CN113893892B - Nanopore based on microfluidic chip and preparation method thereof - Google Patents

Nanopore based on microfluidic chip and preparation method thereof Download PDF

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CN113893892B
CN113893892B CN202111328503.8A CN202111328503A CN113893892B CN 113893892 B CN113893892 B CN 113893892B CN 202111328503 A CN202111328503 A CN 202111328503A CN 113893892 B CN113893892 B CN 113893892B
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CN113893892A (en
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王成法
高琪
潘新祥
李冬青
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Dalian Maritime University
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Abstract

The invention provides a nanopore based on a microfluidic chip and a preparation method thereof, and relates to the technical field of manufacturing of nano devices. The nano-holes are prepared on the micro-fluidic chip; the micro-fluidic chip is formed by bonding a PDMS chip and a glass slide, a micro-channel is arranged in the PDMS chip, the middle section of the micro-channel is a zooming channel, the middle section of the zooming channel is a trapezoidal micro-channel, micro-scale spherical particles are arranged in the trapezoidal micro-channel, and four micro gaps formed between the micro-scale spherical particles and the inner wall of the trapezoidal micro-channel are nano holes. The preparation method of the nanopore is very simple and low in cost; the nanopore can be applied to an RPS detection device based on a microfluidic chip to realize the rapid detection of nano-scale particles or cells.

