G45.
  • the ring RG4647 is arranged so as to provide a distance between the lens G46 and the lens G47.
  • the aperture diaphragm 444 is provided between the front lens group 441F and the rear lens group 441R, and adjusts the amount of passing light.
  • the lens G41 and the lens G42 are positioned by the screw N42 and the protrusion 442Fd of the front frame body 442F.
  • the lens G41 and the lens G42 are fitted into the front frame body 442F from the opening 442Fb of the front frame body 442F so that the edge portion of the lens G41 abuts on the protrusion 442Fd.
  • the screw N42 fixes the lens G42 and the edge portion of the lens G41 by pressing them against the protrusion 442Fd in the positive direction of the Z axis.
  • the lenses G43 to G48 are positioned by the screw N43, the ring RG4445, the ring RG4647, and the protrusion 442Rc of the rear frame body 442R.
  • the lens G43 to the lens G48, the ring RG4445, and the ring RG4647 are fitted into the rear frame body 442R from the opening 442Ra of the rear frame body 442R so that the edge portion of the lens G48 abuts on the protrusion 442Rc.
  • the screw N43 attaches the edges of the lens G43 and the lens G44, the ring RG4445, the edges of the lens G45 and the lens G46, the ring RG4647, and the edges of the lens G47 and the lens G48 to the protrusion 442Rc in the negative direction of the Z axis. Press and fix.
  • the lens G49 is positioned by the adhesive AD49 and the protrusion 442Rd of the rear frame body 442R.
  • the lens G49 is fitted into the rear frame body 442R from the opening 442Rb of the rear frame body 442R so that the edge portion of the lens G49 abuts on the protrusion 442Rd.
  • the adhesive AD49 fixes the lens G49 to the rear frame 442R on the outside of the rear frame 442R.
  • the rear frame body 442R is fitted into the front frame body 442F from the opening 442Fb of the front frame body 442F, and a lens structure 44 in which a plurality of lenses 441 are positioned in the lens frame 442 is obtained.
  • the light As shown in FIG. 13, as the light travels in the negative direction of the Z axis, the light is generally diffused in the front lens group 441F, and the light is generally focused in the rear lens group 441R.
  • FIG. 14 is a diagram showing an example of lens design of the first embodiment and the fourth embodiment.
  • a plurality of lenses 441 are designed so as to satisfy the above-mentioned first design conditions to the fourth design conditions.
  • the lens frame 442 holds a plurality of lenses 441 in the lens frame 442 without using an adhesive.
  • the adjacent lens G41 and lens G42 form a joining division group whose position in the lens frame 442 is determined by abutting against each other.
  • Adjacent lenses G43 and G44, as well as G45 and lens G46, lens G47 and lens G48 also form a junction division group.
  • the lens G49 is a positive lens located closest to the particle 51 and having positive power.
  • the lens G42, the lens G43, the lens G45, and the lens G47 are positive lenses (three or more positive lenses).
  • the lens G41, the lens G44, the lens G46, and the lens G48 are negative lenses (two or more negative lenses).
  • the lenses G41 to G44 are front lens groups 441F having a negative refractive index or substantially non-power.
  • Lenses G45 to G49 are rear lens groups 441R having a positive refractive index.
  • ft / f1 0.004.
  • ft / f2 0.489. Therefore, (8) ⁇ 0.24 ⁇ ft / f1 ⁇ 0.08 and (9) 0.35 ⁇ ft / f2 ⁇ 0.7 are satisfied.
  • FIGS. 15 and 16 are diagrams showing an example of a schematic configuration of the lens structure of the second embodiment and the fifth embodiment.
  • the illustrated lens structure 45 includes a plurality of lenses 451 and a lens frame 452, a fixing member 453, and an aperture diaphragm 454.
  • the focal length is 10 mm
  • the number of lenses is 8
  • the numerical aperture NA is 0.85
  • the objective field of view ⁇ is 0.6 mm.
  • the plurality of lenses 451 include lenses G51 to G58.
  • the lens G53 and the lens G54 form a junction division group.
  • the lens G56 and the lens G57 also form a joint division group.
  • the lens frame 452 includes an opening 452a, an opening 452b, a protrusion 452c, and a protrusion 452d.
