PCR detection chip, associated test apparatus, and analysis system for implementation

Abstract

Disclosed is a system for testing a microfluidic sample chip for testing biological samples, in particular for PCR and / or fluorescence analysis, having the form of a hollow block comprising at least one chamber which is delimited by an upper wall and a lower wall and into which a sample to be tested can be introduced and which is associated with thermalization means and with fluorescence measurement means. According to the invention, the system further comprises a tab, preferably opaque and / or rigid or semirigid, positioned in the continuation of, and preferably in the same plane as, the walls.

Description

DESCRIPTION TITLE: PCR Detection Chip, Associated Testing Device and Implementation Analysis System

[0001] The invention relates to a microfluidic sample chip for the analysis of biological samples by PCR and / or fluorescence using a compatible testing device. It relates more particularly to the field of rapid point-of-care diagnostic tests or self-tests, the speed of the test and the ease of use of the device being sufficient for an untrained user, particularly the general public, to use it and obtain a rapid result simply by reading the instructions for use.

[0002] To perform a rapid, real-time PCR (Polymerase Chain Reaction), it is necessary to change the temperature of a mixture containing the sample and its reagent as quickly as possible, preferably while measuring its fluorescence (hereinafter referred to as rapid sample "cycling"). A method and device of this type are described, for example, in patent application WO2011 / 138748.

[0003] The PCR reaction is generally performed in a disposable container because, following the reaction, the large-scale amplification of the target DNA contaminates the surface of the container with the target DNA, preventing its reuse. PCR reaction containers are therefore considered consumable.

[0004] These PCR consumables must be easy, straightforward, and quick to handle and insert into the sample testing device to facilitate use, particularly in contexts where operational efficiency is critical. Furthermore, the PCR consumables must be completely leak-proof during and after the PCR reaction to prevent contamination of the environment and subsequent tests by the reaction products.

[0005] WO 2018 / 1 14625 describes in particular a microfluidic sample chip containing a chamber that can be sealed with a sealing film, preferably flat and / or of low thickness formed between a face that is at least partially transparent and a heat-conducting face, for example made of aluminum, the transparent face allowing the PCR reaction to be followed using an optical means, for example a fluorescent probe imaging technique, while allowing a very rapid variation of the sample temperature thanks to its heat-conducting face.This chip can be used for example in combination with a microfluidic temperature control system as described in this application WO2018 / 114625, allowing the temperature of the sample to be changed very quickly between its two extreme values ​​typical of PCR technology, in a time on the order of a second while being simple to introduce into the device and ensuring perfect sealing of the consumable during and after reaction.

[0006] However, known sample chips generally require the use of a micropipette to introduce the sample to be analyzed into the chip. Using a micropipette requires a trained operator and should not be entrusted to an untrained user, as this could compromise the test result. The openings are usually small to ensure a tight seal between the micropipette tip and the chip when injecting the sample. The drop-based action simplifies chip use but necessitates that at least one of the two openings be larger. Increasing the diameter of the opening makes sealing more difficult by reducing the adhesive area, particularly between the edge of the sealing adhesive and the opening.

[0007] W02020123957 describes a PCR diagnostic cartridge integrating sample pretreatment and the PCR chamber. In this device, the sample is transferred into a receptacle in solid form (e.g., a swab) or using a transfer tool that requires no special technical skill and can be performed by a non-specialist. However, these cartridges have the disadvantage of being bulky, expensive to produce, and have a speed limited reaction: they are therefore not suitable for decentralized use, at the patient's bedside or for self-testing.