Description

Nanopore based on microfluidic chip and preparation method thereof
Technical Field
The invention relates to the technical field of manufacturing of nano devices, in particular to a nanopore based on a microfluidic chip and a preparation method thereof.
Background
The micro-fluidic chip can integrate basic operation units of sample preparation, reflection, separation, detection and the like in the biological, chemical and medical analysis process on a chip with a plurality of square centimeters, automatically complete the whole analysis process, and has great application potential in the fields of biology, chemistry, medicine and the like. At present, the materials for manufacturing the microfluidic chip mainly comprise: PDMS, PMMA, and inorganic glass, and the like. Different methods for preparing microstructures by chips of different materials are different, and soft lithography is a common technology for preparing microchannels on PDMS materials.
The particle resistance pulse detection technology (RPS) based on the microfluidic chip is an effective method for realizing micro-nano particle or cell detection by using a narrow detection channel, and has been widely concerned in many fields such as biology, chemistry, medicine and the like at present. Because the amplitude of the electric pulse signal generated at the two ends of the detection channel is closely related to the size of the detected particles or cells, the information such as the size of the detected particles or cells can be obtained by analyzing the amplitude of the electric pulse signal. Generally, the smaller the particle size to be detected, the smaller the RPS detection channel size required. However, the above soft lithography technology has difficulty in processing channels of nanometer scale in PDMS, and cannot prepare RPS detection channels capable of detecting nanoparticles or cells. In view of the above, a simple method for preparing a nanopore is to be proposed to improve the microfluidic chip based RPS detection technology.
Disclosure of Invention
The invention provides a novel nanopore based on a microfluidic chip and a preparation method thereof, and solves the problem that the existing RPS detection technology cannot accurately detect nanoparticles or cells.
In order to achieve the above purpose, the invention adopts the technical scheme that:
the invention provides a nanopore based on a microfluidic chip, which is prepared on the microfluidic chip; the microfluidic chip is formed by bonding a PDMS chip and a glass slide, a microchannel is arranged in the PDMS chip, and two ends of the microchannel are respectively communicated with an inlet sample liquid storage tank and an outlet sample liquid storage tank; the microchannel sequentially comprises a first main channel, a zooming channel and a second main channel from an inlet to an outlet, and the zooming channel is positioned in the middle section of the microchannel; the middle section of the zooming channel is of a trapezoidal structure to form a trapezoidal micro-channel, circular arc grooves are symmetrically arranged on two side wall surfaces of the trapezoidal micro-channel, and the micro-scale spherical particles are embedded into the trapezoidal micro-channel through the grooves and mutually contacted with the groove wall surfaces on two sides of the trapezoidal micro-channel and the upper horizontal wall surface and the lower horizontal wall surface; four tiny gaps are formed between the micro-scale spherical particles and the inner wall of the trapezoidal micro-channel, and the four tiny gaps are the nano-pores.
Preferably, the length of the micro-channel is 1-2cm, the height is 2-5 μm, the specific height is determined by the diameter of the micro-scale spherical particles, and the width of the first main channel and the second main channel of the micro-channel is 20-50 μm.
Preferably, the length of the zoom channel is 50-70 μm.
Preferably, the length of the trapezoidal microchannel is 10-15 μm, the width of the inlet of the trapezoidal microchannel is 5-10 μm, and the width of the outlet of the trapezoidal microchannel is 1-4 μm.
Preferably, the width of the inlet of the trapezoidal microchannel is larger than the diameter of the micro-scale spherical particles, and the width of the outlet of the trapezoidal microchannel is smaller than the diameter of the micro-scale spherical particles, and the specific size is determined by the diameter of the selected micro-scale spherical particles.
Preferably, the inlet sample reservoir and the outlet sample reservoir are both provided with reservoirs.
Preferably, the diameter of the micro-scale spherical particles is 2-5 μm.
In another aspect, the present invention provides a method for preparing the nanopore, the method comprising the following steps:
s1, soft lithography processing: preparing a microchannel on a PDMS chip by using a soft lithography technology;
s2, bonding: preparing an inlet sample liquid storage tank and an outlet sample liquid storage tank on a PDMS chip by using a puncher, and then bonding the PDMS chip with a glass slide to complete the processing of the microfluidic chip;
s3, adding particles: fully mixing and diluting the micro-scale spherical particles with a buffer solution to form a particle mixed solution, adding the particle mixed solution into an inlet sample liquid storage tank, conveying the particles into a trapezoidal microchannel under the action of pressure difference, and slightly extruding the surface of a PDMS chip by using tweezers to clamp the micro-scale spherical particles in the trapezoidal microchannel through a groove;
s4, removing the solution: and (3) sucking the redundant particle mixed solution in the liquid storage tank by using a liquid-transfering gun, flushing the microchannel by using an aqueous solution, wherein four micro gaps formed between the micro-scale spherical particles fixed in the trapezoidal microchannel and the wall surface of the channel are nanopores, and the sizes of the nanopores can be flexibly adjusted by changing the sizes of the micro-scale spherical particles and the trapezoidal microchannel.
Preferably, the buffer is a phosphate buffered saline solution.
Compared with the prior art, the invention has the following beneficial effects:
the preparation method of the nanopore is very simple and low in cost; the invention provides a novel nano-pore based on a micro-fluidic chip and a preparation method thereof.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions of the prior art, the drawings needed to be used in the description of the embodiments or the prior art will be briefly described below, it is obvious that the drawings in the following description are only some embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to the drawings without creative efforts.
FIG. 1 is a schematic diagram of a microfluidic chip according to the present invention;
FIG. 2 is a schematic top view of a microfluidic chip according to the present invention;
fig. 3 is a schematic cross-sectional view of a nanopore based on a microfluidic chip according to the present invention.
In the figure: 1. a PDMS chip; 2. glass slide; 3. an inlet sample reservoir; 4. an outlet sample reservoir; 5. a microchannel; 6. a trapezoidal microchannel; 7. micro-scale spherical particles; 8. a nanopore; 9. a liquid storage pipe; 10. zooming the channel; 11. a trapezoidal microchannel inlet; 12. a trapezoidal microchannel outlet; 13. a first main channel; 14. and a second main channel.
Detailed Description
