EP4314775A1 - Appareil, dispositif et procédé d'irradiation d'un échantillon biologique particulier, avec composant optique holographique - Google Patents

Appareil, dispositif et procédé d'irradiation d'un échantillon biologique particulier, avec composant optique holographique

Info

Publication number
EP4314775A1
EP4314775A1 EP22716224.5A EP22716224A EP4314775A1 EP 4314775 A1 EP4314775 A1 EP 4314775A1 EP 22716224 A EP22716224 A EP 22716224A EP 4314775 A1 EP4314775 A1 EP 4314775A1
Authority
EP
European Patent Office
Prior art keywords
light
light source
holographic
optical component
hoe
Prior art date
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.)
Pending
Application number
EP22716224.5A
Other languages
German (de)
English (en)
Inventor
Reinhold Fiess
Eugen BAUMGART
Ingo Ramsteiner
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Robert Bosch GmbH
Original Assignee
Robert Bosch GmbH
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Priority claimed from DE102022202634.9A external-priority patent/DE102022202634A1/de
Application filed by Robert Bosch GmbH filed Critical Robert Bosch GmbH
Publication of EP4314775A1 publication Critical patent/EP4314775A1/fr
Pending legal-status Critical Current

Links

Classifications

    • 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
    • G01N21/645—Specially adapted constructive features of fluorimeters
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6844—Nucleic acid amplification reactions
    • C12Q1/686—Polymerase chain reaction [PCR]
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B5/00—Optical elements other than lenses
    • G02B5/32—Holograms used as optical elements
    • 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
    • G01N2021/6417—Spectrofluorimetric devices
    • G01N2021/6419—Excitation at two or more wavelengths
    • 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
    • G01N21/645—Specially adapted constructive features of fluorimeters
    • G01N2021/6463—Optics
    • 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
    • G01N21/645—Specially adapted constructive features of fluorimeters
    • G01N2021/6463—Optics
    • G01N2021/6471—Special filters, filter wheel
    • 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
    • G01N21/645—Specially adapted constructive features of fluorimeters
    • G01N21/6456—Spatial resolved fluorescence measurements; Imaging

