A quasi-two-dimensional cell trajectory magnetic regulation method and system

Abstract

This invention discloses a quasi-two-dimensional cell trajectory magnetic control method and system. The method involves introducing a fluid containing magnetically labeled cells into the microchannel of a quasi-two-dimensional cell trajectory magnetic control chip through a sample inlet. The chip has soft magnetic tape parallel to the microchannel on its upper and lower sides of the microchannel plane. The control boundary of the soft magnetic tape forms an angle θ with the non-magnetic fluid flowing from the sample inlet towards the residual liquid outlet in the microchannel, where 0° < θ ≤ 90°. An external uniform magnetic field is activated to magnetize the soft magnetic tape on both sides. Then, based on the magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid entering the microchannel... f The regulation was adjusted so that the magnetically labeled cells moved along the regulatory boundary and parallel to the soft magnetic tape at the regulatory boundary; where k = 95403.3 mT·s 2 / mm 2 0°<θ≤90°, B0=21.6mT. This method achieves precise control of the anti-adherent quasi-two-dimensional magnetic bead-cell trajectory.

Description

Technical Field

[0001] This invention relates to a quasi-two-dimensional cell trajectory magnetic control method and system, belonging to the field of microfluidics technology. Background Technology

[0002] Biomagnetic sorting is commonly used in biological fields such as virus detection, cell manipulation, and drug screening, and has important applications in clinical medicine. Medical products derived from its industrial applications are also frequently used for health monitoring. Magnetophoresis antibody detection technology typically involves first modifying the surface of magnetic beads with antibodies corresponding to surface antigens of certain viruses or cell proteins. This allows the magnetic beads-antibodies to specifically bind to biomolecules and even cell surface proteins. Finally, a magnetic field is used to regulate and control the movement of the magnetic beads in a flow field, dragging biomolecules or cells for transport, thereby achieving biofluidic sorting.

[0003] Currently, the combination of magnetophoresis technology and microfluidics typically involves placing a magnet on one side of the microfluidic channel. The magnet attracts and alters the trajectory of the magnetic beads, achieving sorting. However, since unilaterally controlled microfluidic chips rely primarily on adsorption, they suffer from significant bead adhesion, leading to channel blockage and affecting sorting accuracy and flow rate. To minimize bead adhesion and ensure effective magnetic field control, magnetically modulated microfluidic chips are typically used for low-velocity magnetic sorting. Because the magnetic field around the magnet dissipates rapidly, the magnetic flux density and magnetic gradient in the direction of the external magnetic field normal decrease quickly. This results in significant differences in the magnetic force applied to beads originally located at different positions in the channel, making it impossible to achieve uniform, stable, and precise control of the magnetic particles. Furthermore, existing magnets apply a fixed magnetic field shape and size in the flow field, requiring adjustments only by changing the magnet's material composition, which limits magnetic field adjustment and makes magnet manufacturing processes cumbersome. Summary of the Invention

[0004] Therefore, the technical problem to be solved by the present invention is to overcome the defect that existing microfluidic chips cannot achieve stable and precise trajectory control of magnetic beads in the flow channel, thereby providing a quasi-two-dimensional cell trajectory magnetic control method and a quasi-two-dimensional cell trajectory magnetic control system.

[0005] The technical solution of the present invention:

[0006] A quasi-two-dimensional cell trajectory magnetic modulation method includes the following steps:

[0007] The fluid to be sorted, containing cells labeled with magnetic beads, is introduced into the microchannel of the quasi-two-dimensional cell trajectory magnetic control chip from the sample inlet. In the quasi-two-dimensional cell trajectory magnetic control chip, soft magnetic materials parallel to the microchannel are respectively arranged on the upper and lower sides of the plane where the microchannel is located. The control boundary of the soft magnetic material forms an angle θ with the non-magnetic fluid flowing in the microchannel along the sample inlet towards the residual liquid outlet. Wherein, 0°<θ≤90°.

[0008] An external uniform magnetic field is activated to magnetize the soft magnetic tape on both sides, and then according to... The magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid entering the microchannel. f The regulation was adjusted so that the magnetically labeled cells moved along the regulatory boundary and parallel to the soft magnetic tape at the regulatory boundary; where k = 95403.3 mT·s 2 / mm 2 B0 = 21.6mT.

[0009] When adjusting the external uniform magnetic field at a fixed flow rate, the process also includes a step of adjusting within the range of magnetic flux density B ± 15%; preferably, the adjustment is performed within the range of magnetic flux density B ± 10%.

[0010] The flow velocity v f The flow rate is 0.012–0.035 mm / s, and the magnetic flux density B is 25–80 mT; preferably, the flow velocity v f The magnetic flux density B is 32-55 mT, and the magnetic flux density is 0.015-0.025 mm / s.

[0011] The control boundary is any one or a combination of several of the following: oblique line, arc, straight line, and broken line.

[0012] The included angle θ is 20° to 45°.

[0013] The preparation process of the magnetic bead-labeled cells is as follows: the magnetic bead solution and the cell solution to be labeled are incubated in a buffer solution environment to obtain magnetic bead-labeled cells after the cells to be labeled are magnetically labeled.

