Device for applying an electromagnetic wave to a sample of biological material

Abstract

A device is disclosed for applying an electromagnetic wave (EMW) to a biological sample (12) comprising biological cells in an in vitro medium, with an improved coupling effect. The coupling of the EMW to the biological cells is enhanced by strengthening the electric field on the sample through the use of electromagnetic resonance modes in a resonator block (20) made of dielectric material. The sample (12) is placed in a housing (21) positioned and sized to correspond to an area where there is a resonance peak in the volume of the resonator (20) at a determined frequency of interest. This results in overcoupling of the incident EMW at the frequency of interest, which improves the level, homogeneity, and dosimetry of the electric field in the sample. Applications include, but are not limited to, the experimental study of the effects of electromagnetic waves on biological cells. Figure to be published with the abstract: Figure 9

Description

Title of the invention: Device for applying an electromagnetic wave to a sample of biological material Technical field of the invention

[0001] The present invention relates generally to bio-electromagnetism implemented in, and more particularly to a device for the application of an electromagnetic wave (EMW) on a sample of biological material comprising biological cells in an in vitro environment.

[0002] The invention finds applications, in particular, for carrying out experimental studies concerning the effects of electromagnetic waves on biological cells in the medical field, public health, industrial processes (sterilization, food processing, etc.). These may be studies with a normative purpose relating to the determination of thresholds of harmfulness and harmlessness (telephony, transmissions, electromagnetic defense applications, electrical equipment, etc.), with a medical purpose (cancer treatment, neuronal stimulation, etc.) or even with an industrial purpose (decontamination, sterilization, food processing, etc.). Technological background

[0003] The field of bioelectromagnetism aims to study the effects of electromagnetic fields on living organisms (see, for example, the book by J. Malmivuo and R. Plonsey: "Bioelectromagnetism - Principles and applications of bioelectric and biomagnetic fields," Oxford University Press, January 1995). In particular, it concerns the investigation of effects, whether deleterious or not, reversible or irreversible, associated with a waveform, a repetition frequency, an amplitude, and a spectral range of an electromagnetic field. For example, the device of the invention can be used to apply strong electromagnetic fields to biological tissues in order to evaluate the effect of a treatment on pathogenic tissues by the application of electromagnetic radiation.

[0004] In vitro exposure systems allow the illumination of biological cells placed in a specific in vitro medium by RF waves. Generally, this biological material is contained in Petri dishes, flasks, microtubes, or wells. The cells under study may be suspended in a solution or adherent to the bottom of the dishes.

[0005] Experiments conducted in the field of bioelectromagnetism require experimental systems capable of exposing various biological materials, of varying sizes and natures, to the electromagnetic fields likely to be encountered. in various real-world situations. The amplitudes of the electric fields that are used can be high (several tens of kV / cm). And the spectral content of the electromagnetic waves concerned extends from the continuous to very high frequencies (beyond the Gigahertz) in the radio frequency (or RF, from the English "Radio Frequency") spectrum, that is to say in the upper part of the ultra-high frequency (or UHF, from the English "Ultra High Frequency") band which is the band of the radio spectrum between 300 MHz and 3 GHz (i.e. the wavelengths from 1 m to 0.1 m) or even in the lower part of the super-high frequency (or SHF, from the English "Super High Frequency") band, namely the radio frequency band which extends from 3 GHz to 30 GHz (wavelength from 10 cm to 1 cm) and which belongs to the domain of microwaves extending from 1 GHz to 1000 GHz.

[0006] To date, a wide variety of techniques exist for the application of electromagnetic waves such as, for example, and in a non-exhaustive manner: - "plane wave" type exposure systems; - reverberation chambers, for example Mode Mixing Reverberation Chambers (MMRC); - guided wave type exposure systems such as rectangular or cylindrical waveguides; - wire-plate cells; - radial transmission lines; - flat conductor transmission lines such as Crawford cells and TEM cells (TEM stands for "Transverse Electro-Magnetic"); and, - coaxial load resistance devices.

[0007] On the other hand, in the field of biology there is a wide variety of commercial containers on the market to allow experiments carried out on biological cells, such as the following biological containers: - flasks; - culture plates; - multi-well plates forming biological basins; - microtubes, for example Eppendorf™ type microtubes (brand of the eponymous company); - Petri dishes; and, - strips.

[0008] These containers are suitable for the handling, culture, observation, and transport of biological media, as well as for equipment used in biological laboratories. However, they are poorly suited to the requirements of the world of electromagnetism.

[0009] In particular, quantifying the effects of the aforementioned type on living organisms requires specific control of the electromagnetic wave (EMW) applied to the cells. To ensure the validation and reproducibility of experiments, particular care must be taken to ensure that all cells exposed to the electromagnetic field are exposed uniformly, by knowing as precisely as possible the exact level of the applied field as a function of the emitted EMW. This control is achieved by the user of an EMW application device, also called an electromagnetic wave applicator, which allows the transmission of an EMW from the source to the cellular medium under test, with particular control over the spatial distribution of the electromagnetic field.

[0010] Document FR2998813A1 discloses a coaxial load resistance applicator characterized in that the electrical resistance defines an internal volume in which the biological material sample is placed.

[0011] Document FR3034521A1 discloses a device for exposing an object under test (or OAT, denoted as "Object under Test") to an electromagnetic field. The device consists of a section of a guided electromagnetic wave device of the waveguide or transmission line type, extending along a longitudinal axis and having a hollow tubular electrical conductor inside which the electric field is established, or several electrical conductors between which the electric field is established. The device also includes a heterogeneous matching structure in mechanical contact with the electrical conductor(s). This matching structure comprises at least one extension element and at least one housing for the OAT, contained within or adjacent to the extension element. The extension element is made of a material having a relative dielectric permittivity (εr) as close as possible to that of the object or each object under consideration.The heterogeneous matching structure has a dimension (L), along the longitudinal axis of the waveguide or transmission line, which is equal to a non-zero multiple of half-wavelengths of the electromagnetic background that must propagate in the exposure device.

[0012] Document FR3021743A1 discloses a method for preparing a test object (TO) intended to be illuminated by an incident electromagnetic wave, the object having a given relative permittivity (er). This method improves the homogeneity and increases the intensity of an electric field induced in the TO subjected to an incident electric field. To this end, the method comprises: - providing a part comprising a cavity for housing the object and at least one extension element made of a material having a relative permittivity equal to the relative permittivity of the object to within ± 50%, said extension element partially delimiting the cavity and extending on either side of the cavity in a direction referred to as the cavity penetration direction, over a length that is at least equal, on either side of the cavity, at one-third of the cavity's length along the direction of passage; and - the placement of the object in the cavity, so that the object is in contact with said at least one extension element along the direction of passage. Summary of the invention

[0013] The present 'invention aims to improve the coupling factor of the electromagnetic wave (EMW) between a source and a sample of biological material as an object under test (OUT), to make the distribution of the electromagnetic field in said sample more uniform, and / or to improve dosimetry, i.e. knowledge of the function that gives the value of the field E in the sample as a function of the power P of the EMW sent, according to the formula E = P.

[0014] This objective is achieved by means of an applicator device for applying an incident electromagnetic wave at a predetermined frequency of interest when said wave is generated by an exposure system comprising an electromagnetic wave source disposed outside the applicator, to a sample of biological material disposed inside the applicator, said applicator comprising a resonator block which • is made of at least one specific solid dielectric material, • has the general shape of a closed volume of determined dimensions, and • is a solid volume except in that it includes at least one hollow compartment, called a sample compartment, associated with a removable cap of the resonator block, said hollow compartment being adapted to receive, and preferably to be filled with, the sample of biological material, in which, furthermore, • The resonator block, by design, exhibits a particular spatial distribution of the electric field when subjected to illumination by the incident electromagnetic wave. This distribution has discrete geometric configurations that define electromagnetic resonance modes, each associated with a natural resonance frequency. These modes can be determined by simulation and adjusted based on constitutive parameters of the applicator, including the relative dielectric permittivity (er) of the dielectric material, in combination with electromagnetic parameters of the biological material sample intended to be housed inside the sample holder, including the relative dielectric permittivity of said sample. • the constituent parameters of the resonator block are defined such that a resonance frequency in said resonator block corresponds to the frequency of interest; and, • the shape of the sample housing and its position in the volume of the resonator block are defined so that said housing occupies an area in said volume of the resonator block which corresponds to a resonance peak of the electromagnetic field at the frequency of interest.

[0015] The quality of the device for exposure to an electromagnetic field, evaluated in terms of the coupling rate of the electric field in the object to be tested and in terms of the homogeneity of the electric field inside said object, is significantly improved.

[0016] The "overcoupling" of the OEM in the OST, achieved through the exploitation of the resonance phenomenon, also allows for the testing of larger samples (in terms of the quantity of illuminated biological solution). The use of the resonance phenomenon, which provides an amplification of the electromagnetic field in certain areas within the applicator, makes it possible to obtain high field values ​​in these areas, where the OST containing the cell sample to be exposed is positioned. This maximizes the coupling ratio of the electric field induced within the volume of the OST, and even achieves overcoupling (K > 100%), which increases the intensity of the electric field induced within the OST. The proposed embodiments also increase the homogeneity of the electric field induced within the volume of the OST and improve dosimetry.

[0017] In some embodiments, the constitutive parameters of the applicator include not only the permittivity (er) of the dielectric material(s) from which the resonator block is made, but also the dimensions and geometry of the resonator block, as well as the boundary conditions of said resonator block. Similarly, the electromagnetic parameters of the biological material sample housed inside the cavity include the complex permittivity, that is, not only the permittivity (er) but also the conductivity (θ), of said sample.

[0018] In embodiments, the resonant block has the form of a bar extending along a determined longitudinal direction (which is the direction of propagation of electromagnetic background), and having a section in a plane orthogonal to said longitudinal direction which is of determined shape, constant along the longitudinal direction of the bar, and of determined dimensions which are less than the dimension of the bar along said longitudinal direction.

[0019] For example, the resonating block may have the shape of a cylindrical bar. A bar is a structure whose geometric shape has a longitudinal dimension (or length) substantially greater than its dimensions transversals (width and height). The term "cylindrical bar" refers to a bar that has the shape of a cylinder, a cylinder being a figure generated by a straight line rotating around another straight line to which it is parallel. The cylindrical bar therefore has a circular cross-section.

[0020] Alternatively, the resonator block may have the shape of a bar with a polygonal cross-section, for example with a hexagonal cross-section, or else square, octagonal, pentagonal or triangular.

[0021] In some embodiments, the electromagnetic field applicator device may further comprise a metallic waveguide section in which the resonator block is adapted to be placed, and which is adapted to guide the incident electromagnetic field generated by the exposure system. A closed system with well-controlled boundary conditions can then be obtained.

[0022] In this case, the waveguide section may include a resonant cavity in which the resonator block is placed, with the biological material sample housed in the closed housing cavity of said resonator block.

