WO2013060478A1

WO2013060478A1 - Miniaturised microscopy system for determining the state of cells

Info

Publication number: WO2013060478A1
Application number: PCT/EP2012/004507
Authority: WO
Inventors: Thomas Velten
Original assignee: Fraunhofer-Gesellschaft zur Förderung der angewandten Forschung e. V.
Priority date: 2011-10-28
Filing date: 2012-10-25
Publication date: 2013-05-02
Also published as: DE102011117228A1

Abstract

The present invention relates to a miniaturised microscopy system for determining the condition of biological cells, said system having an optical image capturing unit with an optical sensor (6) and an optional illumination unit (20). The optical image capturing unit is enclosed by a water- and moisture-tight casing (8) that is formed by an optical transparent layer (5) over a sensor surface of the optical sensor (6) or at least one region of the sensor surface. The surface of the optically transparent layer (5) that faces away from the sensor surface is arranged at a distance from the sensor surface such that a shadow projection of cells lying upon the surface onto the sensor surface allows for an optical resolution which is determined by a pixel size of pixels of the optical sensor (6). The transmission of captured image data of cells that adhere to the optically transparent layer to an external receiver unit allows the microscopy system for determining the condition of cells to be used directly in a bioreactor.

Description

MINIATURIZED MICROSCOPY SYSTEM FOR CONDITIONAL ASSESSMENT

OF CELLS

Technical application

The present invention relates to a

Microscopy system for determining the state of biological cells comprising an optical imaging unit with an optical sensor and an optional illumination unit. The invention also relates to a use of such a microscopy system in a cell-populated sealed environment, for example in a bioreactor.

Bioreactors can only be used efficiently for bacterial growth and / or metabolite synthesis if optimum conditions for the cells are present everywhere in the bioreactor. However, especially in large bioreactors, the same conditions do not prevail everywhere (eg temperature, nutrient concentrations, etc.). It would therefore be desirable to be able to detect the state of the cells at different locations within the bioreactor. So far, however, no technique is known with the state of cells in the bioreactor quasi-continuous and can be detected on-line. State of the art

Nowadays, the efficiency of the processes taking place in the bioreactor is assessed on the basis of the resulting products. If the desired products were produced in sufficient quantity, then it is assumed that the from outside set and kept constant parameters (such as temperature) an optimal state of the cells. The state of the cells themselves is not monitored. A today's established method around the state of the

To measure cells themselves is the observation of cells with the help of a microscope. One can e.g. from the

Divide rate to close the vitality of the cells. In addition, the cell morphology or the adhesion behavior of the cells can be used as a criterion. However, these investigations are only possible outside a bioreactor, since one is not a complete one

Microscope can bring in a bioreactor. Even more impossible it is several microscopes in one

Bioreactor so as to detect the state of cells in different locations of the bioreactor.

A concept for the realization of a very compact microscopy system was developed by Li et al. (Wei Li;

Knoll, T .; Thielecke, H .; On-chip integrated

lensless microscopy module for optical monitoring of adherent mammalian cells ", Engineering in

Medicine and Biology Society (EMBC), 2010 Annual

International Conference of the IEEE, Aug. 31, 2010- Sept. 4, 2010, pages 1012-1015). The

Microscopy system includes an optical sensor chip and a lighting unit. For the determination of

Condition of cells is a heavily miniaturized bioreactor or incubator placed on the sensor chip. So the microscopy system is located

outside the bioreactor / incubator. Through an optically transparent thin silicon nitride membrane that covers the bottom surface of the bioreactor /

Incubators can be added to the bioreactor / incubator look inside and observe cells adhering to the silicon nitride membrane. However, the described system is not suitable for use in a bioreactor.

The object of the present invention is to provide a microscopy system for determining the

State of biological cells that can also be used in a bioreactor.

Presentation of the invention

The object is achieved with the microscopy system according to claim 1. Advantageous embodiments of the microscopy system are the subject of the dependent claims or can be the following

Description and the exemplary embodiments.

