Optical cartridge for analyzing a biological liquid and method of manufacturing an optical cartridge

Abstract

The present application relates to an optical cartridge for analyzing a biological liquid and to a method of manufacturing an optical cartridge. The analysis cartridge comprises an array of channels (4, 4') defining a plurality of analysis paths. Each channel is defined by at least two channel walls facing each other and defining a channel height. According to the invention, at least one wall of a channel has a step (M) so as to define, on either side of the step: - a first section (S1) of the channel, in which the wall has a first surface energy (E1) and a first elevation (e1) defining a first height (h1) of the channel; - a second section (S2) of the channel, in which the wall has a second surface energy (E2) and a second elevation (e2) defining a second height (h2) of the channel. The first height (h1) of the channel and the first surface energy (E1) of the structured wall are respectively greater than the second height (h2) of the channel and the second surface energy (E2).

Description

Technical Field

[0001] The technical field of this invention is bioanalysis, specifically for detecting the presence and / or concentration of analytes in biological fluid samples. More specifically, this invention relates to a cartridge comprising multiple analytical chambers for receiving biological fluids. This cartridge is preferably used in portable immunoassay devices of the "point of care" type, that is, allowing for on-site implementation and interpretation of tests to facilitate immediate clinical decisions at the patient's bedside rather than in a central laboratory. The cartridge can also be used for any other type of bioanalysis, such as molecular or cellular analysis. Background Technology

[0002] Document EP3447492 discloses a method for capturing and detecting molecules (commonly referred to as "analytes") in biological liquid samples. The principle of patterned capture and detection implemented by this method is also described in the article "Magnetophoretic induced convective capture of highly diffusive superparamagnetic nanoparticles" by Fratzl et al., Soft Matter, 14. 10. 1039 / C7SM02324C. According to this method, the sample is mixed with magnetic particles of nanoscale or more generally submicron scale, each magnetic particle being coupled to a capture element capable of binding to or quantifying the molecules to be detected. The molecule to be detected (analyte) can be an antigen, and the element can be an antibody, but the reverse configuration is also possible. Detection elements are also introduced into the sample, such as detection antibodies carrying photoluminescent labels (e.g., fluorescent labels). Thus, a complex consisting of the capture element, analyte, and detection element is formed in solution, and this complex is then immobilized on a scaffold comprising magnetic microsources arranged according to a defined spatial pattern. The pattern is defined by regions of strong and weak magnetic fields that induce large magnetic field gradients. Complexes entrained by magnetic particles are intended to accumulate on the scaffold at the region of maximum magnetic field norm. Photoluminescent (and particularly fluorescent) markers can make the defined spatial pattern apparent, indicating the presence of the analyte in solution. The average intensity (spatially) of this optical pattern is often referred to as the “specific signal.”

[0003] In most cases, and especially when the analyte is not present in the sample or when its amount in the sample is limited, unbound detection elements carrying photoluminescent labels remain suspended in the solution. They contribute to the formation of a relatively uniform light background. The average intensity (spatially) of this light background forms a signal known as the "supernatant signal." This light background, in addition to the unbound photoluminescent labels, is also composed of the light intensity emitted by all photoluminescent materials in the sample. Capture elements not bound to the analyte and detection element are also fixed to the support but do not carry labels; they do not contribute to the light pattern or light background.

[0004] The spatial arrangement of the magnetic field microsources within the scaffold plane and the light intensity of the pattern exposed by the photoluminescent markers allow for the detection and quantification of analytes in the sample without washing, i.e., without removing the liquid solution after immobilizing the complex on the scaffold surface, which is particularly advantageous. To enable this detection, the surfaces of the sample and the scaffold are irradiated to allow detection of the photoluminescent markers, and a digital image is acquired. This digital image thus possesses an inflowable spatial intensity (within the image plane) dependent on the strength of the magnetic field generated by the scaffold. Image processing identifies this spatial variation and determines the specific signal and supernatant signal; the specific signal / supernatant signal ratio allows for the conclusion of the presence of the analyte in the sample, and even allows for concentration estimation.

[0005] The simplicity of this method, particularly the elimination of a washing step, allows for its integration into autonomous immunoassay devices that are portable or “bedside” transportable, on-site, and require no pumps or valves, whereas this type of analysis is traditionally performed in a central laboratory.

[0006] To allow for the application of detection methods, the biological liquid is introduced into a cartridge comprising multiple analytical chambers, which is intended to be inserted into the analytical device. The multiple analytical chambers enable various analyses to be performed from the biological liquid sample, each analysis being performed independently on the sample held separately in each chamber.