[0023] Alternatively or in addition, the waveguide section may comprise a plurality of parallel arms in which a plurality of resonator blocks may be placed so that respective biological material samples placed in their respective sample housing cavities are simultaneously illuminated by the incident electromagnetic field. Advantageously, this illumination of the different samples occurs independently and symmetrically.

[0024] When the electromagnetic field applicator device includes a metallic waveguide section as described in the three paragraphs above, and when it includes a bar-shaped resonator block as defined above, one and / or the other of the longitudinal end faces of said resonator block are in contact with respective walls of the metallic waveguide.

[0025] In embodiments, at least the internal walls of the sample housing (21) which are intended to be in contact with the sample of biological material, are made of a biocompatible material or are coated with a biocompatible material.

[0026] In embodiments, the sample housing is adapted to receive the biological material sample in the form of a biological solution containing biological cells to be illuminated by the incident electromagnetic field.

[0027] In embodiments, the dielectric material or materials of which the resonator block is made may have a relative permittivity (er) greater than one, and equal to the relative permittivity (er) of the biological sample to within ±50%.

[0028] For example, the dielectric material or materials of which the resonator block is made may have a relative permittivity (er) greater than one, and equal to the relative permittivity (er) of the biological sample to within ±40%, preferably to within ±30%, preferably still to within ±25%, preferably to within ±20%, and even more preferably to within ±10%.

[0029] In some embodiments, the dielectric material(s) from which the resonator block may be made include ceramic. Ceramic has the advantage of being a biocompatible material and having high permittivity.

[0030] In embodiments, the resonator block may comprise a plurality of closed compartments for housing respective biological samples, the shape of said sample compartments and their respective positions in the resonator block being chosen so that said sample compartments each occupy an area in the volume of said resonator block which corresponds to a resonance peak of the electromagnetic field.

[0031] A second aspect of the invention relates to a system comprising at least one electromagnetic wave applicator device according to the first aspect above, defined for the application, to a sample of biological material disposed inside said device, of an electromagnetic wave incident at a determined frequency of interest, the system further comprising an exposure device which is configured to generate the electromagnetic wave incident at the frequency of interest for which the electromagnetic wave applicator device is defined and which comprises at least one guided electromagnetic wave device adapted to guide the electromagnetic wave incident generated by the exposure system and in which the resonator block of the electromagnetic wave applicator device is placed.

[0032] For example, the guided electromagnetic wave device of the system can be a waveguide section in which the resonator block is placed.

[0033] In embodiments of the system, the waveguide section may include a drawer adapted for the arrangement of the resonator block and its placement in said waveguide section.

[0034] In embodiments of the system, the guided electromagnetic wave device comprises a plurality of parallel waveguide sections in which a plurality of electromagnetic field applicator devices are respectively placed so that respective samples of biological material placed in the sample housing of the respective resonator blocks of said electromagnetic field applicator devices are simultaneously illuminated by the incident electromagnetic wave as it propagates in the guided electromagnetic wave device.

[0035] When the electromagnetic field applicator device(s) include a bar-shaped resonator block as defined above, one and / or the other of the longitudinal end faces of said resonator block may be in contact with respective walls of the guided wave device of the system.

[0036] A third aspect of the invention relates to yet another method for designing an electromagnetic wave applicator device comprising the steps for applying an incident electromagnetic wave at a determined frequency of interest, when said wave is generated by an exposure system comprising an electromagnetic wave source disposed outside the applicator, to a sample of biological material disposed inside said applicator, comprising the following steps: - choose a geometry and dimensions for a resonator block made of a specific dielectric material; - determine by numerical simulation the natural modes of electromagnetic resonance in the resonator block defined by the geometry and dimensions chosen, giving different electric field distributions in said resonator block when it is exposed to the incident electromagnetic wave; - retain an electric field distribution corresponding to a specific resonance mode; - deduce the position and dimensions of a hollow housing in the resonator block, accessible from outside said resonator block, which correspond to an area of ​​the volume of the resonator block in which the simulation indicates a peak of electromagnetic resonance, said hollow housing being intended to receive the sample of biological material; - to tune the resonance frequency associated with the selected resonance mode to the frequency of interest, by modifying constituent parameters of the resonator block, including the value of the relative dielectric permittivity of the material from which said block is made, in combination with electromagnetic parameters of the biological material sample intended to be housed inside the sample housing, including the relative dielectric permittivity of said sample.

[0037] Finally, a fourth and final aspect of the invention relates to a method of using an electromagnetic wave applicator device according to the first aspect, comprising the following steps: - Insert a sample of the biological material to be tested into the sample compartment of the resonator block, then close the compartment using a stopper, - place the resonator block in a device exposing it to an electromagnetic wave incident at a specific frequency of interest, - to control an electromagnetic wave source from the exposure device in order to illuminate the sample of biological material present in the resonator, at a given power, for a determined duration - remove the resonator block cap and retrieve the biological cell sample after illumination, in order to submit it to analyses to evaluate the effect on biological cells of exposure to the electromagnetic wave at the frequency of interest. Presentation of the drawings

[0038] Other features and advantages of the invention will become apparent from the following description. This description is purely illustrative and should be read in conjunction with the accompanying drawings, in which: [Fig.l] is a schematic representation of the illumination of an object under test in radiated mode, in an anechoic chamber; [Fig.2] is a schematic representation of the illumination of an object under test in guided mode, in a waveguide; [Fig.3A] and [Fig.3B] are representations of a known biological container, namely an Eppendorf™ type microtube before and after, respectively, filling with a solution comprising the biological cells to be tested; [Fig.4] is a diagram illustrating the distribution of the electric field in a sample of cells in solution in a Petri dish within a TEM (Transverse Electric Magnetic) cell according to prior art wave application systems; [Fig.5] is a diagram illustrating the distribution of the electric field in a sample of cells in solution in a Petri dish within a waveguide in accordance with other OEM wave application systems according to the prior art. [Fig.6] is a cross-sectional representation of electric field configurations in a rectangular section waveguide having conductive walls as a function of the frequency of the EM in the waveguide; [Fig.7A] to [Fig.7D] are representations of the electric field distribution for various resonance modes in a resonator block made of dielectric material; [Fig.8A] to [Fig.8C] are representations of the location, and of the geometry and dimensions, of the areas in a resonator block where there is a maximum of the electric field, and where consequently placing the object under test makes it possible to obtain a coupling factor greater than one with respect to the incident wave and good homogeneity of the electric field by taking advantage of the electromagnetic resonance effect; [Fig.9] is a schematic representation of a resonator block and the housing of a sample of biological material in this resonator block, according to embodiments of the invention; [Fig. 10] is a representation of an electromagnetic system comprising the resonator block of [Fig. 9] placed in an electromagnetic wave exposure device of the guided wave type, namely a waveguide; [Fig. 11] is a cross-sectional view of the system in figure [Fig. 10] at the level of the resonator block, illustrating in particular the definition of the propagation conditions at the boundaries of said resonator block; [Fig.12] is an isometric perspective view of an embodiment of a waveguide section provided with a lateral drawer adapted for the arrangement of a resonator block and its placement in the waveguide; [Fig. 13] is a diagram illustrating the distribution of the electric field in a sample of cells in solution in the sample housing of the resonator block of [Fig. 9], placed within the exposure device of [Fig. 10] and [Fig. 11], according to an EMF wave application system according to the invention; and, [Fig. 14] is a schematic representation of an electromagnetic system comprising several resonators according to embodiments placed in a multi-branch electromagnetic wave exposure device for the simultaneous illumination of these resonators by the same electromagnetic wave in guided mode. Description of the implementation methods

[0039] In the description of embodiments that follows and in the figures of the attached drawings, the same or similar elements bear the same reference signs.

[0040] With reference to [Fig. 1] and [Fig. 2], there are two ways of sending an electromagnetic wave (EMW) onto a sample to be tested 12, or object under test (OUT), such as a sample of biological material in the applications which are considered here: - either in a radiated manner, when the electromagnetic wave source 11 emits an electromagnetic field E which propagates, in the form of a plane wave with a frequency in the radio or microwave range, in an open medium (often air) in which the sample is located; - either in a guided manner when the electromagnetic wave source 11 emits a constrained electromagnetic field for propagation of the EM in guided space, in an electrically conductive structure in which the sample to be tested 12 is placed.

[0041] In the first case above, which is schematically illustrated in [Fig. 1], the electromagnetic wave source 11 can be a single antenna, such as a horn antenna, The source 11 is placed in an anechoic chamber 10 or HF chamber. Absorbers are placed on the internal walls of the chamber to minimize reflections and prevent interference with the outside. The source 11 emits a plane electromagnetic wave at a given frequency Fo in the radio frequency or microwave spectrum. The dimensions of the chamber are chosen to be larger than the corresponding wavelength X in order to incorporate far-field conditions. In this context, the far field is a function of the wavelength X and the diagonal dimension D of the antenna, and is equal to 2D² / X. Thus, the object under test 12 can be illuminated by a plane wave. The electromagnetic wave propagates radiatedly in an open medium corresponding to the space inside the anechoic chamber 10. This medium is typically air, or a vacuum with a permittivity e₀ equal to unity (e₀ = 1).

[0042] For the record, the dielectric permittivity er of any propagation medium is a complex quantity that describes the response of said medium to an applied electric field, concerning the organization of electric charges in the medium, in particular the displacement of charges and the reorientation of electric dipoles. In the context of the applications considered here, the dielectric permittivity er is equal to: [Math.l] p — p j'-Ff..... where: &r -If-J e0 is the permittivity of free space, eO is equal to unity: (e0 = 1), er is the real part of the relative permittivity of the propagation medium ("relative" being understood with respect to the permittivity of free space e0), and is also called, in this document, the real part of the permittivity of the propagation medium or simply "permittivity", oE is the equivalent electrical conductivity of the propagation medium (it represents the phenomena of both electrical conduction and dielectric polarization), f is the electromagnetic background frequency propagating in the propagation medium, and j is the complex number such that j2 = -1.

[0043] The dielectric properties of a propagation medium are defined either by the value θE defined above in the formula for the relative dielectric permittivity εr, or by the pair {εr, Tanô], where Tanô is the tangent of the dielectric loss angle. The term Tanô takes into account simultaneously the losses by conduction and the losses due to polarization and relaxation phenomena, according to the formula: [Math.2] tanô = œ =

[0044] In the following description, both notations are used. Reference will essentially be made to the value er of the real part of the relative dielectric permittivity of a propagation medium formed by the material constituting a considered element of the device, which is independent of the electromagnetic background frequency f, to characterize said material in relation to the other elements of said described device.

[0045] The relative permittivity of the object under test (OUT) must also be taken into account to assess the coupling phenomenon between the propagation medium of an electromagnetic wave (EMW) and the OUT that is illuminated by this EMW. For example, when the biological cells to be tested are suspended in an aqueous solution, the relative permittivity of the sample is on the order of 80 (er ~ 80).