The proposed microscopy system for determining the state of biological cells, which can also be regarded as a cell microscope, has a

optical image capture unit with an optical sensor or detector and an optional lighting unit on. The optical image acquisition unit is enclosed by a water-tight and moisture-tight envelope, which extends over the sensor surface of the optical sensor or at least one region used for the state determination of this sensor surface by an optical

transparent layer is formed. In this microscopy system, the state determination is carried out by evaluating an image recorded with the image acquisition unit, which is caused by the interaction of the illumination unit

emitted light is obtained with cells on the optically transparent layer lie over the sensor surface or adhere to this layer. In

Dependence on the light irradiation by the

Lighting unit can through the generated image

Reflection of the light can be obtained at the cells or in the form of a shadow projection of the cells. Secondary radiation can also be used for image recording, for example luminescence light emitted by the cells or substances associated therewith.

Due to the water- and moisture-tight enclosure or encapsulation, the microscopy system can remain for several days in the cell-populated environment, in particular in a bioreactor, and the recorded image data to an outside of this

Send the surrounding receiving unit. The images can then be displayed and / or appropriately evaluated for the condition of the cells

for example, to determine their size or shape. For this purpose, the microscopy system is preferably equipped with a wireless transmission unit, which is likewise located within the enclosure and can transmit the image data recorded by the image generation unit wirelessly to the external reception unit. A power supply for the individual components of the microscope system, in particular for the electronics of the image acquisition unit, the illumination unit and the transfer unit, which is arranged within the envelope, is also preferably also

arranged inside the enclosure. The power supply unit can be designed to be rechargeable and designed such that the recharging by inductive coupling of energy from outside the Serving is made possible. The charging can then be continuous during the operation of the

Microscopy system or after each use. In another embodiment of a

rechargeable power supply unit electrical contacts are led to the outside, which can be connected to a charging electronics. The power supply unit can then be charged after each use via these contacts. The sensor surface of the optical sensor is composed in a known manner from a two-dimensional grid or array of individual detector elements, also referred to as pixels.

The optically transparent layer over the

Sensor surface must be arranged at a small distance to the sensor surface and be made sufficiently thin to an evaluation of the state such as the cell morphology, the division behavior or the

To allow adhesion of the overlying cells from the generated image. For this purpose, the outwardly directed surface of the optically transparent layer, which faces away from the sensor surface, is at a distance from the sensor surface, in which the optical resolution of the cells resting on this surface is limited by the size of the pixels of the sensor surface and not by diffraction effects in the case of shadow projection , Thus, for example, the optical resolution R _{c of} a camera chip, as it can be used as an optical sensor in the present microscopy system, is given by the double pixel size S _P , ie R _c =

2S _P. The resolution for a shadow _projection R _s is given by R _s = d _Lufl ). Here denote d _s

the thickness of the optically transparent layer, n _s the Refractive index of this layer, d _air the thickness of the

Air gap between the optically transparent layer and the sensor surface and λ the wavelength of the incident and used for the shadow projection light. The thickness of the optically transparent layer and the air gap should be chosen so that the resolution of the selected camera chip through the pixels of the chip and not by the diffraction effects in the

Shadow projection can be determined or limited. The following applies: R _s ^ Rc- This results in the following d 2

Condition: + d _Lufl <- S /

n _s λ

Of course, the optically transparent layer can also be applied directly or via one or more intermediate layers on the sensor surface

lie, wherein a similar relationship results, in which only the term d _LU ft is omitted or replaced by terms d _ZS i / n _zsi for the individual intermediate _layers i with thicknesses d _ZS i and refractive indices n _zsi . Under the optical transparency is present in the

Patent application to understand that the layer is transparent in an optical wavelength range. This does not have the entire wavelength range of

optical wavelengths, but can be one

concern smaller area of it. Under the

Wavelength range of the optical wavelengths is to be understood as the range from the infrared via the visible wavelengths to the ultraviolet. The proposed microscopy system has

preferably no additional lenses between the sensor surface and the optically transparent layer, so that the image of the cells preferably as pure Shadow projection is displayed on the sensor surface. This makes the microscopy system very compact.

The optically transparent layer can rest directly on the sensor surface via one or more intermediate layers. The one or more intermediate layers may in turn form or include an optical filter that filters out unwanted wavelength ranges for imaging. Different application examples of such a filter can be found in the following embodiments. The one or more intermediate layers may also include a leveling layer if the sensor surface is not planar. This may be the case with a sensor chip with integrated microlenses, in which the surface is due to the external shape of the

Microlenses is structured. By a leveling layer of a suitable material that compensates for the bumps, these bumps do not disturb the planarity of the optically transparent layer and possibly an intermediate filter layer or filter layer sequence.