  • the fixing member 453 includes a screw N51, a ring RG5152, a ring RG5253, a ring RG5455, a ring RG5556, and an adhesive AD58.
  • the ring RG5152 is arranged so as to provide a distance between the lens G51 and the lens G52.
  • the ring RG5253 is arranged so as to provide a distance between the lens G52 and the lens G53.
  • the ring RG5455 is arranged so as to provide a distance between the lens G54 and the lens G55.
  • the ring RG5556 is arranged so as to provide a distance between the lens G55 and the lens G56.
  • the aperture diaphragm 454 is provided between the lens G52 and the lens G53 to adjust the amount of passing light.
  • the lenses G51 to G57 are positioned by the screw N51, the ring RG5152, the ring RG5253, the ring RG5455, the ring RG5556, and the protrusion 452c of the lens frame 452.
  • the lenses G51 to G57, the ring RG5152, the ring RG5253, the ring RG5455, and the ring RG5556 are fitted into the lens frame 452 from the opening 452a of the lens frame 452 so that the edge portion of the lens G57 abuts on the protrusion 452c.
  • the screw N51 has the edge of the lens G51, the ring RG5152, the edge of the lens G52, the ring RG5253, the edge of the lens G53 and the lens G54, the ring RG5455, the edge of the lens G55, and the ring RG5556 in the negative direction of the Z axis. Further, the edges of the lens G56 and the lens G57 are pressed against the protrusion 452c to be fixed.
  • the lens G58 is positioned by the adhesive AD58 and the protrusion 452d of the lens frame 452.
  • the lens G58 is fitted into the lens frame 452 through the opening 452b of the lens frame 452 so that the edge portion of the lens G58 abuts on the protrusion 452d.
  • the adhesive AD58 fixes the lens G58 to the lens frame 452 on the outside of the lens frame 452.
  • the lens frame 452 is a single frame body, and the lens structure 45 in which a plurality of lenses 451 are positioned in the lens frame 452 can be obtained by the above fitting and fixing.
  • light is generally focused by the plurality of lenses 451 as it travels in the negative direction of the Z axis.
  • FIG. 17 is a diagram showing an example of lens design of the second embodiment and the fifth embodiment.
  • a plurality of lenses 421 are designed so as to satisfy the above-mentioned first design condition to fourth design condition and sixth design condition.
  • the lens frame 452 holds a plurality of lenses 451 in the lens frame 452 without using an adhesive.
  • the adjacent lens G53 and lens G54 form a joining division group whose position in the lens frame 452 is determined by abutting against each other.
  • Adjacent lenses G56 and G57 also form a junction division group.
  • the lens G58 is a positive lens located closest to the particle 51 and having positive power.
  • the lens G53, the lens G55, and the lens G56 are positive lenses (three or more positive lenses) corresponding to the third design condition.
  • the lens G51, the lens G54, and the lens G57 are negative lenses (two or more negative lenses).
  • the outer shape of the plurality of lenses 451 becomes smaller from the side where the excitation light EL1 or the like is incident to the side where the excitation light EL1 or the like is emitted (as the Z-axis is negative).
  • FIGS. 18 and 19 are diagrams showing an example of a schematic configuration of another lens structure of the second embodiment.
  • the illustrated lens structure 46 includes a plurality of lenses 461, a lens frame 462, and a fixing member 463.
  • the focal length is 10 mm
  • the number of lenses is 6
  • the numerical aperture NA is 0.75
  • the objective field of view ⁇ is 0.5 mm.
  • the plurality of lenses 461 include lenses G61 to lenses G66.
  • the lens G62 and the lens G63 form a joint division group.
  • the lens G64 and the lens G65 also form a joint division group.
  • the lens frame 462 includes an opening 462a, an opening 462b, a protrusion 462c, a protrusion 462d, an air hole 462e, an air hole 462f, and an air hole 462g.
  • the air hole 462e is a hole for letting out the air inside when the lens G61 is fitted into the lens frame 462.
  • the air hole 462f is a hole for letting out the air inside when the lens G63 is fitted into the lens frame 462.
  • the air hole 462g is a hole for letting out the air inside when the lens G65 and the lens G66 are fitted into the lens frame 462.
  • the fixing member 463 includes a screw N61, a ring RG6162, and a ring RG63664.
  • the ring RG6162 is arranged so as to provide a distance between the lens G61 and the lens G62.