[0008] The microfluidic sample chip according to the invention for testing biological samples, in particular for PCR analysis, having the form of a block (10), preferably parallelepiped-shaped, comprising at least one hollow chamber (4) for receiving a sample, delimited in particular by a first wall (or lower wall) (3) made of a heat-conducting material with a thermal conductivity greater than 0.1 wm-1.K-1, preferably greater than 15 wm-1.K-1, preferably metallic, and a second wall (or upper wall) (2), parallel to the first, made at least partially of a transparent and preferably hydrophilic material, the block (10) having an inlet orifice (50) allowing the introduction of the sample into the chamber (4) and an outlet orifice (51) allowing the evacuation of the atmosphere present in the chamber during its introduction, the inlet orifice (50) extending into a cavity (52) for receiving the sample,is characterized in that the inlet orifice has a surface area greater than 4 mm2, in that the cavity (52) is combined with a first capillary bridge allowing the sample to penetrate by capillary action into the chamber (4) and preventing it from remaining in the cavity (52), and in that the chip also comprises a sealing film (6), configured to be in contact with the block (10), this sealing film (6) having a surface area sufficient to cover the inlet (50) and outlet (51) orifices simultaneously, said sealing film being configured to adhere to said block (10) and to close the inlet (50) and outlet (51) orifices of the block in a hermetic manner after the chamber (4) has been filled with the sample.

[0009] Preferably the chip will also include a tab (9) at least partially opaque and rigid or semi-rigid, preferably arranged in the extension and preferably in the same plane as the two walls (2,3), the block (10) being preferably integral with the tab.

[0010] Preferably, the chip will include a protective film (60) disposed on the second wall (2), having two openings (81) whose sides are adjacent to the perimeter of the orifices (50, 51) of the chamber (4) allowing protect the areas in contact with the sealing film (6) from particulate or liquid contamination which may limit the adhesion capacity of the sealing film (6).

[0011] Preferably, the distance between the lower and upper walls of the chamber will also be less than 1 mm, preferably less than 500 pm.

[0012] Preferably, the first capillary bridge will be characterized according to a first alternative by at least one projection in the block positioned on the rim of the cavity (52) facing the first wall (3), this projection having at least one vertical surface contiguous to at least one of the flanks of the inlet orifice (50) on the one hand, and to the first wall (3) on the other hand, or according to a second alternative by an element of hydrophilic porous material in contact on one of its faces with at least one of the flanks of the inlet orifice (50) on the one hand and on another of its faces with the first wall (3) on the other hand.

[0013] Preferably, the chip will be characterized in that the cavity (52) has a shape chosen between that of a funnel (52) such as a truncated cone, a truncated pyramid or a trapezoidal prism having an angle at the apex between 20 and 90 degrees, preferably between 20 and 50 degrees, and that of a straight hole with an elliptical or polygonal cross-section, passing through the second wall (2).

[0014] According to a first variant, the first capillary bridge may include at least one projection located on the upper face of the chamber (4), the size of the height of the chamber (4) and having a vertical surface located in the extension of a part of the sides of the inlet orifice (50).

[0015] According to another variant, a second capillary bridge located on the upper part of the second wall (2), positioned on one side of the cavity (52), may include at least one projection the depth of the cavity (52), taking the form of a fin (53) or a cross-shaped element, perpendicular to the upper surface of the chamber (4), the longitudinal direction of which points towards the center of the inlet orifice (50), the fin extending into the cavity (52) in the direction from its center over a distance of between 0.1 mm and the edge-center distance of the inlet orifice. According to these two variants, the sample, upon reaching the bottom of the cavity, becomes wetted by the protrusions of the first wall, allowing it to enter the chamber by capillary action.

[0016] Alternatively, the first capillary bridge can be a hydrophilic fiber element, such as a sheet of cellulose paper or a sheet of sponge material, held between the first wall and the upper face of the chamber, such that the element at least partially overlaps the edge of the cavity facing the first wall. In this case, the sample, upon reaching the bottom of the cavity, is absorbed by the paper or sponge and transported to the first wall, allowing it to then enter the chamber by capillary action.

[0017] Preferably the chamber (4) of the chip shall have on its second wall (2) at least one, preferably at least three capillary guides corresponding to a protrusion or groove collinear with the flow of the liquid when it fills the chamber and preferably having a semi-elliptical cross-section.