Definitions

  • Device for irradiating a sample, in particular a biological one, with a holographic-optical component
  • fluorescence measurements are also widely used in medical technology, for example in flow cytometry, in quantitative real-time PCR, in antibody tests or in histopathology.
  • the light source is of particular importance, because every fluorescence measurement requires excitation of the sample with a selected wavelength band.
  • the task of the light source is to deliver light of high spectral density in the range of the absorption band of the respective fluorophore.
  • the light must be limited to a precisely defined wavelength interval in order not to outshine the fluorescence wavelengths, which are usually not far away spectrally. These requirements are exacerbated when several dyes that have different, usually overlapping, excitation and fluorescence bands are to be addressed separately.
  • the light source must then be able to switch between different excitation bands, which are spectrally characterized by precisely defined wavelength bands with steep slopes.
  • such light sources are realized by either providing a very broadband light source (e.g. white light-emitting diode, incandescent lamp or gas discharge lamp) with exchangeable dielectric band-pass filters, which are arranged, for example, on a slider or wheel, or uses multiple light sources that either have intrinsically limited spectral ranges (laser, SLD) and/or are broader (e.g. colored LEDs), but are spectrally defined by a fixed bandpass filter. If several light sources are used, the beam paths are usually brought into line with dichroic mirrors before the light hits the sample.
  • a very broadband light source e.g. white light-emitting diode, incandescent lamp or gas discharge lamp
  • exchangeable dielectric band-pass filters which are arranged, for example, on a slider or wheel, or uses multiple light sources that either have intrinsically limited spectral ranges (laser, SLD) and/or are broader (e.g. colored LEDs), but are spectrally defined by a fixed band
  • the invention relates to a device for irradiating a sample.
  • the device comprises a first light source, a second light source and at least one holographic-optical component.
  • the first light source, the second light source and the holographic-optical component are arranged relative to one another such that first light from the first light source and second light from the second light source are directed via the holographic-optical component onto a common sample area for irradiating the sample.
  • the sample preferably comprises a material which can be excited to fluoresce with suitable optical excitation, in particular due to fluorophores contained in the sample or added to the sample.
  • the sample can in particular be a biological sample.
  • the sample can include parts of a body fluid from an animal or a human, for example components from blood, urine, sputum or a swab or from a tissue sample.
  • the sample can contain nucleic acids or segments of nucleic acids, preferably amplified parts of nucleic acids from an isothermal or polymerase chain reaction (“PCR”)-based amplification of nucleic acid segments.
  • PCR polymerase chain reaction
  • the sample can include a product from a detection method for detecting pathogens, in particular a product from an isothermal or PCR-based DNA amplification, with the detection of the presence of certain pathogens in particular via a fluorescence-based selection of DNA samples marked with fluorophores should take place.
  • the invention is fundamentally suitable for any application and for any device used for this purpose, in which fluorescent examination objects have to be illuminated with several wavelength bands, in particular in the field of life science, forensics and also in protecting and verifying the authenticity of products, in particular documents against product piracy.
  • the light sources can be, for example, light-emitting diodes (“LED”), superluminescent or laser diodes, incandescent lamps, gas discharge lamps or also lighting means excited by primary sources such as, for example, phosphorus-converted sources.
  • the first light source preferably has a first emission spectrum which differs from a second emission spectrum of the second light source.
  • the wavelength spectrum of the first light can preferably differ from that of the second light.
  • the first or second light source comprises beam-shaping elements (e.g. concave mirrors, Fresnel or refractive lenses) and/or spectrally filtering elements (e.g. dielectric filters or colored glasses).
  • the first or the second light source can include a bandpass filter for defined limitation of the emitted light.
  • bandpass filters can advantageously limit the spectral range of the light sources to a narrower band and steeper edges than is possible solely through the intrinsic spectral width of the holographic optical component.
  • scattered light with undesired wavelengths can be reduced.
  • beam-shaping and/or spectrally filtering elements can be arranged at other points of the device, in particular between the holographic-optical component and the sample area or the sample.
  • the holographic optical component (also referred to as "HOE" for short) is in particular an element whose holographic properties can be used for the optics of devices, for example to replace conventional lenses, mirrors and prisms.
  • the HOE preferably comprises a substrate for mechanical stabilization (eg glass or plastic) and at least one or more optically active layers in which one or more holograms are inscribed.
  • the HOE is designed to direct light from the first and second light sources at least partially onto the sample area.
  • the intrinsic wavelength selectivity of the holograms can preferably be used as a desired filter function in order to avoid a specific wavelength band to illuminate the sample area and the sample.