  • the ring RG6364 is arranged so as to provide a distance between the lens G63 and the lens G64.
  • the lenses G61 to G65 are positioned by the screw N61, the ring RG6162, the ring RG6364, and the protrusion 462c of the lens frame 462.
  • the lens G61 to the lens G65, the screw N61, the ring RG6162, and the ring RG6364 are fitted into the lens frame 462 from the opening 462a of the lens frame 462 so that the edge portion of the lens G65 abuts on the protrusion 462c.
  • the screw N61 presses the edge of the lens G61, the ring RG6162, the edge of the lens G62 and the lens G63, the ring RG6364, and the edge of the lens G64 and the lens G65 against the protrusion 462c to fix the screw N61 in the negative direction of the Z axis. do.
  • the lens G66 is positioned by the protrusion 462d of the lens frame 462.
  • the lens G66 is fitted into the lens frame 462 from the opening 462b of the lens frame 462 so that the edge portion of the lens G66 abuts on the protrusion 462d.
  • the lens G66 may be fixed to the lens frame 462 with an adhesive or the like on the outside of the lens frame 462.
  • the lens frame 462 is a single frame body, and the lens structure 46 in which a plurality of lenses 461 are positioned in the lens frame 462 can be obtained by the above fitting and fixing.
  • the light is generally focused by the plurality of lenses 461 as the light travels in the negative direction of the Z axis.
  • the lens structure 46 may also be designed so as to satisfy at least a part of the above-mentioned first design condition to fourth design condition and sixth design condition.
  • FIG. 20 is a flowchart showing an example of processing (particle analysis method) executed in the particle analyzer.
  • step S1 the excitation light is focused on the spot using the lens structure.
  • the light source 1 emits the excitation light EL1 and the like, and the objective lens 4 (lens structure 41 to the lens structure 46) focuses the excitation light EL1 and the like on the spot 51s.
  • the particles 51 located at the spots 51s are excited and light such as fluorescent FL is emitted.
  • step S2 the light emitted from the particles is detected.
  • the detection unit 7 detects light such as fluorescent FL emitted from the particles 51 by the focused irradiation of the excitation light EL1 in the previous step S1.
  • step S3 the detection data is processed.
  • the processing control unit 120 processes the detection data obtained by the detection in the previous step S2, and analyzes, for example, the component, size, and the like of the particle 51.
  • an open cell sorter has been described as an example.
  • the disclosed technique may be applied to a closed cell sorter, an analyzer type flow cytometer, a microscope for acquiring an image of particles on a flow path, and the like.
  • microchip method a method using a microchip 5 (microchip method) has been described as an example.
  • various methods such as a droplet method, a cuvette method, and a flow cell method may be adopted.
  • the excitation light EL1 (wavelength 349 nm), the excitation light EL2 (wavelength 405 nm), the excitation light EL3 (wavelength 488 nm), the excitation light EL4 (wavelength 561 nm), and the excitation light EL5 (wavelength 637 nm).
  • An example of being irradiated with light was described. However, the number and wavelength of excitation light are not limited to this example.
  • the particles 51 may be irradiated with various excitation lights (combinations) depending on the type of the particles 51, the content of the analysis thereof, and the like.
  • the mirror group 2, the mirror 3, and the imaging lens 6 are used as an optical system for guiding the excitation light EL1 or the like to the objective lens 4 and guiding the scattered light BSC or the like from the objective lens 4 to the detection unit 7.
  • the detection unit 7 was explained as an example.
  • any optical system capable of achieving the same purpose may be adopted.
  • the particle analyzer 100 includes excitation light EL1 containing light having a wavelength of 400 nm or less (may be 350 nm or less) (including light having a wavelength longer than 400 nm).
  • a light source 1 that emits light such as A detection unit 7 that detects the light (fluorescent FL and scattered light BSC) emitted from the particles 51 by being excited by the light EL1 or the like, and a processing unit (processing control) that processes the detection data acquired by the detection unit 7. Part 120) and.
  • the lens structure 41 includes a plurality of lenses 411 arranged along the optical axis of the excitation light EL1 and the like (along the negative direction of the Z axis), and a lens frame 412 holding the plurality of lenses 411. At least one of the plurality of lenses 411 (lens G12, lens G13, lens G15, and lens G16) is positioned in the lens frame 412 by abutting on a lens adjacent to the lens.