[0007] The cartridge includes a liquid inlet opening, multiple outlets located downstream of the analytical chamber, and an array of channels for fluidly connecting the opening to the analytical chamber. The biological liquid sample in the inlet opening propagates through the channel array via capillary action to fill the chamber. However, it has been observed that the propagation of liquid in the channel array is not uniform from one channel to another. Flow may favor certain channels, potentially causing liquid to overflow from the cartridge. More generally, some chambers may fill more slowly or not at all. As a result of this phenomenon, analysis is delayed, and sometimes even impossible, to be performed in at least some analytical chambers of the cartridge.

[0008] Document US2004 / 028566 relates to a microfluidic device and aims to control the flow of fluid within the device by combining the displacement of the fluid with pressure applied by an external pump. More precisely, this document aims to enable the injection and control of multiple fluids into an analytical cartridge in multiple directions. One flow is achieved through capillary action, while other flows are forced by external pressure. Summary of the Invention

[0009] Therefore, the object of the present invention is to provide at least a partial solution to this problem. More precisely, the object of the present invention is to provide a cartridge for a portable immunoassay device of the "point of care" type, comprising multiple analytical chambers capable of being filled with biological fluid in a reliable, repeatable manner and over a controlled filling period via capillary action. Therefore, one object of the present invention is to better control the propagation of the biological fluid within the cartridge via capillary action. Advantageously, the immunoassay is magnetic, using magnetic particles to label the presence of analytes in the biological fluid sample.

[0010] The claimed features are used to control the propagation of biological fluids in the fluid array of the box, and in particular to facilitate the loading of biological fluids into the chamber of the array in a reliable, repeatable manner and during filling periods of controlled duration.

[0011] Other beneficial and non-limiting features of the invention, whether used alone or in any feasible combination of technologies: The wall facing the structured wall is planar and has a third surface energy greater than the first and second surface energies; The step has sides, and the surface energy of the side of the step is closer to the first energy than to the second energy. The main surfaces of the bracket and the top cover (7) are hydrophilic; The channel array includes an upstream channel for fluidly connecting the opening to the analysis chamber, and an exhaust channel for fluidly connecting the analysis chamber to the exhaust port; A step is provided in at least one discharge channel, and is intended to reduce the height of that channel in the direction of fluid flow; The analysis kit includes at least one culture chamber fluidly disposed upstream of the analysis chamber in the analysis path; The analytical and / or culture chambers include at least one reagent; The reagents include photoluminescent markers, and at least for the portion suspended above the analytical chamber, the cover is formed of a material that is transparent in the wavelength range of the photoluminescent markers; The top cover has at least a portion of an optically polished outer surface; The opening is surmounted by the reservoir, and the outlet is surmounted by the surrounding walls, the height of which is at least equal to the height of the reservoir.

[0012] The interlayer membrane is an adhesive membrane, advantageously a double-sided adhesive membrane; The magnetic layer includes a magnetically polarized region that defines the determined detection pattern.

[0013] According to another aspect, the present invention relates to a method for manufacturing the analytical cartridge, the method comprising providing a support and a top cover, and assembling the support to the top cover by making the respective main surfaces of the support and the top cover face each other, thereby forming facing walls defining a channel. Attached Figure Description

[0014] Other features and advantages of the present invention will be set forth in specific embodiments of the invention with reference to the accompanying drawings, wherein: [ Figure 1a ] [ Figure 1b ] Figure 1a and Figure 1b The box according to the invention is shown in perspective view and exploded view, respectively; [ Figure 1c ] Figure 1c An example of a fluid array of a cartridge according to the present invention is shown schematically; [ Figure 2a ] Figure 2a A longitudinal section of a channel with features that allow for slowing or accelerating the flow of biological fluids is shown; [ Figure 2b ] Figure 2b A longitudinal section is shown, including an exhaust channel with a height restriction in the form of steps. [ Figure 3 ] Figure 3 A cross-section of the box at the analysis chamber is shown; [ Figure 4 ] Figure 4 A top view schematically illustrates the magnetization generated by the magnetic layer integrated in the holder of the box, the magnetic field present in the analysis chamber, and the detection pattern defined by the norm of the magnetic field. [ Figure 5 ] Figure 5 An analytical apparatus for using a cartridge according to the invention is shown; [ Figure 6 ] Figure 6 Other views of the elements constituting the box according to the invention are shown schematically. Detailed Implementation

[0015] Figure 1a and Figure 1b A cartridge 1 is shown for receiving biological liquid samples, which may contain analytes whose concentration is to be detected or determined. The cartridge 1 includes a clamping end 1a that allows manipulation of it. The clamping end of the cartridge is labeled and positioned on the upper surface side of the cartridge, specifically allowing identification of the cartridge by means of an identification mark (e.g., a QR code) or for carrying any other type of information.

