Cell for a microfluidic analysis system

Abstract

A cell for a microfluidic analysis system is described wherein the cell is adapted to be traversed by a flow of a reagent fluid and comprising a hollow main container having an outer side wall defining an inner chamber, a lower base having a flow port and a well formation, a lower outlet port for inlet of the reagent fluids, a vertical outlet port for outlet of the reagent fluids from the cell with a lower inlet port arranged in the inner chamber adjacent to an upper edge of the well formation and an upper portion comprising an upper inlet port and an upper outlet port.

Description

[0001] Cell for a microfluidic analysis system

[0002] Technical Field

[0003] The present invention is generally placed, in general, in the field of microfluidic systems; in particular, the invention refers to a cell for a microfluidic analysis system.

[0004] Background art

[0005] Microfluidics is an interdisciplinary field that deals with the manipulation and control of liquids at micro and nanometric scales. It has had a significant impact on biosensor technology and in the analysis of biological samples through the development of miniaturized devices with applications in different fields.

[0006] The precise control of fluids through such applications allows the creation of controlled and reproducible environments.

[0007] Furthermore, through these applications, the consumption of samples and reagents is reduced, allowing precious materials or those with limited availability to be exploited more efficiently.

[0008] Microfluidic technologies have made automation possible and improved analytical capacity, allowing numerous experiments to be performed quickly in parallel. This allows the creation of portable and low-cost devices, suitable for use in the field or in healthcare facilities with limited resources. Such devices can significantly improve access to medical care in remote or disadvantaged areas and provide rapid results for timely and accurate diagnosis.

[0009] Currently, multi-well plates are among the most used for the simultaneous analysis of different samples. These plates are flat plates with a multiplicity of wells (usually 6, 12, 24, 48, 96, or 348 depending on the applications) where the samples on which to perform the tests can be housed. However, this type of plate presents limitations due to long incubation times and high consumption of reagents. In the publication R. Rauti et al., "Transforming a well into a chip: A modular 3D-printed microfluidic chip," APL Bioeng., vol. 5, no. 2, Jun. 2021, DOI: 10.1063 / 5.0039366, a single microfluidic unit, or cell, is disclosed, which can be connected in series with other units, offering greater control over flows in the culture medium and more uniform reagent distribution. The possibility of connecting multiple units in series allows the use of a single culture medium, reducing the use of reagents.

[0010] However, this solution is not easily miniaturizable and has low tolerance to high pressures.

[0011] In Z. He et al., "Composable microfluidic plates (cPlate): A simple and scalable fluid manipulation system for multiplexed enzyme-linked immunosorbent assay (ELISA)," Anal. Chem., vol. 93, no. 3, pp. 1489-1497, 2021, DOI: 10.1021 / acs.analchem.0c03651., specific microfluidic pathways are disclosed in a plate made with PDMS (polydimethylsiloxane) to distribute reagents in various wells. The loading of the wells, in this case, takes place through the use of a pipette and not a pump, disadvantageously not easily automatable.

[0012] It is also important to note that microfluidics in PDMS used in the known art presented, while having excellent qualities in terms of flexibility, does not always allow good sealing in the wells due to its mechanical characteristics. These limitations of this material may lead to liquid leaks in the case of high pressures, a necessary condition for feeding multiple cells with a single fluid flow.

[0013] Summary of the invention

[0014] It is the purpose of the present invention to create an independent cell for a microfluidic analysis system capable of overcoming the disadvantages of the known technique cited above, in particular a cell that allows efficient management of a reagent flow through a system that includes a plurality of cells, fluidically connected, that is modular, easily customizable according to experimental needs, and that meets the operational parameters necessary for applications in the field of interest.

[0015] The aforementioned and other purposes and advantages, which will be better understood later, are achieved, according to the present invention, by a cell for a microfluidic system having the characteristics defined in the appended claim 1.

[0016] Particular embodiments form the subject of the dependent claims, the content of which is to be understood as an integral part of this description.

[0017] In summary, the cell for the microfluidic system comprises within a vertical inlet duct of reagent fluids and a vertical outlet duct of reagent fluids. The cell also includes a well formation for channelling the fluid introduced by the vertical inlet duct of reagent fluids towards the vertical outlet duct.