[0014] The concentration of the cell solution to be labeled is 1–2.5 × 10⁻⁶. 5 The magnetic beads are in a 4:10 ratio to the number of cells to be labeled; or the magnetic beads are incubated at 35:40°C for 30:40 min; or the buffer solution is in a 5:1 to 8:1 ratio to the cell solution to be labeled.

[0015] A quasi-two-dimensional cell trajectory magnetic control system, used in the aforementioned quasi-two-dimensional cell trajectory magnetic control method, includes a quasi-two-dimensional cell trajectory magnetic control chip, an external uniform magnetic field, and a fluid injection device;

[0016] The quasi-two-dimensional cell trajectory magnetic control chip includes: flexible magnetic tapes parallel to the microchannel are respectively disposed on the upper and lower sides of the plane where the microchannel is located, and the upper and lower flexible magnetic tapes are the same size and shape and are aligned in position; the flow field of the microchannel includes a liquid inlet, a control region and a liquid outlet in sequence along the flow direction of the fluid; the liquid inlet includes a sample inlet; the liquid outlet includes a magnetic separation outlet and a residual liquid outlet; the flow direction of the fluid between the sample inlet and the residual liquid outlet is the non-magnetic fluid flow direction in the flow field, and there is an angle between the sample inlet and the magnetic separation outlet and the non-magnetic fluid flow direction; the flexible magnetic tape is disposed at the location of the control region, and the flexible magnetic tape has a control boundary extending from the adjacent sample inlet to the adjacent magnetic separation outlet, and the angle between the control boundary and the non-magnetic fluid flow direction is θ, 0°<θ≤90°;

[0017] The fluid injection device is connected to the sample inlet and is used to introduce the fluid containing magnetically labeled cells into the microchannel for sorting; the external uniform magnetic field is used to magnetize the soft magnetic tape on the upper and lower sides.

[0018] The straight-line distance d between the two flexible magnetic tapes is ≤500μm; wherein the straight-line distance between the flexible magnetic tape and the surface of the microchannel on the same side is 190-215μm, and the height of the microchannel is 50-70μm; and / or, the ratio of the height to the width of the microchannel is 1:(20-40); and / or, the flexible magnetic tape is an etched amorphous metal tape with a thickness of 20-50μm; more preferably, the amorphous metal tape is made of at least one of iron oxide, cobalt oxide, and nickel oxide; and / or, the microchannel is formed by stacking two templates, one side of which is provided with a microchannel pattern, the microchannel pattern being located between the two templates after stacking and forming a microchannel; preferably, the template is a soft template, more preferably, the soft template is made of PDMS (polydimethylsiloxane) or PMMA (polymethyl methacrylate).

[0019] The fluid injection device is a thrust injection pump; and / or, the external uniform magnetic field is a double-coil Helmholtz coil, the chip is located at the center of the two coils, and each of the soft magnetic tapes is located at the axial center of its corresponding Helmholtz coil.

[0020] Beneficial technical effects of the present invention

[0021] 1. A quasi-two-dimensional cell trajectory magnetic control method of the present invention, wherein the angle θ between the control boundary of the flexible magnetic material and the flow direction of the non-magnetic fluid in the flow field is fixed in the quasi-two-dimensional cell trajectory magnetic control chip, according to... Adjusting the magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid in the microchannel f The force exerted on the magnetic beads in the control chip causes the magnetic force (F) at the control boundary to increase.磁 ) and fluid drag (F 曳 The combined force approximates the regulatory boundary, thereby controlling the trajectory of the magnetic beads, ultimately causing the magnetic beads to drag the cells (V). 细胞 It moves along a set trajectory (the control boundary of the soft magnetic tape), such as... Figure 6 As shown, the magnetic adsorption and adhesion of the magnetic beads is weakened by the competitive capture of the magnetic magnetic material on both the upper and lower sides by the magnetized soft magnetic material. This results in the magnetically labeled cells in the microchannel moving very little along the channel height, but rather moving along the flow field direction parallel to the soft magnetic material, thus preventing the magnetic beads from adhering to the wall and achieving high-throughput magnetic sorting and efficient sorting functions. In summary, the control method of this invention makes the force direction of the magnetic beads tend to be quasi-two-dimensional through spatial adjustment of the magnetic field, and realizes a precise magnetic control method for the quasi-two-dimensional magnetic bead-cell trajectory to prevent adhesion, providing a better solution for the next generation of flow cytometry detection based on magnetic sorting.

[0022] 2. It also includes the step of adjusting the magnetic flux density of the external uniform magnetic field within the range of B±15% under a fixed flow rate, which can ensure that the magnetic bead moves along the control boundary at the control boundary of the soft magnetic tape.

[0023] 3. Optimal flow velocity v f The flow rate is 0.012–0.035 mm / s, and the magnetic flux density B is 25–80 mT; more preferably, the flow velocity v f The magnetic flux density (B) ranges from 0.015 to 0.025 mm / s and from 32 to 55 mT. This enables the regulation of magnetic bead-cell trajectories under high-intensity magnetic fields and high flow rates, achieving high-throughput, high-efficiency, and stable magnetic sorting.