[0018] Preferably, the function of holding the sealing film (6) on the inlet orifice (50) will be ensured by fins, preferably by at least four fins (53) forming a cross or a grid located on the outside side of the second wall (2) of the chamber (4) to prevent the sealing film (6) from penetrating deeply into the inlet orifice during the sealing step.

[0019] Preferably, the cavity surface has a polished surface finish of a quality ranging from SPI B2 to SPI A1 (according to the standard of the "Society of Plastics Industry") to promote the movement of the liquid in the direction of the capillary bridge.

[0020] According to one embodiment, the chip will comprise a funnel-shaped cavity (52) characterized in that the surface of the funnel (52) has undulations consisting of at least 6 periods, forming grooves extending from the center of the funnel to its periphery, these undulations promoting the movement of the liquid in the direction of the first capillary bridge located in the center of the funnel, which prevents the sample, if it is not well placed in the center of the inlet orifice, from remaining stuck on a periphery of the orifice without reaching the capillary bridge.

[0021] Preferably the adhering sealing film (6), located on the side of the block at least partially transparent (2), attached to the tab (9), extends towards the opening (16) without being attached to the block (2), the part not attached to the block being protected by a protective cover (26).

[0022] According to one variant, the chip will include an identification label (80) preferably at the level of the tab (9).

[0023] Preferably, the chip according to the invention will include error-proofing means consisting of a notch (8) located on the chip, preferably located on the tab (9).

[0024] Surprisingly, after sealing, the device according to the invention maintains perfect sealing despite temperature variations which induce pressure variations on the inner walls of the consumable and on the film, pressure due to the degassing of the reagent which can be greater than 1 bar, and this despite the large size of the inlet orifice.

[0025] The invention also relates to a testing system for a microfluidic sample chip for PCR-type testing of biological samples, the chip being as described above.

[0026] The test system according to the invention, in addition to the chip described above and comprising error-proofing means (8), shall comprise a test apparatus having a side wall (23) provided with an opening (22) allowing the chip to pass through to thermalization means (21), complementary error-proofing means intended to cooperate with error-proofing means present on the chip, and means for maintaining the chip in contact with the thermalization means after its introduction through the opening, which maintain the chip in contact with the thermalization means. preference generated by the cooperation of the error-proofing means located on the chip and the complementary error-proofing means located in the device, in particular to start the sample analysis sequence, this system also including means for measuring the fluorescence of the sample.

[0027] According to a preferred mode, the chip testing system according to the invention will include fluorescence imaging means.

[0028] Preferably, the strip should be at least partially opaque to prevent external light from reaching the sample once the chip is inserted into the associated test device. Furthermore, the strip should preferably be rigid or semi-rigid, positioned in line with, and preferably in the same plane as, the walls.

[0029] According to a preferred embodiment, the sample chip according to the invention includes error-proofing means intended to cooperate with complementary error-proofing means located in the test device to analyze the sample contained in the chamber in order in particular to start the sample analysis sequence automatically when the chip is inserted into the PCR analysis device and to detect if the chip has been inserted backwards.

[0030] These error-proofing methods consist, for example, of a notch located on the chip, preferably located on the tab.

[0031] While in general the tab is preferably flat and arranged in the same plane as the walls, a variant of the invention consists of providing a tab having for example the shape of a protrusion, preferably arranged in the extension of the walls, allowing better gripping of the chip to introduce it into the test device.

[0032] The invention also relates to a method for performing a test with a chip as described above, in which the chip is positioned in a test apparatus as described above, thanks to the cooperation of the error-proofing means (8) and the complementary error-proofing means, and a holding force of at least 25N using for example a glass block (20) driven by a stepper motor compressing the sample chip against the thermalization element (21) at the level of the sealing film (6), so as to seal the chamber (4) by favoring greater compression of the film thanks to the edges of the inlet (54) and outlet (58) openings, and ensure good thermal contact minimizing the air gap between the thermalization means (21) and the metal plate (3), then the sample is subjected to PCR thermal cycling, then to fluorescence detection and finally the holding means release the chip.