  • the holographic-optical component can comprise a reflection hologram or a transmission hologram, which is designed as a surface hologram or preferably as a volume hologram.
  • the holographic-optical component is thus preferably designed, with a predetermined placement relative to the HOE of the first light source with the first emission spectrum and the second light source with the second emission spectrum, at least some of the light from the first light source and some of the light from the second light source to steer a given sample area.
  • the (common) sample area means a specific spatial area relative to the HOE and preferably relative to the two light sources, in particular a specific solid angle area in which a sample can be placed for illumination according to the invention.
  • the invention thus advantageously provides excitation optics for fluorescence spectroscopy, wherein excitation light from different excitation channels can be directed onto the sample to be examined via the holographic-optical component.
  • the holographic-optical component can advantageously fulfill a number of functions, in particular due to holographic structures designed for this purpose in one or more holographic layers of the HOE.
  • the HOE is preferably designed to geometrically combine several beam paths and thus to deflect light beams incident on the HOE from different directions into a common area, in particular a common direction, in particular via reflection or transmission caused by the HOE.
  • the HOE can be designed to direct only light beams of specific wavelengths in a specific direction, preferably onto the sample.
  • the HOE can be designed to shape an intrinsic radiation distribution of one or more of the light sources, in particular to filter, diffract, focus/bundle, collimate and/or expand without the usual filters, diffraction gratings, lenses or curved mirrors to need.
  • the HOE can be set up Bending light of predetermined wavelengths, in particular at the short and/or long-wave edge of a predetermined wavelength band.
  • the HOE can preferably be designed to fulfill one or more of these functions at the same time.
  • the HOE can have a beam-shaping effect for light of predetermined wavelengths and/or from predetermined directions.
  • the HOE can preferably be permeable to light of other wavelengths and/or from other predetermined directions, ie only slightly, if at all, influence such light.
  • the invention thus advantageously enables the sample to be illuminated with spectrally different light from the same direction via the HOE.
  • the invention enables a wavelength-sensitive, deterministic deflection of light, as described above. This is particularly advantageous when different fluorophores in the sample are to be excited with different excitation spectra under the same geometric illumination conditions, ie when the sample is to be exposed to the same illumination situation regardless of the spectral distribution of the light.
  • the same illumination made possible by the device according to the invention has the particular advantage that inhomogeneities in the sample are illuminated in the same way by different excitation light and different fluorescence signals do not affect different ones.
  • the device according to the invention has the advantage that no moving parts and an associated mechanism that is susceptible to errors and wear and tear are required. Furthermore, it is avoided that a large part of the light generated by the broadband light source remains unused, possibly being shielded by further measures, and the associated waste heat having to be dissipated.
  • the invention also has particular advantages over the solutions with a plurality of light sources explained at the outset.
  • the solution according to the invention can be implemented much more cost-effectively than using a plurality of light sources. Because for light sources with intrinsically limited spectral ranges are often Laser or superluminescent diodes and for broadband light sources comparatively expensive dielectric bandpass filters are required.
  • the device according to the invention does not require any additional optical components such as, for example, dichroic mirrors for combining the beam paths of the various light sources.
  • the invention thus provides resource-saving, compact and comparatively inexpensive excitation optics which are particularly suitable for use in low-cost molecular diagnostics, in particular for use in lab-on-a-chip platforms, preferably with quantitative real-time PCR with multiplex functionality, for example in point-of-care solutions.
  • the HOE and the two light sources are arranged relative to one another in such a way that first light from the first light source and second light from the second light source are directed via the HOE onto a common light path or beam path for irradiating the sample, preferably via a reflection the HOE, for which the HOE includes a reflection hologram.
  • the common light path or common beam path can be realized in particular by at least partially, preferably completely, overlapping beam paths or beam profiles.
  • the device according to the invention enables an at least partially, preferably completely congruent wavefront of the light beams deflected by the HOE.
  • the device can also comprise more than two light sources as described above, with the further light sources also being able to have beam-shaping and/or spectrally filtering elements as explained above.
  • the device comprises in particular between two and six, preferably between two and four light sources, in particular light sources suitable for molecular diagnostic applications.
  • the sample area can advantageously be illuminated in the same way with many different light spectra.