[0024] 4. The control boundary in this invention is any one or a combination of several of the following: oblique line, arc, straight line, and broken line. By changing the boundary morphology of the soft magnetic material, the movement trajectory of the magnetically labeled cells is controlled, thereby achieving precise control of the cell trajectory pattern.

[0025] 5. The included angle θ in this invention is preferably 20° to 45°. Within this angle range, it is convenient for the quasi-two-dimensional magnetic field to fully control the particles in the flow field.

[0026] 6. This invention provides a quasi-two-dimensional cell trajectory magnetic control system. The soft magnetic materials on the upper and lower sides of the chip, under the magnetization of an external uniform magnetic field, regulate the magnetic field within the microchannel. The competitive capture of the magnetic beads by the two soft magnetic materials weakens the vertical force on the magnetic beads in the microchannel, preventing them from adhering to the wall and controlling the force on the magnetic beads to be primarily horizontal. The magnetic field distribution is uneven at different locations in the microchannel, with the strongest magnetic force pointing inwards at the control boundary of the soft magnetic materials. Therefore, by adjusting the external uniform magnetic flux density and the fluid velocity, the force on the magnetic beads is adjusted, causing the magnetic beads and cells to move along the control boundary. Thus, the control system of this invention can achieve precise control of the magnetic bead-cell movement trajectory on a quasi-two-dimensional plane.

[0027] 7. The straight-line distance d between the two flexible magnetic tapes is ≤500μm; wherein, the straight-line distances d1 and d2 between the flexible magnetic tape and the surface of the microchannel on the same side are 190-215μm, and the height d3 of the microchannel is 50-70μm. The distances d1 and d2 between the flexible magnetic tape and the surface of the microchannel are adjusted according to the characteristics of the flexible magnetic tape, thereby adjusting the distance d. Flexible magnetic tapes with high permeability and thinness require larger distances d1 and d2, while flexible magnetic tapes with low permeability and thickness require smaller distances d1 and d2. d ≤500μm and d1 and d2 of 190-215μm ensure precise trajectory control of the magnetic beads in the microchannel by the magnetized flexible magnetic tape, even achieving stable high-throughput magnetic bead guidance within a range of several millimeters at micron resolution in the flow field. Furthermore, a microchannel height of 50-70μm concentrates the magnetic beads in the efficient control area while meeting the high throughput of particles in the microchannel, avoiding any impact on control efficiency. In summary, by comprehensively setting the above three distances, the magnetic control zone formed by the soft magnetic tape on both sides can achieve stable adjustment in both the vertical and horizontal directions, thereby enabling high-throughput, high-velocity, and highly stable anti-wall-adhesion quasi-two-dimensional magnetic bead trajectory control.

[0028] Commonly used flexible magnetic tape models include MATS-2010S, ZC600, 1J46, and VAC17. The aforementioned flexible magnetic tape is an amorphous metal tape with a thickness of 20–50 μm, which reduces the difficulty of forming the flexible magnetic tape, ensures its rigidity, and prevents deformation; simultaneously, it weakens the attenuation of magnetic flux density at the control boundary of the flexible tape, ensuring the magnetic control effect at the control boundary.

[0029] Both templates forming the microchannel are soft templates, which makes it easier to control the straight-line distance between the soft magnetic material and the surface of the microchannel on the same side. In addition, the soft templates are tough, not easy to break, and have good biocompatibility.

[0030] 8. The system in this invention uses a Helmholtz coil as a magnetic field source, and the field strength value is flexible and controllable to meet the different magnetic field strength requirements of chips with different angles. Attached Figure Description

[0031] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0032] Figure 1 This is a schematic diagram of a quasi-two-dimensional cell trajectory magnetic control system according to the present invention;

[0033] Figure 2 This is a schematic diagram of a quasi-two-dimensional cell trajectory magnetic control chip according to Embodiment 1 of the present invention;

[0034] Figure 3 yes Figure 2 AA diagram;

[0035] Figure 4 This is a schematic diagram of the quasi-two-dimensional cell trajectory magnetic control chip structure in Example 2;

[0036] Figure 5 This is a schematic diagram of the quasi-two-dimensional cell trajectory magnetic control chip structure in Example 3;

[0037] Figure 6 This is a top view of the force analysis of magnetically labeled cells in the microchannel at the control boundary of the flexible magnetic material in the quasi-two-dimensional cell trajectory magnetic control chip of the present invention;

[0038] Figure 7 These are diagrams showing the fluid sorting results in the microchannels in Embodiments 4-11 of the present invention; Figure 7 (a) Fluid microscopy observation of the magnetic separation outlet, with black particles representing magnetically labeled cells; Figure 7 (b) Fluid microscopy observation of the residual liquid outlet;

[0039] Figure 8 This is the fluid distribution in the microchannel in Comparative Example 1 of the present invention;

[0040] Figure 9 This is a diagram showing the fluid sorting results in the microchannel of Comparative Example 2 of this invention;

[0041] Figure 9 (a) is a fluid microscopy observation of the residual liquid outlet, where black particles represent magnetically labeled cells; Figure 9 (b) is a fluid microscope observation of the magnetic separation outlet.