It should be noted that the embodiments and features of the embodiments of the present invention may be combined with each other without conflict. The present invention will be described in detail below with reference to the embodiments with reference to the attached drawings.
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. The following description of at least one exemplary embodiment is merely illustrative in nature and is in no way intended to limit the invention, its application, or uses. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
It is noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of exemplary embodiments according to the invention. As used herein, a singular form of a schematic drawing also includes the plural form unless the context clearly dictates otherwise, and furthermore, it should be understood that when the terms "comprises" and/or "comprising" are used in this specification, it indicates the presence of the features, steps, operations, devices, components and/or combinations thereof.
The relative arrangement of the components and steps, the numerical expressions and numerical values set forth in these embodiments do not limit the scope of the present invention unless specifically stated otherwise. Meanwhile, it should be understood that the sizes of the respective portions shown in the drawings are not drawn in an actual proportional relationship for the convenience of description. Techniques, methods, and apparatus that are known by one of ordinary skill in the relevant art may not be discussed in detail but are intended to be part of the specification where appropriate. In all examples shown and discussed herein, any particular value should be construed as merely illustrative, and not limiting. Thus, other examples of the exemplary embodiments may have different values. It should be noted that: like reference numbers and letters refer to like items in the following figures, and thus, once an item is defined in one figure, further discussion thereof is not required in subsequent figures.
In the description of the present invention, it is to be understood that the directions or positional relationships indicated by the directional terms such as "front, rear, upper, lower, left, right", "lateral, vertical, horizontal" and "top, bottom", etc., are generally based on the directions or positional relationships shown in the drawings for the convenience of description and simplicity of description, and that these directional terms, unless otherwise specified, do not indicate and imply that the device or element so referred to must have a particular orientation or be constructed and operated in a particular orientation, and therefore should not be considered as limiting the scope of the invention: the terms "inner and outer" refer to the inner and outer relative to the profile of the respective component itself.
A novel nanopore based on a microfluidic chip, as shown in fig. 1-3, a nanopore 8 is prepared on the microfluidic chip; the micro-fluidic chip is formed by bonding a PDMS chip 1 and a glass slide 2, a micro-channel 5 is arranged in the PDMS chip 1, the length of the micro-channel 5 is 1-2cm, the height of the micro-channel is 2-5 mu m, and two ends of the micro-channel 5 are respectively communicated with an inlet sample liquid storage tank 3 and an outlet sample liquid storage tank 4; the micro-channel 5 comprises a first main channel 13, a second zooming channel 10 and a second main channel 14 from an inlet to an outlet in sequence, the zooming channel 10 is positioned at the middle section of the micro-channel 5, the widths of the first main channel 13 and the second main channel 14 are 20-50 micrometers, and the length of the zooming channel is 50-70 micrometers; the middle section of the zooming channel 5 is in a trapezoidal structure to form a trapezoidal micro-channel 6, the length of the trapezoidal micro-channel 6 is 10-15 mu m, the width of the trapezoidal micro-channel inlet 11 is 5-10 mu m, and the width of the trapezoidal micro-channel outlet 12 is 1-4 mu m; arc-shaped grooves are symmetrically arranged on two side wall surfaces of the trapezoidal micro-channel 6, the micro-scale spherical particles 7 are embedded into the trapezoidal micro-channel through the grooves and are in mutual contact with the groove wall surfaces on two sides of the trapezoidal micro-channel and the upper horizontal wall surface and the lower horizontal wall surface, the diameter of the micro-scale spherical particles 7 is 2-5 micrometers, the diameter of the micro-scale spherical particles 7 determines the processing height of the micro-channel 5 and the size of the trapezoidal micro-channel 6, the width of an inlet 11 of the trapezoidal micro-channel is larger than the diameter of the micro-scale spherical particles 7, and the width of an outlet 12 of the trapezoidal micro-channel is smaller than the diameter of the micro-scale spherical particles 7; four tiny gaps are formed between the micro-scale spherical particles 7 and the inner wall of the trapezoidal micro-channel 6, and the four tiny gaps are the nano-pores 8.
A preparation method of a nanopore based on a microfluidic chip comprises the following steps:
s1, soft photoetching: preparing a microchannel 5 as shown in FIGS. 1-3 on a PDMS chip 1 by using a soft lithography technique;
s2, bonding: preparing an inlet sample liquid storage tank 3 and an outlet sample liquid storage tank 4 on a PDMS chip 1 by using a puncher, and then bonding the PDMS chip 1 and a glass slide 2 to complete the processing of the microfluidic chip;
s3, adding particles: fully mixing and diluting the micro-scale spherical particles with Phosphate Buffered Saline (PBS) solution to form a particle mixed solution, adding the particle mixed solution into an inlet sample liquid storage tank 3 through a liquid storage pipe 9, conveying the particles into a trapezoidal microchannel 6 under the action of pressure difference, and slightly extruding the surface of a PDMS chip 1 by using tweezers to clamp the micro-scale spherical particles 7 at the arc-shaped groove of the trapezoidal microchannel 6;
s4, removing the solution: after the micro-scale spherical particles 7 are fixed at the arc-shaped groove in the trapezoidal micro-channel 6, sucking out redundant particle mixed solution in the liquid storage tank by using a liquid-transferring gun, washing the micro-channel 5 by using aqueous solution, and taking four micro gaps formed between the micro-scale spherical particles 7 fixed in the trapezoidal micro-channel 6 and the wall surface of the channel as nano holes 8; the size of the nanometer hole 8 can be flexibly adjusted by changing the sizes of the micro-scale spherical particles 7 and the trapezoidal micro-channel 6.
The invention provides a nanopore preparation method based on a microfluidic chip, and a novel nanopore with a very small size can be obtained by the method. Meanwhile, the size of the novel nanopore can be flexibly adjusted, the preparation principle is very simple, and the operability is strong. The novel nano-pore is applied to the RPS detection device based on the microfluidic chip, and can accurately detect nano-scale particles or cells in a short time.
The above description is only for the preferred embodiment of the present invention, but the scope of the present invention is not limited thereto, and any person skilled in the art should be considered to be within the technical scope of the present invention, and the technical solutions and the inventive concepts thereof according to the present invention should be equivalent or changed within the scope of the present invention.