[0018] Advantageously, the geometry described here allows the surface of the culture table associated with this type of cell to be wetted uniformly. Furthermore, the geometry allows the management of the reagent fluid flow using peristaltic or continuous pumps.

[0019] The claimed system, comprising a multi-well plate that allows a high degree of customization, is efficient for various numbers of cells and allows the type of culture table to be changed according to the specific requirements.

[0020] Brief description of the drawings

[0021] The functional and structural characteristics of some embodiments of a microfluidic cell and system according to the invention will now be described. Reference is made to the accompanying drawings, in which: figure 1, is a perspective view of a cell for a microfluidic system according to an embodiment of the present invention; figure 2, is a top view of a cell for a microfluidic system according to an embodiment of the present invention; figure 3, is a bottom view of a cell for a microfluidic system according to an embodiment of the present invention; and figure 4, shows two perspective views of a multi-well plate for a microfluidic system according to an embodiment of the present invention. figure 5, is a bar graph reporting the experimental results of the absorption peak of cells arranged in series according to an embodiment of the present invention; and figure 6, shows the resulting data from theoretical simulations on the pressure of cells arranged in series according to an embodiment of the present invention.

[0022] Detailed description

[0023] Before explaining in detail, a plurality of embodiments of the invention, it must be clarified that the invention is not limited in its application to the construction details and the configuration of the components presented in the following description or illustrated in the drawings. The invention can assume other embodiments and of being implemented or practically realized in different ways. It must also be understood that the phraseology and terminology have a descriptive purpose and should not be understood as limiting. The use of "include" and "comprise" and their variations are to be understood as encompassing the elements listed below and their equivalents, as well as additional elements and their equivalents.

[0024] In Figure 1, a perspective view of a cell 10 for a microfluidic analysis system according to an embodiment of the present invention is shown.

[0025] The cell 10 is designed to be traversed by a reagent flow to be supplied to a culture table 80 where biological samples, for example, may be present.

[0026] The cell 10 comprises a hollow main container 20 with an external side wall 30 that defines an internal chamber 40, a lower base 70 having a flow port 50, and a well formation 60 that extends vertically from the base 70 into the internal chamber 40, circumscribing the flow port 50. This well formation 60, when the cell 10 is placed vertically on the culture table 80, delimits an analysis volume 90 having as its base a subsurface of the culture table 80.

[0027] In Figure 2, a bottom view of the cell 10 is shown, making the flow port 50 visible. The flow port, in a preferred embodiment, is connected to a lower outlet port 91 of a vertical inlet duct 90 of reagent fluids to introduce the reagent fluids into the analysis volume 90. The well formation 60, which has a height less than the height of the external side wall 30, is arranged to channel the fluid, introduced by the lower outlet port 91 of the vertical inlet duct 90 for introducing reagent fluids connected to the flow port 50, from the analysis volume 90 to the internal chamber 40 of the hollow container 20.

[0028] In further embodiment, the well formation 60 is coaxially internal to the external side wall 30 of the hollow container 20 and defines with it a cylindrical annular interspace.

[0029] The cell 10 also includes a vertical outlet duct 100 for discharging reagent fluids from the cell 10 with a lower inlet 101 arranged in the internal chamber 40 adjacent to an upper edge 61 of the well formation 60.

[0030] The cell 10 also includes an upper part 110 of the hollow main container 20 comprising an upper inlet port 92 of the vertical inlet duct 90 and an upper outlet port 102 of the vertical outlet duct 100 of the reagent fluids.

[0031] In Figure 3, a top view of the cell 10 is shown in an embodiment of the present invention that shows the upper inlet port 92 of the vertical inlet duct 90 and the upper outlet port 102 of the vertical outlet duct 100.

[0032] The cell 10 can be, for example, 4 cm high with the upper inlet port 92 having a diameter of 0.9 mm and the upper outlet port 102, instead, a diameter of 0.6 mm.

