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
  • G01N21/6452—Individual samples arranged in a regular 2D-array, e.g. multiwell plates
• • 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
  • G01N21/6452—Individual samples arranged in a regular 2D-array, e.g. multiwell plates
  • G01N21/6454—Individual samples arranged in a regular 2D-array, e.g. multiwell plates using an integrated detector array
• • 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
  • G01N2021/6417—Spectrofluorimetric devices
  • G01N2021/6421—Measuring 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
  • 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

• the invention relates to a device for supporting chromophoric elements.
• the chromophoric elements are chemical or biological molecules which are generally fixed on a substrate after a hybridization or affinity reaction in a liquid, or else elements dyes added or grafted to these molecules or certain types of nanostructures semiconductor such as son or quantum boxes, each chromophore element being adapted to emit light spontaneously (it is the or bio- or chemiluminescence) or in response to a light excitation (this is fluorescence) over a determined wavelength which depends on the nature of this chromophore element.
• Support devices for chromophoric elements are described in particular in application WO-A-02/16912 in the names of Claude WEISBUCH and Henri BENISTY and include means making it possible to reinforce the light intensity of excitation of the chromophoric elements and to increase the light intensity emitted by these elements, by interference effects produced by stacks of layers of judiciously chosen materials and by effects of light extraction guided by lateral structures having dimensions of the same order of magnitude as the wavelength of guided light, such as photonic crystals.
• the chromophoric elements are fixed on areas of the substrate (called “spots” in English terminology) separated from each other and arranged regularly, in particular in rows and columns. These ranges have dimensions, for example of a few tens or hundreds of ⁇ m approximately, which are clearly greater than the wavelengths considered.
• the ranges where the chromophoric elements are fixed for example during the hybridization step are delimited for example by a deposition technique of the "spotting" type which comprises a physicochemical treatment of the surface of the overall substrate, the "spot” then being determined by the area wetted by the fluid deposition, or by a selective spatial treatment for example by selective silanization, the "spot” being determined by this treatment.
• the ranges can comprise chromophoric elements of different types, in general two, for example Cy3 and Cy5 which emit on different wavelengths.
• the signals emitted are picked up by suitable photodetectors, in particular by arrays or arrays of photodetectors of the CCD type which also pick up an overall background noise formed by an excitation light which is incompletely filtered, by a fluorescence coming from chromophoric elements of ranges. neighbors according to grazing or guided rays, etc., this background noise being difficult to eliminate with precision for each wavelength considered and can represent a significant part of the intensity of the signals picked up.
• the information carried by the light emitted by the chromophores can be read, depending on the type of apparatus used, on one side or the other of the support.
• the measurement can be made on dried biological material or else in the liquid phase, for which the chromophoric elements carrying the information are those specifically hung on pads of the support, the liquid comprising in its volume a certain quantity of chromophores in non-informative suspension and forming a source of background noise at the emission wavelengths of chromophores as well as diffusing particles, also sources of background noise.
• the object of the present invention is in particular to provide a simple and effective solution to the problem of determining and eliminating this background noise.
• the invention also aims to optimize the structure of the substrate for its use at several different wavelengths corresponding to different types of chromophores.
• a device for supporting chromophoric elements which allows a reliable, precise and automatic determination of the above-mentioned background noise.
• a device for supporting chromophoric elements capable of emitting light by chemiluminescence or by fluorescence in response to a light excitation, the wavelength emitted by each chromophore element depending on the nature of this element, these chromophoric elements being fixed on areas separated from each other, characterized in that the surface of the support is structured in several zones having different optical properties in amplitude and in phase of the reflectivity and in transmission, these properties resulting from the presence or the absence, on the substrate areas, of at least one set of layers chosen from the following:
• At least one layer absorbing at least partially at least one of the emission and / or excitation wavelengths, - at least one layer transparent to all the emission and excitation wavelengths, said layers being determined to produce at least one of the following effects:
• the device according to the invention therefore comprises different optical environments, which will mix the useful signal differently with the background noise, which makes it possible, thanks to appropriate digital processing, to reconstruct the useful signal.
• a first type of zone can produce the measurement
• Measurement (1) ai " Signal + b1 " noise (A) while a second type of zone will produce the measurement
• Measure (2) a2vSignal + b2 » noise
• B the coefficients ai and b1 being the transfer parameters of zone 1 at the wavelength considered, a2 and b2 being those of zone 2. These parameters are known by construction or by calibration. It then suffices to solve the system of two equations with two unknowns (A, B) to deduce therefrom the sought after values "Signal” and "noise".
• the value of the coefficients ai and a2 can be varied by using, for example, amplifier layers for the emitted light and / or the excitation light, as described in application WO-A-02/16912 in the names of Claude WEISBUCH and Henri BENISTY, on the basis of destructive or constructive interference effects.
• the dimensions of these zones can, depending on the applications, be greater, less than or equal to those of the above-mentioned ranges. In the case where the zone is smaller than the range, the resolution of the Signal-Noise system is done locally on the same range including the different zones. Otherwise, the Signal-Noise system is resolved by comparing the measurements made on different ranges located in different areas.
• the background noise corresponding to the liquid phase can have an intensity of the same order of magnitude as that of the useful signal, or even greater. This is one of the reasons why measurements in the presence of the liquid phase are not usually made.
• the invention makes it possible to effectively solve this problem, which in this case represents more than a simple improvement in performance and corresponds to a new type of measurement and apparatus.
• This solution also makes it possible to provide a temporal resolution to the measurement since the process of attachment or hybridization of the chromophoric elements can thus be analyzed throughout its course.
• the structuring of the support can be carried out by usual techniques of lift-off lithography and / or dry or wet etching.
• a particular embodiment corresponds to a relatively simple structure (no absorption layer) produced by modulating the thickness of a transparent layer located above a reflective layer.
• the device comprises a plane mirror covered with a layer of material transparent to the lengths of emitted waves and on which the chromophoric elements are distributed in ranges separated from each other and having lateral dimensions greater than the wavelengths of the emitted fluorescence, characterized in that said layer of transparent material has a thickness of the same order of magnitude as the wavelengths of the fluorescence emitted and comprises, for each range of chromophoric elements, at least two zones having different thicknesses, the thickness of a first of these zones being determined to generate by a phenomenon d 'destructive interference a minimum intensity of the fluorescence emitted over a wavelength by chromophoric elements of said zone.
• the fluorescence emitted on the surface of this zone is at a minimum value which is zero or substantially zero. Consequently, the light signal picked up on the surface of this zone represents the overall noise at the wavelength considered.
• the two above conditions are close and correspond to thicknesses which typically differ by less than 30 nm, at any intermediate condition between the two above conditions.
• the two effects can also coincide if the angle of incidence of the excitation light is chosen for this purpose.
• the transparent material layer comprises, at least one other zone having a thickness different from that of the first zone, such as the difference of the optical paths in these two zones for a wavelength considered is equal to about an odd multiple of a quarter of this wavelength.
• This zone allows maximum amplification of the fluorescence emitted.
• the wavelength considered for determining the difference in thickness between the two zones can be, as indicated above, either the wavelength of the fluorescence emitted, the excitation wavelength, or both the wavelength of the emitted fluorescence and the excitation wavelength (to obtain a maximum of excitation intensity and a maximum of emitted fluorescence intensity on said other zone), i.e. a further intermediate condition when the thicknesses of the two regions calculated for the two wavelengths are very "close to one another.
