METHOD AND FLOW CELL FOR SEPARATING BIOMOLECULES FROM LIQUID MEDIUM

Abstract

The present invention relates to a method for separating biomolecules from a liquid medium. The method comprises adding magnetic nanoparticles to the liquid medium comprising the biomolecules, the biomolecules each adapted to bind to respective surfaces of the magnetic nanoparticles; bringing the liquid medium to which the magnetic nanoparticles have been added into contact with a collector; applying a magnetic field to the liquid medium in contact with the collector to attract the magnetic nanoparticles ound with the biomolecules to a surface of the collector; and applying an electric potential to the surface of the collector to release the biomolecules from the magnetic nanoparticles.

Claims

1. A method for separating biomolecules from a liquid medium, comprising: adding magnetic nanoparticles to the liquid medium comprising the biomolecules, the biomolecules each adapted to bind to respective surfaces of the magnetic nanoparticles; bringing the liquid medium to which the magnetic nanoparticles have been added into contact with a collector; applying a magnetic field to the liquid medium in contact with the collector to attract the magnetic nanoparticles bound with the biomolecules to a surface of the collector; and applying an electric potential to the surface of the collector so that the surfaces of the magnetic nanoparticles are positively or negatively charged to release the biomolecules from the magnetic nanoparticles.

2. The method of claim 1, wherein the biomolecules comprise proteins fused with peptide tags.

3. The method of claim 2, wherein the peptide tags comprise charged peptide tags.

4. The method of claim 3, wherein an electrostatic repulsion is created between the magnetic nanoparticles and the charged peptide tags upon the application of the electric potential.

5. The method of claim 2, wherein the peptide tags form charge transfer complexes with the respective surfaces of the magnetic nanoparticles, the complex formation being breakable by a change of electrostatic interactions.

6. The method of claim 1, further comprising collecting the biomolecules released from the magnetic nanoparticles.

7. The method of claim 1 , wherein bringing the liquid medium to which the magnetic nanoparticles have been added into contact with the collector comprises: allowing the liquid medium to which the magnetic nanoparticles have been added to pass through a flow cell comprising a chamber and the collector, the collector being disposed in the chamber.

8. The method of claim 1, further comprising, before the application of the electric potential, separating the collector from the liquid medium; and bringing at least part of the collector into contact with another liquid medium.

9. The method of claim 1, wherein the magnetic nanoparticles comprise iron oxide nanoparticles.

10. The method of claim 2, wherein the peptide tags comprise negatively charged peptide tags and the method comprises applying a negative electric potential to the collector to repulse the proteins fused with the negatively charged peptide tags, thereby releasing the biomolecules from the magnetic nanoparticles.

11. The method of claim 1, further comprising removing the magnetic field to release the magnetic nanoparticles from the collector.

12. A flow cell for separating biomolecules from a liquid medium, the flow cell comprising: a chamber comprising an inlet and an outlet which define therebetween a fluid path for the liquid medium, and a volume for containing the liquid medium provided along said fluid path, the volume comprising a collecting area for biomolecules; a magnetic source proximal to the volume of the chamber, the magnetic source arranged and configured to generate a magnetic field extending at least between the collecting area and the remaining volume for containing the liquid medium; and a working electrode proximal to the volume for containing the liquid medium, the working electrode arranged and configured to generate an electric field at the collecting area, wherein the flow cell is an HGMS-based device, and a volume of the chamber ranges from 1,000 mm.sup.3 to 10 m.sup.3.

13. The flow cell of claim 12, wherein the magnetic source is a permanent magnet movably coupled to the working electrode and/or to the chamber or an electromagnet.

14. The flow cell of claim 12, further comprising a first collecting unit in fluid communication with the outlet of the chamber for collecting the biomolecules from the liquid medium, a second collecting unit in fluid communication with the outlet of the chamber for collecting a magnetic material from the liquid medium, and/or a supply unit in fluid communication with the inlet of the chamber for supplying the liquid medium to the chamber.

15. The flow cell of claim 12, wherein the working electrode is a matrix of the HGMS-based device, the matrix being disposed substantially along a central axis of the chamber and comprising a ferromagnetic material, and the HGMS-based device further comprises a counter electrode disposed on the chamber.

16. The method of claim 2, further comprising attaching the peptide tags to the proteins.

17. The method of claim 2, wherein the peptide tags comprise negatively charged peptide tags.

18. The method of claim 1, wherein the magnetic nanoparticles comprise superparamagnetic iron oxide nanoparticles (SPION).

