MICROFLUIDIC DEVICE FOR THE TRAPPING, TRANSPORT AND / OR RESUSPENSION OF MAGNETIC NANOPARTICLES

Abstract

Microfluidic device for trapping, transporting and / or resuspension of magnetic nanoparticles. The present invention relates to a microfluidic device for trapping, transporting, and / or resuspensioning magnetic nanoparticles, comprising: (i) a microfluidic chip with one or more chambers adapted for housing magnetic nanoparticles; (ii) a first magnet (6), arranged adjacent to a first chip wall and adapted to be displaced relative to said first wall; and (iii) a second magnet, arranged adjacent to a second chip wall facing the first wall, and adapted to be displaced relative to the second wall and to the first magnet, such that the resulting magnetic force to which the magnetic nanoparticles are subjected varies as a consequence of the relative distance between the first and second magnets.Thanks to this configuration, not only is the transport of nanoparticles between chip chambers achieved without channel obstructions due to the backflow of nanoparticles to the starting chamber, but the agglomeration of the nanoparticles is also avoided.

Description

MICROFLUIDIC DEVICE FOR TRAPPING, TRANSPORT AND / OR RESUSPENSION OF MAGNETIC NANOPARTICLES FIELD OF INVENTION The present invention falls within the field of microfluidics. More specifically, the object of the invention relates to a microfluidic device capable of adapting the intensity and gradient of the magnetic field it generates to control the movement of magnetic nanoparticles within it. BACKGROUND OF THE INVENTION Magnetic nanoparticles (MNPs) have garnered increasing interest in biomedicine, primarily due to their small size (1-100 nm), the physical properties of their magnetic core, their high surface area to volume ratio, and the possibility of surface functionalization with biologically relevant molecules. Specifically, they have become a highly useful nanomaterial for use as a contrast agent in magnetic resonance imaging (MRI), as a drug delivery and release system for controlled delivery to specific areas of the body, and in cancer treatment through intracellular hyperthermia in response to an external magnetic field. In addition to their in vivo applications, NPMs can also be used in in vitro assays for the direct detection of biologically important analytes, such as DNA, proteins, or bacteria present at very low concentrations in biological fluids. Although their implementation has been carried out in various biosensors or integrated devices, the transport and handling of these nanoparticles within the microfluidic chip continues to pose a technological challenge, either because their entrapment is achieved but with particle aggregation, or because complex actuation systems are required. Efficient transport of nanoparticles (NPs) in solutions requires strong magnetic fields, which are determined by the size of the magnet used to attract and move them. In the case of microfluidic chips, a magnet comparable in size to the chambers is needed to transfer NPs between them. The relatively large size of the magnet means that, during its movement, it attracts not only the NPs from the initial chamber but also those already transferred to the target chamber, moving them in the opposite direction to the intended flow. This can lead to clogging of the chip channels by particle agglomeration, thus reducing transport efficiency. Furthermore, particle agglomeration prevents direct contact between the particles and the solution, hindering their interaction with its components.This interaction is necessary for the efficient washing of NPMs or for their reaction with certain solutes present in the solution. To date, all technologies developed for the controlled movement of NPMs in a fluid within a microfluidic chip or a "lab-on-chip" device are based on the dragging of the NPMs by pumping the fluid and subsequent manipulation of their trajectory with a magnetic field [Khizar, S. et al. Electrophoresis, 2020, vol. 41, no. 13-14, pp. 1206-1224], or in the action of a magnet between the chip chambers with a simple sweep [Berr and , SM et al. Lab Chip, 2011, vol. 11, no. 10, pp. 1747-1753]. This is insufficient in the case of viscous fluids. Regarding the mixing or resuspension of NPMs in miniaturized systems using a magnetic field, the solutions described in the state of the art include permanent magnets fixed to rotating disks located in planes parallel to the microfluidic chip [Berenguel-Alonso, M. et al. Anal. Bioanal. Chem., 2014, vol. 406, p. 6607-6616; Zhong, R. et al. RSC Adv., 2020, vol. 10, no. 49, p. 29311-29319]. The rotation