• the light signal picked up on the surface of this other zone corresponds to the sum of the maximum intensity of the fluorescence emitted at the first length of wave on the surface of this other area and overall background noise.
• the layer of transparent material comprises, for each range of chromophoric elements, a plurality of the aforementioned zones of different thicknesses, making it possible to sample between a minimum value and a maximum value the intensity of the fluorescence emitted on said first wavelength by chromophoric elements of said range
• - the layer of transparent material comprises, for each range chromophoric elements, a plurality of the aforementioned zones of different thicknesses, making it possible to vary the intensity of the fluorescence emitted on different wavelengths by chromophoric elements of different types from said range.
• said zones are arranged in lines or in strips parallel to the surface of said layer of transparent material.
• these zones may have a matrix arrangement in rows and columns on the surface of the layer of transparent material. This then comprises, over its entire surface, a plurality of zones of different thicknesses with preferably regular distribution which form a structure of the paving type or the like.
• these zones of different height can be formed on the aforementioned reflecting layer or on an intermediate layer of different index interposed between the transparent layer and the reflecting layer.
• the means for capturing the fluorescence emitted by the chromophoric elements can be located above the device for supporting these chromophoric elements, or even below, as already described in the aforementioned international application WO-A-02/16912.
• these capture means may comprise a matrix of photodetectors of the CCD or CMOS type which is fixed under the device, the latter comprising a first layer of material highly reflecting at the excitation wavelength and a second layer of material selectively absorbing the excitation radiation, the first layer being deposited on the second, so that the emitted fluorescence easily reaches the detectors, but not the excitation radiation.
• the reflection of the excitation radiation on the first layer can then possibly provide the aforementioned effect of reinforcing the emitted fluorescence.
• a weakly resonant cavity is formed between the lens on the upper surface of the layer carrying the chromophoric elements and said first reflective layer (at the excitation wavelength) when the latter also has a non-negligible reflectivity at length. of the fluorescence emitted. This effect can be used to increase the intensity of the fluorescence channeled to the photodetectors.
• the upper layer of this device is made of a material with a high refractive index. This promotes the formation of said weakly resonant cavity and therefore good detection of fluorescence by the photodetectors fixed under the device.
• the accuracy of the measurement can also be improved. performed following the resolution of the system of equations (A) and (B) above.
• the device can be designed to have the same useful signal level on the two types of zone.
• the background noise at the emission wavelength is negligible.
• Measure (2) a * Signal + noise (ex.)
• the useful signal free of noise due to excitation light can be easily found by subtracting the measurement carried out on the first type of zone from twice the measurement carried out on the second:
• One zone maximizes the emission of a first type of chromophore and the other zone minimizes this emission.
• the first type of chromophore is treated in a targeted manner, the second type of chromophore being treated generically; or one zone maximizes the emission of a first chromophore and the other zone maximizes the emission of a second type of chromophore.
• the two types of chromophores are processed in a targeted manner for the signal, the noise being processed generically.
• the invention also applies to chemiluminescent compounds.
• the structuring of the surface of the support only takes into account the emission wavelengths.
• the invention also applies also to the format of microplates ("SBS" format) (for example 24, 96, 384 or 1536 wells), the structure of the surface of the support then being adapted to the geometry wells of the microplates so as to present one or more zones per well.
• SBS format of microplates
• the structured support constitutes the bottom common to all the wells of a microplate. In another embodiment, individual supports are deposited at the bottom of each well of a monolithic microplate.
• the invention also applies to microplates in the format of microscope slides with micro-wells produced by depositing a layer with a thickness of several tens of microns with orifices forming these wells (for example HTC treatment of "Teflon” type Cel-Line brand sold by Erie Scientific Corp., Portsmouth, NH).