  • the device comprises at least one third light source and the holographic-optical component is designed to be transparent to light of predetermined wavelengths from the third light source.
  • the HOE is located between the third light source and the two other light sources and the sample.
  • the HOE is designed to deflect light from the first and second light sources onto the sample and to let light from the third light source pass through, so that preferably light from all three light sources hits the sample area and in particular the sample from the same direction .
  • the HOE thus preferably has no significant effect on light of predetermined wavelengths from the third light source, with the exception of effects due to interface and material properties that cannot be completely avoided (Fresnel reflection and slight scattering and absorption).
  • further light sources can be placed on the same side of the HOE as the first and second light sources or on the same side as the third light source, so that selected light from these light sources is also deflected via the HOE or guided through the HOE.
  • the HOE can be set up to allow light of predetermined wavelengths to pass through the HOE from a predetermined direction and to change the direction in the process, so that an effect similar to the diffraction of light occurs.
  • the device comprises one or more additional optical elements, in particular diaphragms, lenses and/or shutters or other beam-shaping and/or spectrally filtering elements as described above.
  • additional optical elements in particular diaphragms, lenses and/or shutters or other beam-shaping and/or spectrally filtering elements as described above.
  • These further optical elements can be arranged with respect to the holographic-optical component in such a way that light from the light sources deflected by the HOE is modified, in particular bundled, focused and/or deflected. This has the advantage that the light deflected by the holographic-optical component can be further adjusted before impinging on a sample.
  • the first light source and the second light source are on a shell of a cone or a Truncated cone, in particular on a circle, arranged around the exit axis of the holographic-optical component.
  • the exit axis is to be understood in particular as the direction in which the HOE, due to its design, emits or deflects light as intended.
  • the holographic-optical component can preferably be arranged at or in the tip of the cone or the truncated cone.
  • the jacket can only be understood to mean the imaginary geometric shape of the geometric object of a cone or a truncated cone.
  • the first and/or the second light source can actually be arranged on or in a casing of a preferably at least partially conical, pyramidal, truncated cone-shaped or truncated pyramid-shaped component.
  • This has the advantage that rotationally symmetrical molded parts can be produced cost-effectively and precisely, and that the adjustment effort is significantly reduced due to the utilization of the geometry of the molded part.
  • the first light source and the second light source are arranged on a circle around the exit axis of the holographic optical component.
  • this component can also be a hollow body.
  • the light sources and/or the HOE can be attached inside, in particular on or in an inner wall, of the hollow body.
  • the device has one or more mirrors.
  • the mirrors are arranged with respect to the light sources and the holographic-optical component in such a way that light emitted by the light sources is directed via the mirrors onto the holographic-optical component.
  • the mirrors can preferably be arranged on or on the jacket of the component described above. If the device has a hollow body as described above, one or more mirrors can be attached to an inside of the hollow body, in particular to an inside of the casing of the hollow body. A part of the casing can also be designed as a mirror, for example due to a reflecting area of a surface of the casing, for example a reflecting metallic surface.
  • the one or more mirrors may be part of a tapered cladding, in particular part of a cladding of a preferably designed as a hollow cone, truncated cone, pyramid or truncated pyramid.
  • the device has a transparent body which is arranged in such a way that light from at least one light source can be guided through the transparent body, preferably using total reflection within the body.
  • the transparent body can be arranged between at least one light source, preferably between both light sources, and the HOE.
  • the transparent body can consist at least partially of glass or transparent plastic.
  • the HOE can be attached or arranged on or in the transparent body. This has the advantage that no separate substrate is required for the placement of the HOE.
  • At least one light source and the HOE adjoin the transparent body, which body can be shaped as a plate.
  • This has the advantage that light coupled into the body by the light source can be guided through the body to the HOE, preferably using total reflection.
  • the HOE can have, for example, a reflection hologram or a transmission hologram for coupling the light out of the body.
  • the light sources are arranged on a common circuit board. This has the advantage that only one component is required for a well-defined placement of the light sources.
  • the holographic-optical component is designed to collimate or focus incident radiation.
  • the incident radiation can be a plane wave, diverging radiation or converging radiation.
  • the diverging radiation can in particular be part of a radiation with a spherical wave front, such as is emitted by an approximately point light source, for example.
  • the invention also relates to a device for examining a sample, in particular a biological one, the device comprising a device according to the invention and a measuring device for detecting light emitted by a sample taken, in particular fluorescent light.