[0042] 1-Glass slide, 2-Template, 3-Soft magnetic material, 4-Microchannel, 5-Encapsulation structure, 6-Magnetic bead, 7-Buffer solution inlet, 8-Sample inlet, 9-Magnetic separation outlet, 10-Residual liquid outlet, 11-Control region, 12-Control boundary, 13-Quasi-two-dimensional cell trajectory magnetic control chip, 14-External uniform magnetic field, 15-Fluid injection device, 16-In-situ observation platform, 17-CCD, 18-Microscope objective, 19-Cell, 20-Computer. Detailed Implementation

[0043] Example 1

[0044] A quasi-two-dimensional cell trajectory magnetic control system, such as Figure 1 As shown, the device includes a quasi-two-dimensional cell trajectory magnetic control chip 13, an external uniform magnetic field 14, and a fluid injection device 15. The fluid injection device 15 provides power to the fluid in the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13, causing the fluid containing magnetically labeled cells in the syringe to flow into the microchannel 4. In this embodiment, the fluid injection device 15 is a thrust injection pump. The external uniform magnetic field 14 provides a locally uniform magnetic field when energized to magnetize the soft magnetic tape material 3 on both the upper and lower sides of the quasi-two-dimensional cell trajectory magnetic control chip 13. In this embodiment, the external uniform magnetic field 14 is a double-coil Helmholtz coil, with the quasi-two-dimensional cell trajectory magnetic control chip 13 located at the center between the two coils, and the soft magnetic tape material 3 located at the axis of the Helmholtz coil.

[0045] like Figure 2 As shown, in the quasi-two-dimensional cell trajectory magnetic control chip 13, flexible magnetic materials 3 parallel to the microchannel 4 are respectively arranged on the upper and lower sides of the plane where the microchannel 4 is located. The upper and lower flexible magnetic materials 3 are the same size and shape and are aligned. The microchannel 4 is composed of two templates 2 stacked together. The surface of one side of the template 2 located on the lower side has a microchannel pattern. The microchannel pattern is located between the two stacked templates 2 and constitutes the microchannel. The microchannel 4 is located at the center position between the two flexible magnetic materials 3. The straight-line distance d between the two flexible magnetic materials 3 is 450 μm, the height d3 of the microchannel 4 is 70 μm, and the straight-line distances d1 and d2 between the two flexible magnetic materials 3 and the surface of the microchannel on the same side are both 190 μm. The height to width ratio of the microchannel 4 is 1:20. The material of both templates 2 is PDMS (polydimethylsiloxane).

[0046] like Figure 3As shown, the flow field of the microchannel 4, along the fluid flow direction, sequentially includes a liquid inlet, a control region 11, and a liquid outlet; the liquid inlet includes a buffer inlet 7 and a sample inlet 8; the liquid outlet includes a magnetic separation outlet 9 and a residual liquid outlet 10. The control region 11 is a rectangular region. The buffer inlet 7 and the sample inlet 8 are spaced apart along the width direction of the microchannel 4 on the inlet side of the control region 11, and the magnetic separation outlet 9 and the residual liquid outlet 10 are spaced apart along the width direction of the microchannel 4 on the outlet side of the control region 11. The line connecting the sample inlet 8 and the residual liquid outlet 10 is a straight line, and the fluid flow direction between them is the non-magnetic fluid flow direction in the flow field. Figure 3 As indicated by the arrows, the non-magnetic fluid flows in a straight line along the flow field of the microchannel 4, i.e., the non-magnetic fluid flows from the sample inlet 8 through the control region 11 to the residual liquid outlet 10 in a straight line. The sample inlet 8 and the magnetic separation outlet 9 are positioned intersectingly, and there is an angle between the sample inlet 8 and the magnetic separation outlet 9 and the non-magnetic fluid flow direction. The flexible magnetic tape 3 is located at the control region 11, and the flexible magnetic tape 3 has a control boundary 12 extending from the adjacent sample inlet 8 to the adjacent magnetic separation outlet 9. The boundary 12 is oblique in shape, and the angle θ between the control boundary 12 and the non-magnetic fluid flow direction in the flow field is 30°. The size of the control region 11 is 5mm × 5mm. A large number of flexible magnetic tape units constitute the overall flexible magnetic tape 3, and the size of the overall flexible magnetic tape 3 is slightly smaller than that of the control region 11. Any flexible magnetic tape unit can be present or etched away as needed to form a control boundary of a specific shape. The minimum size of each flexible magnetic tape unit is 50μm × 50μm.

[0047] To enable real-time observation of the fluid in the microchannel 4, the control system includes an in-situ observation platform 16. The in-situ observation platform 16 has a hollow structure, which can accommodate the microscope objective 18 and the CCD 17 (charge-coupled device, also known as an image controller). The CCD 17 is located at the top of the in-situ observation platform 16 and connected to the computer 20; the microscope objective 18 is located at the bottom of the in-situ observation platform 16, which is situated above the quasi-two-dimensional cell trajectory magnetic control chip 13.

[0048] The flexible magnetic tape 3 is an amorphous metal tape with a thickness of 50 μm. The amorphous metal tape is bonded to the substrate and etched into shape. The flexible magnetic tape model is MATS-2010S, and the substrate is a glass plate with a thickness of 1 mm.