The water and moisture proof enclosure is preferably formed of one or more biocompatible materials or coated with a thin layer of biocompatible material. Through this biocompatible encapsulation or wrapping is the use within a bioreactor or another

Environment, eg. Within a human or animal body, easily possible to observe there the state of cells, for example. Cell growth. The wrapper can also be designed so that they open and close again several times leaves, for example, to allow a battery change. In one embodiment, the envelope for this purpose can be formed from two halves screwed together with an intermediate seal.

In the proposed method of detecting the state of cells in a cell-populated environment, several of the proposed miniaturized microscopy systems are located within the environment and generated by the optical sensors

Image data evaluated. Due to the small size and self-contained design of the microscopy systems, these can be used without problems in a bioreactor. For this purpose, an embodiment with integrated wireless data transmission unit and integrated power supply is preferably used, so that no cables run from the microscopy systems through the bioreactor, since these would represent a potential leak for both the microscopy systems and for the wall of the bioreactor. With the proposed microscopy system and the proposed method, cells in a bioreactor can be observed or microscoped over a long period of time virtually quasi-continuously. The method thus allows on-line monitoring of the condition of cells in a bioreactor or other open or closed environment.

Brief description of the drawings

The proposed method and the associated miniaturized and self-contained microscopy system will be briefly explained again below with reference to an exemplary embodiment in conjunction with the drawings. Hereby show: Fig. 1 shows an example of an encapsulated microscope system according to an embodiment of the present invention, in which the imaging of the cells on the optical sensor means

Shadow projection takes place;

FIG. 2 shows an example of an encapsulated microscope system according to a further embodiment of the present invention, in which the light of the illumination unit strikes the optically transparent layer; FIG.

3 shows an example of an encapsulated microscopy system according to another embodiment of the present invention, in which the optically transparent layer simultaneously functions as an optical waveguide;

4 shows an example of an encapsulated microscope system according to another embodiment of the present invention in which between the optically transparent layer and optical

Sensor uses an optical filter; and

5 shows an example of an encapsulated microscope system according to a further embodiment of the present invention, in which an indicator layer which can be used for spatially resolved detection of the concentration of one or more substances is immobilized on the optically transparent layer.

Ways to carry out the invention

In the exemplary embodiments described below, an extremely highly miniaturized microscope consisting of a lighting unit 20, an optical detector or sensor 6, a

wireless data transmission unit 7 and a

Power supply unit 10 constructed and in a suitable water and moisture sealed as well

sterilizable housing 8 introduced. Such a system can remain in a bioreactor for several days and transmit the recorded image data wirelessly to a reception unit 9 located outside the bioreactor. The external receiving unit 9 can either display the images themselves on a screen and / or the image data can be sent to a

Computer be forwarded. The microscopy system according to the invention in this example thus consists of an extremely highly miniaturized microscope, a wireless data transmission unit, a power supply unit, a suitable water and moisture proof and sterilizable housing and a receiving unit located outside the bioreactor which receives the wirelessly transmitted image data. The compactness of the system is preferably achieved by constructing the microscope lens-free. The image of the cells is therefore not imaged by means of lenses on the optical detector. Instead, it is located directly on the detector, e.g. a CCD chip, a very thin optically transparent layer on which the cells adhere.

In a first embodiment that consists

System according to the invention from the basic components shown in FIG. 1: a lighting unit 20,

consisting of light source 1 and a collimator 2, an optically transparent thin layer as

Slide 5, an optical detector 6, a

Data transmission unit 7, a power supply unit 10, a housing 8, at least the

Components 5, 6, 7 and 10 encloses one

Fixing device 3, which connects the lighting unit 20 to the housing 8 and an external

Receiver unit 9. The electrical parts of the

Illumination device 20 can also be used together with the fastening device from the housing 8

enclosed or even be housed in a separate water-and moisture-tight enclosure, in which case the electrical connection with the power supply unit 10 must also be suitably sealed.