[0046] In the radiated approach, only the coupling surface of the object under test 12 is exposed, so that little energy (relative to the total energy emitted by the source 11) reaches the biological sample concerned. Furthermore, the field distribution in the biological sample is not spatially homogeneous. If expressed (as a percentage) as the inverse of the difference between the maximum value Emax and the minimum value Emin of the electric field in the sample, H(Emax - Emin), the field homogeneity is less than 5% for a biological sample diluted in 1.5 milliliters (ml) of aqueous solution contained in a conical Eppendorf™-type microtube made of polypropylene with a thickness of 1 mm, for example. The main drawback of the radiated approach, in this case, is the permittivity contrast, on the order of 1 / 80, between the permittivity of the vacuum (e0 = 1) and that of the sample (er ~ 80).Furthermore, the conical shape of the micro-tube 14 implies small dimensions at the bottom of this tube (at its end opposite the inlet of the micro-tube), where the field is concentrated.

[0047] Although the implementation of the invention can accommodate a free-space exposure device (radiated approach), the embodiments proposed below are based on the principle of the guided approach, which is schematically illustrated in [Fig. 2]. This approach is more controllable than the radiated approach. In the guided approach, the sample 12 forming the object under test (OUT) is placed inside a guided wave structure 13 of an exposure device that includes said structure and the source 11. This structure is, for example, a section of waveguide, for example, a metallic waveguide with a square or rectangular cross-section.It has at least two electrically conductive elements made of electrically conductive material, for example metallic elements, which are arranged substantially parallel to each other and parallel to a longitudinal direction X-X' of the structure (which is the direction of propagation of the EM wave), and which are opposite each other, so that the OST can be positioned between these two elements. In the example shown schematically in [Fig. 2], these two plates are, for example, the upper and lower walls of a waveguide. Rectangular 13. Waveguided structures are single-mode structures, in that only one mode of the electromagnetic wave propagates. Thus, a metallic waveguide filled with a homogeneous and isotropic dielectric material (usually air) supports an electric transverse mode (TE) and a magnetic transverse mode (TM), but not an electromagnetic transverse mode (TEM). Unwanted modes, generated by reflections at the OST for example, do not propagate in the waveguide.

[0048] The exposure device may also be, in other examples, a resonant cavity or a reverberation chamber (CRBM). It may also be a section of transmission line having at least two coplanar electrical conductors, including and without limitation a two-wire line, a two-plate line such as a strip-line, a coaxial line, a wire-plate cell (or WPC, from the English "wire patch ceU"), a three-plate line, or a Transverse Electrical Magnetic (TEM) cell.

[0049] As a reminder, TEM cells are rectangular transmission lines capable of generating uniform and perpendicular electric and magnetic fields (TEM mode). They contain a partition separating two cavities, formed by a metallic plate called a "septum," parallel to the electric field and narrower than the outer metallic plates. This cell is capable of generating a uniform and perpendicular electric and magnetic field (TEM mode) within a so-called test area, also known as the test volume. The frequency band in which the cell is capable of creating this TEM wave depends directly on its transverse dimensions. The plates confine the electromagnetic field in order to characterize the electromagnetic radiation to which the OST is subjected according to a polarization determined by their geometry.The use of TEM cells relies on the propagation of a transverse electromagnetic mode contained between the "septum" and the outer walls of the metallic enclosure. The "septum" is a large-diameter planar conductor, placed within the internal volume of the enclosure and connected to the outside of the cell via two connectors passing through the enclosure walls. The OST is thus placed in an enclosure that protects it from the external environment, and it is exposed to a grazing electromagnetic wave of the transverse electromagnetic (TEM) type.

[0050] An electromagnetic wave source emits an EM wave at a given frequency in the radio frequency or microwave spectrum, which is made to propagate, preferably in guided mode, inside the exposure device. The coupling between the electromagnetic field emitted by the source and the biological cell sample exposed in the applicator is better than in the radiated approach because the emitted electromagnetic energy is essentially, or even entirely, contained within the exposure device. Other advantages of the guided approach compared to the radiated approach include: • improved electromagnetic compatibility (EMC) because the system can be naturally shielded (as in the case of a metallic waveguide, coaxial cable, etc.), • a smaller footprint, and • the fact that the user is not exposed to the emitted electromagnetic field since there is no, or very little, electromagnetic leakage to the outside of the exposure device.

[0051] The conventional approach for guided exposure consists of inserting a conventional biological container, for example a Petri dish or an Eppendorf™-type microtube as illustrated in [Fig. 3A] and [Fig. 3B], into the exposure device in which the electromagnetic field emitted by the source propagates in the radio and microwave ranges. The source 11 is integrated into, or coupled to, the exposure device, for example at a vent in a conductive wall in the case of a waveguide 13 as schematically represented in [Fig. 2].

[0052] For in vitro exposure of biological cells, these cells are generally biological cells suspended in a liquid medium, for example, an aqueous solution. Such a solution is usually contained in a device such as an Eppendorf™-type microtube or a Petri dish, for manipulation in the laboratory. Alternatively, the cells can be deposited directly (sometimes said to be "glued") between two glass slides or in an incubator, such as a Petri dish. In the case of conventional solutions used for exposure to an EM (Electromagnetic Element), the EM in question therefore consists of these slides or this incubator, along with the biological cells.

[0053] Figure 3A shows an open and empty Eppendorf™-type microtube 14, and Figure 3B shows the same microtube 14 closed by its cap after being filled with a solution 15, for example an aqueous solution, containing the biological cells to be tested in suspension. In the case of conventional solutions used for exposure to an EM, the EM in question therefore comprises the microtube 14 as well as the aqueous solution 15 with the biological cells in suspension contained therein. The actual part er of the relative dielectric permittivity of such test objects is high, greater than 20, or even very high, often greater than 50, in the radio frequency and microwave ranges.

[0054] The embodiments of the present invention provide, for an incident EM at a given frequency in the radio frequency or microwave spectrum, a better energy balance from an electrical point of view than conventional implementations which are carried out on Petri dishes or Eppendorf™ type microtubes, for example. More Specifically, the performance in terms of electromagnetic field application obtained according to the teachings of the invention is improved with respect to three factors. These three factors are: - the coupling factor (denoted K in this description) of electromagnetic background in the OST; - the spatial homogeneity of the electromagnetic field in the OST; and, finally, - the dosimetry of the IEI electromagnetic field.

[0055] The coupling factor K can be defined as the ratio between the incident electromagnetic field level and the electric field level E actually emitted into the sample under test. In what follows, the coupling factor of the exposure device is expressed as a percentage (%). If K is less than or greater than 100%, the electromagnetic wave is undercoupled or overcoupled, respectively. Of course, by virtue of the general principle of conservation of energy, any overcoupling can only be limited to a specific area of ​​the propagation medium and compensated by undercoupling in other areas of said medium so that, overall, the distribution of the IEI values ​​of the electromagnetic field in the volume considered complies with this general principle.

[0056] The spatial homogeneity of the electromagnetic field in the OST accounts for the contrast between the maximum and minimum field values ​​in different parts of the sample under test. This is also referred to, more simply, as the homogeneity or uniformity of the electromagnetic field. In what follows, and unless explicitly stated otherwise, the uniformity of the electromagnetic field in the OST is expressed by the Coefficient of Variation (CV), defined as the standard deviation (denoted by the Greek letter sigma, 0) of the mean (Emoy) value of the electromagnetic field. This coefficient more precisely measures the dispersion of field values ​​in the sample relative to the mean. Ideally, there should be no variation in the IEI amplitude of the electric field within the volume of the sample, to ensure that each cell is subjected to the same electric field value.In practice, it is desirable to minimize the standard deviation of the electric field values ​​in the sample under test in order to best assess the thresholds of effects of exposure of the biological cells of the sample to the electromagnetic field.

[0057] Dosimetry, for its part, is the function that gives the IEI value of the electric field in the sample as a function of the power P of the emitted electromagnetic wave, according to the relation IEI = F(P). Control of this function (and therefore control of the electromagnetic operation of the system) allows direct access to the value of the electric field in the sample by knowing the emitted power.

[0058] In the context of the invention, what is called an electromagnetic wave (EMW) application device (or applicator) is a device that provides the interface between the OST and the device exposing this OST to an electromagnetic field at the frequency of interest. Its function is to improve the transition, within the biological cells contained within the OST, of the electromagnetic field propagating (preferably in a guided manner) within the exposure device. More specifically, an application device according to the proposed embodiments aims to provide good homogeneity of the electromagnetic field in the sample under test to allow for tests with satisfactory experimental value, and to use resonance modes to obtain coupling factors greater than unity (≥1) specifically at the OST. This therefore makes it possible to obtain strong electric field levels in the biological cells under test in order to observe the effects of exposure to the electromagnetic field on these cells, which are to be studied.Improving the dosimetric aspect of electromagnetic coupling, combined with good homogeneity of electric field values ​​in the OST, also proves useful for researching and studying the thresholds at which the effects in question appear.

[0059] The drawback of conventional applicators known in the prior art is that biological containers are not electrically optimized for use with such applicators. These containers, for example Eppendorf™-type microtubes or Petri dishes, are mostly made of polypropylene (with a permittivity er of 2.2) or similar materials. They are not effective in ensuring a good transition of the OEM in a biological medium (for example, an aqueous solution with a permittivity er of approximately 80). Indeed, the sometimes complex shapes and the significant heterogeneity of permittivities and geometries of the various elements and materials involved severely mismatch the applied OEM. Furthermore, the more permittivity breaks and uncontrolled geometric planes there are, the more the system generates inhomogeneity in the field values ​​within the OST.This drawback is compounded by insufficient coupling of the OEM within the cellular environment. In other words, direct exposure of conventional biological containers presents serious limitations. The invention is based on the observation that these limitations stem from a problem related to the interface between media with strong contrasts in dielectric permittivities, and addresses this problem.

[0060] A study presented in the article V. VAN EEGHEM et al., "The impact of millimeter waves on the enzymatic activity of succinate dehydrogenase" - Abstract from The Joint Annual Meeting of The Bioelectromagnetics Society and the European BioElectromagnetics Association co-organized with the European COST EMF-MED Action BMI309, Ghent, Belgium - 2016, suggests that coupling is improved if a polypropylene container containing OST is placed in a favorable medium, i.e., one having the same electrical characteristics as OST, rather than in a vacuum (whose permittivity eR0 is equal to 1). However, the high variability of distribution The field makes a quantitative study difficult. The interface with the polypropylene container is a limiting factor. Indeed, a permittivity contrast of 2.2 / 80 remains at this interface.

[0061] With reference to diagram 101 in [Fig. 4], it follows that, for example, illuminating biological cells in solution in a conventional Petri dish exposed in a TEM cell, with a delivered power of 1 W generating an incident field on the sample estimated at 156 V / m, results in a field distribution within the sample that is not very uniform. This is reflected in diagram 101, which is a diagram of the distribution of IEI field values ​​(in V / m) represented on the horizontal axis between 0 and 1600 V / m, at a number of points on the OST that are substantially regularly distributed (spatially) within this object. In [Fig. 4], the number of points (expressed as a percentage of the total number of points recorded) for each field value among a given number of discrete values ​​is indicated on the left-hand vertical axis.Curve 102 shows, between the values ​​0 and 100% indicated on the right-hand vertical axis, the cumulative field values ​​recorded in the interval [0 V / m - 1600 V / m]. In this example, the average value (Emoy, average value over the number of points) of the field transmitted in the OST is 41 V / m, i.e., a coupling rate of 26% (i.e., K=26%) with a maximum value (Emax) equal to 359 V / m, a minimum value (Emin) equal to 41 V / m, and a standard deviation (o) of 16.2 V / m on said average value.