[0033] In this patent application, the following terms shall have the following meanings:

[0034] The term "PCR-compatible material" means a material that does not contain PCR inhibitors, does not degrade enzymes, fluorescent probes, dNTPs, oligonuclear nucleotides, and does not absorb oligonucleotides and other RNA or DNA sequences and does not diffuse molecules that can interact with the functioning of enzymes or change the characteristics of the PCR reaction.

[0035] The term "opaque material" means a material whose optical transmission in the visible spectrum, i.e. between 300 nm and 800 nm, is less than 20%, preferably less than 1%, for a material thickness of one millimeter.

[0036] The term "transparent material" means a material whose optical transmission, particularly in the visible spectrum, is greater than 80%.

[0037] The term "high thermal conductivity material" means a material whose thermal conductivity is greater than 15 wm-1 .K-1 .

[0038] The term "rigid or semi-rigid material" means a material whose Young's modulus is greater than 10 MPa.

[0039] The terms apparatus, test apparatus, sample testing apparatus, PCR analysis apparatus are used interchangeably to refer to the PCR sample analysis apparatus.

[0040] The term hydrophilic material means a material whose contact angle of advance of a drop of pure water on a flat surface made of that material is less than 90°.

[0041] The invention will be better understood with the aid of the following embodiments, given by way of non-limiting example, together with the figures which represent:

[0042] [Fig. 1 a] [Fig. 1 b] Figures 1 a and 1 b, a top and bottom view of the sample chip according to the invention.

[0043] [Fig. 2] Figure 2, a top view of the sample chip from Figures 1a and 1b without the protective and sealing films.

[0044] [Fig. 3a] Figure 3a, a cross-sectional view of the active part of the chip along axis AA of Figure 2.

[0045] [Fig. 3b] Figure 3b, a detail view C of the inlet orifice of Figure 3a.

[0046] [Fig. 4] Figure 4, a cross-sectional view of the active part of the chip along the BB axis of Figure 2.

[0047] [Fig. 5] Figure 5, an exploded view of the chip according to the invention before the elements of the active part of the chip were glued together.

[0048] [Fig. 6a] [Fig. 6b] [Fig. 6c] Figures 6a-c show the different stages of bonding the window and the metal part to create the active part of the chip.

[0049] [Fig. 7a] [Fig. 7b] [Fig. 7c] Figures 7a-c show a schematic view of the chip in combination with the test device.

[0050] [Fig. 8] Figure 8, curves representing the fluorescence signal of different samples (RFU on the ordinate) as a function of the number of cycles (on the abscissa).

[0051] [Fig. 9a] [Fig. 9b] [Fig. 9c] [Fig. 9d] [Fig. 9e] Figures 9a-e, the method of using the chip according to the invention.

[0052] [Fig. 10] Figure 10, rear view of the chip without the metal plate (3)

[0053] In these figures, the same elements bear the same references.

[0054] In Figures 1a and 1b, the sample chip according to the invention is shown as a preferably parallelepiped-shaped block, which, in a preferred embodiment, has the shape of a rectangular plate 10. This parallelepiped-shaped plate 10, made of opaque material, has a length equal to approximately five times its width in the figure (preferably between 50x10 mm (and a thickness of at least 1 mm) and 200x100 mm (and a thickness of at most 10 mm)). Its active part 13 (left part of the figure) has a substantially square opening 16 (see Figure 5) surrounded by an opaque frame 1 in which a transparent window 2 is housed (see Figure 5).This chip is provided with two orifices 50 and 51, respectively an inlet orifice 50 for introducing the liquid sample and an outlet 51 for venting the atmosphere present in chamber 4. These orifices are located diagonally across window 2 and communicate with chamber 4 (see Figures 3 and 4), which is preferably equipped with capillary guides 12 (Figure 5). Extending from and in the same plane as frame 1 is tab 9, which has a notch 8 on one side. This notch serves as a means of preventing incorrect insertion of the rectangular wafer 10 into the test device, which is not shown in the figure. Figures 1a and 1b show the top and bottom views of the sample chip, respectively, and Figure 2 shows an enlarged view of the active part 13 of the chip without the protective film 60 and the sealing film 6.In Figure 1b, a metal plate 3 is housed in the opening 16 (Figure 5), thus closing the active part 13 of the sample chip. This metal plate 3, in line with the tab 9, will be slightly raised (above) relative to the tab 9 in order to achieve good thermal and mechanical contact with the thermalization element 21 (figure 7a-c).