After introducing this system into a bioreactor, cells in the bioreactor settle on the system, but especially on the slide 5. The image of the cells on the optical detector 6 is preferably formed by shadow projection. For this purpose, the light of a nearly punctiform light source 1 (for example LED) is parallelized by means of a suitable device, a so-called collimator 2, which may be, for example, a thin tube with a blackened inner wall a few centimeters long and placed on the cells located on the slide 5. The light is preferably incident

perpendicular to the surface of the slide 5, i.

preferably parallel to the normal on the slide 5. The shadows of the cells are from the located directly below the slide 5 optical

Detector 6 detected. In order to minimize the influence of diffraction effects on the image quality, the distance between the cells and the optical detector 6 is kept very low. This is preferably done by

ensures that the slide 5 is very thin. The slide 5 can be designed, for example, as approximately 1 μm thin silicon nitride layer. This can be realized, for example, by using silicon dioxide (PECVD) (plasma enhanced chemical vapor deposition) of the desired thickness (for example, about 1 μm) on a substrate, for example on a silicon wafer.

is deposited. In the area in which the optical detector 6 is to be located later, the

Substrate selectively removed by means of an etching process (in the case of a silicon substrate, for example by wet chemical etching with potassium hydroxide solution). The result is a silicon nitride membrane of the desired size, which is surrounded by a silicon frame or a frame of another material. This frame is designated in FIG. 1 as substrate 4. The lateral

Dimensions of the substrate 4 (e.g., a silicon frame) may be measured by a suitable device, e.g. be brought to the desired size by means of a wafer saw. The substrate 4 facilitates that

Handling the extremely thin slide 5 and can be used to connect the slide 5 with the housing 8 of the overall system. Another one

preferred substrate material is metal. Preferably, the metal substrate is made of the same material as the housing 8 of the miniaturized microscopy system.

In a further embodiment according to the invention and outlined in FIG. 2, the incidence of light is not perpendicular to the slide 5. Instead, the light source 1 of the illumination unit is arranged such that the light beam 21 strikes the slide 5 in a grazing manner. The light beam 21 can furthermore be formed by a suitably shaped diaphragm, which, for example, can be split. is executed shaped (not shown), limited. In grazing incidence, the reflectivity of the slide 5 has almost the value 1, ie the light coming directly from the light source can not practically pass through the slide 5 and therefore can not reach the optical detector 6. Only the light which is reflected or scattered on the cells located on the slide 5 is able to reach the optical detector 6. The from the

Optical image 6 recorded image thus contains information about the adhering to the slide 5 cells. By a spatial delimitation of the light beam, only the cells sitting directly on the slide 5 are irradiated, so that only light from this can scatter or reflect in the direction of the optical detector 6.

This method of grazing illumination has the advantage that the overall system can be made even more compact than the system shown in FIG.

In a further embodiment according to the invention and shown in Fig. 3, the light of

Light source 1 of the illumination unit coupled in a suitable manner in the very thin slide 5. The slide 5 fulfills the purpose of an optical waveguide and conducts the light. The light coupling, which is carried out with the aid of a coupling-22, can be done by known methods. This includes, for example, the

Coupling by means of prisms or by means of suitable grating structures, so-called grating couplers. It is known that the electromagnetic field of the light guided in a waveguide is not completely in the waveguide. Part of the electromagnetic The field also projects out of the waveguide, eg into the bioreactor. This is known as that

This evanescent wave field decays exponentially with distance and typically protrudes only fractions of a micron out of the waveguide, and in the bioreactor it hits either parts of a cell adhering to the slide 5 or liquid

Light is scattered or reflected at the cells and parts of the scattered or reflected light

reach the optical detector 6 and there give an image that allows a statement about the adhesion behavior of the cells on the slide 5. A

Observation of the adherence behavior over a longer period of time continues to provide information about the movement of the cells on the slide 5. Based on the attachment pattern, a cell division is still recognizable. So you can also gain a statement about the proliferative behavior of the cells.

In a further preferred embodiment of the microscopy system according to the invention according to FIG. 4, an optical filter 23 is located between the cells to be observed and the optical detector 6 in order to filter out or transmit light of specific wavelengths. For example, such filters can consist of a plurality of superimposed thin dielectric layers (interference filter) and be produced by means of suitable thin-film technology.

Preferably, the optical filter 23 is located directly on one side of the thin slide 5. The filter layers are preferably deposited directly on the slide 5. In another preferred arrangement, the filter layers are deposited directly on the optical detector 6.