  • the invention also relates to a method for irradiating a sample, in particular a biological one, with the device according to the invention, i.e. a method in which first light from a first light source and second light from a second light source are directed via a holographic-optical component onto a common sample area, in particular onto a common light path, are directed to irradiate the sample.
  • the method according to the invention is particularly suitable for use in lab-on-a-chip platforms, preferably for fluorescence measurements in (quantitative) real-time PCR with multiplex functionality, for example in point-of-care solutions.
  • FIG. 7 shows a flow chart of an exemplary embodiment of the method according to the invention.
  • FIG. 1 shows a first exemplary embodiment of the device 100 according to the invention.
  • the device 100 comprises a holographic-optical component 150 ("HOE"), a first light source 110 and a second light source 120 in order to illuminate a sample 210 placed in a sample area 200.
  • the sample comprises, as described above, DNA sections amplified via the polymerase chain reaction, which are to be detected by exciting fluorescent probes in the sample.
  • FIG. 1 also shows an exemplary embodiment of the device 1000 according to the invention, comprising the exemplary embodiment of the device 100 according to the invention and a measuring device 300, the measuring device 300 being set up, for example as a camera, in order to capture fluorescent light emitted by the sample 210.
  • the HOE 150 includes one or more holograms, such as volume holograms, which may be embedded in one or more photopolymer layers.
  • the photopolymer layers can be applied to a suitable support, for example a glass plate, or embedded between two supports, for example in laminated form.
  • the hologram can be exposed into the layer(s) according to the desired arrangement of the light sources relative to the HOE and the sample area.
  • the hologram can be embossed into the layer, for example with the aid of an embossing template (also called “master”).
  • the first light source 110, the second light source 120 and the holographic-optical component 150 are arranged in relation to one another in such a way that first light 10 from the first light source 110 and second light 30 from the second light source 120 via the HOE 150 a common sample area 200 for the irradiation of the sample 210 can be directed can.
  • the HOE 100 is designed in such a way that it directs planar light waves, in particular collimated radiation, of a first wavelength from a first direction onto a common sample area 200 .
  • "light waves of a ... wavelength” means in particular light waves with wavelengths from a wavelength interval of finite width, ie in particular light waves with wavelengths from a specific wavelength spectrum of finite width.
  • the HOE 150 is specified in this example to redirect planar light waves (collimated rays) of a first wavelength from a first direction 10 to a second direction 20, preferably perpendicular to the surface of the HOE 150 as shown in FIG.
  • the HOE 150 is also designed in such a way that planar light waves of a second wavelength are also deflected from a third direction 30 into the second direction 20, ie both light paths are combined.
  • a reflection hologram is preferably used, as shown, because in practice it is usually possible to achieve better quality with regard to diffraction efficiency and light scattering.
  • the device 100 can also be configured with a transmission hologram.
  • the HOE 150 can in particular (additionally) be configured to carry out a wave selection depending on the direction of the light incident on the HOE 150 .
  • two light sources 110, 120 are mounted in such a way that they can illuminate the HOE 150, with the wavelength and direction of incidence having to correspond to the design of the HOE 150 hologram.
  • the first light source 110 may be an LED that emits light having a center wavelength of approximately 472 nanometers (nm) and a FWHM of 15 nm.
  • the second light source 120 could also be an LED, for example, which emits light with a central wavelength of approximately 530 nm and a full width at half maximum of 32 nm.
  • the HOE 150 is designed to deflect light with a wavelength of 472 nm coming from the first light source 110 and light with a wavelength of 530 nm coming from the second light source 120 in a common direction 20 towards the sample region 200 .
  • the efficiency of the HOE 150 hologram typically follows one Lorentz distribution with a width of 15 nm, for example, around the respective central wavelength, which does not necessarily have to match the central wavelengths of the LEDs 110, 120 exactly.
  • the HOE 150 preferably restricts the deflected radiation to the range given by its own FWHM and keeps it dependent on operation, in particular dependent on temperature and current supply, and despite the central wavelengths of the LEDs that vary due to manufacturing tolerances.
  • the HOE 150 thus combines the two from different directions 10,
  • the exemplary embodiment shown in FIG. 1 for two light sources 110, 120 and two directions 10, 30 with two different wavelength spectra can be expanded to include additional light sources if required.
  • the symmetry of the light sources 110, 120 shown in FIG. 1 in relation to the HOE 150 is not mandatory.
  • the light sources 110, 120 can be arranged with respect to the HOE, in contrast to what is shown, in such a way that light emitted by them forms different angles of incidence to a surface normal of the HOE 150.
  • the HOE 150 can also be designed so that, at predetermined angles of incidence, the light incident from the light sources 110, 120 is not deflected along the normal to the surface of the HOE150, as shown, but at a different angle to the normal to the surface.