[0055] Figure 3a shows a cross-sectional view along axis AA (diagonal of the active part 13) defined by the two orifices 50 and 51 (or inlet and outlet orifices, respectively). The transparent window 2 rests, by means of its peripheral end 17, on a rim 18 integral with the frame 1, creating a groove 7 into which an adhesive (silicone or other) will be injected all around the window 2, allowing the metal plate 3 to be fixed tightly to the window 2 and to the frame 1. The metal plate 3 thus delimits the chamber 4 into which the sample is introduced through the injection orifice 50. A sealing film 6 covers the orifices 50 and 51 to hermetically seal these openings after injection of the sample to be analyzed into the chamber 4. Figure 3b shows a detailed view of a part of the funnel showing the angle at the top between the wall of the funnel 52 and its edge 54.Figure 3a shows the second capillary bridge formed by the projections (53).

[0056] Figure 4 shows a cross-sectional view along the BB axis (Figure 2) of the active part of the sample chip.

[0057] Figure 5 shows an exploded view of the sample chip of Figures 1a and 1b, before the assembly of the window 2 and the metal plate 3 inside the opening 16 in the rectangular wafer 10, and the protective and sealing films 60 and 6 respectively, as well as an identification label 80. Several longitudinal capillary guides 12, facilitating the circulation of the liquid sample and its distribution within chamber 4, are shown in chamber 4. The protective film 60 has an opening 81 corresponding to the inlet orifice 50. The opening in this protective film 60 corresponding to the outlet orifice 51 is not shown in Figure 5.

[0058] Figures 6a, 6b, and 6c illustrate the assembly of the window 2 and the metal plate 3. Using an injection device 19, liquid or semi-liquid adhesive is used to fill the groove 7 (Figure 6c) located all around the window 2. The semi-liquid adhesive 14, such as silicone or another PCR-compatible material (Figure 6b), is then applied to the window 2 to seal the resulting chamber 4. The adhesive is then polymerized (cured) using a suitable UV source 15. Once the sample chip is prepared, the sample to be analyzed can be introduced into the chip, and the ports 50 and 51 are then sealed with a PCR-compatible adhesive film. The sample analysis is then performed as shown in Figures 7a, 7b, and 7c.

[0059] For this purpose, a test apparatus is available, left part of figures 7a-c (interior). This includes means for holding, for example, a transparent (preferably glass) movable block 20 through which fluorescence measurement can be made and a thermalization element 21 for the sample; (for more details on a thermalization element, see application WO2018 / 114625 in the name of the Applicant). On the other side of the wall 23 of the test device (on the right in figures 7a-c: "exterior") the rectangular plate 10 is introduced through the opening 22 in this wall, using the tab 9, so as to bring the active part 13 of the chip above and in contact with the thermalization element: the proper positioning of the chip on the thermalization element 21 is achieved thanks to the keying notch 8 which cooperates with additional keying means located in the test device.This cooperation then triggers a movement (downwards in Figure 7a) of the glass block 20, which presses against the active part 13 of the sample chip (Figure 7b), ensuring the sealing of the chamber 4 by applying overpressure to the rim of the inlet orifice 54, the rim of the outlet orifice 58, and the flatness pads 55 (Figure 2). The pressing action on the active part 13 also ensures good thermal contact of the chip (whose metallic part is in contact with the thermalization element), which allows the thermal cycling of the sample to begin. During this cycling, the optical signal 25 from the sample's fluorescence is collected on the sensor 24, enabling real-time monitoring of the fluorescence in the chip. After a predetermined time (approximately 40 cycles), the thermal cycling is stopped, and the glass block 20 is raised (Figure 7b). 7c) in its resting position, the chip being pulled outwards by means of the tab 9.