  • the inside of the lens frame 412 can have an adhesive-less structure. Therefore, the burning of the adhesive and the burning of the outgas adhering to the lens surface emitted from the adhesive may occur due to the excitation light in the ultraviolet region having a short wavelength such as 400 nm or less passing through the plurality of lenses 411. Etc. can be prevented. Therefore, it is possible to suppress deterioration of optical characteristics.
  • the plurality of lenses 411 have a positive lens (lens G18) on the most object side (Z-axis positive direction side), and the glass material of the positive lens is Assuming that the refractive index on the d-line is nd and the Abbe number on the d-line is ⁇ d, (1) 1.73 ⁇ nd ⁇ 1.83 and (2) 45 ⁇ d may be satisfied. As a result, it is possible to secure the transmittance in the ultraviolet region and correct the chromatic aberration in a wide band.
  • the plurality of lenses 411 have at least three or more positive lenses (lens G12, lens G14, lens G15 and lens G17), and the plurality of lenses 411 have three or more positive lenses.
  • the refractive index at the d-line is nd
  • the Abbe number at the d-line is ⁇ d
  • the partial dispersion ratio between the g-line and the F-line is PgF
  • the glass material of the lens is (3) nd ⁇ 1.52
  • (4). 79 ⁇ d and (5) PgF ⁇ 0.54 may be satisfied. This also makes it possible to secure the transmittance in the ultraviolet region and correct chromatic aberration in a wide band.
  • the plurality of lenses 411 have at least two or more negative lenses (lens G11, lens G13, and lens G16), and the glass of the two or more negative lenses.
  • the material may satisfy (6) nd ⁇ 1.78 and (7) 29 ⁇ d, where nd is the refractive index on the d-line and ⁇ d is the Abbe number on the d-line. This also makes it possible to secure the transmittance in the ultraviolet region and correct chromatic aberration in a wide band.
  • the plurality of lenses 411 are arranged in order from the side where the excitation light EL1 or the like is incident to the side where the excitation light EL1 or the like is emitted (Z-axis negative direction), and negative refraction. It may be composed of a front lens group having a force or substantially non-power and a rear lens group 411R having a positive refractive power.
  • the focal length of the front lens group 411F is f1
  • the focal length of the rear lens group 411R is f2
  • (8) -0.24 ⁇ ft / f1 ⁇ 0.08 and (9) 0.35 ⁇ ft / f2 ⁇ 0.7 may be satisfied.
  • the focal length of the objective lens can be shortened as compared with the case where the telephoto configuration is provided, for example.
  • the particle analyzer 100 may include an aperture diaphragm 414 provided between the front lens group 411F and the rear lens group 411R. Thereby, the amount of passing light can be adjusted between the front lens group 411F and the rear lens group 411R.
  • the lens structure 41 may condense the light (fluorescent FL and scattered light BSC) emitted from the particles 51 toward the detection unit 7. As a result, the light to be detected can be efficiently guided to the detection unit 7.
  • the outer shape of the plurality of lenses 451 may be substantially reduced from the side where the excitation light EL1 or the like is incident to the side where the excitation light EL1 or the like is emitted (the Z-axis positive direction).
  • a telephoto configuration there is an increased possibility that the number of lenses can be reduced and the cost can be reduced as compared with the case where, for example, a retrofocus configuration (reverse telephoto configuration) is provided.
  • a retrofocus configuration reverse telephoto configuration
  • the particle analyzer 100 includes an optical system (mirror 3) arranged between the light source 1, the lens structure 41 (objective lens 4), and the detection unit 7. good.
  • the optical system (mirror 3) may include a central portion 31 that reflects the excitation light EL1 and the like, and a peripheral portion 32 that transmits the light (fluorescent FL and scattered light BSC) emitted from the particles 51.
  • the excitation light EL1 or the like from the light source 1 can be guided to the lens structure 41, and the fluorescent FL or the like from the objective lens 4 can be guided to the detection unit 7. .
  • the particle analysis method described with reference to FIGS. 1, 3, 20 and the like is also one form of the present disclosure.