[0016] Box 1 also includes a microfluidic portion 1b. This portion extends along a main plane intended for horizontal positioning. Figure 1c As schematically shown, it includes an injection opening 2 that allows biological fluid to be introduced into the cartridge 1, for example, via a pipette. The opening 2 leads to channel arrays 4, 4' that extend in the main plane of the cartridge 1 and allow the biological fluid to flow and be distributed in the multiple analysis chambers 5 through the so-called "upstream" channels 4 of the channel arrays.

[0017] The channel array of box 1 also includes an exhaust channel 4' that fluidly connects the analysis chamber 5 to the exhaust port 3, which allows air to be discharged from the fluid array of box 1 as the biological liquid moves through the array.

[0018] The sample being analyzed is formed from the biological liquid filling chamber 5, and thus the illustrated cartridge 1 allows for multiple analyses of the biological liquid, which can be performed independently on the samples held separately in chamber 5. Opening 2, vent 3, and channel arrays 4 and 4' connecting opening 2 to vent 3 define multiple channels of the analysis cartridge 1.

[0019] exist Figure 1a and Figure 1b In the example shown, opening 2 is covered by a reservoir 2' protruding from the upper surface of box 1. The reservoir has sufficient capacity to hold a fluid array at least equal to that of box 1 (i.e., such as...). Figure 1c The volume of the bio-liquid in the channel array 4, analysis chamber 5, and discharge channel 4' shown is typically 5 mm. 3 up to 500mm 3 Between, and more precisely, at 20mm 3 Up to 100mm 3 between.

[0020] Similarly, the discharge ports 3 are each covered by surrounding walls to retain excess volume of biological liquid according to the principle of communicating vessels. Advantageously, the height of these walls is at least equal to the height of the reservoir 2' to prevent liquid from escaping from the container, which could lead to hygiene problems or even damage to the analytical device into which the container is intended to be inserted.

[0021] For example, box 1 may have a width and length between 2cm and 10cm, and a thickness between 4mm and 10mm. Each chamber 5 may typically have a thickness of 1mm. 3 up to 50mm 3 The volume between them is designed to accommodate the sample, advantageously within 5 mm. 3 Up to 25mm 3 between.

[0022] Box 1 is formed by a support 6 and a cover 7 that covers the support. The support 6 and the cover 7 are assembled together by placing their so-called "main" surfaces facing each other. The fluid array of box 1 is defined by recesses formed on the main surfaces of the support 6 and / or the cover 7 (i.e., on the surfaces of these two elements used for assembly with each other). Each channel of arrays 4, 4' is defined by two channel walls that face each other and define the channel height, and by two side walls that define the channel width. The walls are formed by the main surfaces of the support 6 and the cover 7 at their recesses. This also applies to the analytical chamber 5 of box 1 and any other element of the fluid array of box 1.

[0023] When the cartridge is used for the immunoassays described in the introduction of this application, at least for the portion suspended above the analytical chamber 5, the cover 7 is formed of a material transparent in the emission wavelength range of the photoluminescent marker. It can be, for example, a plastic material based on polycarbonate, cyclic olefin copolymers, or polystyrene. It can also be glass. The outer surface of the cover 7 is optically polished, at least on the right side of the analytical chamber 5. These features allow and facilitate optical analysis of biological liquid samples contained in the chamber 5, as will be explained later in this specification.

[0024] Therefore, as Figure 1c The fluid array shown extends in the main plane of the housing. It has millimeter-scale dimensions, meaning the widths of the channel arrays 4, 4' and the analysis chamber 5 are typically between 0.1 mm and 10 mm. The heights of these elements, i.e., their range in the direction perpendicular to the main plane of housing 1, are also millimeter-scale, between 0.1 mm and 10 mm. The biofluid propagates in this array via capillary action.

[0025] According to a key feature of Box 1, one of the walls defining the height of at least one channel, called a "structured wall," has steps, and the energy of the upstream and downstream surfaces of the steps is controlled to accelerate or decelerate the propagation of fluid in the channel, depending on the choice. Simply put, "surface energy" refers to the surface energy density between the surface under consideration and the biological fluid.

[0026] The cartridge 1 according to the invention utilizes this feature to facilitate the loading of biological liquid into the chamber 5 of the fluid array, and more generally to control the propagation of the liquid within the fluid array of the cartridge.

[0027] More precisely, and referring to Figure 2a At least one channel of box 1 includes a step M defining a first segment S1 of the channel, wherein a structured wall (formed herein by the main surface of support 6) has a first surface energy E1 and a first elevation e1 defining a first height h1 of the channel. Step M also defines a second segment S2 of the channel, wherein a structured wall has a second surface energy E2 and a second elevation e2 defining a second height h2 of the channel. The first height h1 of the channel and the first surface energy E1 of the structured wall are respectively greater than the second height h2 of the channel and the second surface energy E2 of the second segment S2.

[0028] Changes in the height of the channel, along with energy changes on the upstream and downstream surfaces, allow for influence on fluid flow within the channel.