[0062] It can be noted that the incident field for the TEM cell is related to its height. It would be possible to have the same incident field with a smaller gap between the conductors. However, those skilled in the art will appreciate that what matters is understanding the concept of coupling between the incident field and the field in the sample, as well as the dispersion of electromagnetic field values ​​in the sample of cells included in the OST.

[0063] Similarly, in a conventional Petri dish exposed in a waveguide with a power output of 1 W generating an incident field on the sample estimated at 376 V / m, the distribution of the field within the sample is also not very uniform. This is reflected in diagram 201 of [Fig. 5], which is a diagram of the distribution of values ​​(in V / m, represented on the horizontal axis, between 0 and 300 V / m) of the electromagnetic field deposited at a number of points on the OST that are substantially regularly distributed (spatially) within this object. In [Fig. 5], the number of points (as a percentage of the total number of points recorded) for each field value is indicated on the left-hand vertical axis. Curve 202 shows the cumulative total, between 0 and 100% (right vertical axis), of the points recorded in the interval [0 V / m - 300 V / m] of the recorded values ​​of the electromagnetic field.In this example, the average value (Emoy) of the electromagnetic field deposited in the OST is 25 V / . m i.e. a coupling of 7% (i.e., K=7%), with a maximum value (Emax) equal to 289 V / m, a minimum value (Emin) equal to 9 V / m, and a standard deviation (o) of 23.4 V / m on said average value.

[0064] In both cases above, it follows, for the applications envisaged, on the one hand, that it is therefore difficult for users to conclude on a field level from which effects of the electromagnetic field on the biological cells of the sample are observed, and on the other hand, that the repeatability of the experiments carried out is strongly impacted.

[0065] Furthermore, a simulation of the electromagnetic field intensity in the TEM cell shows that most of the EM is not coupled in the OST. At best, only 10% to 20% of the field penetrates the biological cells of the OST. In other words, the coupling K of the EM in the OST is limited to a low value, between 10% and 20%. It is therefore necessary to oversize the radiofrequency or microwave wave source, for example by using an oversized radiofrequency (RF) amplifier, which is significantly more expensive, to achieve the desired field level in the biological cell sample for the purposes of the scientific experiment. Indeed, in some use cases, the electric field values ​​expected to observe an impact on biological cells are on the order of several kilovolts per meter (kV / m), up to a few hundred kilovolts per meter.

[0066] This is why electromagnetic wave application devices have already been designed specifically for bioelectromagnetic analysis. The most common of these known applicators is the biological cuvette made of plastic of variable thickness (on the order of a millimeter) held between two metallic electrodes. Applying a voltage to these electrodes induces an electromagnetic field in the device. The electromagnetic field thus applied to the samples is directly the ratio of the potential difference across the electrodes to the inter-electrode distance. However, the change in interface at the OST creates a lack of homogeneity in the field to which the cells in the sample being tested are exposed.Other devices utilize the electrical properties of biological solutions to insert them into adapted electromagnetic transmission lines; that is, they employ the principle of transmission line matching. The devices can be planar, allowing for real-time microscopic observation of the samples.

[0067] Devices of this type have been the subject of the following theses, as well as a report by ASSFET: • Sophie KOHLER: "Contribution to the development and multi-scale dosimetry of devices for the exposure of biological cells to high intensity nanosecond and sub-nanosecond electrical pulses" - University of Limoges - Thesis no. 43-2013 - November 2013. • Maïak SOUEID: "Contribution to the development and characterization of applicators for bio-electromagnetic studies involving radiofrequency ounces and high-intensity nanosecond electrical pulses" - Thesis - University of Limoges - November 2016. • Aude SILVE: "New devices for the controlled application of nanosecond electrical pulses and for the detection of their effects on cells. New results and hypotheses on the parameters controlling the electro-permeabilization of biological cells" - Thesis - University Paris Sud XI - November 2011. • Report from the French Agency for Environmental and Occupational Health Safety (AFSSET) "Radiofrequencies", Annex 3.2: "Review of existing experimental exposure systems", October 2009.

[0068] The applicators disclosed in the prior art documents already cited in the introduction to this description, namely documents FR2998813A1, FR3034521A1, and FR3021743A1, were also specifically designed for bioelectromagnetic analysis. These applicators are optimized from the point of view of radio frequency (RF) electromagnetism. Indeed, they make it possible to achieve high electric field values ​​with good homogeneity, defined in these documents and therefore in this paragraph as the ratio of the difference between the maximum intensity (Emax) and the minimum intensity (Emin) to the maximum intensity, i.e., the ratio (Emax-Emin) / Emax, and expressed as a percentage. In fact, this homogeneity can exceed 95%. However, they only involve extremely small volumes of liquid solution (including the cell sample to be tested), on the order of a few microliters.Furthermore, their handling is delicate. And although the electrical coupling factor K between the object and the electric field is excellent, it remains below 100%.

[0069] As will now be described, embodiments of the invention allow the in vitro exposure of biological cells, benefiting, for an incident EM at a given frequency of interest Fo, from an electrically improved energy balance compared to prior art implementations conventionally performed on biological cells in Petri dishes or Eppendorf™ tubes, for example, and also compared to known applicators disclosed in documents FR2998813A1, FR3034521A1, and FR3021743A1. This improved energy balance at the OST level results from the phenomenon of electromagnetic resonance. In other words, the embodiments improve the coupling ratio K of the incident EM in the OST, by allowing for local overcoupling (> 100%). This avoids the cost of oversizing electromagnetic wave sources (and more specifically the cost of high-performance amplifiers required in these sources), or being less limited in terms of coupled field values ​​in the sample under test for a given source.

[0070] Furthermore, with the embodiments proposed here, the biological cells of the sample under test are exposed to a relatively more uniform field level than in the case of conventional implementations or implementations with applicators known from documents FR2998813A1, FR3034521A1 and FR3021743A1 already discussed. In other words, the homogeneity of the spatial distribution of the electromagnetic field to which the biological cells of the sample are exposed is improved compared to everything known of this prior art.

[0071] Finally, thanks to the proposed embodiments, the electromagnetic field level applied to the OST is better controlled. This improves dosimetry, in that the function F, which gives the IEI value of the electromagnetic field in the sample as a function of the power P applied by the EM source, is more deterministic; that is, the field value actually available in the OST is more precise and reproducible. More specifically, this means that it is more consistent with the expected values ​​based on the relationship IEI = F(P).

[0072] As indicated above, the idea behind the embodiments of the invention consists of placing the object under test (OST), namely the biological cells to be illuminated and their support, at the heart of an electromagnetic resonance mode of the electromagnetic wave produced by the exposure device and which propagates in the applicator device and is coupled in the OST, while ensuring a significant improvement in the spatial homogeneity of the field level in the OST compared to existing applicators, as well as better control (dosimetry) of the I El value of the field which illuminates the cells.For this purpose, the biological cells to be illuminated are placed in a resonator block of the electromagnetic field applicator, which is specially designed and adapted so that the OST is placed inside a housing provided in said resonator, the position and dimensions of which correspond to an area where, during illumination, there is a peak of electromagnetic Fonde resonance at the frequency of interest Fo. .

[0073] To this end, the resonator block is made of at least one specific dielectric material (i.e., an electrically insulating material). It has the general shape of a closed volume of specific dimensions. It is a solid (not empty) volume, except that it includes at least one compartment, called the sample compartment, to receive the sample containing the biological cells to be tested. For example, the sample may be an aqueous solution in which the biological cells are suspended, with a The relative dielectric permittivity er is close to that of water, around 80 (er = 80). The sample chamber can be sealed with a stopper after the sample to be tested has been inserted in a biological manner, for example, after filling the chamber with the aforementioned solution. The stopper includes at least one portion made of the same dielectric material as the resonator block, in order to limit dielectric permittivity discontinuities. In practice, it is also advantageous for the chamber to be completely filled with the sample. This avoids leaving a void in the chamber, which would cause dielectric permittivity discontinuities within the resonator that could alter the electromagnetic resonance mode(s) in the resonator block.

[0074] By design, this resonator block is adapted to exhibit, within itself, a particular spatial distribution of an electromagnetic field at a given frequency when it is subjected to illumination by said field. This spatial distribution of the field has discrete geometric configurations defining electromagnetic resonance modes. In other words, areas of determined location and extent are identified within the volume of the resonator block, in which the field resonates at the frequency of the wave injected into the resonator. These areas are associated with a natural resonance frequency and are determinable by numerical simulation and adjustable based on constitutive parameters of the applicator. These parameters include the relative permittivity er of the dielectric material from which the resonator block is made, as well as the geometry and dimensions of said resonator block, and the boundary conditions imposed.The identification of electromagnetic resonance zones is further performed in combination with electromagnetic parameters of the biological material sample (Le.. of the OST) housed within the sample compartment, including the relative permittivity of the sample, but also preferably the electrical conductivity ν of said sample. The aforementioned constitutive parameters of the resonator block are chosen, during the design of the applicator, such that a resonance frequency in said resonator when it is exposed to an electromagnetic field at that frequency corresponds to the frequency of interest ΔF₀. In particular, the shape of the sample compartment and its position within the resonator block are defined so that said compartment occupies an area within the solid volume of said resonator block that corresponds to a resonance peak of the electromagnetic field at said frequency of interest ΔF₀.

[0075] In practice, the applicator design process can largely include numerical simulation operations to verify the suitability of design choices, and to modify, refine, etc., them according to the results predicted by the simulations. The use of numerical simulation is a good alternative to the direct ("in concrete") measurement of the electromagnetic field distribution, in the extent to which the electromagnetic phenomena involved are well modeled in simulation software since they essentially proceed from Maxwell's equations of electromagnetism which are well understood for structures with a simple and determined geometry (parallelepiped, cylinder, etc.)

[0076] A person skilled in the art will appreciate that direct field measurement is inherently intrusive, in that the presence of any measuring tool necessarily influences the polarization of the OEM at the measurement site. Therefore, direct measurement using conventional means is not suitable for generating the readings and diagrams referred to in this disclosure to illustrate the performance of the various configurations considered, both with regard to the prior art discussed and the proposed implementation methods.However, since all the constituent parameters of the devices under consideration are controlled (nature of the source and intensity of the emitted electromagnetic wave, geometric shapes, dimensions, nature of the materials constituting the devices), and since we are dealing with a closed propagation system (the applicator preferably being placed in a guided wave device of an exposure unit), the field values ​​at all points of the system can be determined by software simulation. In the examples considered here, the simulation software used belongs to the CST Studio Suite™ design environment from the simulation software publisher Dassault Systèmes™. This includes, in particular, the CST Microwave Studio™ software.Regarding the design of electromagnetic field applicator devices according to the embodiments of the invention, the comparison between the numerical simulation and the measurement carried out with sophisticated means made it possible to validate the response of the applicator with respect to the initial assumptions.