19. The method of claim 11, further comprising collecting the magnetic nanoparticles released from the collector.

Description

[0044] The general aspects of the present invention being described above, specific, non-limiting embodiments for further illustrating the present invention will be described below with reference to the respective drawings, in which:

[0045] FIG. 1 is a schematic diagram of a flow cell in accordance with a first embodiment of the present invention; and

[0046] FIG. 2 is a schematic diagram of a flow cell in accordance with a second embodiment of the present invention.

[0047] Referring to FIG. 1, a flow cell 100 in accordance with a first embodiment of the present invention is a microfluidic chip for separating biomolecules 120 from a liquid medium (not shown). The biomolecules 120 have bound to magnetic nanoparticles 130 before they are introduced together into the flow cell 100.

[0048] The flow cell 100 comprises a chamber 102. The chamber 102 comprises an inlet 104 and an outlet 106 disposed at respective ends along a length direction thereof. The inlet 104 and the outlet 106 define therebetween a fluid path 108. When a liquid medium is introduced into the chamber 102, it flows generally along the fluid path 108. The chamber 102 comprises a volume for containing the liquid medium, which is defined by a wall 103 of the chamber 102 and further by the inlet 104 and the outlet 106.

[0049] The flow cell 100 further comprises a working electrode 112 and a counter electrode 114. The working electrode 112 is disposed on the wall 103 of the chamber 102. The counter electrode 114 is disposed also on the wall 103 of the chamber 102, opposite to the working electrode 112. An electric potential difference may be established across the working electrode 112 and the counter electrode 114.

[0050] A magnetic source 110 is provided in proximity of the chamber 102, adjacent to the working electrode 112. The magnetic source 110 is a permanent magnet movably coupled to the chamber 102.

[0051] As shown in FIG. 1, the working electrode 112 is in contact with the volume containing the liquid medium. Therefore, the working electrode 112 defines a collecting area roughly at an interface between the working electrode 112 and the volume containing the liquid medium, i.e., the space occupied by the magnetic nanoparticles 130 in combination with the biomolecules 120.

[0052] The magnetic source 110, when placed in proximity of the chamber 102 as shown in FIG. 1, generates a magnetic field B extending at least between the collecting area and a rest of the volume of the chamber 102. Accordingly, when the liquid medium is contained in the chamber 102, the magnetic nanoparticles 130 bound with the biomolecule 120 are attracted by the magnetic field B toward the collecting area (or toward the working electrode 112) and immobilized in the collecting area.

[0053] The working electrode 112 is capable of generating an electric field extending at the collecting area. Thereby, after the magnetic nanoparticles 130 bound with the biomolecule 120 are collected on the working electrode 112 (or within the collecting area), an electric potential is applied at the surface of the working electrode 112 to break up the binding between the biomolecules 120 and the magnetic nanoparticles 130, releasing the biomolecules 120 whilst maintaining the magnetic nanoparticles 130 on the working electrode 112. The biomolecules 120 flowing out from the chamber 102 are collected, for example, in a collecting unit (not shown) in fluid communication with the outlet 106.

[0054] Referring to FIG. 2, a flow cell 200 in accordance with a second embodiment of the present invention is an HGMS-based device. The HGMS-based device comprises a chamber 202 and a matrix 212 disposed in the chamber 202, substantially along a central axis thereof. The chamber 202 comprises a volume, defined by the wall 203 of the chamber, for containing a liquid medium from which biomolecules are to be separated. The HGMS-based device further comprises a magnetic source 210 disposed in a proximity of the chamber 202, configured to apply a magnetic field to the liquid medium contained in the chamber 202.

[0055] The chamber 202 comprises an inlet 204 and an outlet 206. The inlet 204 is in fluid communication with a supply unit 250 for supplying the liquid medium. The supply unit 250 comprises a container 251 for containing the liquid medium and a stirrer 252. Here, the liquid medium comprises cell lysate 240 into which magnetic nanoparticles 230 are added. The cell lysate 240 comprises proteins that are fused with peptide tags having strong affinity with the magnetic nanoparticles 230. The stirrer 252 is used to stir the liquid medium to achieve a uniform dispersion. The stirring further helps the combination of the proteins with the magnetic nanoparticles 230.

[0056] The cell lysate 240 together with the magnetic nanoparticles 230, i.e. the liquid medium, are introduced into the chamber 202 via the inlet 204. In the meantime, the outlet 206 is closed so that the chamber 202 contains the liquid medium in its volume. The magnetic source 210 then applies a magnetic field across the chamber 202. High-gradient magnetic fields are created in the vicinity of the matrix 212. These fields extend at least from the volume of the chamber to the matrix 212. Therefore, the magnetic nanoparticles 230 will be attracted to the matrix 212. That is, a surface of the matrix 212 defines a collecting area for the magnetic nanoparticles 230 (or for the biomolecules).