[0049] like Figure 2 As shown, in order to improve the overall strength of the chip, the chip also includes a packaging structure 5. The packaging structure 5 is disposed on the upper and lower sides of the plane where the microchannel 4 is located. The soft magnetic tape 3 on each side is located between the packaging structure 5 and the surface of the microchannel 4 on the same side. The material of the packaging structure 5 is PDMS, and the total thickness from the top of the upper packaging structure 5 to the bottom of the lower packaging structure 5 is 3.5mm.

[0050] To ensure the rigidity of the core components of the magnetic control chip, the chip also includes a support sheet, which is a 1mm thick glass sheet 1, respectively disposed on the lower packaging structure 5 and the upper packaging structure 5. The overall thickness of the chip structure is 5.50mm. Thus, the chip forms a multi-layer structure of glass sheet-PDMS packaging structure-flexible magnetic tape-PDMS-microfluidic channel-PDMS-flexible magnetic tape-PDMS packaging structure-glass sheet.

[0051] Example 2

[0052] This embodiment differs from Embodiment 1 in that it adjusts the angle of boundary 12. For example... Figure 4 As shown, in this embodiment, the angle θ between the control boundary 12 of the soft magnetic tape 3 and the flow direction of the non-magnetic fluid in the flow field is 45°.

[0053] Example 3

[0054] This embodiment differs from Embodiment 1 in that it adjusts the angle of boundary 12. For example... Figure 5 As shown, in this embodiment, the angle θ between the control boundary 12 of the soft magnetic tape 3 and the flow direction of the non-magnetic fluid in the flow field is 60°.

[0055] Example 4

[0056] A quasi-two-dimensional cell trajectory magnetic modulation method, employing the quasi-two-dimensional cell trajectory magnetic modulation system described in Example 1, includes the following steps:

[0057] (1) Magnetic labeling of tumor cells

[0058] The concentration used is 4×10 8 5 μl of EpCAM-labeled magnetic beads per ml were added to 1000 μl of 50 mol / L PBS buffer solution and reacted with a concentration of 2 × 10⁶. 5 K562 cells were incubated at 37°C for 30 min at a concentration of 1.5 × 10⁶ / ml and a volume of 800 μl, respectively, to achieve a cell concentration of 1.5 × 10⁶ / ml. 5 / mL.

[0059] (2) Regulation of quasi-two-dimensional trajectory of tumor cells

[0060] The magnetically labeled tumor cells obtained in (1) were introduced into the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13 through the sample inlet; the Helmholtz coil was activated to magnetize the soft magnetic tape 3 on both sides, according to... The magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid entering the microchannel 4 fThe parameters were adjusted so that the magnetically labeled tumor cells moved along the control boundary 12 at the control boundary 12 and parallel to the soft magnetic tape 3, and flowed out along the magnetic separation outlet 9; where k = 95403.3 mT·s 2 / mm 2 θ = 30°, B0 = 21.6 mT. The magnetic flux density at the center of the Helmholtz coil is 30 mT, and the fluid velocity v f The calculated value is 0.0188 mm / s, the fluid velocity is 0.0170 mm / s, and the error is -9.4%.

[0061] Example 5

[0062] A quasi-two-dimensional cell trajectory magnetic modulation method, employing the quasi-two-dimensional cell trajectory magnetic modulation system described in Example 1, includes the following steps:

[0063] (1) Magnetic labeling of tumor cells

[0064] The concentration used is 4×10 8 6 μl of EpCAM-labeled magnetic beads per ml were added to 1000 μl of 50 mol / L PBS buffer solution and reacted with a concentration of 2 × 10⁶. 5 K562 cells were incubated at 37°C for 30 min at a concentration of 2×10⁶ / ml at a volume of 800 μl. 5 / mL.

[0065] (2) Regulation of quasi-two-dimensional trajectory of tumor cells

[0066] The magnetically labeled tumor cells obtained in (1) were introduced into the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13; the Helmholtz coil was activated to magnetize the soft magnetic tape 3 on both sides, according to... The magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid entering the microchannel 4 f The parameters were adjusted so that the magnetically labeled tumor cells moved along the control boundary 12 at the control boundary 12 and parallel to the soft magnetic tape 3, and flowed out along the magnetic separation outlet 9; where k = 95403.3 mT·s 2 / mm 2 θ=30°, B0=21.6mT, the magnetic flux density of the magnetic field at the center of the Helmholtz coil is 40mT, the calculated fluid velocity is 0.0278mm / s, the actual fluid velocity is 0.0260mm / s, and the error is -6.4%.

[0067] Example 6

[0068] A quasi-two-dimensional cell trajectory magnetic modulation method, employing the quasi-two-dimensional cell trajectory magnetic modulation system described in Example 1, includes the following steps:

[0069] (1) Magnetic labeling of tumor cells

[0070] The concentration used is 4×10 8 6 μl of EpCAM-labeled magnetic beads per ml were added to 1000 μl of 50 mol / L PBS buffer solution and reacted with a concentration of 2 × 10⁶. 5 K562 cells were incubated at 37°C for 30 min at a concentration of 2×10⁶ / ml at a volume of 800 μl. 5 / mL.