[0029] Therefore, when the fluid encounters an "ascending" step, that is, when the fluid flows from the first segment S1 to the second segment S2 of the channel through capillary action, its flow slows down due to the height limitation of the step and the minimum surface energy of the second segment. Conversely, when the fluid encounters a "descending" step, that is, when the fluid flows from the second segment S2 to the first segment S1 of the channel through capillary action, its forward movement accelerates.

[0030] It's important to note that surface energy density is a positive quantity characterizing an interface, in this case, the interface between the structured wall surface and the biological fluid. It is well known to be determined by measuring the contact angle of a water droplet placed on a surface on which energy is being measured. A high contact angle greater than 90° indicates a weak surface energy density, and this surface is called a hydrophobic surface. Conversely, a contact angle less than 90° indicates a strong surface energy density, and this surface is called a hydrophilic surface. A relatively more hydrophobic surface will be intended to slow the fluid's advance through capillary action, compared to the advance of the fluid on a relatively more hydrophilic surface.

[0031] To explain these principles, Figure 2bA “rising” step is shown in the discharge channel 4' of the cartridge, i.e., the channel between the analysis chamber 5 and the discharge port 3. Fluid thus advances towards the discharge port 3 in this channel. The step formed on the support 6 defines the upstream portion of the discharge channel (in the general terminology of the invention, the first section of the channel), which has a greater surface energy on the support 6 than the downstream portion of the channel 4' (the second section on the discharge port 3 side). Advantageously, such a step can be provided in each discharge channel 4' of the cartridge 1, in each analysis path of the cartridge.

[0032] These features of Box 1 have the effect of slowing the advance of the biological liquid when it encounters a step after it has advanced along the channel array 4 and through the analysis chamber 5 via capillary action. When this slowing effect occurs in the discharge channel 4', which fills faster than other channels, it forces liquid to flow into the other channels of the array (i.e., supplying the other analysis chambers 5 of Box 1) and balances the advance of the liquid in each of these channels. Therefore, these features of the channels or discharge channels 4' ensure that the analysis chamber 5 is filled with biological liquid in a reliable, repeatable manner and within a controlled filling period. It can be predicted that the "rising" step may be located in the upstream channel 4 or multiple upstream channels 4, rather than in the discharge channel 4'. Some of these steps may be located in the upstream channels 4, while others may be located in the downstream discharge channel 4'.

[0033] More generally, the cartridge 1 according to the invention may have one or more ascending and / or descending steps in each analysis path. Thus, a “descending” step may be provided in the upstream channel 4 of the chamber 5 supplying the analysis path, and an “ascending” step may be provided in the discharge channel 4' connecting the chamber 5 to the discharge port 3 in the same analysis path or in another analysis path.

[0034] Favorably, and as Figure 2a and Figure 2b As shown, the step causes a sudden change in height within the passageway, with its side forming a 90° angle with the wall. More generally, however, this side can form an angle between 60° and 90°. For example... Figure 2a and Figure 2b As shown, the step can be formed on the support 6, but it is conceivable that it can be formed entirely or partially on the opposite wall of the upper cover 7. It can have a height between 0.1 mm and 0.5 mm to effectively decelerate or accelerate the forward movement of the liquid. Advantageously, the side of the step has a surface energy closer to the first surface energy E1 than to the second surface energy E2.

[0035] The main surfaces of the support 6 and the cover 7 (and therefore the walls of the channels 4, 4' and chamber 5 of the box 1) are preferably selected or treated to be hydrophilic. This property generally allows for the advancement of fluid in the fluid array via capillary action. Surprisingly, the difference between the first surface energy E1 of the first segment S1 and the second surface energy E2 of the second segment S2 does not need to be very large. In particular, the second surface energy E2 on the “higher” segment of the step does not need to be hydrophobic to retain the advancement of the liquid. Measured by contact angle, the difference between these two energies E1, E2 can be 5° or greater, or 10° or greater, and these energies impart hydrophilicity to the surface, i.e., the contact angle remains less than 90°. For example, the first segment S1 can have a contact angle characterizing the first energy E1, which is between 50° and 80°, while the second segment S2 has a contact angle between 65° and 89° (while maintaining a difference of at least 5°). In the remainder of this specification, a detailed example of the fabrication of the scaffold 6 will be given, which allows control of the first surface energy E1 and the second surface energy E2, as just presented.

[0036] Advantageously, the wall opposite the structured wall has a third surface energy greater than the first surface energy E1 and the second surface energy E2. This energy can be between 15° and 65° (advantageously maintaining a greater than both the first and second surface energies), and advantageously less than 50°, or even close to or less than 35°. This energy results in the formation of a more hydrophilic surface on the opposing wall than on the structured wall, and generally allows liquid to move into the fluid array via capillary action. The variable energy of the structured walls in different segments S1, S2 of the channel allows for the regulation of this forward movement. When the wall having this third energy E3 is provided by the main surface of the cover 7, the wall can be treated with, for example, a poloxamer-based surfactant intended to increase the hydrophilicity of the surface.