• the different wells can be used as separate hybridization zones for different test samples.
• each well may include one or more areas where the chromophoric elements are fixed.
• FIG. 1 is a schematic representation from above of a part of a device according to the invention.
• FIG. 2 is a partial view on a larger scale of Figure 1;
• FIG. 3A, 3B and 3C are views corresponding to Figure 2, for alternative embodiments of the invention;
• FIG. 4 is a partial schematic perspective view of another alternative embodiment
• FIGS. 5A and 5B are partial schematic sectional views of two embodiments of a device according to the invention.
• - Figure 6 is an enlarged view of part of the section shown in Figure 5A;
• FIG. 7 is a graph schematically representing the intensities of the fluorescence emitted on the surface of three different zones of the device of Figure 6;
• the device shown diagrammatically in FIG. 1 comprises a support 10 of generally rectangular shape, the upper face 12 of which comprises a plurality of pads 14 on which chromophoric elements are fixed, these pads 14 forming an assembly where they are distributed in regular lines and in columns for example.
• the dimensions of the areas 14 are of the order of 30 to 400 ⁇ m, and the spacing D between adjacent areas is of the order of 40 to 500 ⁇ m. As indicated above, the dimensions of the ranges are determined by a fluid deposition or by a selective spatial treatment.
• the support can be made of glass, silicon, silicon carbide, sapphire (AI203), metal or plastic.
• the upper part of the support 10 comprises a layer 12 of a material which is transparent to the wavelengths of the fluorescence emitted by the chromophoric elements of the pads 14 in response to a light excitation, the layer 12 comprising at least dielectric material such for example a semiconductor material, an oxide, a glass, a nitride, a fluoride, a chalcogenide, an organic polymer or a mineral or organometallic compound obtained by sol-gel route.
• This material preferably has a relatively high refractive index and is for example formed of TiO2 having a refractive index of between 2.2 and 2.5 depending on the crystalline form used.
• the layer 12 is made of SiO 2 to optimize the quality of the chemical functionalization of the surface of the support.
• the transparent layer (12) can also be made of organic polymer with a flat or rough surface ("3-D" effect). In the latter case, the 3-D effect increases the effective surface of the device.
• the transparent layer (12) can also be porous.
• This layer 12 has a thickness which is of the same order of magnitude as the wavelengths of the fluorescence emitted by the chromophoric elements and covers a plane mirror which can be reflective for the excitation wavelength, this plane mirror being above all reflecting for the wavelengths of the fluorescence emitted.
• the free surface or upper surface of the layer 12 is structured for example as shown diagrammatically in FIGS. 2, 3 and 4.
• this layer 12 comprises a plurality of parallel bands 16 of different thicknesses, these bands having a width in the plane of the layer 12 which is greater than the wavelengths of the fluorescence emitted by the chromophoric elements and less than dimensions in the plane of the ranges 14.
• the thicknesses of the different bands 16 are determined so that one of these thicknesses produces, in each range 14, a destructive interference effect on the surface of the layer 12 for a wavelength excitation and / or for a given wavelength of the fluorescence emitted by the chromophoric elements.
• the other bands 16 have different thicknesses, one of which corresponds to a constructive interference effect on the upper surface of the layer 12, for the excitation wavelength and / or for the fluorescence wavelength issued.
• the bands 16 of different thicknesses are formed alternately in the layer 12, their thicknesses being determined to produce the aforementioned effects of destructive interference and constructive interference for one or preferably for several wavelengths emitted by the different elements chromophores present in ranges 14 and / or for the corresponding excitation wavelengths. It is therefore possible, considering a given wavelength emitted by chromophoric elements of a range 14, to determine two thicknesses corresponding to the two aforementioned interference effects and one or more intermediate thicknesses, which makes it possible to sample the light intensity emitted at this wavelength between a minimum value and a maximum value.
• each area 14 comprises a plurality of adjacent plates 20 of square or rectangular shape which have different heights.
• the dimensions of these plates 20 at the upper surface of the layer 12 can be identical from one plate to another or different.
• the structuring of the support can also be carried out as shown diagrammatically in FIGS. 3B and 3C with zones 16 having dimensions greater than those of the pads 14 of chromophoric elements.