  • the excitation light EL1 or the like containing light having a wavelength of 400 nm or less is focused on a predetermined position (spot 51s) in the flow path 53 by using the lens structure 41 (objective lens 4) (step S1). ),
  • the light (fluorescent FL and scattered light BSC) emitted from the particles 51 by being excited by the excitation light EL1 or the like of the particles 51 flowing at a predetermined position is detected (step S2), and the detection data is obtained. Processing (step S3) and the like.
  • the lens structure 41 includes a plurality of lenses 411 arranged along the optical axis of the excitation light EL1 and the like (along the negative direction of the Z axis), and a lens frame 412 holding the plurality of lenses 411. At least one of the plurality of lenses 411 (lens G12, lens G13, lens G15, and lens G16) is positioned in the lens frame 412 by abutting on a lens adjacent to the lens. As described above, such a particle analysis method also makes it possible to suppress deterioration of optical characteristics.
  • the optical measuring device 110 described with reference to FIGS. 1 and 3 and the like is also an embodiment of the present disclosure.
  • the optical measuring device 110 includes a light source 1 that emits excitation light EL1 or the like containing light having a wavelength of 400 nm or less, and a lens structure 41 that focuses the excitation light EL1 or the like at a predetermined position (spot 51s) in the flow path 53. (Objective lens 4) and a detection unit 7 that detects light (fluorescent FL and scattered light BSC) emitted from the particles 51 when the particles 51 flowing at a predetermined position are excited by the excitation light EL1 or the like. Be prepared.
  • the lens structure 41 includes a plurality of lenses 411 arranged along the optical axis of the excitation light EL1 and the like (along the negative direction of the Z axis), and a lens frame 412 holding the plurality of lenses 411. At least one of the plurality of lenses 411 (lens G12, lens G13, lens G15, and lens G16) is positioned in the lens frame 412 by abutting on a lens adjacent to the lens.
  • Such an optical measuring device 110 also makes it possible to suppress deterioration of optical characteristics as described above.
  • the disclosed technique may have the following configurations.
  • a light source that emits excitation light containing light with a wavelength of 400 nm or less
  • a lens structure that collects the excitation light at a predetermined position in the flow path
  • a detection unit that detects the light emitted from the particles when the particles flowing through the predetermined position are excited by the excitation light.
  • a processing unit that processes the detection data acquired by the detection unit, and With The lens structure is A plurality of lenses arranged along the optical axis of the excitation light, A lens frame that holds the plurality of lenses and With At least one of the plurality of lenses is positioned in the lens frame by abutting on a lens adjacent to the lens. Particle analyzer.
  • the plurality of lenses have a positive lens on the most object side, and the glass material of the positive lens has a refractive index on the d-line of nd and an Abbe number on the d-line of ⁇ d. (1) 1.73 ⁇ nd ⁇ 1.83 And (2) 45 ⁇ d Satisfy The particle analyzer according to [1].
  • the plurality of lenses have at least three positive lenses, and the glass material of the three or more positive lenses has a refractive index at the d-line of nd, an Abbe number at the d-line of ⁇ d, and a g-line.
  • the particle analyzer according to [1] or [2].
  • the plurality of lenses have at least two negative lenses, and the glass material of the two or more negative lenses has a refractive index at the d-line of nd and an Abbe number at the d-line of ⁇ d.
  • (6) nd ⁇ 1.78 And (7) 29 ⁇ d Satisfy The particle analyzer according to any one of [1] to [3].
  • the plurality of lenses have a group of front lenses having a negative refractive power or substantially non-power, which are arranged in order from the side where the excitation light is incident to the side where the excitation light is emitted, and after having a positive refractive power. Consists of a lens group, The particle analyzer according to any one of [1] to [4]. [6] For the plurality of lenses, assuming that the focal lengths of the plurality of lenses are ft, the focal lengths of the front lens group are f1, and the focal lengths of the rear lens group are f2.
  • the outer shape of the plurality of lenses becomes substantially smaller from the side where the excitation light is incident to the side where the excitation light is emitted.
  • the particle analyzer according to any one of [1] to [9].
  • the optical system is The central part that reflects the excitation light and Peripheral parts that transmit the light emitted from the particles and including, The particle analyzer according to [10].
  • the light source emits excitation light including light having a wavelength of 350 nm or less.
  • the light source also emits excitation light containing light having a wavelength longer than 400 nm.