[0077] The theoretical aspects related to resonance modes in a cavity are well known. Reference can be made, for example, to the work by D. KAJFEZ and P. GUILLON entitled "Dielectric Resonators", Artech House, 1986. It is known that in a waveguide or a resonant cavity with conductive walls, the electromagnetic field can only take on specific configurations. Indeed, the conductive walls impose a boundary condition such that the electric field is necessarily zero at the level of said conductive walls. Any propagation configuration not respecting these conditions cannot exist in such a system.

[0078] Thus for example, as illustrated on [Fig.6] which shows the cross-section, in a plane (Y,Z), of a waveguide of rectangular section having infinitely conductive walls (i.e., walls of a material whose conductivity θ tends towards infinity), the field can only take certain configurations including, for example, configuration (a) on the left and configuration (b) in the center of the figure. These configurations impose specific guided wavelengths Xg, that is, guided wave frequencies Fg, which can only exist. The third case (c), shown on the right of the figure, is not a possible configuration because it does not satisfy the boundary conditions under which the value of the IEI field is zero on the conducting walls (which is a consequence of Maxwell's equations of electromagnetism).

[0079] In electromagnetics, resonant cavities are widely used for filtering electromagnetic interference (EMI), with very low losses, especially at high power levels. In a waveguide such as the one shown on the left of [Fig. 6], configuration (a) represents the minimum frequency configuration (in Xg / 2) of the electromagnetic field that can exist, which gives a cutoff frequency. All higher frequencies can propagate in this waveguide. Indeed, a waveguide has no boundary conditions in the third direction (i.e., in the example shown, the X direction, which is orthogonal to the cross-sectional plane of the representations in [Fig. 6]), so the electromagnetic field is not constrained along this direction. However, if conductive walls are added in this third X direction, a cavity is obtained in which the field is constrained in all three directions (i.e.(the three degrees of freedom), allowing only certain field configurations. These are the resonance modes of the cavity, at the corresponding natural frequency / g of the guided wave.

[0080] In electromagnetics, the term "cavity" refers to a finite dielectric medium, bounded in three-dimensional space by a closed envelope, and within which an electromagnetic field exists. This electromagnetic field must satisfy Maxwell's equations, taking into account any boundary conditions imposed on the surface of the closed envelope. In practice, three main types of cavities are used: parallelepiped cavities (with rectangular cross-sections), cylindrical cavities (with circular cross-sections), and coaxial cavities, for which Maxwell's equations can be solved exactly (which is not the case for resonant volumes of arbitrary shape). The solutions found to these equations constitute the natural modes of oscillation of the cavity. Those skilled in the art will appreciate that the concept of "cavity" is not limited to empty volumes (containing air, or a vacuum with a permittivity e0 equal to one).In other words, in electromagnetics, cavities may or may not be filled with a homogeneous dielectric material.

[0081] For example, Figures [Fig. 7A] to [Fig. 7D] show different electric field distributions for four distinct resonance modes in a resonant cavity 20, also called a resonator block in the context of the invention, which is shown here in isometric perspective (in three dimensions, or 3D). It is a solid block made of one (or more) dielectric material(s), and of the form A parallelepiped with dimensions determined along each of the three directions X, Y, and Z of a three-dimensional orthogonal coordinate system {X,Y,Z}, which were mentioned earlier with reference to [Fig. 6]. By convention, the X direction is oriented along the longest side of the parallelepiped and is therefore called the longitudinal direction. Similarly, by convention, the Z direction is oriented along the direction of gravity and is therefore called the vertical direction. In the following description, the terms "vertical" and "vertically," "up" and "down," "lower" and "upper" are understood to refer to an orientation substantially parallel to the Z direction, and the terms "horizontal" and "horizontally" to refer to an orientation substantially parallel to the (X,Y) plane. Finally, the (Y,Z) plane, which is orthogonal to the longitudinal direction, is called the transverse plane.In the context of this description, the term "transverse" means orthogonal to the longitudinal direction.

[0082] The resonance modes shown in the examples in Figures [Fig. 7A] to [Fig. 7D] each correspond to a specific electromagnetic field (EMF) of a given frequency, called the natural frequency of the resonance mode, or resonance frequency / r. These figures provide a visualization of different resonance modes, each corresponding to a certain distribution of the field within the volume of the resonator block 20, with the electric field values ​​being higher at some points than at other points in this volume. This creates a localized reinforcement of the electromagnetic field at certain points in the volume of the resonating cavity compared to other points in this volume, these reinforcements corresponding to resonance peaks. In the figures, the darkest points are thus points where the IEI value of the electric field is equal to the maximum value Emax, and the lightest points are points where the IEl value of the electric field is equal to the minimum value Emin.

[0083] The distribution of electrical energy in the volume of the resonator block 20 depends on:

[0084] the frequency of the OEM,

[0085] of the relative dielectric permittivity of the dielectric medium, i.e. of the constituent material(s) of the resonator block or present in said block,

[0086] of the geometry and dimensions of the resonator block, and

[0087] boundary conditions of this block.

[0088] The invention teaches how to position biological cells within the OST in a region corresponding to a maximum (i.e., a peak) of the electric field distribution within the dielectric material of a resonator. Taking into account the dielectric properties of biological cells suspended in an aqueous solution (having a permittivity close to that of water, therefore around 80), the objective is to associate a certain volume of these solutions with one or more dielectric materials and to provide specific boundary conditions so that the OST is located in an area corresponding to a resonance peak.

[0089] Thus, by controlling the arrangement of the elements and their geometries and dimensions, the cells suspended in the solution are located in the maximum of the electric field, ensuring both a coupling factor greater than unity (100%) with respect to the incident wave and good homogeneity of the field in the OST.

[0090] Three cases are presented in cross-sectional views in [Fig. 8A], [Fig. 8B], and [Fig. 8C] to visualize the ideal position of the OST to take advantage of the resonance effect. In these figures, the areas within the volume of the resonator block 20 that correspond to a resonance peak at the considered OEM frequency are identified by two white circles 81 in [Fig. 8A], by a white rectangle 82 in [Fig. 8B], and by a white ellipse 83 in [Fig. 8C]. It should be noted, in particular with reference to [Fig. 8A], that it is entirely possible for there to be two (or more) areas within the resonator block corresponding to distinct resonance peaks at the same OEM frequency. The distribution of the electric field in the resonator block depends only on known parameters, which are the frequency of the OEM, the relative dielectric permittivity of the dielectric material from which it is made, its geometry, its dimensions, and the boundary conditions.

[0091] The design of a resonator block for manufacturing an electromagnetic field application device (applicator) according to embodiments of the invention, is therefore carried out taking into account all these parameters, and taking into account, in addition, the relative dielectric permittivity er and the electrical conductivity o of the OST, with regard to the frequency of interest Fo of the OEM to which the biological cells are to be subjected for the purposes of the experiment considered.More specifically, taking these parameters into account in the software tools used to perform the simulations leads to defining the position and dimensions of one or more housings in which it will be possible to place the OST so that, when exposed to an EM at the planned frequency Fo, the biological cells in this OST are illuminated by an electric field of maximum value, with in any case a coupling coefficient K greater than 100%, and this in a homogeneous manner and with good precision and reproducibility (dosimetric aspect).

[0092] Those skilled in the art will appreciate that applicators manufactured according to embodiments of the invention are single-frequency, that is, they function as expected only under illumination by an OEM at the frequency for which they were specifically designed. This implies that, for a given type of OST, an applicator is manufactured for each frequency Fo of interest, by working on the permittivity of the material(s), the geometry, or The dimensions of the resonator and / or the boundary conditions. In some embodiments, however, one can start with a readily available resonator block designed for a frequency higher than the frequency of interest and encase this existing block (after inserting the OST into the sample chamber and sealing the chamber) in a different dielectric material. For example, one can fabricate a container made of dielectric material into which an existing resonator block is inserted. This amounts to adding an envelope to the existing resonator block, which lowers the resonant frequency of the device, depending on the dimensions and permittivity of the envelope. We will return to such an embodiment later.

[0093] In what follows, we will detail the manufacturing methods of an example of an OEM applicator, considering, without limitation, the case of a resonator block which is a parallelepiped-shaped rod of dielectric material as illustrated by Figures [Fig. 7A] to [Fig. 7D] in which, at the frequency Fo of interest, the distribution of the IEI values ​​of the electric field conforms to the example in [Fig. 8B]. In this example, the area 82 of the volume of the resonator block 20 which corresponds to a resonance peak is a central area inside the rod, and extends along the principal direction of this rod.

[0094] In this example, the object under test is a volume of distilled water whose real part of the relative dielectric permittivity er is 78.4 and the equivalent electrical conductivity oE is 5.6.10-6 S / m at the frequency Fo of 1.5 GHz.

[0095] With reference to the isometric perspective diagram in [Fig. 9], the resonator block 20 can be a rectangular bar made of a dielectric material such as a ceramic. Composite materials with high relative and real dielectric permittivities, greater than, for example, 70, can also be considered. Preferably, the materials in contact with the biological solution are also chosen for their biocompatibility, such as ceramics, for example, barium (meta)titanate (BaTiO3). Preferably, the resonator is made of a ceramic with a so-called "high relative real permittivity." In one example, the relative real permittivity of such a ceramic is approximately 78.

[0096] Each geometric structure has as many natural frequencies and associated vibrational modes (oscillation patterns) as there are degrees of freedom for vibration propagation, and these frequencies can be calculated for structures with defined geometry. In the case of the parallelepiped-shaped dielectric resonator block 20 shown in [Fig. 9], for example, the resonance frequency fr for the m,n,p resonance mode is given by the following mathematical equation, called the resonance equation, which is solved by commercially available simulation software: [Math 3] Or : It is the speed of light; er denotes the relative permittivity of the dielectric material from which the resonator is made; A, B, and C denote the dimensions of the resonator block along the three axes of the resonator block, respectively; and, m, n, and p are many positive integers or nuis that characterize the resonance mode along the three directions X, Y, and Z of the resonator block, respectively, and correspond to the number of half-wavelengths along these three directions, respectively.

[0097] The method for designing the applicator device according to embodiments of the invention comprises the following steps: • for a given resonator block (nature of the dielectric material, geometry and dimensions of said resonator block), for example the parallelepiped of [Fig.9], the calculation of the eigenmodes gives different distributions of the IEI value of the electromagnetic field (see the figures from [Fig.8A] to [Fig.8C] for the case of a parallelepiped resonator block in the example considered here); • we retain the distribution corresponding to a determined resonance mode which is considered promising (in this case that of [Fig.8B] in the case of the example considered here); • The position and dimensions of a hollow cavity 21 in the resonator block 20 are determined, namely a recess accessible from outside the block (in this case, from the top of the resonator block as shown in [Fig. 9]), which corresponds to a region of the resonator volume in which the simulation indicates a peak, or resonance peak, in the distribution of the IEI electric field values ​​in the resonator block. The sample of biological cells to be tested will be placed in this cavity, at the center of the resonator region corresponding to the resonance mode in question, where the field level will be relatively homogeneous and of high value, higher than that of the incident field due to the resonance phenomenon; then, • The resonance frequency / r associated with this resonance mode is set to the frequency of interest Fo (i.e., the one at which we want to perform a test). illumination of biological cells) by modifying the value er of the permittivity by changing the nature of the dielectric, and / or by modifying the geometry or dimensions of the resonator, and / or by modifying the boundary conditions to bring the resonance frequency to the frequency of interest Fo.