[0057] The matrix 212 also functions as a working electrode. As shown in FIG. 2, a counter electrode is set on the chamber wall 203. Thereby, the working electrode and the counter electrode are capable of generating an electric field directing from the collecting area to the rest of the volume of the chamber (or vice versa). After the magnetic nanoparticles 230 are collected on the matrix 212, an electric potential is applied to the matrix 212 to repulse the biomolecules from the magnetic nanoparticles 230. The outlet 206 is then opened to allow the biomolecules to flow out and then be collected.

[0058] A skilled person will appreciate that many changes may be made to the details of the above-described embodiments and implementations without departing from the underlying principles thereof. The scope of the present invention should, therefore, be determined by the following claims.

[0059] The following aspects concern some embodiments of the invention:

1. A method for separating biomolecules from a liquid medium, comprising:

[0060] adding magnetic nanoparticles to the liquid medium comprising the biomolecules, the biomolecules each adapted to bind to respective surfaces of the magnetic nanoparticles;

[0061] bringing the liquid medium to which the magnetic nanoparticles have been added into contact with a collector;

[0062] applying a magnetic field to the liquid medium in contact with the collector to attract the magnetic nanoparticles bound with the biomolecules to a surface of the collector; and applying an electric potential to the surface of the collector to release the biomolecules from the magnetic nanoparticles.

2. The method of aspect 1, further comprising collecting the biomolecules released from the magnetic nanoparticles.
3. The method of aspect 1 or 2, wherein bringing the liquid medium to which the magnetic nanoparticles have been added into contact with the collector comprises:

[0063] allowing the liquid medium to which the magnetic nanoparticles have been added to pass through a flow cell comprising a chamber and the collector, the collector being disposed in the chamber.

4. The method of any of the preceding aspects, further comprising, before the application of the electric potential,

[0064] separating the collector from the liquid medium; and

[0065] bringing at least part of the collector into contact with another liquid medium.

5. The method of any of the preceding aspects, wherein the magnetic nanoparticles comprise iron oxide nanoparticles, preferably superparamagnetic iron oxide nanoparticles (SPION).
6. The method of any of the preceding aspects, wherein the biomolecules comprise proteins fused with peptide tags, preferably the method further comprising attaching the peptide tags to the proteins.
7. The method of aspect 6, wherein the peptide tags comprise charged peptide tags, preferably negatively charged peptide tags.
8. The method of any of aspects 6-7, wherein the peptide tags comprise negatively charged peptide tags and the method comprises applying a negative electric potential to the collector to repulse the proteins fused with the negatively charged peptide tags, thereby releasing the biomolecules from the magnetic nanoparticles.
9. The method of any of the preceding aspects, further comprising removing the magnetic field to release the magnetic nanoparticles from the collector, the method preferably further comprising collecting the magnetic nanoparticles released from the collector.
10. A flow cell for separating biomolecules from a liquid medium, the flow cell comprising:

[0066] a chamber comprising [0067] an inlet and an outlet which define therebetween a fluid path for the liquid medium, and [0068] a volume for containing the liquid medium provided along said fluid path, the volume comprising a collecting area for biomolecules;

[0069] a magnetic source proximal to the volume of the chamber, the magnetic source arranged and configured to generate a magnetic field extending at least between the collecting area and the remaining volume for containing the liquid medium; and

[0070] a working electrode proximal to the volume for containing the liquid medium, the working electrode arranged and configured to generate an electric field at the collecting area.

11. The flow cell of aspect 10, wherein the magnetic source is a permanent magnet movably coupled to the working electrode and/or to the chamber or an electromagnet.
12. The flow cell of aspect 10 or 11, further comprising

[0071] a first collecting unit in fluid communication with the outlet of the chamber for collecting the biomolecules from the liquid medium,

[0072] a second collecting unit in fluid communication with the outlet of the chamber for collecting a magnetic material from the liquid medium, and/or

[0073] a supply unit in fluid communication with the inlet of the chamber for supplying the liquid medium to the chamber.

[0074] 13. The flow cell of any of aspects 10 to 12, wherein the flow cell is a microfluidic chip, and a volume of the chamber ranges from 1μm.sup.3 to 1,000 mm.sup.3, wherein the microfluidic chip preferably further comprises a counter electrode, and the working electrode and the counter electrode are respectively disposed on opposite walls of the chamber.

[0075] 14. The flow cell of any of aspects 10 to 12, wherein the flow cell is an HGMS-based device, and a volume of the chamber ranges from 1,000 mm.sup.3 to 10 m.sup.3.

[0076] 15. The flow cell of aspects 14, wherein the working electrode is a matrix of the HGMS-based device, the matrix being disposed substantially along a central axis of the chamber and comprising a ferromagnetic material, and the HGMS-based device further comprises a counter electrode disposed on the chamber.