[0033] In a further embodiment, the external side wall 30 has one or more projections 31 that are designed to be housed in corresponding locking seats 121 present in a plurality of wells 120 of a multi-well plate 130 associated with the culture table 80. The locking seats 121 are configured to cooperate with one of the projections 31 to lock and support a cell 10 in a vertical position.

[0034] Now referring to Figure 3, a perspective view of a multi-well plate 130 with the corresponding wells 120 according to an embodiment of the present invention is shown. In a further embodiment, the locking seats 121 of the well 120 are shaped to cooperate with the projections 31 of the external side wall 30 locking the cell 10 in a detachable manner according to a bayonet configuration.

[0035] In a further embodiment, the base 70 also includes a sealing element 140 in a closed-ring shape that surrounds a lower part of the well formation 60 so that, when the cell 10 is locked through the locking seats 121 as described above, the analysis volume 90 is sealed. The sealing element 140 can be, for example, a toroidal O-ring.

[0036] The sealing element 140 may have an internal diameter of 6.2 mm and an external diameter of 9.6 mm.

[0037] The present invention also relates to a microfluidic analysis system for implementing the flow of reagent fluids to a culture table 80, which will now be illustrated.

[0038] In Figure 4, two perspective views of a multi -well plate 130 according to an embodiment of the present invention are shown.

[0039] The system includes a multi-well plate 130 with a plurality of wells 120 configured for the temporary support and locking of a corresponding plurality of cells 10 as described in the previous embodiments.

[0040] In this system, the cells 10 are arranged to be fluidically connected and traversed in series by a reagent fluid flow through a plurality of connecting channels 150 that connect each upper outlet port 102 of the vertical outlet duct 100 of a preceding cell 10 with the upper inlet port 92 of the vertical inlet duct 90 of a subsequent cell 10. Through this configuration, it is possible to provide the reagent fluid into the analysis volumes 90 defined by the respective well formations 60 of the respective cells 10 through a single reagent flow. This configuration in series is not to be considered limiting; the system also foresees the possibility of different connections in series in parallel with each other. The system also foresees the possibility of changing the culture table, as needed, with culture tables of different materials such as, for example, glass, PDMS, or other biocompatible materials.

[0041] The flow management of the system claimed here can be carried out, for example, by a peristaltic pump or a continuous pump.

[0042] In a further embodiment, the microfluidic analysis system can be made through an additive manufacturing process such as 3D printing. For example, this additive manufacturing process may be carried out using an acrylonitrile butadiene styrene filament.

[0043] In a further embodiment of the microfluidic system, when the cells 10 include a sealing element 140 as illustrated above, the locking seats 121 of each well 120 are arranged in such positions that, in a locked condition in which the projections 31 of the cells 10 are engaged in the well seats 121, the respective closed-ring sealing elements 140 are vertically compressed and tightly surround the bases of the analysis volumes 90.

[0044] Without departing from the principle of the invention, the forms of implementation and the particular details may be widely varied with respect to what has been described and illustrated purely by way of non-limiting example, without thereby departing from the scope of protection of the invention defined by the appended claims.

[0045] EXAMPLES

[0046] Example 1: Peak absorption experiment with 8 cells in series

[0047] From an experimental point of view, the uniform and highly efficient coating of a nanostructured surface with gold nanoparticles was demonstrated. In particular, the nanostructure was used to exploit a physical phenomenon called "localized surface plasmon resonance" to monitor the coating of the surface through spectrophotometric measurements in a simple and fast way. In particular, the shift of the absorption peak was observed. The greater the measured peak shift, the greater the surface coverage by the biological element (mouse-produced standard IgG antibody at a concentration of 25 pg / mL). The concentration and volumes of the antibody solution used for this test were excessive, with an amount of material that in principle would have been enough to saturate approximately 200 wells.

[0048] Fixing the antibody concentration and flow time, the solution was passed through 8 single cells connected to each other through silicone tubes. The results shown in Figure 5 demonstrate that the same absorption shift was obtained for each of the 8 wells, evidence that all wells have a surface saturated with antibodies.

[0049] Example 2: Theoretical cell pressure simulations

[0050] As for theoretical simulations, a cell having the same dimensions and geometry was reproduced, and the uniformity of the wells and the pressures to which the individual cells are subjected when placed in series were verified.