• the layer 12 of transparent material carrying the chromophoric elements C is formed on a plane mirror 22 which is highly reflective at least for fluorescence emitted by the chromophoric elements.
• the plane mirror is formed from one or more layers of a reflecting metal or a dielectric material such as for example a semiconductor material, an oxide or a glass, a nitride, a fluoride, a chalcogenide. organic or a compound obtained by sol-gel route from mineral or organometallic compounds.
• the plane mirror 22 is made of silicon.
• the plane mirror 22 comprises at least one metallic layer deposited on the support and for example aluminum, gold, silver or chrome.
• Metallic mirrors are generally completely opaque in the visible range of the light spectrum.
• this plane mirror comprises at least two layers of oxides such as, for example, SiO2 and TiO2.
• TiO2 can be replaced by Nb2O5, Ta205 or Hf2O5.
• the plane mirror 22 comprises at least one layer of SiO2 and at least one layer of amorphous silicon.
• the determination of the thicknesses of the bands 16 leading to constructive and destructive interference respectively must take account of the length of penetration (and therefore of the phase change at reflection) of the excitation or fluorescence at the length of wave considered in the mirror 22, as well as the reflectivity of this mirror and the index of the transparent layer 12.
• the mirror 22 can for example be a dielectric mirror (Bregg mirror), well known to those skilled in the art, characterized by a reflectivity greater than 70% and by a Bragg wavelength (on which the Bragg mirror is centered).
• the Bragg mirror can thus be centered on the excitation wavelength or on the emission wavelength of a type of chromophoric element or else on a wavelength intermediate between these wavelengths .
• the Bragg mirror can be centered around 655nm.
• the mirror can be centered on an intermediate wavelength between the emission and / or excitation wavelengths of the different types of chromophores.
• the length of centering wave of the mirror can be chosen at 605nm.
• a stack of the dielectric layers Si02 and TiO2 or else SiO2 and Nb205 is used, having particularly high index differences.
• the reflective layer 22 is an optical microcavity which comprises two mirrors (dielectric or metallic) separated by a transparent layer ("the cavity") having an optical thickness 2 * n * ⁇ c / 4, where n is an integer and the wavelength ⁇ c is chosen in the spectral range where the reflectivity of the two mirrors is high.
• the structuring of the device can be obtained for example by the modulation of the thickness between different zones of the transparent layer covering the stack or else by the modulation of the thickness of the layer of the cavity.
• the reflective layer 22 is a structure with multiple optical microcavities, for example having three mirrors and two cavities.
• Bragg mirrors or microcavity structures are generally semi-transparent stacks in the visible range of the spectrum.
• the signals which can be picked up above the zones of different thicknesses of a range 14 have been shown diagrammatically in FIG. 7, the intensity I being represented on the ordinate and a dimension in the plane in the layer 12 being represented by abscissa.
• the curve of FIG. 7 comprises a first part 24 of minimum intensity corresponding to a destructive interference effect, a part 26 of maximum intensity corresponding to a constructive interference effect, and a part 28 of average intensity which corresponds for example to the signal which would be obtained in the absence of plane mirror 22, that is to say say in the absence of any interference phenomenon.
• a variant of the invention consists in structuring the mirror 22 into zones by eliminating the reflective layer in certain zones, that is to say by creating orifices in this layer.
• pairs of signals 26 and 28, or else 24 and 28, will be used as modulation of the signal to be detected, depending on the thickness chosen for layer 12, with a flat upper face.
• FIG. 8 shows an alternative embodiment of the invention, in which a set of photodetectors 30 of the CCD type or the like is located under the support 10, on the face opposite to that which carries the chromophoric elements C.
• This embodiment has the advantage of not requiring an image forming objective on the photodetectors 30.
• the different layers (transparent, reflective and / or absorbent) of the support can be deposited on a matrix of CCD elements.
• the emission of fluorescence downwards towards the photodetectors 30 is modulated by the same interference effects as those described above, but the amplitude of these effects is determined by a different physical mechanism and is in lower general: this is multi-wave interference linked to the fact that the layer