of the disks causes the magnets to periodically move closer to and further from the chip chambers along the chip's major axis, thus inducing the movement of the NPMs through the channels that connect the mixing chamber to adjacent chambers. However, this movement is uncontrolled, resulting in unwanted NPM losses. Based on this state of the art, the need arises to develop a microfluidic device for the trapping, transport and / or resuspension of NPMs capable of adapting the intensity and gradient of the magnetic field it generates to control the movement of said NPMs, avoiding the obstruction of the chip channels, as well as the agglomeration of the same if said nanoparticles are in suspension in a fluid. DESCRIPTION OF THE FIGURES Figure 1a shows a traditional microfluidic device for trapping and transporting magnetic nanoparticles (NPMs) comprising a microfluidic chip with three chambers and a magnet of comparable size to the chambers. This magnet generates a magnetic field that attracts not only the NPMs from the first chamber but also those already transferred to the second chamber, obstructing the junction channel. Figure 1b shows the device of the invention in one of its preferred embodiments. By using a second magnet, the NPMs transferred to the second chamber by the first magnet are held in place, allowing as many sweep cycles as necessary between the first and second chambers to be performed to transport all the NPMs without risk of clogging the bonding channel. Figure 2 shows a preferred embodiment of the method of the invention where the variation of the relative distance between the first and second magnets is repeated alternately until the desired resuspension, washing, or mixing of the NPMs is achieved. This variation comprises displacing the first and second magnets substantially perpendicular to the first and second chip walls, respectively. Figure 3 shows another preferred embodiment of the method of the invention where, in addition to displacing the first and second magnets substantially perpendicular to the first and second walls of the chip, respectively, both magnets are simultaneously displaced substantially parallel to the first and second walls, respectively. Figure 4 shows another preferred embodiment of the method of the invention, which includes performing the following steps: (a) trapping at least a portion of the NPMs located in the first chamber of the microfluidic chip with the second magnet; (b) moving the second magnet substantially parallel to the second wall of the chip with the trapped NPMs from the first chamber to a second chamber of the microfluidic chip; (c) positioning the first and second magnets facing each other; (d) varying the relative distance between the first and second magnets, facilitating the trapping of the NPMs by the first magnet; (e) moving the second magnet substantially parallel to the second wall of the chip from the second chamber to the first chamber; (f) repeating steps a) to e) until no magnetic nanoparticles remain in the first chamber; (g) trapping the magnetic nanoparticles located in the second chamber with the second magnet;(h) moving the first magnet substantially parallel to the first chip wall to the third chamber; (i) moving the second magnet substantially parallel to the second chip wall with the trapped magnetic nanoparticles from the second chamber to a third chip chamber. NUMERICAL REFERENCES USED IN THE FIGURES In order to aid a better understanding of the technical characteristics of the invention, the aforementioned figures are accompanied by a series of numerical references where, for illustrative and non-limiting purposes, the following is represented: DETAILED DESCRIPTION OF THE INVENTION In light of the prior art problems discussed above, the object of the present invention relates, firstly, to a device for trapping, transporting, and / or resuspensioning magnetic nanoparticles (MNPs), capable of adapting the intensity and gradient of the magnetic field it generates to control the movement of said MNPs. Advantageously, the device of the invention comprises: - a microfluidic chip (1) comprising one or more chambers (2, 3, 4) adapted for housing NPMs (5), wherein said microfluidic chip (1) is defined by a plurality of walls (10, 10), and wherein at least two of said walls (10, 10) are substantially opposite each other; - a first magnet (6), arranged adjacent to a first wall (10) of the two substantially opposing walls (10, 10), wherein said first magnet (6) has a magnetic field configured to capture the NPMs (5) located in one or more of the chambers (2, 3, 4) of the microfluidic chip (1), and wherein the