[0098] In the last step above, the constitutive parameters of the applicator (in particular the er of the dielectric material, the geometry and dimensions of the resonator block, as well as the boundary conditions) are adjusted so that the resonance frequency fr corresponds to the frequency of interest Fo considered, and also as a function of the permittivity of the medium containing the biological cells to be tested, i.e., the OST. This can be achieved, by successive approaches, through numerical simulations carried out using simulation software such as, for example, the CST Microwave Studio™ software already mentioned above.

[0099] As a person skilled in the art will appreciate, when designing a new applicator for testing biological cells by illumination under an EM at a given frequency of interest Fo, one can start from the design file of an applicator previously made for testing a similar OST by illumination under an EM at another frequency close to said frequency of interest Fo, and proceed to modify all or part of the constitutive parameters of the initial applicator to arrive at the intended applicator.

[0100] Alternatively, the designer's experience will allow them to define an initial applicator design that provides one or more resonances defining peak areas of the electric field whose position, general shape, and / or approximate dimensions appear probable. Then, using successive simulations and an incremental process (through successive approximations), the designer can refine the parameters of their applicator design to arrive at a satisfactory final version.

[0101] Preferably, the dimensions of the resonator can be worked on in a "homothetic" manner, that is, by changing all the dimensions by the same ratio, without modifying its geometry. This makes it possible to simply shift the resonance frequency / r in the frequency spectrum without moving the resonance zone(s) within the volume of the resonator block corresponding to a natural resonance mode.

[0102] As an alternative or in addition, one can also work on the geometry of the resonator. Alternatively or in addition, one can impose new boundary conditions on the resonator. These modifications may, in some cases, result in establishing a different resonance mode. Therefore, these two options are probably less suitable for making minor adjustments to the resonance regime in the resonator block 20, when it is only a matter of tuning a resonance frequency / r already obtained and identified to the frequency of interest Fo, without having to redefine a new position and new dimensions for the sample housing 21.

[0103] As an alternative or complement, one can also work on the permittivity er of the dielectric material constituting the resonator block 20. However, this option is not preferred for applications in bioelectromagnetism, where the dielectric material is preferably chosen from among ceramics, due to the biocompatibility of ceramics. Indeed, changing the ceramic, or even radically changing the nature of the dielectric material, can entail significant manufacturing costs. Moreover, not all materials that can be considered based on their permittivity er value are necessarily biocompatible. The method based on selecting a specific dielectric material for each new applicator, chosen for its relative dielectric permittivity er value, is therefore more complicated than in other applications, even if these complications are not insurmountable.Furthermore, working solely on permittivity without simultaneously addressing the geometry and / or dimensions of the resonator block may prevent the design of the desired resonator because, in effect, only a limited number of dielectric materials are available. Therefore, only discrete permittivity values ​​are possible, and intermediate values ​​cannot be obtained through this method.

[0104] Therefore, in other variations, and as briefly mentioned above, one can start with a readily available ceramic block and encase this block (after inserting the OST into the sample compartment and closing said compartment) in another dielectric material, for example, plastic (with a dielectric permittivity er of approximately 3 or 4). The nature and dimensions of this encasement can be adapted to modify the resonant frequency / r to match the frequency of interest Fo. This method gives good results when it is necessary to lower the resonant frequency / r relative to that given by the permittivity er of the ceramic and by the geometry and dimensions of the initial resonator. In some cases, this avoids having to manufacture a new ceramic piece to create another resonator at a slightly different frequency of interest.The ceramic can also be replaced by a dielectric gel contained within the casing, for example agar agar, which is a polymer of galactose (galactan) with a relative permittivity er of 76 and an electrical conductivity oE of 0.37 S / m at a frequency of 1.5 GHz, for example.

[0105] The invention makes it possible to illuminate a volume of biological solution on the order of one milliliter (1 ml), which corresponds substantially to the volume of an Eppendorf™ type microtube, and is substantially greater than the volume of solution that can be tested with applicators conforming to the prior art known from documents FR2998813A1, FR3034521A1 and FR3021743A1. Furthermore, the applicator is easy to handle. It defines a hermetic system with respect to external contamination, thus preventing any contamination of the biological cells to be tested during handling (in particular when placing it in the OEM exposure device).

[0106] Those skilled in the art will appreciate that, particularly when the biological cells are not suspended in a solution but are supported by another medium (e.g., lamellae), other dielectric materials can be used instead of ceramics to make the resonator block. For example, polymers can be used, notably Plexiglas with a permittivity er of 2.72 or polypropylene with a permittivity er of 2.2 and a tan θ of approximately 0.001, or polytetrafluoroethylene (better known by its acronym PTFE).

[0107] In all cases, the best results are obtained when the dielectric material(s) from which the resonator block is made have a relative permittivity er greater than one, and equal to the relative permittivity er of the biological sample to within ±50%, or even to within + / -40%, preferably to within + / -30%, preferably still to within + / -25%, preferably to within + / -20%, and even more preferably to within + / -10%. By setting a frequency of interest, the shape and dimensions of the resonator block are then optimized to obtain a useful resonance mode, as desired.

[0108] With further reference to [Fig. 9], the hollow housing 21 opens from the top to the outside of the resonator block, in order to make said housing accessible from the outside to allow the insertion of the OST. In some embodiments, the resonator 20 includes a removable plug 24 adapted to close the sample housing 21 after insertion, and preferably after the housing 21 has been completely filled by the OST (in order to fill any empty space in the hollow housing 21 and thus avoid a discontinuity in dielectric permittivity). It should be noted that, in the example considered, the object to be tested 12 (OST) is a sample of biological material, in this case a volume of aqueous solution containing biological cells, on the order of 1 milliliter (1 ml).

[0109] The plug 24 may include a lower portion 24a made of dielectric material, preferably the same material as that of which the resonator block 20 is made. This lower portion ideally has a height corresponding to the upper portion of the housing 21, which is not included in zone 82 (see [Fig. 8B]), which corresponds to a resonance peak and in which the electric field is at its maximum. It is undesirable for the OST to occupy this zone, and therefore it is desirable that it be filled by the dielectric material constituting the resonator 20.

[0110] The plug 24 may also include, in addition, an upper portion 24b made of metallic material, preferably the same material as that of which the walls of the waveguide 100 are made. This upper portion ideally has a height at least equal to, and preferably greater than, the thickness of the upper wall of the waveguide 100 along the vertical direction Z (see [Fig. 11]). Thus, this portion 24b contributes to establishing the propagation conditions at the boundaries of the resonator block 20 when the plug 24 is put in place to seal the hollow housing 21. Indeed, this metallic portion is then in contact and therefore in electrical continuity with the upper wall of the waveguide 100. [YES] In the embodiment shown in Figures [Fig. 9], [Fig. 10], and [Fig. 11], the dimensions of the cross-section of the plug 24, in section in the horizontal XY plane, are smaller than those of the resonator block 20 in section in said plane. They correspond to the dimensions of the cross-section of the hollow housing 21 in this plane. Other embodiments are nevertheless possible. In another embodiment, for example, the dimensions of the cross-section of the plug 24, in section in the horizontal XY plane, may correspond to those of the resonator block 20 in section in said plane. The construction of the plug is thus simpler.Indeed, this amounts to manufacturing a resonator block in two superimposed portions with respective cross-sectional dimensions, in section in the horizontal plane XY, which are the same and constant along its entire height in the vertical direction Y: a lower portion comprising the hollow housing 21 up to its open upper edge, on the one hand, and an upper portion extending above it, i.e., directly above this upper edge, which corresponds to the plug 24, the latter being a solid block (i.e., without a hollow part). In this embodiment, the plug 24 is always made of two stacked materials: its lower part 24a is made of the same dielectric material as the lower portion of the resonator block and is surmounted by its upper part 24b made of metallic material as described above.

[0112] Those skilled in the art will appreciate that, in the example shown in Figures [Fig. 9], [Fig. 10], and [Fig. 11], the upper portion 24b of the plug 24 protrudes upwards outside the waveguide 100 when the resonator block 20 is placed in the waveguide. This allows the user to grasp the plug 24 to open or close the housing 21 while the resonator block is in place in the waveguide (possibly in a non-removable manner). In other words, the portion of the upper portion 24b of the plug 24 that protrudes beyond the thickness of the upper wall of the Waveguide 100 can be used by the user to grip the plug when handling said plug. In other words, the system formed by the resonator block 20, the waveguide 100, and the removable plug 24 can be delivered to users with said block 20 attached to said waveguide 100, and a user can fill the hollow housing 21 with the OST by temporarily removing the removable plug 24.

[0113] In the embodiments considered here, the electromagnetic wave (EMW) exposure device is based on a waveguide. A waveguide makes it possible to control the field level sent to the resonator and therefore to the OST. Furthermore, being closed, it prevents the experimental personnel from being exposed to the emitted EMWs, which, it should be noted, can be of a relatively high level, such that they could have undesirable and potentially harmful effects on their bodies.

[0114] With reference to [Fig. 10], the guided-wave structure of the exposure device comprises a waveguide segment 100, for example, a waveguide made of conductive material (e.g., a metallic material such as copper), with a rectangular cross-section defined by a longer side and a shorter side. One end of the waveguide segment is intended to be connected to an electromagnetic wave source (not shown in [Fig. 10], but arranged in the same way as the source 12 visible on the waveguide 13 of [Fig. 2] described above). In practice, there is an aperture (iris) or a coupling mechanism (e.g., an antenna or a radiating wire) to supply energy from outside the waveguide 100. The electromagnetic wave source can be a radio frequency source or a microwave source.

[0115] The waveguide section 100 is intended to carry, in a guided manner, an electromagnetic wave delivered by the electromagnetic wave source. Propagation occurs along the longitudinal axis XX' of the waveguide section 100. During this propagation, the electric field lines extend between the two conductors of the transmission line formed by the lower and upper faces of the waveguide 100, substantially perpendicular to the longitudinal axis XX' of said waveguide, which is the propagation axis of the guided EMF.