For example, commercially available CCD chips can be used as optical detectors 6. These are usually with a variety

microscopic lenses (microlenses). This results in a very uneven surface of the CCD chip. If the optical filter layers are deposited directly on the microlenses, a light beam incident perpendicularly to the CCD chip impinges on the filter layers at different locations due to the curved surfaces of the microlenses. This is effective in the

Filter layers traversed optical path length

also different from place to place. This results in a filter effect different from place to place. In a preferred embodiment of the microscopy system according to the invention, therefore, a thin optically transparent is first applied to the microlenses of the CCD chip

Layer 24 applied, which has a leveling effect. This results in a relatively flat surface. The thickness of this leveling layer 24 should preferably be in the range of a few micrometers. In a

preferred embodiment of the invention

Microscopy system, the filter layers 23 are applied to the leveling layer 24. Preferably, the leveling layer 24 is made of a relatively soft polymer material which conforms to the object carrier 5 when pressed against it. In this way, small cavities are prevented between the leveling layer 24 and the slide 5, which fill with a medium having a different optical refractive index than the planar end Alternatively, these voids may be filled with a material, eg a suitable liquid, having the same refractive index as the planarizing material or as used

Slide 5 (e.g., silicon nitride layer). In the

Optically, such liquids are known as "immersion oil." If the leveling layer 24 is omitted, the small cavities forming between the microlenses and the thin slide 5 (eg, silicon nitride layer) should be filled with a material, such as a suitable liquid, which contains the same refractive index as the material has the

Microlenses or the slide 5 used. In another possible embodiment, the optical filter 23 consists of a self-contained thin film (polymer or glass) and is placed between slide 5 and CCD chip or between slide 5 and leveling layer 24, e.g. inserted, clamped or glued. Also in this case, the between the filter 23 and the leveling layer 24 and between the filter 23 and the

Microlens forming cavities with a material, e.g. a suitable liquid, which has the same refractive index as the planarizing material or like the microlenses.

The use of the described optical filter 23 can advantageously be used if, in addition to or as an alternative to the image of the cells, further optical effects, such as luminescence, are to be detected. For example, at least some of the cells in the bioreactor can be provided with so-called fluorescent markers, which are present either from the outset are, or only when a certain

Situation in the cell are formed. The

Fluorescence radars do not emit light but are usually excited by photons (photoluminescence). In this case, the above-described optical filters 23 can be used to filter out the excitation light which may originate from a light source 1 of the microscopy system according to the invention or also from a light source independent thereof, and only the resulting one

Luminescence to the optical detector 6 to pass. If a light source 1 of the microscopy system

is to be used, then all the above configurations (light falls perpendicular to the slide 5, light falls grazing on the slide 5, slide acts as an optical waveguide, in which the excitation light is coupled) usable.

Furthermore, the microscopy system according to the invention can be used to detect chemiluminescence or bioluminescence occurring in the cells. In these latter cases, no excitation light is necessary for exciting the luminescence. One

Microscopy system according to the invention for detecting chemo- or bioluminescence can thus be constructed without its own light source 1 or illumination unit. Of course, when using a lighting unit in addition to the chemo- or bioluminescence also an image of the cells can be detected.

In a further inventive embodiment according to FIG. 5, the slide 5 is provided with a thin layer of an indicator molecule whose

Luminescence behavior of the presence of certain Substances or depends on the pH or temperature of the immediate environment. Preferably, the indicator molecules are embedded in a polymer matrix in order to ensure a fixed retention of the indicator molecules on the slide surface, even over a longer period

Period to ensure. For application as a thin layer, for example, the polymer material is dissolved together with the indicator molecules in a solvent and the resulting liquid by methods such spin-on, spray, print or any other method known in the art on the

Surface of the slide 5 is applied. After evaporation of the solvent, a solid forms

Polymer film in which the indicator molecules are embedded. Such a method is described, for example, in M. Staal, EI Prest, JS Vrouwenvelder, LF Rickelt, M. Kuhl: "A simple optode-based method for imaging 0 ₂ distribution and dynamics in tap water biofilms", waterresearch 45 (2011) pp. 5027 -5037 for the case of oxygen-sensitive indicator molecules

Ruthenium (II) tris-4, 7-diphenyl-1, 10-phenanthroline (Ru-dpp) was in a

Polystyrene matrix immobilized. The solvent used was chloroform.