first magnet (6) is adapted to be displaced relative to the first wall (10); and - a second magnet (7), arranged in a position adjacent to a second wall (10) of the two substantially opposing walls (10, 10), wherein said second magnet (7) has a magnetic field configured to trap the NPMs (5) located in the one or more chambers (2, 3, 4) of the microfluidic chip (1), and wherein the second magnet (7) is further adapted to be displaced relative to the second wall (10) and to the first magnet (6), such that the resulting magnetic force to which the NPMs (5) are subjected varies as a consequence of the relative distance between the first (6) and second (7) magnets. For the purposes of this invention, "magnet" means any permanent magnet or body capable of generating a persistent magnetic field. "Substantially opposing walls" means walls on opposite sides of the chip, regardless of whether they are parallel, although they are preferably parallel. "Assembled in a contiguous position" means arranged such that the chip wall closest to the first magnet is the first of the two opposing walls, or such that the chip wall closest to the second magnet is the second of the two opposing walls. Finally, "resultant magnetic force" means the force experienced by the NPMs when subjected to the magnetic field generated by the first and second magnets in combination. Thanks to this configuration, as many sweep cycles as necessary can be performed between chambers (2, 3, 4) of the chip (1) with the second magnet (7) without the NPMs (5) already transferred to the destination chamber (2, 3, 4) being attracted, since they are trapped in the magnetic field created by the first magnet (6). This prevents the obstruction of the channels (9) of the chip (1) that connect the chambers (2, 3, 4) to each other (see Figs. 1a and 1b). In addition, the agglomeration of NPMs (5) is avoided when they are in suspension in a fluid. In a preferred embodiment of the device of the invention, the first magnet (6) is adapted to move closer to or further away from the first wall (10). In this way, the intensity of the magnetic field generated by said first magnet (6) is modulated on the NPMs (5) located in one or more of the chambers (2, 3, 4) of the microfluidic chip (1), trapping or releasing them, respectively. In another preferred embodiment of the device of the invention, the second magnet (7) is adapted to move substantially parallel to the second wall (10). In this way, the second magnet (7) can transport the NPMs (5) trapped in its magnetic field from one chamber to another (2, 3, 4) of the microfluidic chip (1). In another preferred embodiment of the device of the invention, the second magnet (7) is adapted to move closer to or further away from the second wall (10). In this way, the intensity of the magnetic field generated by said second magnet (7) is modulated on the NPMs (5) located in one or more of the chambers (2, 3, 4) of the microfluidic chip (1), trapping or releasing them, respectively. In another preferred embodiment of the device of the invention, the first magnet (6) is adapted to move substantially parallel to the first wall (10). In this way, the first magnet can move to different chambers (3, 4) of the chip (1). In another preferred embodiment of the device of the invention, the chip (1) comprises a plurality of chambers (2, 3, 4) separated by partitions (8) and connected by one or more channels or openings (9). More preferably, one or more of the channels (9) are arranged adjacent to the second wall of the chip (10). A second object of the invention relates to a method for trapping, transporting, and / or resuspending NPMs, preferably suspended in a fluid. Advantageously, the method of the invention comprises placing a plurality of NPMs (5) inside a chamber (2, 3, 4) of a device according to any of the embodiments described herein and, furthermore, performing the following steps: a) arrange the first (6) and second magnet (7) substantially opposite each other; and ) vary the relative distance between the first (6) and second (7) magnets, promoting the trapping of the NPMs (5) by one of said first (6) and second magnet (7). Preferably, the variation of the relative distance between the first (6) and second (7) magnets is repeated alternately, until the desired resuspension, washing or mixing of the NPMs is achieved. In another preferred embodiment of the method of the invention, varying the relative distance of the magnets (6, 7) comprises displacing the first magnet (6) and / or the second magnet (7) substantially perpendicular to the first (10) and / or second wall (10). As can be seen in Fig. 2, this