[0116] In the example considered, since the resonator block 20 is parallelepiped in shape, its two smaller faces 22 and 23 can be brought into contact with the walls of the waveguide section 100, as shown in [Fig. 11], when it is arranged vertically in the waveguide 100. Thus, the conductive walls of the waveguide 100 impose boundary conditions for the resonator block 20 (zero electric field value on these walls, which are metallic). Preferably, the lower face 22 and the upper face 23 of the resonator 20 come into electrical contact with the inner face of the lower wall and the inner face of the upper wall, respectively, of the metallic waveguide 100. To this end, these faces 22 and 23 can be metallized to improve electrical coupling with the conductive walls of the waveguide, thereby effectively defining the boundary conditions by imposing a zero IEI value for the electromagnetic field at these faces 22 and 23. This metallization of the faces 22 and 23 of the resonator can be achieved, for example, by depositing a layer of tin. Those skilled in the art will appreciate that the same provisions can be made if the resonator block 20 is a bar with a cross-section other than a quadrilateral, for example, if its cross-section is circular (i.e., cylindrical), hexagonal, octagonal, etc.In all cases indeed, when the resonator block 20 is arranged vertically along its main extension direction, it presents a lower face and an upper face like the face 22 and the face 23, respectively, of the parallelepiped bar 20 shown in figures [Fig.9], [Fig.10], and [Fig.11]. .

[0117] The isometric perspective view of [Fig. 12] shows an embodiment of a waveguide section 100 provided with a lateral drawer 110 which is adapted for the arrangement of the resonator block 20 and its placement in the waveguide 100.

[0118] In this embodiment, the waveguide 100 has a lateral opening 120, that is, an opening in a vertical face of the rectangular waveguide extending in a plane XZ, in this case the right-hand face in the example shown (but it could also be the left-hand face). The axis of this opening 120 extends transversely along the Y direction. The waveguide section 100 of the electromagnetic system includes the slide 110, which can slide transversely within the waveguide 100 along the Y direction. This slide has the shape of a frame with geometry and dimensions corresponding overall to the cross-section of the waveguide 100, namely a rectangular frame in the illustrated example. This frame thus comprises a right-hand vertical post 111, a left-hand vertical post 112, a lower horizontal post 113, and an upper horizontal post 114.

[0119] More specifically, those skilled in the art will appreciate that, in the example shown, the external dimensions of this frame at the uprights 112, 113, and 114 are slightly larger than the internal dimensions of the cross-section of the waveguide segment 100, so that the drawer can fit into conforming grooves provided on the opposing internal faces of said cross-section of the waveguide. Regarding the right-hand vertical upright 111, its shape and dimensions allow it to close the lateral opening 120 when the drawer 110 is fully retracted into the waveguide segment 100. More particularly, this upright 111 fits into a conforming recess provided around the periphery of the opening 120 in the thickness of the right-hand vertical wall of the waveguide segment 100. on the outer side of said wall. The three aforementioned grooves, which cooperate with the three other uprights 112, 113, and 114 of the frame of the slide 110, are recessed grooves provided on the inner side of the corresponding walls of the waveguide section 100. Thus, when the slide 110 is fully retracted into the waveguide section 100, its four uprights fit snugly into the recess and grooves of the corresponding walls of said section. In this way, the geometric characteristics of the waveguide section 100 remain unchanged compared to embodiments without the lateral slide 110. Furthermore, the slide 110 is made of the same conductive material, i.e., the same metal, for example, as the waveguide section 100.Thus, the characteristics of the guided Fonde propagation in the waveguide are also unchanged, compared to embodiments without the side slider 110, when the slider is housed in place (i.e., is fully retracted laterally) in the waveguide section 100 via the side opening 120.

[0120] As further shown in [Fig. 12], the inner face of the lower upright 113 of the drawer 110 includes a non-through recess 113'. Furthermore, the lower portion of the resonator block 20 (which includes the hollow housing opening upwards into which the solution 12 containing the biological cells to be tested can be inserted, as symbolized in the figure by a downward-pointing vertical arrow) can be mounted on a base 113” made of the same conductive material, i.e., the same metal in this example, as the waveguide section 100. The shape and dimensions of this conductive base 113” correspond to those of the recess 113' provided in the lower upright 113 of the drawer 110. Thus, the assembly consisting of the resonator block (and in this case its lower portion) and its conductive base 113” can be placed in the recess 113', the resonator block 20 then extending vertically inside the frame of the sliding drawer 110.Those skilled in the art will appreciate that, for this purpose, the height along the Z direction of the resonator block is less than the distance along this direction which separates the internal faces opposite the lower upright 113 and the upper upright 114 of the drawer 110. In some embodiments, the resonator block 20 can be fixed to the conductive base 113”, for example by mechanical “force” fitting, by gluing, or by any other equivalent means.

[0121] Finally, and as also shown in [Fig. 12], the upper upright 114 of the drawer 110 includes an opening 113', i.e., a through-window, allowing the plug 24 to be inserted. The plug 24 has a cross-section in the horizontal plane XY of the same shape and dimensions as the cross-section of the lower portion of the resonator block 20. The lower part 24a of the plug 24 (made of the same dielectric material as the block 20) ​​can thus be inserted, as symbolized in [Fig. 13] by a downward-pointing vertical arrow, via the opening 114', to come against the upper edge of the lower portion of the resonator block 20. The upper part 24b of the plug (made of the same conductive material as the waveguide 100) is shaped like a cap attached to the lower part 24a by mechanical interlocking, bonding, or any other means. Those skilled in the art will understand that the plug 24 is inserted while the drawer is pulled outwards from the waveguide 100. The drawer 110 can be inserted into the section of waveguide 100 with the sample to be tested 12 inserted into the hollow housing of the resonator block 20 and after closing said resonator block by inserting the plug 24 as described above.

[0122] It goes without saying that, while the drawer 110 of the example shown in [Fig. 12] is a lateral drawer opening and closing by sliding along the transverse direction Y, it could just as well be a vertical drawer opening and closing by sliding along the vertical direction Z.

[0123] Diagram 301 of [Fig. 13] is comparable to diagrams 101 and 201 of [Fig.4] and [Fig.5], respectively, which were presented above for prior art systems, and illustrates the distribution of the incident field on the sample under test for a system according to the example of figures [Fig.9], [Fig.10] and [Fig.11] according to the invention, considering an OST which is the same biological solution as that in the Petri dish within the TEM cell ([Fig.4]) and the waveguide ([Fig.5]), respectively.

[0124] Diagram 301 of [Fig. 13] shows the distribution of IEI field values ​​(in V / m) represented on the horizontal axis between 550 and 650 V / m, at a number of points on the OST which are substantially regularly distributed (spatially) within this object. The number of points (expressed as a percentage of the total number of points recorded) for each field value among a determined number of discrete values ​​is indicated on the left-hand vertical axis.

[0125] Curve 302 shows, between the values ​​0 and 100% indicated on the right vertical axis, the summation of the points recorded in the interval [550 V / m - 650 V / m] of the field values ​​recorded.

[0126] For a power delivered by the OEM source of 1 W (i.e., as in the case of diagrams 101 and 201 of [Fig. 4] and [Fig. 5], respectively), the incident field in the waveguide 100 of the exposure device is estimated at 376 V / m. The average value of the field deposited on sample 12 is 601 V / m, or approximately 160% coupling (i.e., K=160%) with a standard deviation of 22.2 V / m, which is excellent and clearly represents overcoupling of the field in the object under test. It can also be noted that the homogeneity of the electric field in this OST is good, in that the Coefficient of Variation of the electric field is less than 10% (it is given by the ratio θ / Emoy). This evaluation is based on the simulations performed.

[0127] This therefore demonstrates the significant potential of the invention for a rigorous quantitative study, in vitro, of the impact of exposure to an electromagnetic field (EMF) on a sample of biological cells, given the low standard deviation of the IEI values ​​of the electric field, on the one hand, and the control (dosimetry) of the field level, on the other. Furthermore, it makes it possible to obtain field values ​​at equal power and comparable solution volume that are significantly higher than those obtained in the existing art, due to the exploitation of the resonance phenomenon.

[0128] A summary of the three cases that were taken as examples in the preceding description is given in the following table: [Tables 1] Petri dish + TEM cell Petri dish + waveguide Invention Power delivered by the source (W) 1 Incident electric field (V / m) 156 376 Mean field in the sample Emoy (V / m) 41 25 601 Standard deviation θ (V / m) 16.2 23.4 22.2 Coupling factor K 26% 6.6% 159.8% Ratio θ / Emoy 0.40 0.97 0.037

[0129] It should also be noted that the same result can be obtained with a resonator of different geometry, for example, a bar with a hexagonal cross-section or a bar with a circular cross-section (cylinder), instead of a bar with a rectangular cross-section, all other things being equal. The resonator can also be a coaxial structure. In all these alternative cases, the resonance equation given above by formula [Math.3] is modified, but the design principle remains identical and the resonance frequency / r remains determinable by a corresponding resonance equation, also well known to those skilled in the art.

[0130] In the preceding description of embodiments, an electromagnetic wave exposure device in the form of a rectangular section of waveguide 100 was considered. As described, the resonator block 20 is placed in this waveguide, which is adapted to guide the incident electromagnetic field generated by the EM source. In other embodiments, however, and instead of the waveguide 100 of [Fig. 10], the field exposure device electromagnetic may include a section of electromagnetic wave transmission line, for example of coaxial type, or of tri-plate line type, for example.

[0131] Another aspect of the invention relates to a method of using an electromagnetic wave applicator to apply an electromagnetic wave to a sample to be tested, in particular a sample of biological cells in the context of experiments intended to study the effect thresholds of cell exposure to an EMW of a given frequency.

[0132] The biological material sample 12 is inserted into the sample compartment 21 of the resonator block 20. Then the compartment is closed using the plug 24.

[0133] The resonator block, for example the resonator block 20 conforming to the example in [Fig. 9], is then placed in an exposure device for an electromagnetic field, such as a guided wave type exposure device. In the case of a system formed in the embodiment of [Fig. 12] in which the waveguide section 100 is equipped with a lateral drawer 110, this step consists of sliding the drawer 110 through the opening 120 so as to fully retract the drawer 110 into the waveguide 100. This results in a closed electromagnetic system with controlled boundary conditions. This can be a rectangular metallic waveguide as in the example considered above with reference to the example in Figures [Fig. 10] to [Fig. 12]. But we can also use the OEM applicator with a plane wave exposure device (according to the radiated approach which was discussed above with reference to [Fig.1]).

[0134] The electromagnetic wave source 11 of the exposure device is then controlled to illuminate the OST present in the resonator 20, under a given power, for a determined duration desired for the needs of the experiment to be carried out.

[0135] The resonator block 20 can then optionally be removed from the exposure device (by sliding the drawer 110 outwards from the waveguide 1000 in the case of the embodiment of [Fig. 12]), and in any case the cap 24 can be removed and the biological cell sample recovered after illumination, in order to submit them to the desired analyses in order to evaluate the effect on biological cells of exposure to the EM at the frequency of interest Fo.

[0136] With reference to [Fig. 14], the incident EM application device, which is again a portion of a waveguide, can comprise a plurality of parallel waveguide sections or arms 101 and 102, in which a plurality of electromagnetic field applicator devices 201 and 202 are respectively placed. This allows respective samples of biological material placed in the sample compartment of the resonator blocks of said applicators 201 and 202 to be simultaneously illuminated by the incident electromagnetic wave, independently and symmetrically, when it is emitted by the source 11 and propagates in the waveguide 100.