• the layer 12 forms a weakly resonant cavity, one of the mirrors of which is formed by the diopter with the air or the surrounding medium on the upper surface of the layer 12 and the other mirror of which is formed of a layer 32 which is semi reflecting at the fluorescence wavelength which it is desired to detect and which is very reflecting for the excitation wavelength of the chromophoric elements.
• the strength of the resonance of such a cavity and the amplitude of the modulation of the collected fluorescent signals are all the stronger the higher the product of the amplitude reflectivities of the two mirrors. It is therefore advantageous to use an upper layer 12 of high index, for example of TiO2 having an index of between 2.2 and
• the amplitude reflectivity of the TiO2 / air diopter is approximately 0.4.
• the radiative emission towards the medium on which the chromophoric elements are found is all the more favored the higher the index of this medium.
• a photodetector 30 is located under each zone or strip 16 of height different from a range 14. It is therefore known directly which photodetector 30 is located opposite a particular zone or strip 16 corresponding to a maximum or a minimum resignation. Of course, several photodetectors 30 can be provided under each zone or strip 16 of different height.
• the upper layer 12 of transparent material has a flat upper surface and the mirror 22 is formed on a structured upper face 34 of the support 10.
• the abrupt discontinuities between the zones can form parasitic channels for the excitation wavelengths and possibly for the fluorescence emitted.
• a more continuous variation (with a triangular or wavy profile for example) of the thickness of the substrate overcomes this drawback.
• the zones are defined by the fact that the desired interference condition is substantially achieved there.
• the substrate 10 of the device comprises a plane mirror 22 and a planar upper layer 12 ′ , with an intermediate layer 35 of different index, which is structured with zones of different thickness, corresponding to the zones 16 above and giving rise to variations in intensity of fluorescence by phase shift.
• the device comprises one or more upper layers deposited on a transparent intermediate layer 37 to form a waveguide 36 for the excitation radiation, for example with propagation along the above-mentioned bands formed by the mirror 22, so as not to bring out the excitation radiation by the discontinuities between bands and diffuse it undesirably towards the photodetectors. It is also possible, to avoid this drawback, to give a sufficiently large thickness to the layer of material which covers the structured mirror 22, which makes it possible to distance the profile of the guided mode and its evanescent part of the structured surface of the mirror 22.
• Another variant of the invention consists in depositing in holes formed in the mirror 22 at least one layer of another material, reflecting, transparent or else absorbent.
• a variant of the invention consists in structuring in zones not the transparent layer carrying the chromophoric elements but the intermediate layer 37 by changing its thickness or even eliminating it in certain zones (FIG. 11B).
• the orifices thus created can then be deposited in a transparent layer 37 ′ of a material other than that of the layer 37 in which the orifices were created (FIG. 11C).
• the device comprises an opaque layer 38, 40 respectively, for example metallic, which limits to the useful surfaces (corresponding to the areas 14) the lighting by the excitation radiation or the transmission of fluorescence towards photodetectors.
• this opaque layer 38 covers the layer 12 of transparent material and has orifices corresponding to the pads 14.
• this opaque layer 38 is inside the layer 12, between the upper face of the latter and the reflective layer 22 and its orifices are aligned with the pads 14 or with the aforementioned zones 16.
• the opaque layer 40 is located inside the substrate 10 and has orifices facing the photodetectors 30 fixed under the substrate.

Landscapes

• Health & Medical Sciences (AREA)
• Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
• Physics & Mathematics (AREA)
• Life Sciences & Earth Sciences (AREA)
• Chemical & Material Sciences (AREA)
• Analytical Chemistry (AREA)
• Biochemistry (AREA)
• General Health & Medical Sciences (AREA)
• General Physics & Mathematics (AREA)
• Immunology (AREA)
• Pathology (AREA)
• Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)

Applications Claiming Priority (3)

Publications (1)

Family

ID=30775985

Family Applications (1)

Country Status (4)

Family Cites Families (5)

• 2002
  • 2002-08-13 FR FR0210285A patent/FR2843634B1/fr not_active Expired - Fee Related
• 2003
  • 2003-08-11 AU AU2003285360A patent/AU2003285360A1/en not_active Abandoned
  • 2003-08-11 WO PCT/FR2003/002510 patent/WO2004017055A2/fr not_active Application Discontinuation
  • 2003-08-11 EP EP03756540A patent/EP1529209A2/de not_active Withdrawn

Non-Patent Citations (1)

Also Published As

Similar Documents

Legal Events