variation causes the NPMs (5) to move slowly and in a cloud-like fashion through the solution toward the side of the nearest magnet (6, 7). Once displaced, but before regrouping, this variation of distance is repeated. Repeating this process allows modulation of the magnetic field in such a way as to favor the resuspension of the NPMs (5) in the solution. Additionally, the first magnet (6) and / or the second magnet (7) can be displaced substantially parallel to the first (10) and / or second wall (10). As can be seen in Figure 3, applying this type of extra displacement to the magnets induces a diagonal or circular movement of the NPMs that further favors their resuspension, mixing, or washing in the solution. In another preferred embodiment of the method of the invention, said method comprises, prior to or subsequent to any of steps a) and b), the performance of the following steps: c) to capture at least a portion of the NPMs (5) located in a first chamber (2) of the microfluidic chip (1) with the second magnet (7); d) displacing the second magnet (7) with the trapped NPMs (5) from the first chamber (2) to a second chamber (3) of the microfluidic chip (1); and, e) trapping the displaced NPMs (5) to the second chamber (3) with the first magnet (6). In the event that the microfluidic chip contains more than two chambers (2, 3, 4), the method of the invention may additionally comprise performing the following steps after step e) (see Fig. 4): f) move the first magnet (6) to a third chamber (4) of the chip (1); g) repeat steps a) -e) from the second (3) to the third (4) camera. These steps will be repeated between successive chambers of the chip (regardless of whether they are contiguous or not) until the desired NPMs are transferred from the initial chamber to the destination or target chamber. For the purposes of this invention, "initial chamber" means the chamber of the microfluidic chip where a plurality of NPMs are initially housed, and "destination chamber" or "target chamber" means the chamber where the NPMs housed in the initial chamber are ultimately to be transferred, preferably for use in in vitro assays. As can be seen in Figure 4, the first magnet (6) acts as a locking magnet; that is, trapping the NPMs carried by the second magnet (7), thus preventing them from being attracted again by the magnetic field generated by the second magnet (7) and, consequently, from obstructing the chip channels (see Fig. 1). Additionally, in order to transport all the NPMs contained in the initial chamber of the microfluidic chip to the destination chamber, the method of the invention may further comprise performing the following steps after step e): h) move the second magnet (7) from the second chamber (3) to the first chamber (2); i) repeat steps c) ae) and h) until there are no more NPMs (5) in the first chamber (2). A third object of the present invention relates to the use of the device according to any of the embodiments described herein for: - the transport of NPMs in a solution; or - the resuspension, washing or mixing of NPMs in a solution. Example of implementation The invention has been implemented and validated in a device for extracting DNA from samples of patients sick with COVID-19. Both the sweep and clamping magnets consist of two stacked neodymium magnets. A small cylindrical magnet, 3 mm in diameter and 6 mm high, is attached to a larger cubic magnet, 10 mm long. The assembly achieves a magnetic field of 470 mT on the circular face of the cylindrical magnet while simultaneously reducing the contact surface area of ​​the magnet stack. The circular faces of the two magnet stacks face each other on the first and second walls (10 and 10) of the chip, oriented towards its interior. The sweep magnet moves some of the NPMs from the initial chamber to the next chamber beneath the clamping magnet. In a second cycle, the sweep magnet collects the remaining NPMs from the initial chamber and moves them to the destination chamber, leaving the initial chamber free of NPMs. Tests were performed with magnetite core NPMs with nonspecific SiO2 coating, as well as with NPMs with specific oligo (dT) coating. Both types of NPMs had diameters between 10 nm and 40 nm. To demonstrate the efficiency of mixing the nanoparticles with the biological samples, DNA extractions were performed with and without mixing the nanoparticles, and the presence of extracted DNA was determined using PCR. No DNA was detected in the extractions without magnetic mixing. With magnetic mixing, the amounts of extracted DNA were comparable to those obtained with typical techniques using Eppendorff tubes and vortex mixing.