[0060] The chip is handled from outside the device, and its insertion does not require opening the device. No panels, drawers, or access doors are needed on the test unit, hence its simplicity and speed of use.

[0061] The chip is inserted through a fixed slot, then removed in the same way once the detection operation is complete.

[0062] The correct positioning of the chip is determined by the notch 8 on the edge of the chip. Once in position, it engages a contactor (not shown in figure 7a) which allows the system to be locked.

[0063] In this variant, the holding and thermalizing means form a movable jaw that allows the chip to be held in position; it is composed of: - a 20cm glass block mounted on a motorized platform allowing its vertical movement for the upper part, - a thermalization element 21 (metallic for example), allowing rapid temperature change during PCR thermal cycles for the lower part.

[0064] The pressure applied to the chip is preferably between 100 g and 10 kg distributed more particularly on the edges of the inlet and outlet ports, respectively 54 and 58, as well as the flatness pads 55 allowing preferential sealing on the ports of chamber 4.

[0065] Once the PCR is complete, the jaw opens automatically (under the control of an automated system programmed for this purpose, in a manner known per se) releasing the chip, which can be removed by the user (figure 7c).

[0066] The pressure-locked position (Figure 7b) ensures maximum contact between the chip's metal plate and the thermalization element, and prevents the sample liquid inside the chip from boiling when the temperature approaches 100°C. But it It also helps to prevent potential leaks of PCR products into the device that could contaminate it.

[0067] The transparent block, preferably made of glass, allows the fluorescence signal from the sample to be collected during PCR amplification.

[0068] Figure 8 shows the RFU fluorescence signal as a function of the number of cycles to which the samples are subjected. TARGET 1 and TARGET 2 curves correspond to two targets of the SARS-CoV-2 coronavirus DNA, while the CONTROL curve represents the internal control. Amplification is clearly detectable from the 35th cycle onward, corresponding to rapid detection in less than 20 minutes.

[0069] Figures 9a-c represent the method of using the chip according to the invention.

[0070] In figure 9a, using a pipette 70, a drop of liquid 71, containing the sample and the reagents allowing PCR, is deposited into the inlet orifice 50.

[0071] In Figure 9b, the liquid enters the chamber through the inlet orifice 50 by capillary action until it reaches the outlet orifice 51.

[0072] In Figure 9c, the protective film 60 is removed from the chip using the tab 61 to remove any liquid from the inlet hole.

[0073] In figure 9d, the protective cover 26 of the sealing film 6 is removed from the film by the tab 28, leaving the adhesive surface of the sealing film exposed to the air.

[0074] In Figure 9e, the sealing film 6 is applied to the second wall 2 or upper wall of the chamber 4, blocking the inlet 50 and outlet 51 ports, after the sample has been injected into the chamber.

[0075] The sealing film preferably consists of a heat-resistant PET backing with a thickness between 20 µm and 500 µm, and a silicone adhesive mass compatible with the PCR process. between 20 µm and 500 µm. Due to the relative viscosity of silicone adhesives during thermal cycling, the presence of crossbars (53) for film support is preferably necessary and in some cases essential. Also due to the relative viscosity of silicone adhesives during thermal cycling, the presence of rims (54, 58) around the inlet and outlet ports, respectively, provides a means of sealing the ports by local compression of the sealing film around the perimeter of these ports.

[0076] Figure 10 shows the lower face of the chamber (4) and the second capillary bridge formed by the two lateral projections (90) allowing the liquid to create a bridge facilitating its entry into the chamber.

[0077] The protective cover 26 of the sealing film 6 is removed at the time of sealing the chip, after injection of the sample and the sealing film 6 is folded back on the upper wall of the chip in order to hermetically cover the holes 50 and 51.