A thus prepared indicator film 25 has

preferably a layer thickness of less than 10 μπι, preferably less than 5 μπι to the spatially resolved

Detection of the substance for which the used

Indicator molecule is sensitive, with a high

to enable spatial resolution. It should be pointed out once again that the application is not limited to the detection of oxygen. If a corresponding indicator molecule is used, indicator layers 25 can also be produced. which are suitable for the detection of other substances, such as glucose, lactate or other substances. Furthermore, the indicator layer does not have to be applied as a necessarily homogeneous layer, but can also be applied to the slide 5 as a structured layer. For this purpose, for example, photolithographic methods can be used, as they are known from microsystems technology. When using a

Printing technique for applying the indicator layer, the indicator layer can be realized as a structured layer already during printing. Printing techniques falling within this invention include e.g. Gravure, high pressure, offset printing, pad printing and the

so-called "micro contact printing", which was developed especially for the realization of microstructures, but the invention is not limited to the mentioned printing techniques

In the simplest case, the structure consists of a single coated strip and an adjacent uncoated strip. Further preferred is the

Structure of individual circles or rectangles.

Preferably, the circles or rectangles are about the size of a single light-sensitive pixel of the optical detector 6 (e.g., CCD chip) or a

integer multiple of this size. Not

Coated bodies can be used, for example, as

Serve reference, i. the intensity of the luminescence coming from the coated sites and the luminescence coming from the uncoated sites can be detected separately. The actual measurement signal can then be separated from each other

measured individual values are calculated. In the Calculation may be advantageous, the ratio between the in the coated area and the

Reference range measured luminescence intensities.

For simultaneous measurement of several substances or

Sizes (e.g., temperature, pH), the indicator layer 25 may consist of regions of various indicator substances. Again, in addition to the for the

various substances sensitive areas, further areas are arranged, which contain no indicator molecules and thus as a reference can.

In the previous statements, it was assumed that the concentration of the substance to be detected should be detected in a spatially resolved manner. By means of an alternative detection method, a value averaged over the entire slide 5 can also be determined

Concentration of substance can be detected. It is averaged over all areas coated with the respective indicator molecule and also averaged over all reference ranges. In this case it is advantageous to nest the reference areas and the measuring areas. When using an indicator molecule for substance A, an indicator molecule for substance B and a reference region (R), when using a stripe pattern, the strips should be printed in the continuous order ABRABRABR, etc. on the

Slide 5 may be arranged. In this way the

Arrangement, influences of inhomogeneities of the substance concentration or the temperature distribution can be reduced to the mean values of the substance concentrations.

For optically exciting a luminescence, all the configurations of the light source described in FIGS. 1, 2 and 3 can be used. In addition, the optical filters 23 shown in Figs. 4 and 5 can be used for Blocking the excitation light can be used advantageously.

For the wireless transmission of the image data recorded by the optical detector 6 as an image acquisition unit are in principle all known today

Data transmission methods usable. This includes e.g. the use of radio or optical methods. Corresponding standardized transmission modules and protocols are present in large numbers. When transmitting by radio, for example, ZigBee, Bluetooth, WLA or MICS standard can be used. In an optical transmission, for example, the IrDA (Infrared Data Association) protocol can be used. The commercially available data transmission modules are usually equipped with a microcontroller. This can be used to control the flow of an image acquisition and transmission sequence

be used.

To protect the microscope system from liquids and moisture, this and the data transmission unit 7 must be encapsulated in a suitable manner. Suitable methods, packaging techniques and materials are, for example, in the field of active

medical implants (e.g., pacemakers). The housing 8 is preferably made of metal, preferably a metal that meets the requirement

Biocompatibility (eg according to ISO 10993 1-20). Preferably, all materials contacting the cells in the bioreactor should have biocompatibility according to ISO 10993 1-20. A suitable metal is eg titanium. Since the slide 5 come directly into contact with cells and thus a part of Housing must form, the slide 5 and the substrate 4, on which the thin slide 5 is deposited, to connect in a suitable manner with the rest of the housing 8. This can be methods like

Welding (e.g., laser welding), soldering or gluing. To ensure biocompatibility, for example, the entire system can be coated with a thin layer of an optically transparent and biocompatible material. A suitable material is, for example, Parylene C. A suitable thickness of the protective layer may be, for example, 1-5 μπι.