  • the device 100 can preferably have two to six light sources, particularly preferably four light sources.
  • the two or more light sources can be arranged on a circle around an axis perpendicular to the HOE, in particular around a surface normal of the HOE, which lies in the second direction 20 according to FIG. 1, as explained further below.
  • This has optical, structural and manufacturing advantages.
  • the maximum number of light sources is only limited for practical reasons, in particular due to limited installation space and a decreasing efficiency of the HOE 150 with an increasing degree of multiplexing.
  • optical collecting elements 111, 121 collimated or bundled depending on the optical design of the hologram of the HOE 150 and beam path.
  • Refractive or diffractive optical elements known from the prior art can be used here, for example simple lenses, parabolic converging lenses, Fresnel lenses, HOEs or combinations of these elements.
  • the device 100 can have bandpass filters 112, 122, as shown in FIG.
  • bandpass filters 112, 122 are placed after the collection elements 111, 121 where the rays emitted by the light sources 110, 120 have as narrow an angular distribution as possible.
  • the space of light source 110, 120 and collecting optics 111, 121 is optically separated from the space of HOE 150 with bandpass filters 112, 122 (e.g. by shielding surfaces or in a tube, as shown in Figure 1), so that no light components from light source 110, 120 to HOE 150 can pass bandpass filter 112, 122.
  • the optional bandpass filters 112, 122 have the advantageous effect of restricting the spectral range of the light sources 110, 120 in a narrower band and with steeper edges than is usually possible due to the intrinsic spectral width of the hologram of the HOE 150. Furthermore, scattered light with undesired wavelengths can be reduced in this way.
  • the device 100 can have a further collecting element 220, in particular a collecting lens, for focusing the beams 20 onto the sample 210.
  • the device 100 can include further optical elements such as diaphragms, lenses and shutters in order to further shape the combined beam path 20 after the HOE.
  • the exemplary embodiment shown in FIG. 1 is particularly suitable for deflecting plane waves or collimated beams. This is particularly advantageous if, in addition to cleaning up scattered light, interference filters 112, 122 (bandpass or edge filters) are to be used, for example in front of a single light source or also a multibandpass filter downstream of the HOE 150.
  • interference filters 112, 122 bandpass or edge filters
  • the device 100 also makes it possible not only to implement a wavelength-selective deflection with the HOE 150, but also to shape the wave front of the incident light.
  • FIG. 2a shows an exemplary embodiment in which the spherical wave 11, 31 emanating from the respective light source 110, 120 is deflected into a spherical wave 21 running towards the sample 210.
  • the divergent light of the light sources 110, 120 is collected by the HOE 150 over a large cross-sectional area and focused on the sample 210.
  • exemplary embodiments of the device 100 can also be realized in which the HOE 150 provides a comparatively plane wave either on the sample or light source side and enables the integration of a bandpass filter on this side, but has a collecting or focusing effect on the other side.
  • the symmetry of the light sources 110, 120 shown in FIG. 2a in relation to the HOE 150 is also not mandatory.
  • FIG. 2b shows a development of the exemplary embodiment from FIG. 2a, which has an additional third light source 130 in addition to the first two light sources 110, 120, light from the third light source 130 preferably irradiating the sample without being influenced by the HOE.
  • the third light source 130 can be arranged on the side of the HOE 150 facing away from the first two light sources 110 , 120 .
  • the HOE 150 can preferably be designed to be transparent to light 40 of a predetermined wavelength range from the third light source 130 .
  • the HOE 150 can be implemented in such a way that it redirects light of certain wavelengths from certain directions to a desired target direction while allowing light of certain other wavelengths and/or from certain other directions to pass in the same target direction substantially unhindered.
  • the light sources 110, 120, 130 could have peak wavelengths of 470 nm, 530 nm, 590 nm, 630 nm, 405 nm or 385 nm. If the holographic material is not suitable for one of the wavelengths desired for the application, for example for 385 nm, it is possible to provide the source 130 with that wavelength according to FIG. 2b.
  • the device 100 can have further lenses 131, 132 and/or bandpass filters 133 arranged in front of the third light source 130 in order to collect/scatter or filter light from the third light source as required.
  • the HOE 150 can be designed to shape, in particular to filter, bundle, light of predetermined wavelengths from the third light source 130 and in special configurations, as already explained above, also light of predetermined wavelengths from the first and/or second light source 110, 120. scatter and/or bend.
  • Figure 2c illustrates a beam shaping of light emitted by the third light source 130 by the HOE 150.
  • the HOE 150 is designed to diffract a predetermined wavelength spectrum 300 at the short-wave edge 310 and/or at the long-wave edge 320 (and thereby effectively cropping the spectrum 300) and/or condensing light 330 passing through the HOE 150 analogously to a converging lens.
  • An intrinsic spectrum of the light source 130 can thus be effectively tailored to a narrower, preferably steeper shape, which advantageously leads to a spectrum incident on the sample 210 which is more robust to fluctuations in the temperature, operating current or component tolerance of the light source 130 .