[0078] A tracking tag with a "datamatrix" type code, for example, allows for the automated referencing of each chip, enabling sample tracking and connection to the hospital's Laboratory Information Management System (LIMS) for medical applications. The tag can also display information regarding the chip's use, for example, for research or in vitro diagnostic purposes.

[0079] This chip structure, with its sealing film, tab, and keying system, allows the chip to be inserted into the test device through slot 22 without needing to open the device.

Claims

CLAIMS A microfluidic sample chip for testing biological samples, particularly for PCR analysis, having the form of a block (10), preferably parallelepiped-shaped, comprising at least one hollow chamber (4) for receiving a sample, delimited in particular by a first wall (or lower wall) (3) made of a heat-conducting material with a thermal conductivity greater than 0.1 Wm⁻¹.K⁻¹, preferably greater than 15 Wm⁻¹.K⁻¹, preferably metallic, and a second wall (or upper wall) (2), parallel to the first, made at least partially of a transparent and preferably hydrophilic material, the block (10) having an inlet orifice (50) allowing the introduction of the sample into the chamber (4) and an outlet orifice (51) allowing the evacuation of the atmosphere present in the chamber during its introduction, the inlet orifice (50) extending into a cavity (52, 54) allowing the receipt the sample,characterized in that the inlet orifice has a surface area greater than 4 mm2, in that the cavity (52) is combined with a first capillary bridge allowing the sample to penetrate by capillarity into the chamber (4) and prevent it from remaining in the cavity (52) and in that the chip also includes a sealing film (6), configured to be in contact with the block (10), this sealing film (6) having a surface area sufficient to cover the inlet (50) and outlet (51) orifices simultaneously, said sealing film being configured to adhere to said block (10) and to close the inlet (50) and outlet (51) orifices of the block in a hermetic manner after filling the chamber (4) with the sample. Chip according to claim 1, characterized in that the chip further comprises a tab (9) at least partially opaque and rigid or semi-rigid, preferably disposed in the extension and preferably in the same plane as the two walls (2,3),the block (10) being preferably integral with the tab. Chip according to any one of the preceding claims characterized in that a protective film (60) disposed on the second wall (2), comprising two, Openings (81) whose sides are adjacent to the perimeter of the orifices (50, 51) of the chamber (4) allow the areas in contact with the sealing film (6) to be protected from particulate or liquid contamination that could limit the adhesion capacity of the sealing film (6). Chip according to any one of the preceding claims, characterized in that the distance between the lower and upper walls of the chamber is less than 1 mm, preferably less than 500 pm.Chip according to any one of the preceding claims, characterized in that the first capillary bridge is characterized according to a first alternative by at least one projection in the block positioned on the rim of the cavity (52) facing the first wall (3), this projection having at least one vertical surface contiguous to at least one of the flanks of the inlet orifice (50) on the one hand, and to the first wall (3) on the other hand, or according to a second alternative by an element of hydrophilic porous material in contact on one of its faces with at least one of the flanks of the inlet orifice (50) on the one hand and on another of its faces with the first wall (3) on the other hand.A chip according to any one of the preceding claims, characterized in that the cavity (52) has a shape selected from that of a funnel (52) such as a truncated cone, a truncated pyramid, or a trapezoidal prism having an apex angle between 20 and 90 degrees, preferably between 20 and 50 degrees, and that of a straight hole with an elliptical or polygonal cross-section, passing through the second wall (2). A chip according to any one of the preceding claims, characterized in that the first capillary bridge comprises at least one projection located on the upper face of the chamber (4), the size of the height of the chamber (4), and having a vertical surface situated in the extension of a portion of the flanks of the inlet orifice (50).Chip according to any one of the preceding claims characterized in that a second capillary bridge located on the upper part of the second wall (2), positioned on a side of the cavity (52) has at least one projection of the depth of the cavity (52), for example the funnel (52) taking the form of a fin (53) or a cross, perpendicular to the upper surface of the chamber (4) whose longitudinal direction points towards the. center of the inlet orifice (50), the fin extending into the cavity (52) towards its center for a distance