[0071] (2) Regulation of quasi-two-dimensional trajectory of tumor cells

[0072] The magnetically labeled tumor cells obtained in (1) were introduced into the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13; the Helmholtz coil was activated to magnetize the soft magnetic tape 3 on both sides, according to... The magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid entering the microchannel 4 f The parameters were adjusted so that the magnetically labeled tumor cells moved along the control boundary 12 at the control boundary 12 and parallel to the soft magnetic tape 3, and flowed out along the magnetic separation outlet 9; where k = 95403.3 mT·s 2 / mm 2 θ=30°, B0=21.6mT, the magnetic flux density of the magnetic field at the center of the Helmholtz coil is 70mT, the calculated fluid velocity is 0.0450mm / s, the actual fluid velocity is 0.0470mm / s, and the error is +4.3%.

[0073] Example 7

[0074] A quasi-two-dimensional cell trajectory magnetic modulation method, employing the quasi-two-dimensional cell trajectory magnetic modulation system described in Example 1, includes the following steps:

[0075] (1) Magnetic labeling of tumor cells

[0076] The concentration used is 4×10 8 6 μl of EpCAM-labeled magnetic beads per ml were added to 1000 μl of 50 mol / L PBS buffer solution and reacted with a concentration of 2 × 10⁶. 5 K562 cells were incubated at 37°C for 30 min at a concentration of 2×10⁶ / ml at a volume of 800 μl. 5 / mL.

[0077] (2) Regulation of quasi-two-dimensional trajectory of tumor cells

[0078] The magnetically labeled tumor cells obtained in (1) were introduced into the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13; the Helmholtz coil was activated to magnetize the soft magnetic tape 3 on both sides, according to... The magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid entering the microchannel 4 f The parameters were adjusted so that the magnetically labeled tumor cells moved along the control boundary 12 at the control boundary 12 and parallel to the soft magnetic tape 3, and flowed out along the magnetic separation outlet 9; where k = 95403.3 mT·s 2 / mm 2 θ=30°, B0=21.6mT, the magnetic flux density of the magnetic field at the center of the Helmholtz coil is 80mT, the calculated fluid velocity is 0.0495mm / s, the fluid velocity is 0.0480mm / s, and the error is -3.0%.

[0079] Example 8

[0080] A quasi-two-dimensional cell trajectory magnetic modulation method, employing the quasi-two-dimensional cell trajectory magnetic modulation system described in Example 2, includes the following steps:

[0081] (1) Magnetic labeling of tumor cells

[0082] The concentration used is 4×10 8 6 μl of EpCAM-labeled magnetic beads per ml were added to 1000 μl of 50 mol / L PBS buffer solution and reacted with a concentration of 2 × 10⁶. 5 K562 cells were incubated at 37°C for 30 min at a concentration of 2×10⁶ / ml at a volume of 800 μl. 5 / mL.

[0083] (2) Regulation of quasi-two-dimensional trajectory of tumor cells

[0084] The magnetically labeled tumor cells obtained in (1) were introduced into the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13; the Helmholtz coil was activated to magnetize the soft magnetic tape 3 on both sides, according to... The magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid entering the microchannel 4 f The parameters were adjusted so that the magnetically labeled tumor cells moved along the control boundary 12 at the control boundary 12 and parallel to the soft magnetic tape 3, and flowed out along the magnetic separation outlet 9; where k = 95403.3 mT·s 2 / mm 2 θ=45°, B0=21.6mT, the magnetic flux density of the magnetic field at the center of the Helmholtz coil is 25mT, the calculated fluid velocity is 0.0084mm / s, the fluid velocity is 0.0080mm / s, and the error is -5.2%.

[0085] Example 9

[0086] A quasi-two-dimensional cell trajectory magnetic modulation method, employing the quasi-two-dimensional cell trajectory magnetic modulation system described in Example 2, includes the following steps:

[0087] (1) Magnetic labeling of tumor cells

[0088] The concentration used is 4×10 8 6 μl of EpCAM-labeled magnetic beads per ml were added to 1000 μl of 50 mol / L PBS buffer solution and reacted with a concentration of 2 × 10⁶. 5 K562 cells were incubated at 37°C for 30 min at a concentration of 2×10⁶ / ml at a volume of 800 μl. 5 / mL.

[0089] (2) Regulation of quasi-two-dimensional trajectory of tumor cells

[0090] The magnetically labeled tumor cells obtained in (1) were introduced into the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13; the Helmholtz coil was activated to magnetize the soft magnetic tape 3 on both sides, according to... The magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid entering the microchannel 4 f The parameters were adjusted so that the magnetically labeled tumor cells moved along the control boundary 12 at the control boundary 12 and parallel to the soft magnetic tape 3, and flowed out along the magnetic separation outlet 9; where k = 95403.3 mT·s 2 / mm 2 θ=45°, B0=21.6mT, the magnetic flux density of the magnetic field at the center of the Helmholtz coil is 70mT, the calculated fluid velocity is 0.0319mm / s, the fluid velocity is 0.0320mm / s, and the error is +0.5%.