  • the HOE can also be designed to convert the light components 310, 320 that are bent away into one internal mode and from there preferably to be converted into an absorber material or a beam trap in order to avoid scattered light.
  • a plurality of light sources 110, 120 are arranged on the jacket 161 of a truncated cone 160 in such a way that their optical axes point to the center of the HOE 150 without further adjustment.
  • the light sources 110, 120 are arranged along a circle 162 about the exit axis 20 of the HOE 150.
  • the truncated cone 160 is preferably arranged rotationally symmetrically about the exit axis 20 of the HOE 150 , the exit axis being perpendicular to the planar HOE 150 , ie congruent with a surface normal of the HOE 150 .
  • the first light source 110 and the second light source 120 are arranged on a common circuit board 170, which is advantageous in terms of manufacturing technology.
  • the printed circuit board 170 can in particular be a substrate that is customary for optical components, for example a circuit board that is customary in electronics.
  • the circuit board can have a core made of a material with good thermal conductivity for dissipating the waste heat, for example comprising aluminum.
  • the collimated beams 10, 30 can then be directed onto the HOE 150, as shown, via an at least partially mirrored surface on the inside of a truncated pyramid 160 or truncated cone 160, or alternatively via mirrors 163 attached to the inside.
  • a mirroring of the inner surface of the stump 160 can be realized, for example, by means of reflective metal surfaces, in particular by means of vapor-deposited aluminum or silver.
  • the reflective surfaces could also be curved and thus also fulfill an optical function.
  • the HOE 150 can likewise be arranged on or in the stump 160 .
  • the device 100 can also have a solid, transparent body 180 in which the beam deflection via total internal reflection.
  • the body 180 can have glass or transparent plastic.
  • Figure 5a shows a further exemplary embodiment of the device 100 according to the invention, comprising a plurality of light sources, in particular light-emitting diodes, in this example four light sources, 110, 120, 130, 140.
  • the LEDs 110, 120, 130, 140 are arranged on a circuit board 171, for example in a two-by-two grid arrangement, as shown in the plan view of circuit board 171 in Figure 5b.
  • the device 100 can also have more or fewer light sources, which are arranged in particular at a minimal distance from one another, as described below.
  • the HOE 150 is arranged opposite a surface of the circuit board 171, which is preferably designed to direct perpendicularly incident light 10, which in this example is approximately spherical, due to the approximately point-shaped light sources 110, 120, 130, 140 into a common sample area, in particular in a common direction 20 , at an angle 175 to the surface normal 174 of the HOE 150.
  • the angle 175 can have a value between 40 and 70 degrees, preferably between 50 and 50 degrees.
  • the angle should preferably be selected in such a way that Fresnel reflections are avoided, which occur in particular when the angle of incidence on the hologram corresponds to the angle of reflection.
  • a distance 126 between circuit board 171 and HOE 150 is preferably at least three times greater, more preferably at least five times greater than a distance 127 between the light sources 110, 120, 130, 140. It is therefore also advantageous if the light sources 110, 120, 130, 140 are placed as close together as possible in order to achieve a good overlap of the emission cones.
  • the device 100 can also have an aperture 124 in order to bring the light deflected by the HOE 150 to a congruent area as far as possible and thus to realize an illumination situation of the sample area 200 that is as identical as possible for all light sources.
  • the light beams are propagated in the carrier medium of the hologram or the HOE, usually a glass plate.
  • Such an arrangement is shown in a further exemplary embodiment of the device 100 according to the invention in accordance with FIG.
  • the device 100 comprises a transparent plate 190 as the transparent body 180, to which the two light sources 110, 120 in the form of LEDs adjoin with as little air gap as possible (possibly mediated by a layer adapted to the refractive index).
  • the transparent plate 190 can be, for example, a glass plate 190 or another transparent carrier medium, for example based on transparent plastic.
  • the light coupled into the transparent plate 190 by the light sources 110, 120 is deflected onto the common sample area 200 by a HOE 150, which is also adjacent to the plate 190 and comprises a reflection hologram.
  • the device can have further HOE 151,
  • the HOE150 could also include a transmission hologram and/or be placed on the opposite side of the plate 190.
  • the first HOE can also extend over a greater width along the plate 190 and thus replace the further HOE 151, 152.
  • FIG. 7 shows a flow chart 600 for an exemplary embodiment of the method 600 according to the invention, which can be carried out, for example, with one of the exemplary embodiments according to FIGS.
  • a first step 601 of the method 600 the device 100 according to the invention is provided.
  • a sample 210 can be placed in the common sample area 200 .
  • a third In step 603 light from the first light source 110 and thereafter or simultaneously light from the second light source 120 is directed via the HOE onto the common sample area in order to excite fluorophores in the sample 210, for example.
  • a fourth step 604 which can take place at the same time as the third step 603, the sample 210 is sent out
  • Fluorescence radiation is detected with the measuring device 300, for example a camera 300 or a photodiode, for a subsequent analysis of the sample 210.