[0037] Of course, cartridges with more complex fluid arrays than the example shown can be provided. Thus, the analytical pathways of the cartridge can include chambers other than analytical chamber 5, such as one or more culture chambers located upstream of analytical chamber 5. These culture chambers can include different reagents, with which the fluid is mixed before being delivered to analytical chamber 5. Therefore, the channel arrays 4, 4' can also be more complex than those shown in the figure, and in each analytical pathway extend from opening 2 to outlet 3, fluidly connecting different chambers according to any possible configuration.

[0038] Reference Figure 1b , Figure 6 and Figure 3 , Figure 3A cross-section of cartridge 1 at analysis chamber 5 is shown, more precisely illustrating an embodiment of cartridge 1 according to the invention. Cartridge 1 of this embodiment allows for the application of magnetic immunoassay, thereby using magnetic particles to label the presence of analytes in biological liquid samples. This embodiment also allows for easy control of surface energy on different upstream and downstream sections of multiple steps (or a single step) present in the fluid array of the cartridge.

[0039] The support 6 is here composed of a rigid substrate 6a including a magnetic layer or region 6b. The substrate 6a may be formed of a plastic material. The magnetic layer / region 6b may be disposed on the substrate 6a, or integrated into the substrate at least at the analysis chamber 5 of the fluid array. It does not necessarily cover the entire surface of the substrate 6a.

[0040] The magnetic layer 6b typically consists of a magnetic composite material (e.g., ferrite) randomly distributed within the polymer or oriented along a predetermined axis. This magnetic layer can be similar to a conventional recording magnetic tape.

[0041] The substrate 6a may also include a non-magnetic film 6c (or multiple such films) covering the magnetic layer 6b, and more generally covering the substrate 6a. The non-magnetic film 6c is optional and its purpose is to keep the magnetic layer 6b away from the bottom of the analytical chamber 5 when the cartridge 1 is formed by assembling the support 6 to the top cover 7. To avoid interfering with measurements, the non-magnetic film 6a has low autofluorescence. For clarity, "non-magnetic" refers to a material with very low magnetic susceptibility, such as paramagnetic or diamagnetic materials. The non-magnetic film 6c may, for example, be formed from a plastic material such as polypropylene.

[0042] In each case, substrate 6a has an exposed surface A1, which can be formed by substrate 6a itself or by a non-magnetic layer 6c (when present). This exposed surface A1 serves to form the first segment S1 of the structured wall of the channel for the fluid array of the cell. The exposed surface A1 is designed or has been treated to exhibit a hydrophilic first surface energy E1. This first surface energy E1 can be caused by the selection of the material forming the exposed surface, or by its texturing or treatment (e.g., plasma or by surfactants), designed to make the surface particularly hydrophilic. As previously described, this first surface energy E1 is characterized by a contact angle between 50° and 80°.

[0043] In addition to the substrate 6a, the support 6 also includes an interlayer film 6d disposed on the exposed surface A1 of the substrate 6a. Figure 1bThe interlayer membrane 6d has cuts that correspond to the upstream channel array 4 and the analytical chamber 5, and advantageously to the opening 2. Typically, the interlayer membrane 6d has cuts D designed to define a portion of the fluid array of the cartridge. When the interlayer membrane 6d is surface-mounted onto the substrate 6a to form a support 6, the support thus has recesses that reproduce the cut pattern D of the membrane 6d. These recesses, combined with complementary recesses formed in the upper cover 7, constitute any other element of the fluid array of the channel arrays 4, 4', the analytical chamber 5, and the cartridge 1. It should be noted that... Figure 1b The cut pattern D of the interlaminar membrane 6d does not use the imprint of the discharge channel 4', and therefore it is formed only by recesses formed in the upper cover 7 rather than in the support 6. This arrangement results in the formation of a rising step at the entrance of the discharge channel 4', as described in the previous example. More generally, the cut pattern D of the interlaminar membrane 6d corresponds only to a portion of the fluid array of the cartridge 1 in order to form the steps of that array.

[0044] The exposed surface A2 of the interlayer film 6d (including the surface opposite to the surface in contact with the substrate 6a) serves as the second segment S2 of the structured wall forming the channel of the fluid array of the housing 1. It is designed or processed to have a second surface energy E2, which is also hydrophilic but less than the first surface energy E1. This second surface energy E2 can be generated by the selection of the material forming the interlayer film or its surface, or by its texturing or specific treatment. As previously described, this second surface energy E2 is characterized by a contact angle between 65° and 89° (while being greater than the first surface energy E1).