[0137] The present invention has been described and illustrated in the present detailed description and in the figures of the accompanying drawings, in possible embodiments that are particularly suitable for studying the effect of an electromagnetic wave of a given frequency on biological cells in liquid media. Its advantage lies in enabling illumination of the sample with good spatial homogeneity, allowing for experiments with a qualitative approach and, above all, the use of resonance modes to obtain coupling factors greater than one. This makes it possible to obtain high electric field levels in the cells under test and thus to study effects of interest on these cells. The dosimetric aspect, combined with good field homogeneity, proves useful for determining the thresholds at which such effects appear.

[0138] The present invention is not limited, however, to the embodiments shown. Other variations and embodiments can be deduced and implemented by a person skilled in the art upon reading this description and the accompanying drawings. Thus, for example, although better suited to guided exposures, for which there are no electromagnetic leaks, the illumination can also be achieved via a radiated field in free space, although this is not the preferred application. Furthermore, there is nothing to prevent the illuminated OST from comprising biological cells contained in solid elements (for example, seeds), without a liquid solution, even if the properties in terms of homogeneity and coupling factor would then be less effective.

[0139] Furthermore, other applications of the proposed applicator, different from illuminating biological cells in vitro to study the effect of an electromagnetic wave of a given frequency on biological cells, can be considered. For example, an applicator based on the proposed embodiments can heat a liquid using the dielectric loss mechanism and also determine its specific heat capacity (a characteristic used in determining the Specific Absorption Rate), after calibration with liquids of known properties. Finally, in the field of microwave resonators and filters, this makes it possible to shift the resonant frequency of the system by modifying the nature of the added liquid, and therefore its permittivity, which is directly related to the system's resonant frequency.

[0140] In the claims, the term "include" or "comprising" does not exclude other elements or other steps. The various features presented and / or claimed may be advantageously combined. Their presence in the The description or in different dependent claims does not exclude this possibility. Reference signs shall not be construed as limiting the scope of the invention. List of documents cited Patent documents

[0141] • FR2998813A1 • FR3021743A1 • FR3034521A1 Non-patent literature

[0142] • Book by J. Malmivuo and R. Plonsey: "Bio-Electromagnetism - Principles and applications of bioelectric and bio-magnetic fields", Oxford University Press, January 1995; • Sophie KOHLER: "Contribution to the development and multi-scale dosimetry of devices for the exposure of biological cells to high-intensity nanosecond and sub-nanosecond electrical pulses" - University of Limoges - Thesis no. 43-2013 - November 2013. • Maïak SOUEID: "Contribution to the development and characterization of applicators for bio-electromagnetic studies involving radiofrequency ounces and high-intensity nanosecond electrical pulses" - Thesis - University of Limoges - November 2016. • Aude SILVE: "New devices for the controlled application of nanosecond electrical pulses and for the detection of their effects on cells. New results and hypotheses on the parameters controlling the electro-permeabilization of biological cells" - Thesis - University Paris Sud XI - November 2011. • Report from the French Agency for Environmental and Occupational Health Safety (AFSSET) "Radiofrequencies", Annex 3.2: "Review of existing experimental exposure systems", October 2009. • D. KAJFEZ and P. GUILLON, “Dielectric Resonators”, Artech house, 1986. • V. VAN EEGHEM et al.: “The impact of millimeter waves on the enzymatic activity of succinate dehydrogenase" - Abstract from The Joint Annual Meeting of The Bio-Electromagnetics Society and the European Bio-Electromagnetics Association co-organized with the European COST EMF-MED Action BMI309, Ghent, Belgium - 2016.

Claims

1. Demands Applicator device for applying an incident electromagnetic wave at a predetermined frequency of interest, when said wave is generated by an exposure system comprising an electromagnetic wave source (11) disposed outside the applicator, to a sample of biological material disposed inside the applicator, said applicator comprising a resonator block (20) which • is made of at least one specific solid dielectric material, • has the general shape of a closed volume of determined dimensions, and • is a solid volume except in that it comprises at least one hollow compartment (21), referred to as the sample compartment, associated with a removable cap of the resonator block, said hollow compartment being adapted to receive, and preferably to be filled with, the sample of biological material, in which, moreover, • the resonator block (20) has within it, by design, a particular spatial distribution of the electric field when it is subjected to illumination by the incident electromagnetic wave, said distribution having discrete geometric configurations which define electromagnetic resonance modes each associated with a natural resonance frequency, which are determinable by simulation and adjustable on the basis of constitutive parameters of the applicator including the relative dielectric permittivity of the dielectric material, in combination with electromagnetic parameters of the biological material sample intended to be housed inside the sample housing, including the relative dielectric permittivity of said sample, • the constituent parameters of the resonator block (20) are defined such that a resonance frequency in said resonator block corresponds to the frequency of interest; and, • the shape of the sample housing (21) and its position in the volume of the resonator block (20) are defined so that said housing occupies an area in said volume of the resonator block which corresponds to a resonance peak of the electromagnetic field at the frequency of interest.

2. Electromagnetic wave applicator device according to claim 1, wherein the constitutive parameters of the applicator include, in addition to the relative dielectric permittivity (er) of the dielectric material(s) of which the resonator block is made, the dimensions and geometry of the resonator block, and the boundary conditions of said resonator block, and / or wherein the electromagnetic parameters of the biological material sample housed inside the sample housing (21) include, in addition to the relative permittivity (er), the conductivity (o) of said sample.

3. Electromagnetic wave applicator device according to claim 1 or claim 2, wherein the resonator block (20) has the form of a bar extending along a determined longitudinal direction (X-X'), and having a section in a plane orthogonal to said longitudinal direction which is of determined shape, constant along said longitudinal direction of the bar, and of determined dimensions (A,B) which are less than the dimension (C) of the bar along said longitudinal direction.

4. Electromagnetic wave applicator device according to claim 3, wherein the resonator block (20) has the shape of a bar with polygonal cross-section, for example with a rectangular, square or hexagonal cross-section.

5. Electromagnetic wave applicator device according to claim 3, wherein the resonator block (20) has the shape of a cylindrical bar.

6. Electromagnetic wave applicator device according to any one of claims 1 to 5, wherein at least the internal walls of the sample housing (21) which are intended to be in contact with the biological material sample, are made of a biocompatible material or are coated with a biocompatible material.

7. Electromagnetic wave applicator device according to any one of claims 1 to 6, wherein the housing The sample housing is adapted to receive the biological material sample in the form of a biological solution containing biological cells to be illuminated by the incident electromagnetic field, said housing being adapted to be closed by the removable cap of the resonator block after insertion, and preferably completely filled by said solution.

8. Electromagnetic wave applicator device according to any one of claims 1 to 7, wherein the dielectric material(s) of which the resonator block is made have a relative dielectric permittivity (er) greater than unity, and equal to the relative dielectric permittivity (er) of the biological sample to within ±50%.

9. Electromagnetic wave applicator device according to claim 8, wherein the dielectric material(s) of which the resonator block is made have a relative dielectric permittivity (er) greater than unity, and equal to the relative dielectric permittivity (er) of the biological sample to within + / -40%, preferably to within + / -30%, preferably still to within + / -25%, preferably to within + / -20%, and even more preferably to within + / -10%.

10. Electromagnetic wave applicator device according to any one of claims 1 to 9, wherein the dielectric material or materials of which the resonator block is made comprise ceramic.

11. Electromagnetic wave applicator device according to any one of claims 1 to 10, wherein the resonator block comprises a plurality of hollow housings for housing respective biological samples, the shape of said sample housings and their respective positions in the resonator block being chosen so that said sample housings each occupy an area (81) in the volume of said resonator block (20) which corresponds to a resonance peak of the electric field.

12. A system comprising at least one electromagnetic wave applicator device according to any one of claims 1 to 11, defined for applying an incident electromagnetic wave at a specified frequency of interest to a sample of biological material disposed within said device, the system further comprising an exposure device that is configured to generate the incident electromagnetic wave at the frequency of interest for which the electromagnetic wave applicator device is defined and which includes at least one guided electromagnetic wave device suitable for guiding the incident electromagnetic wave generated by the exposure system and in which the resonator block of the electromagnetic wave applicator device is placed.

13. System according to claim 12, wherein the guided electromagnetic wave device is a waveguide section in which the resonator block is placed.

14. System according to claim 13, wherein the waveguide section (100) includes a drawer (110) adapted for the arrangement of the resonator block and its placement in said waveguide section (100).

15. A system according to any one of claims 12 and 13, wherein the guided electromagnetic wave device comprises a plurality of parallel waveguide sections (101,102) in which a plurality of electromagnetic field applicator devices (201,202) are respectively placed so that respective samples of biological material placed in the sample housing of the respective resonator blocks of said electromagnetic field applicator devices are simultaneously illuminated by the incident electromagnetic wave as it propagates in the guided electromagnetic wave device.

16. System according to any one of claims 12 to 14, limited in that the electromagnetic field applicator device(s) comprise a bar-shaped resonator block as defined in claim 3, wherein one and / or the other of the longitudinal end faces of said resonator block are in contact with respective walls of the guided wave device.

17. A method for designing an electromagnetic wave applicator device for applying an incident electromagnetic wave at a specified frequency of interest, when said wave is generated by an exposure system comprising an electromagnetic wave source (11) disposed outside the applicator, to a sample of biological material disposed inside said applicator, comprising the following steps:

18. - choose a geometry and dimensions for a resonator block made of a specific dielectric material - determine by numerical simulation the eigenmodes of electromagnetic resonance in the resonator block defined by the chosen geometry and dimensions, giving different electric field distributions in said resonator block when it is exposed to incident electromagnetic field; - retain an electric field distribution corresponding to a specific resonance mode; - deduce the position and dimensions of a hollow housing (21) in the resonator block (20), accessible from outside said resonator block, which correspond to an area of ​​the volume of the resonator block in which the simulation indicates a peak of electromagnetic resonance, said hollow housing (21) being intended to receive the sample of biological material; - to tune the resonance frequency associated with the selected resonance mode to the frequency of interest, by modifying constituent parameters of the resonator block, including the value of the relative dielectric permittivity of the material from which said block is made, in combination with electromagnetic parameters of the biological material sample intended to be housed inside the sample housing, including the relative dielectric permittivity of said sample. A method for using an electromagnetic wave applicator device according to any one of claims 1 to 15, comprising the following steps: - insert a sample of biological material to be tested (12) into the sample compartment (21) of the resonator block (20), then close the compartment using a stopper (24), - place the resonator block in a device exposing it to an electromagnetic wave incident at a specific frequency of interest, - control an electromagnetic wave source (11) from the exposure device to illuminate the biological sample in the resonator (20), at a given power, for a specified duration - remove the cap (24) from the resonator block (20) and retrieve the biological cell sample after illumination, in order to submit it to analyses to evaluate the effect on the cells biological effects of exposure to electromagnetic waves at the frequency of interest.

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