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Abstract

L'invention concerne un appareil (100) et un procédé (600) pour irradier un échantillon notamment biologique (210), comprenant une première source de lumière (110), une seconde source de lumière (110) et au moins un composant optique holographique (150), la première source de lumière (120), la seconde source lumineuse (120) et le composant optique holographique (150) étant positionnés l'un par rapport à l'autre de telle sorte que la première lumière (10, 11) provenant de la première source lumineuse (110) et la seconde lumière (30, 31) provenant de la seconde source lumineuse (120) sont déviées par le biais du composant optique holographique (150) sur une zone d'échantillon commune (200) pour l'irradiation de l'échantillon (210).
EP22716224.5A 2021-03-23 2022-03-18 Appareil, dispositif et procédé d'irradiation d'un échantillon biologique particulier, avec composant optique holographique Pending EP4314775A1 (fr)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
DE102021202794 2021-03-23
DE102022202634.9A DE102022202634A1 (de) 2021-03-23 2022-03-17 Vorrichtung, Einrichtung und Verfahren zur Bestrahlung einer insbesondere biologischen Probe mit einem holografisch-optischen Bauelement
PCT/EP2022/057119 WO2022200202A1 (fr) 2021-03-23 2022-03-18 Appareil, dispositif et procédé d'irradiation d'un échantillon biologique particulier, avec composant optique holographique

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EP4314775A1 true EP4314775A1 (fr) 2024-02-07

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EP22716224.5A Pending EP4314775A1 (fr) 2021-03-23 2022-03-18 Appareil, dispositif et procédé d'irradiation d'un échantillon biologique particulier, avec composant optique holographique

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DE102021212505A1 (de) * 2021-11-08 2023-05-11 Robert Bosch Gesellschaft mit beschränkter Haftung Optische Vorrichtung zum Anregen einer Probe, Analysegerät und Verfahren zum Anregen einer Probe

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US6744502B2 (en) * 2001-09-28 2004-06-01 Pe Corporation (Ny) Shaped illumination geometry and intensity using a diffractive optical element
JP2004340733A (ja) * 2003-05-15 2004-12-02 Minolta Co Ltd 測定装置
US20170343476A1 (en) * 2016-05-31 2017-11-30 Molecular Devices, Llc Imaging system with oblique illumination

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