When using an optical data transmission method, the housing 8 is preferably provided with an optical window. For this purpose, for example, glass can be used, which is connected by means of one of the methods described above with the rest of the housing. In the case of data transmission by electromagnetic waves is usually a

Antenna used. This should preferably be arranged outside of the housing, since a strong attenuation of the electromagnetic waves would result, especially in the case of a metal housing. The

electrical connection between electronics (inside the housing) and antenna (outside the housing), for example by means of so-called glass or

Ceramic bushings can be realized. These are usually designed so that they are directly in one

Metal housing can be welded. Alternatively, the above-mentioned methods soldering and gluing can also be used here. The antenna itself can be coated, for example, by means of a thin Parylene C layer. Alternatively or in addition The antenna can be embedded in biocompatible silicone.

The power supply of the invention

Systems can be done by means of a battery.

For example, a battery can be used, as it is also used in active medical implants, for example. Pacemaker. The invention is not disclosed on the

Embodiments restricted. Others too

Variations can be deduced therefrom by the skilled person without the scope of the invention

leave. For example, the lighting unit need not necessarily be connected to the imaging unit or its housing or enclosure, but may also form a separate unit with its own power supply. The embodiments of the individual embodiments can also be combined with each other.

LIST OF REFERENCE NUMBERS

1 light source

2 collimator

3 fastening device

4 substrate

5 object bearer

6 optical sensor

7 data transmission unit

8 housing

9 external receiving unit

10 power supply unit

20 lighting unit

21 grazing light beam

22 coupling device

23 optical filter

24 leveling layer

25 indicator film

Claims

claims

Microscopic system for determining the state of cells comprising an optical image acquisition unit with an optical sensor (6) and an optional

Lighting unit (20),

characterized,

that the optical image acquisition unit of a water and moisture-tight enclosure (8)

is enclosed, which is formed over a sensor surface of the optical sensor (6) or at least a portion of this sensor surface by an optically transparent layer (5), wherein one of the

Sensor surface facing away from the surface of the optically transparent layer (5) a distance from

Sensor surface, in which a shadow projection of resting on the surface cells on the sensor surface allows an optical resolution, which is determined by the pixel size of pixels of the optical sensor (6).

Microscopy system according to claim 1,

characterized,

the optically transparent layer (5) rests on the sensor surface directly or via one or more optically transparent intermediate layers.

Microscopy system according to claim 2,

characterized,

that the one or more intermediate layers form one or more optical filters (23) or include.

4. Microscopy system according to claim 2 or 3,

characterized,

in that one of the one or more intermediate layers forms a planarization layer (24) for a surface structure of the sensor surface.

Microscopy system according to one of Claims 1 to 4, characterized

a wireless data transmission unit (7) is arranged within the enclosure (8), with the image data generated by the image detection unit being able to be transmitted to an external reception unit (9).

Microscopy system according to one of claims 1 to 5, characterized

in that a power supply unit (10) for the optical image acquisition unit and the

Lighting unit (20) and possibly the wireless data transmission unit (7) within the enclosure (8) is arranged.

Microscopy system according to one of Claims 1 to 6, characterized

the illumination unit (20) is arranged in such a way that a shadow projection of cells resting on the optically transparent layer (5) on the sensor surface takes place.

Microscopy system according to one of claims 1 to 6, characterized

the lighting unit (20) is arranged in this way is that a grazing incidence of light on the optically transparent layer (5).

Microscopy system according to one of claims 1 to 6, characterized.

that the optically transparent layer (5) as

Optical waveguide is formed and the

Lighting unit (20) arranged and

it is formed that they light in the

Coupled optical fiber.

Microscopy system according to one of claims 1 to 9, characterized

that a layer with indicator molecules (25) at least partially on the optical

transparent layer (5) is applied.

Microscopy system according to one of claims 1 to 10, characterized

that the envelope (8) made of biocompatible

Materials formed or coated with a layer of a biocompatible material.

Method for detecting the state of cells in a cell-populated environment, in particular in a bioreactor, in which one or more of the microscopy systems according to one or more of the preceding claims falls within the scope of the invention

Environment arranged and evaluated by the microscopy systems image data to be evaluated.