[0045] In this configuration, when the interlayer film 6d with the cut D has been assembled onto the substrate 6a, the main surface of the support 6 is then formed by the exposed surface A2 of the interlayer film 6d with the second surface energy E2 and the exposed surface A1 of the substrate 6c at the cut D of the interlayer film 6d with the second surface energy E2.

[0046] Advantageously, the interlayer film 6d is an adhesive film, which also allows the top cover 7 and the support 6 to be tightly assembled and held together at their contact surfaces (i.e., around the recess). It can be a double-sided adhesive film, thus ensuring that it is simultaneously assembled to the substrate 6a and the top cover 7. Such films are known to be composed of strips (e.g., plastic strips coated with adhesive material on both sides). Due to its properties, this adhesive material can naturally have a surface energy lower than that of the exposed surface of the substrate 6a, and thus constitutes a second surface energy.

[0047] Regardless of whether it is formed by a double-sided adhesive interlayer film, the box 1 is assembled by providing a substrate 6a having a magnetic region 6b, forming a support 6 by placing an interlayer film 6d on the substrate 6a, and then assembling the assembly to the top cover 7. This assembly is achieved by aligning complementary recesses on the main surface of the cover 7 with recesses defined by the cut pattern D of the interlayer film 6d of the support 6. The fluid array of the box 1 is defined in this way.

[0048] Of course, other membranes with different slit patterns can be formed on the interlayer membrane 6d to create multiple continuous rising or falling steps in the channels of the fluid array. In this case, care should be taken to maintain the relationship that at each step, the surface energy of the channel segment with the maximum height is greater than the surface energy of the channel segment with the minimum height.

[0049] Returning to the description of the magnetic characteristics of the box, magnetic layer 6b includes two different directions ( Figure 3 A series of regions polarized (opposite to the middle). For example... Figure 4 As shown, it shows a top view of the magnetic layer 6b. In the example shown, the magnetically polarized region extends in a straight line along the main direction P.

[0050] At the interface between two different polarization regions, a region of relatively high magnetic intensity is generated, referred to as the attraction region in the remainder of this specification. Therefore, the attraction region is arranged in the form of multiple lines Za oriented along the principal direction P. The specific arrangement of these lines defines the detection pattern.

[0051] It should be understood that the arrangement of lines in the example is only a special case of the detection pattern. Box 1 is more generally provided with a magnetically polarized region and a clearly defined detection pattern, but its configuration can be freely chosen.

[0052] Figure 4 The field Bc generated by the magnetic layer 6b and its norm are also shown. As will be disclosed in the remainder of this disclosure, it may be useful to add an additional external field Bext to the field generated by layer 6b. Figure 4 The external field Bext is shown in combination with the field Bc generated by the layer, and the norm of the combined field is also shown. It is observed that applying the external magnetic field Bext can result in the elimination of a specific attraction region Za that occurs when only the magnetic field provided by the magnetic layer 6b is present. However, in each case, these attraction regions are arranged along a line Za oriented along the principal direction P, or more generally, in a detection pattern perfectly determined according to their characteristics.

[0053] In the case of chamber 5 having the above-mentioned dimensions, it is conceivable to form a detection pattern comprising 2 to 50 lines, the thickness of which is between 1 micrometer and 150 micrometers (advantageously between 5 micrometers and 30 micrometers), and the spacing between them is between 5 micrometers and 300 micrometers, advantageously between 25 micrometers and 150 micrometers.

[0054] return Figure 3 The description describes the preparation of a cartridge 1 to place a controlled amount of nanoscale magnetic particles 9, typically between 25 nm and 500 nm, preferably between 100 nm and 260 nm, in each chamber 5. These particles are typically in the form of beads exhibiting superparamagnetic properties and biocompatibility. Specifically, they may be coated with a polymer (polystyrene type) that has been surface-treated to enable them to be functionalized with Ac or Ag type proteins. The magnetic particles are coupled with trapping elements capable of binding to analytes. The controlled amount of particles is such that, once filled with biological fluid, their concentration in the chamber volume is within 10... 6 Up to 10 11 Between particles / ml. Controlled amounts of nanoparticles and trapping elements are arranged here in the form of dry clusters 9 placed on a support 6 in chamber 5.

[0055] Similarly, each chamber 5 also contains clusters 10 of detection elements. These detection elements are also capable of binding analytes and carrying photoluminescent labels, such as fluorescent labels.

[0056] Clusters 9 and 10 of the capture and detection elements can also be found in Figure 1b As seen in the image. Before the top cover 7 is placed on the support 6, it can be placed on the support 6 at a position corresponding to the location of the analysis chamber 5. These positions can be identified using the recesses of the support 6, which specifically define the cavity of the chamber 5.