[0091] Example 10

[0092] A quasi-two-dimensional cell trajectory magnetic modulation method, employing the quasi-two-dimensional cell trajectory magnetic modulation system described in Example 3, includes the following steps:

[0093] (1) Magnetic labeling of tumor cells

[0094] The concentration used is 4×10 8 6 μl of EpCAM-labeled magnetic beads per ml were added to 1000 μl of 50 mol / L PBS buffer solution and reacted with a concentration of 2 × 10⁶. 5 K562 cells were incubated at 37°C for 30 min at a concentration of 2×10⁶ / ml at a volume of 800 μl. 5 / mL.

[0095] (2) Regulation of quasi-two-dimensional trajectory of tumor cells

[0096] The magnetically labeled tumor cells obtained in (1) were introduced into the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13; the Helmholtz coil was activated to magnetize the soft magnetic tape 3 on both sides, according to... The magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid entering the microchannel 4 f The parameters were adjusted so that the magnetically labeled tumor cells moved along the control boundary 12 at the control boundary 12 and parallel to the soft magnetic tape 3, and flowed out along the magnetic separation outlet 9; where k = 95403.3 mT·s 2 / mm 2 θ=60°, B0=21.6mT, the magnetic flux density of the magnetic field at the center of the Helmholtz coil is 40mT, the calculated fluid velocity is 0.0160mm / s, the fluid velocity is 0.0150mm / s, and the error is -6.5%.

[0097] Example 11

[0098] A quasi-two-dimensional cell trajectory magnetic modulation method, employing the quasi-two-dimensional cell trajectory magnetic modulation system described in Example 3, includes the following steps:

[0099] (1) Magnetic labeling of tumor cells

[0100] The concentration used is 4×10 8 6 μl of EpCAM-labeled magnetic beads per ml were added to 1000 μl of 50 mol / L PBS buffer solution and reacted with a concentration of 2 × 10⁶. 5 K562 cells were incubated at 37°C for 30 min at a concentration of 2×10⁶ / ml at a volume of 800 μl. 5 / mL.

[0101] (2) Regulation of quasi-two-dimensional trajectory of tumor cells

[0102] The magnetically labeled tumor cells obtained in (1) were introduced into the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13; the Helmholtz coil was activated to magnetize the soft magnetic tape 3 on both sides, according to... The magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid entering the microchannel 4 f The parameters were adjusted so that the magnetically labeled tumor cells moved along the control boundary 12 at the control boundary 12 and parallel to the soft magnetic tape 3, and flowed out along the magnetic separation outlet 9; where k = 95403.3 mT·s 2 / mm 2 θ = 60°, B0 = 21.6 mT, the magnetic flux density of the magnetic field at the center of the Helmholtz coil is 70 mT, the calculated fluid velocity is 0.0260 mm / s, the actual fluid velocity is 0.0240 mm / s, and the error is -7.7%.

[0103] Comparative Example 1

[0104] A quasi-two-dimensional cell trajectory magnetic modulation method, employing the quasi-two-dimensional cell trajectory magnetic modulation system described in Example 1, includes the following steps:

[0105] (1) Magnetic labeling of tumor cells

[0106] The concentration used is 4×10 8 5 μl of EpCAM-labeled magnetic beads per ml were added to 1000 μl of 50 mol / L PBS buffer solution and reacted with a concentration of 2 × 10⁶. 5 K562 cells were incubated at 37°C for 30 min at a concentration of 1.5 × 10⁶ / ml and a volume of 800 μl, respectively, to achieve a cell concentration of 1.5 × 10⁶ / ml. 5 / mL.

[0107] (2) Regulation of quasi-two-dimensional trajectory of tumor cells

[0108] The magnetically labeled tumor cells obtained in (1) are introduced into the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13; the Helmholtz coil is activated to magnetize the soft magnetic tape 3 on both sides, and the magnetic flux density of the external uniform magnetic field and the flow rate of the fluid entering the microchannel 4 are adjusted. The magnetic flux density of the magnetic field at the center of the Helmholtz coil is 30mT and the fluid flow rate is 0.0320mm / s.

[0109] Comparative Example 2

[0110] A quasi-two-dimensional cell trajectory magnetic modulation method, employing the quasi-two-dimensional cell trajectory magnetic modulation system described in Example 1, includes the following steps:

[0111] (1) Magnetic labeling of tumor cells

[0112] The concentration used is 4×10 8 6 μl of EpCAM-labeled magnetic beads per ml were added to 1000 μl of 50 mol / L PBS buffer solution and reacted with a concentration of 2 × 10⁶. 5 K562 cells were incubated at 37°C for 30 min at a concentration of 2×10⁶ / ml at a volume of 800 μl. 5 / mL.

[0113] (2) Regulation of quasi-two-dimensional trajectory of tumor cells

[0114] The magnetically labeled tumor cells obtained in (1) are introduced into the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13; the Helmholtz coil is activated to magnetize the soft magnetic tape 3 on both sides, and the magnetic flux density of the external uniform magnetic field and the flow rate of the fluid entering the microchannel 4 are adjusted. The magnetic flux density of the magnetic field at the center of the Helmholtz coil is 40mT and the fluid flow rate is 0.0150mm / s.