of between 0.1 mm and the edge-center distance of the inlet orifice. Chip according to any one of claims 5 to 8 according to the second alternative of the capillary bridge characterized in that the first capillary bridge is a hydrophilic fiber element such as a sheet of cellulose paper or a sheet of sponge material held between the first wall (3) and the upper face of the chamber (4) such that the element at least partially overlaps the edge of the cavity facing the first wall (3). Chip according to any one of the preceding claims characterized in that the chamber (4) has on its second wall (2) at least one, preferably at least three capillary guides corresponding to a protrusion or groove collinear with the flow of the liquid when it fills the chamber and preferably having a semi-elliptical cross-section.A chip according to any one of the preceding claims, characterized in that the function of retaining the sealing film (6) on the inlet orifice (50) is ensured by fins (53), preferably by at least four fins (53) forming a cross-shaped or grid-like structure located on the outer side of the second wall (2) of the chamber (4) to prevent the sealing film (6) from penetrating deeply into the inlet orifice during the sealing step. A chip according to any one of the preceding claims, characterized in that the surface of the cavity has a polished surface finish of a quality ranging from SPI B2 to SPI A1 (according to the Society of Plastics Industry standard) to promote the movement of the liquid in the direction of the capillary bridge.Chip according to any one of the preceding claims comprising a funnel-shaped cavity (52) characterized in that the surface of the funnel (52) is provided with undulations consisting of at least 6 periods, forming grooves starting from the center of the funnel and projecting to its periphery, these undulations promoting the movement of the liquid in the direction of the first capillary bridge located at the center of the funnel, which prevents the sample, if it is not well positioned at the center of the inlet orifice, from remaining stuck on a periphery of the orifice without reaching the capillary bridge. A chip according to any one of the preceding claims, characterized in that the adhering sealing film (6), located on the at least partially transparent side of the block (2), is integral with the tab (9) and extends towards the opening (16) without being integral with the block (2), the portion not integral with the block being protected by a protective cover (26). A chip according to any one of the preceding claims, characterized in that it comprises an identification label (80), preferably located on the tab (9). A chip according to any one of the preceding claims, characterized in that it comprises error-proofing means consisting of a notch (8) located on the chip, preferably located on the tab (9). A testing system for a microfluidic sample chip for PCR-type testing of biological samples, characterized in that it comprises a chip according to any one of claims 1 to 16.A test system according to claim 17, comprising a sample chip provided with error-proofing means (8) and a chip testing apparatus, characterized in that the testing apparatus comprises a side wall (23) provided with an opening (22) allowing the passage of the chip towards thermalization means (21), complementary error-proofing means intended to cooperate with the error-proofing means (8) present on the chip, means for maintaining the chip in contact with the thermalization means after its introduction through the opening which maintain the chip in contact with the thermalization means, contact preferably generated by the cooperation of the error-proofing means (8) located on the chip and the complementary error-proofing means located in the apparatus, in particular to start the sample analysis sequence, this system further comprising means for measuring the fluorescence of the sample.A chip testing system according to claim 17 or 18, characterized in that it comprises fluorescence imaging means (24,25). A method for performing a test with a chip according to any one of claims 1 to 16, characterized in that the chip is positioned in a test apparatus according to any one of claims 17 to 19. Through the cooperation of the error-proofing means (8) and complementary error-proofing means, a force of at least 25 N is applied, preferably using, for example, a glass block (20) driven by a stepper motor, compressing the sample chip against the thermalization element (21) at the level of the sealing film (6), so as to seal the chamber (4). This compression of the film is achieved by means of the edges of the inlet (54) and outlet (58) openings, ensuring good thermal contact and minimizing the air gap between the thermalization means (21) and the metal plate (3). The sample is then subjected to PCR thermal cycling, followed by fluorescence detection, and finally to the holding means. release the chip.

Images