[0057] When the biological fluid to be analyzed is introduced into cartridge 1, it flows into channel array 4 to fill analytical chamber 5 and propagates in discharge channel 4'. At least one step (and the surface energy change upstream and downstream of this step) exists in these channels 4, 4' to ensure that all analytical chambers 5 are properly filled within a defined time, as described in the preceding paragraph. Detection element 10 and trapping element associated with magnetic particles 9 are resuspended in the sample in each chamber 5 for mixing therein. During the subsequent reaction time, and while the analyte is present in the sample, a complex is formed consisting of the trapping element, magnetic particles, analyte, and detection element. These complexes are anchored to the support 6 of each chamber 5, preferably accumulating at the maximum magnetic field strength, and thus arranged according to the detection pattern defined by magnetic layer 6b. Excess detection element remains suspended in the sample.

[0058] It can be specified that each chamber 5 of cartridge 1 is configured to receive different types of capture and detection elements for multiple analyses of the biological fluid introduced into cartridge 1. It can also be specified that the detection pattern encoded by a portion of the magnetic layer 6b disposed in chamber 5 differs from one chamber to another.

[0059] In each case, the presence of the analyte in the sample retained in the analysis chamber 5 results in the formation of a detection pattern defined by the magnetic layer 6b.

[0060] To fully control the phenomena occurring in the analytical chamber during the reaction period, and to detect the presence and intensity of the detection pattern at the end of the period, it is advantageous to place box 1 in... Figure 5 The analytical apparatus E is schematically shown in the middle or on top.

[0061] The apparatus E includes elements for receiving the cartridge 1 to position it as precisely as possible at the analytical location. At this location, at least one chamber 5 of the cartridge 1 is arranged in the field of an imaging device 11, such as an image sensor. The chamber 5 is also positioned in the illumination field of a light source 12 (e.g., a diode-based light source). Optical elements 13, such as separators, filters, and objectives, can also be arranged in the optical path between the light source 12, the chamber 5, and the imaging device 11 to improve image quality and, in particular, to select appropriate magnification and depth of field. With this arrangement, digital images of the sample and the support 6 of the chamber 5 can be acquired to display the light intensity produced by the fluorescent marker on the image. The cartridge 1 is positioned in the analytical apparatus such that a transparent top cover 7, at least perpendicular to the chamber 5, is located in the optical path to enable image imaging.

[0062] During operation, the photoluminescent markers in the solution of the sample or those fixed on the support 6 of the illumination chamber 5 are activated by the light source 12 and made visible in the image plane of the imaging device 11. Therefore, a digital image of the distribution of the photoluminescent markers in the support plane can be acquired.

[0063] continue Figure 3 The analytical apparatus E may also include a mechanical actuator 14, such as a piezoelectric actuator, capable of contacting the holder 6 of the cartridge to apply a vibrational force thereto. The actuator 14 may be activated after the cartridge is introduced into or onto the apparatus E to allow the trapping elements, magnetic particles, and detection elements 9, 10 to be effectively resuspended in the sample.

[0064] Finally, the device E may include a magnetic field source 15, such as an electromagnet, which can be activated to enhance the magnetic field generated by the magnetic layer 6b. The magnetic field generated by the source 15 can be between 1 mT and 400 mT in chamber 5, but in each case, it must be kept below the value of the coercive magnetic field of the magnetic layer 6b in order to maintain its magnetization and the detection pattern defined by that magnetization. The field generated by the source 15 is preferably oriented perpendicular to the surface of the support 6 in order to add to the field generated by the magnetic layer 6b, thereby increasing the magnetic field strength in the attraction region Za and enhancing the detection pattern. As previously mentioned, the presence of this field may lead to a change in the line arrangement Za of the attraction region, or more generally, a redefinition of the detection pattern. The field generated by the source 15 can be continuous or pulsed, in which case it typically has a pulse duration greater than 1 ms. The field generated by the source 15 also enables the superparamagnetic particles of the sample to be magnetized. Therefore, this promotes the migration of these particles and complexes (when they are present) to the surface of the support 6 to fix them.

[0065] To operate the analysis device E and perform digital processing on the acquired images, the device E also includes a computing device 16. This can be a microcontroller, microprocessor, or FPGA circuitry. In addition to the computing device itself, the computing device 16 includes a memory component for storing data and a computer program for operating the device E. The computing device 16 may also include an interface component (USB interface type) for exchanging data or connecting the analysis device E to maintenance equipment. The interface component may also include a screen and control buttons to allow the operator to use the device E. The computing device 16 is connected, for example, via an internal bus to the imaging device 11, the light source 12, the mechanical actuator 14, and the magnetic field source 15 to coordinate their actions and / or collect generated data, such as digital images provided by the imaging device 11.