Claims

1. A microfluidic device for trapping, transporting, and / or resuspensioning magnetic nanoparticles, characterized in that it comprises: - a microfluidic chip (1) comprising one or more chambers (2, 3, 4) adapted for housing magnetic nanoparticles (5), wherein said microfluidic chip (1) is defined by a plurality of walls (10, 10), and wherein at least two of said walls (10, 10) are substantially opposite each other; - a first magnet (6), disposed in a position adjacent to a first wall (10) of the two substantially opposite walls (10, 10), wherein said first magnet (6) has a magnetic field configured to trap the magnetic nanoparticles (5) located in one or more of the chambers (2, 3, 4) of the microfluidic chip (1), and wherein the first magnet (6) is adapted to be displaced relative to the first wall (10); and - a second magnet (7),arranged adjacent to a second wall (10) of the two substantially opposing walls (10, 10), wherein said second magnet (7) has a magnetic field configured to capture the magnetic nanoparticles (5) located in the one or more chambers (2, 3, 4) of the microfluidic chip (1), and wherein the second magnet (7) is further adapted to move relative to the second wall (10) and to the first magnet (6), such that the resulting magnetic force to which the magnetic nanoparticles (5) are subjected varies as a consequence of the relative distance between the first (6) and second (7) magnets.

2. Device according to the preceding claim, wherein the first magnet (6) is adapted to move closer to or further from the first wall (10).

3. Device according to any of the preceding claims,where the second magnet (7) is adapted to move substantially parallel to the second wall (10).

4. Device according to any of the preceding claims, wherein the second magnet (7) is adapted to move towards or away from the second wall (10).

5. Device according to any of the preceding claims, wherein the first magnet (6) is adapted to move substantially parallel to the first wall (10).

6. Device according to any of the preceding claims, wherein the chip comprises a plurality of chambers (2, 3, 4) separated by partitions (8) and connected by one or more channels or openings (9).

7. Device according to the preceding claim, wherein one or more of the channels (9) are arranged adjacent to the second wall of the chip (10).

8. Method for trapping,Transport and / or resuspension of magnetic nanoparticles comprising placing a plurality of magnetic nanoparticles (5) inside a chamber (2, 3, 4) of a device according to any one of claims 1-7 and further comprising performing the following steps: a) placing the first (6) and second magnet (7) substantially opposite each other; and b) varying the relative distance between the first (6) and second (7) magnets, thereby facilitating the entrapment of the magnetic nanoparticles (5) by one of said first (6) and second (7) magnets.

9. Method according to the preceding claim, wherein the magnetic nanoparticles are suspended in a fluid.

10. Method according to the preceding claim, wherein the variation of the relative distance between the first (6) and second (7) magnets is repeated alternately until resuspension is achieved.washing or mixing the desired magnetic nanoparticles.

11. Method according to any of claims 8-10, wherein the variation of the relative distance between the first (6) and second (7) magnets comprises displacing the first magnet (6) and / or the second magnet (7) substantially perpendicular to the first (10) and / or second wall (10).

12. Method according to the preceding claim, further comprising displacing the first magnet (6) and / or the second magnet (8) substantially parallel to the first (10) and / or second wall (10).

13. Method for trapping, transporting, and / or resuspensioning magnetic nanoparticles (5) according to claim 8, wherein the microfluidic chip (1) comprises a plurality of chambers (2, 3, 4) and wherein said method comprises, prior to or subsequent to any of steps a) and b),the performance of the following steps: c) trapping at least a portion of the magnetic nanoparticles (5) located in a first chamber (2) of the microfluidic chip (1) with the second magnet (7); d) displacing the second magnet (7) with the trapped magnetic nanoparticles (5) from the first chamber (2) to a second chamber (3) of the microfluidic chip (1); and, e) trapping the magnetic nanoparticles (5) displaced to the second chamber (3) with the first magnet (6).

14. Method according to the preceding claim, wherein the microfluidic chip (1) comprises at least three chambers (2, 3, 4) and wherein said method further comprises performing the following steps after step e): f) displacing the first magnet (6) to a third chamber (4) of the chip (1); g) repeating steps a)-e) from the second (3) to the third chamber (4).

15. Method according to any of claims 13-14, wherein said method further comprises,Performing the following steps after step e): h) moving the second magnet (7) from the second chamber (3) to the first chamber (2); i) repeating steps c) to e) and h) until no magnetic nanoparticles (5) remain in the first chamber (2).

16. Use of the device according to any one of claims 1-7, for: - transporting magnetic nanoparticles in a solution; or - resuspending, washing, or mixing magnetic nanoparticles in a solution.

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