[0115] Comparative Example 3

[0116] A quasi-two-dimensional cell trajectory magnetic modulation method, employing the quasi-two-dimensional cell trajectory magnetic modulation system described in Example 2, includes the following steps:

[0117] (1) Magnetic labeling of tumor cells

[0118] The concentration used is 4×10 8 6 μl of EpCAM-labeled magnetic beads per ml were added to 1000 μl of 50 mol / L PBS buffer solution and reacted with a concentration of 2 × 10⁶. 5 K562 cells were incubated at 37°C for 30 min at a concentration of 2×10⁶ / ml at a volume of 800 μl. 5 / mL.

[0119] (2) Regulation of quasi-two-dimensional trajectory of tumor cells

[0120] The magnetically labeled tumor cells obtained in (1) are introduced into the microchannel 4 of the quasi-two-dimensional cell trajectory magnetic control chip 13; the Helmholtz coil is activated to magnetize the soft magnetic tape 3 on both sides, and the magnetic flux density of the external uniform magnetic field and the flow velocity of the fluid entering the microchannel 4 are adjusted. The magnetic flux density B of the magnetic field at the center of the Helmholtz coil is 70mT, and the actual flow velocity of the fluid is 0.0140mm / s.

[0121] Results Analysis

[0122] In Examples 4-11, K562 cells did not move continuously along the non-magnetic fluid flow direction from the sample inlet 8 to the residual liquid outlet 10 in the microchannel. Instead, they moved along the control boundary 12 at the control boundary 12 of the flexible magnetic material 3, and their trajectory was parallel to the flexible magnetic material 3. Finally, the magnetic beads 6-cells 19 flowed out from the magnetic separation outlet 9. Figure 6 , Figure 7 of Figure 7 As shown in (a); however, no magnetic particles or magnetically labeled cells were detected at the residual liquid outlet 10, such as Figure 7 As shown in (b).

[0123] In Comparative Examples 1 and 3, no cells or magnetic beads flowed out from the magnetic separation outlet 9 and the residual liquid outlet 10. Because the horizontal control force of the magnetic field is much greater than the fluid drag force, the magnetic particles cannot detach from the magnetic control region. Therefore, after the flow was stopped, K562 cells and magnetic beads remained at high concentrations within the microchannel under a microscope. Figure 8 As shown.

[0124] In Comparative Example 2, no cells or magnetic beads flowed out from the magnetic separation outlet 9, but instead flowed out from the residual liquid outlet 10. Because both K562 cells and magnetic beads in the microchannel were insufficiently regulated, cell trajectory control was not achieved, and the cells still moved along the flow field's trajectory. Therefore, magnetically labeled cells were detected at the residual liquid outlet 10. Figure 9 of Figure 9 As shown in (a); however, no magnetic particles or magnetically labeled cells could be detected at magnetic separation outlet 9, such as Figure 9 As shown in (b).

[0125] In summary, the control method of the present invention can achieve high-throughput, quasi-two-dimensional anti-adherent magnetic beads-cell trajectory magnetic precision control.

[0126] Obviously, the above embodiments are merely illustrative examples for clear explanation and are not intended to limit the implementation. Those skilled in the art will recognize that other variations or modifications can be made based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations here. However, obvious variations or modifications derived therefrom are still within the scope of protection of this invention.

Claims

1. A quasi-two-dimensional cell trajectory magnetic control method, characterized in that, Includes the following steps: The fluid to be sorted, containing cells labeled with magnetic beads, is introduced into the microchannel of the quasi-two-dimensional cell trajectory magnetic control chip from the sample inlet. In the quasi-two-dimensional cell trajectory magnetic control chip, soft magnetic materials parallel to the microchannel are respectively arranged on the upper and lower sides of the plane where the microchannel is located. The control boundary of the soft magnetic material forms an angle θ with the non-magnetic fluid flowing in the microchannel along the sample inlet towards the residual liquid outlet. Wherein, 20°<θ≤45°. An external uniform magnetic field is activated to magnetize the soft magnetic tape on both sides, and then according to... The magnetic flux density B of the external uniform magnetic field and the flow velocity v of the fluid entering the microchannel. f The regulation was adjusted so that the magnetically labeled cells moved along the regulatory boundary and parallel to the soft magnetic tape at the regulatory boundary; where k = 95403.3 mT·s 2 / mm 2 B0 = 21.6 mT; Flow velocity v f The value is 0.015~0.025 mm / s.

2. The control method according to claim 1, characterized in that, The control boundary is any one or a combination of several of the following: oblique line, arc, straight line, and broken line.

3. The control method according to any one of claims 1-2, characterized in that, The preparation process of the magnetic bead-labeled cells is as follows: the magnetic bead solution and the cell solution to be labeled are incubated in a buffer solution environment to obtain magnetic bead-labeled cells after the cells to be labeled are magnetically labeled.

4. The control method according to claim 3, characterized in that, The concentration of the cell solution to be labeled is 1–2.5 × 10⁻⁶. 5 cells / mL; And / or, the ratio of the number of magnetic beads to the number of cells to be labeled is 4 to 10; And / or, incubate at 35–40°C for 30–40 min; And / or, the ratio of the buffer solution to the cell solution to be labeled is 5:1 to 8:1.

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