[0066] Therefore, once the operator places the box 5 in or on the analysis device E, and the receiving element holds the box 5 in the analysis position, the computing device 16 can perform the following sequence of actions, for example, after actuating the device E via a control button: - The mechanical actuator 14 is activated to apply a vibrational force to the cartridge 1 and to cause or facilitate the resuspension of the trapping elements, magnetic particles, and detection elements in the sample. This activation marks the start of the trapping period; - Activate magnetic field source 15 to facilitate the fixation of magnetic particles, whether they are combined with or not with the analyte, onto the support 6 of chamber 5. - The light source 12 is turned on, and then the imaging device 11 is activated at the end of the capture period to continue acquiring images of the sample and the scaffold. The images can be transferred from the imaging device 11 to the storage component of the computing device 16 so that processing operations can be performed therein.

[0067] - Process images to determine detection signals that represent the presence or concentration of analytes in the sample.

[0068] - Process the detection signal to provide information indicating the presence, absence, and / or concentration (e.g., per milliliter) of the analyte in the sample.

[0069] Of course, the present invention is not limited to the described embodiments, and variations thereof may be provided without departing from the scope of the invention as defined by the claims.

Claims

1. An optical cartridge for analyzing biological fluids, the optical cartridge including a liquid inlet opening into a channel array in which liquid permeates via capillary action, the channel array defining a plurality of analytical paths, each analytical path including an analytical chamber, for each analytical path, the channel array including an upstream channel for fluidly connecting the opening to the analytical chamber and an outlet channel for fluidly connecting the analytical chamber to an outlet, the optical cartridge being formed of a support having a main surface and a top cover at least partially formed of a transparent material and also having a main surface, the top cover and the support being assembled to each other via their main surfaces, at least a portion of the channel array being formed by complementary recesses partially formed on the main surface of the support and partially formed on the main surface of the top cover, then each channel being defined by a channel wall, referred to as a "structured wall," provided by the support and an opposing channel wall provided by the top cover, the two walls facing each other and defining a channel height, the structured wall having a step defining, on either side of the step: - A first segment of the channel, in which the structured wall has a first surface energy and a first elevation defining a first height of the channel; - The second segment of the channel, wherein The structured wall has a second surface energy and a second elevation that defines a second height of the channel; The first height of the channel and the first surface energy of the structured wall are respectively greater than the second height and the second surface energy of the channel; The support also includes: - A rigid substrate having the first surface energy, the rigid substrate being bonded to a magnetic layer disposed under a non-magnetic film, the non-magnetic film covering the magnetic layer; - An interlayer film having the second surface energy, the interlayer film disposed on the rigid substrate, the interlayer film defining a portion of the fluid array of the optical cell and having a notch pattern for forming recesses in the support and not covering the exposed surface of the rigid substrate. The main surface of the support is composed of the exposed surface of the interlayer film having the second surface energy and the exposed surface of the rigid substrate having the first surface energy. In this embodiment, at least one discharge channel is formed solely by a recess in the top cover and not by the cut pattern of the interlayer membrane, such that a rising step is formed at the entrance of the at least one discharge channel, and is intended to reduce the height of the at least one discharge channel in the direction of fluid flow.

2. The optical box according to claim 1, wherein, The wall facing the structured wall is planar and has a third surface energy greater than the first surface energy and the second surface energy.

3. The optical box according to claim 1 or 2, wherein, The step has sides, and the surface energy of the sides of the step is closer to the first energy than to the second energy.

4. The optical box according to claim 1 or 2, wherein, The main surface of the bracket and the main surface of the top cover are hydrophilic.

5. The optical box according to claim 1, wherein the optical box comprises at least one culture chamber fluidly disposed upstream of the analysis chamber in the analysis path.

6. The optical box according to claim 5, wherein, The analytical chamber and / or the at least one culture chamber include at least one reagent.

7. The optical box according to claim 6, wherein, The at least one reagent includes a photoluminescent marker, and at least for the portion suspended above the analytical chamber, the cover is formed of a material that is transparent within the wavelength range of the photoluminescent marker.

8. The optical box according to any one of claims 5 to 7, wherein, The top cover has at least a portion of an optically polished outer surface.

9. The optical box according to claim 1 or 2, wherein, The opening is covered by a storage container, and the discharge outlet is covered by a peripheral wall, the height of which is at least equal to the height of the storage container.

10. The optical box according to claim 1 or 2, wherein, The interlayer membrane is an adhesive membrane.

11. The optical box according to claim 10, wherein, The interlayer membrane is a double-sided adhesive membrane.

12. The optical box according to claim 1 or 2, wherein, The magnetic layer includes a magnetically polarized region that defines a specific detection pattern.

13. A method for manufacturing an optical box according to any one of claims 1 to 12, the method comprising: The bracket and the top cover are provided, and the bracket is assembled to the top cover by making the main surfaces of the bracket and the top cover face each other, thereby forming an facing wall defining the channel.

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