[0051] The result of the simulation confirms the uniformity of the wells of the various cells positioned in series.

[0052] Indeed, analysing the individual spots obtained from the simulation of the solution flow on the surface, it is noted that the profiles of the 4 spots are identical to each other.

[0053] Furthermore, the pressure within the entire circuit of 8 cells arranged in series with each other was also simulated, and it was seen that the pressure values obtained are compatible with what was seen experimentally, see Figure 6.

Claims

CLAIMS1. A cell (10) for a microfluidic analysis system with associated culture table, wherein the cell (10) is adapted to be traversed by a flow of a reagent fluid and comprises:- a hollow main container (20) having an outer side wall (30) defining an inner chamber (40), a lower base (70) having a flow port (50) passing through it and a well formation (60) extending vertically from the lower base (70) into the inner chamber (40), circumscribing the flow port (50) and delimiting an analysis volume having as its base a sub-surface of the culture table (80);- said flow port (50) being connected to a lower outlet (91) of a vertical inlet duct (90) of the reagent fluids for introducing the reagent fluids into the analysis volume;- said well formation (60) having a height of less than the height of the outer side wall (30) and being arranged to convey the fluid, introduced by the lower outlet port (91) of the vertical inlet duct (90) of the reagent fluids, from the analysis volume to the inner chamber (40) of the hollow container;- a vertical outlet duct (100) of the reagent fluids for the emission of the reagent fluids from the cell (10) with a lower inlet port (101) disposed in the inner chamber (40) adjacent an upper edge (61) of the well formation (60); and- an upper portion (110) of the hollow main container (20) comprising an upper inlet port (92) of the vertical inlet duct (90) and an upper outlet port (102) of the vertical outlet duct (100) of the reagent fluids.

2. Cell (10) according to claim 1, wherein said well formation (60) is coaxially internal to the outer side wall (30) of the hollow main container (20) and defines with it an annular cylindrical cavity.

3. Cell (10) according to claim 1 or 2, wherein said hollow main container (20) has on the outer side wall (30) one or more protrusions (31) suitable to be accommodated in corresponding locking seats (121) presented by a plurality of wells (120) of a multi-well plate (130) associated with the culture table (80), wherein each locking seat (121) is configured to cooperate with one of said protrusions (31) to lock and support a cell (10) in an upright position.

4. Cell (10) according to claim 3, wherein the locking seats (121) of the well are shaped to cooperate with the protrusions (31) on the outer side wall (30) by releasably locking the cell (10) according to a bayonet configuration.

5. Cell according to claim 3 or 4, wherein said base comprises a closed-loop sealing element (140) surrounding a lower part of the well formation (60) in such a manner that, when the cell (50) is blocked, the analysis volume is tightly bounded.

6. A microfluidic analysis system for implementing reagent fluid flow to a culture table (80), comprising a plurality of cells (10) according to any one of the preceding claims, a multi-well plate (130) with a plurality of wells (120) configured to support and temporarily block a corresponding cell (10) of said plurality of cells (10), wherein the cells (10) are fluidically connected to be traversed in series by a flow of reagent fluid, and comprising a plurality of connecting channels (150) each connecting the upper outlet port (102) of the vertical outlet duct (100) of a preceding cell (10) with the upper inlet port (92) of the vertical inlet duct (90) of a succeeding cell (10) such that the reagent fluid is supplied into the analysis volumes delimited by the respective well formations (60) of the respective cells (10) via a single flow of reagent fluid.

7. Microfluidic analysis system according to claim 6, wherein the multi-well plate (130) is made by an additive manufacturing process.

8. Microfluidic analysis system according to claim 7, wherein the multi-well plate (130) is made using an acrylonitrile butadiene styrene filament.

9. Microfluidic analysis system according to claim 6, 7, or 8 comprising a plurality of cells (10) according to claim 5, wherein the locking seats (121) of each well (120) are arranged in positions such that, in a locked condition in which the protrusions (131) of the cells (10) are engaged in the locking seats of the wells (121), the respective closed-loop seals(140) are vertically compressed and tightly surround the bases of the analysis volumes.

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