  • the particle analyzer according to any one of [1] to [12].
  • the excitation light containing light with a wavelength of 400 nm or less is focused at a predetermined position in the flow path. Detecting the light emitted from the particles by being excited by the excitation light of the particles flowing through the predetermined position, and Processing detection data and Including
  • the lens structure is A plurality of lenses arranged along the optical axis of the excitation light, A lens frame that holds the plurality of lenses and With At least one of the plurality of lenses is positioned in the lens frame by abutting on a lens adjacent to the lens. Particle analysis method.
  • a light source that emits excitation light containing light with a wavelength of 400 nm or less, A lens structure that collects the excitation light at a predetermined position in the flow path, and A detection unit that detects the light emitted from the particles when the particles flowing through the predetermined position are excited by the excitation light.
  • the lens structure is A plurality of lenses arranged along the optical axis of the excitation light, A lens frame that holds the plurality of lenses and With At least one of the plurality of lenses is positioned in the lens frame by abutting on a lens adjacent to the lens.
  • Optical measuring device is A plurality of lenses arranged along the optical axis of the excitation light, A lens frame that holds the plurality of lenses and With At least one of the plurality of lenses is positioned in the lens frame by abutting on a lens adjacent to the lens.
  • Optical measuring device 110 Optical measuring device 120 Processing control unit 1 Light source 11 Light source 12 Light source 13 Light source 14 Light source 15 Light source 2 Mirror group 21 Mirror 22 Mirror 23 Mirror 24 Mirror 25 Mirror 3 Mirror 31 Central part 32 Peripheral part 4 Objective lens 5 Micro Chip 51 Particles 51s Spot 52 Biological sample 53 Flow path 6 Imaging lens 7 Detection unit 8 Detection unit 41 Lens structure 411 Multiple lenses 411F Front lens group 411R Rear lens group G11 Lens G12 Lens G13 Lens G14 Lens G15 Lens G16 Lens G18 Lens 412 Lens frame 412F Front frame 412F Opening 412Fb Opening 412Fc Projection 412Fd Projection 412R Rear frame 412Ra Opening 412Rb Opening 412Rc Projection 412Rd Ring AD18 Adhesive 414 Opening Squeeze

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PCT/JP2021/045300 2021-01-14 2021-12-09 粒子分析装置、粒子分析方法及び光学測定装置 Ceased WO2022153736A1 (ja)

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JP2004219608A (ja) * 2003-01-14 2004-08-05 Kurobane Nikon:Kk 対物レンズ及びこの対物レンズを備える顕微鏡
JP4252447B2 (ja) * 2001-06-22 2009-04-08 カール ツァイス イェナ ゲーエムベーハー 対物レンズ
JP2015038539A (ja) * 2012-10-26 2015-02-26 シャープ株式会社 レンズ素子
WO2016185623A1 (ja) * 2015-05-18 2016-11-24 シャープ株式会社 微小粒子検出装置

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US6704151B2 (en) * 2001-07-10 2004-03-09 Olympus Optical Co., Ltd. Optical unit and method for assembling the same
JP4817442B2 (ja) * 2006-07-31 2011-11-16 シスメックス株式会社 粒子分析装置用光学系、及びそれを用いた粒子分析装置
JP4990746B2 (ja) 2007-12-14 2012-08-01 ベイバイオサイエンス株式会社 液体フローに含まれる生物学的粒子を分別する装置ならびにその方法
JP5720233B2 (ja) 2010-12-17 2015-05-20 ソニー株式会社 マイクロチップ及び微小粒子分取装置

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JP4252447B2 (ja) * 2001-06-22 2009-04-08 カール ツァイス イェナ ゲーエムベーハー 対物レンズ
JP2003090945A (ja) * 2001-07-10 2003-03-28 Olympus Optical Co Ltd レンズ保持装置および顕微鏡用対物レンズ
JP2004219608A (ja) * 2003-01-14 2004-08-05 Kurobane Nikon:Kk 対物レンズ及びこの対物レンズを備える顕微鏡
JP2015038539A (ja) * 2012-10-26 2015-02-26 シャープ株式会社 レンズ素子
WO2016185623A1 (ja) * 2015-05-18 2016-11-24 シャープ株式会社 微小粒子検出装置

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