Claims (6)

1. A nanopore based on a microfluidic chip is characterized in that: the nano-hole is prepared on the microfluidic chip; the microfluidic chip is formed by bonding a PDMS chip (1) and a glass slide (2), a microchannel (5) is arranged in the PDMS chip (1), and two ends of the microchannel (5) are respectively communicated with an inlet sample liquid storage tank (3) and an outlet sample liquid storage tank (4); the microchannel (5) sequentially comprises a first main channel (13), a zooming channel (10) and a second main channel (14) from an inlet to an outlet, and the zooming channel (10) is positioned in the middle section of the microchannel (5); the middle section of the zooming channel (10) is of a trapezoidal structure to form a trapezoidal micro-channel (6), arc-shaped grooves are symmetrically formed in two side wall surfaces of the trapezoidal micro-channel (6), and the micro-scale spherical particles are embedded into the trapezoidal micro-channel (6) through the grooves and mutually contacted with the groove wall surfaces and the upper horizontal wall surfaces and the lower horizontal wall surfaces on two sides of the trapezoidal micro-channel (6); four tiny gaps are formed between the micro-scale spherical particles (7) and the inner wall of the trapezoidal micro-channel (6) and are nano-holes (8);
the length of the trapezoidal micro-channel (6) is 10-15 mu m, the width of the trapezoidal micro-channel inlet (11) is 5-10 mu m, and the width of the trapezoidal micro-channel outlet (12) is 1-4 mu m;
the width of the trapezoid micro-channel inlet (11) is larger than the diameter of the micro-scale spherical particles (7), the width of the trapezoid micro-channel outlet (12) is smaller than the diameter of the micro-scale spherical particles (7), and the specific size is determined by the diameter of the selected micro-scale spherical particles (7);
the diameter of the micro-scale spherical particles (7) is 2-5 mu m.
2. The microfluidic chip-based nanopore according to claim 1, wherein: the length of the micro-channel (5) is 1-2cm, the height is 2-5 mu m, the specific height is determined by the diameter of the micro-scale spherical particles (7), and the width of the first main channel (13) and the second main channel (14) of the micro-channel (5) is 20-50 mu m.
3. The microfluidic chip-based nanopore according to claim 1, wherein: the length of the scaling channel (10) is 50-70 μm.
4. The microfluidic chip-based nanopore according to claim 1, wherein: liquid storage pipes (9) are arranged on the inlet sample liquid storage tank (3) and the outlet sample liquid storage tank (4).
5. A method of making a nanopore according to any of claims 1-4, wherein: the method comprises the following steps:
s1, soft photoetching: preparing a micro-channel (5) on a PDMS chip (1) by using a soft lithography technology;
s2, bonding: preparing an inlet sample liquid storage tank (3) and an outlet sample liquid storage tank (4) on a PDMS chip (1) by using a puncher, and then bonding the PDMS chip (1) and a glass slide (2) to complete the processing of the microfluidic chip;
s3, adding particles: fully mixing and diluting the micro-scale spherical particles with a buffer solution to form a particle mixed solution, adding the particle mixed solution into an inlet sample liquid storage tank (3), conveying the particles into a trapezoidal micro-channel (6) under the action of pressure difference, and slightly extruding the surface of a PDMS chip (1) by using tweezers to clamp the micro-scale spherical particles (7) in the trapezoidal micro-channel (6) through grooves;
s4, removing the solution: and (3) sucking out the redundant particle mixed solution in the liquid storage tank by using a pipette, washing the micro-channel by using aqueous solution, and forming four micro gaps between the micro-scale spherical particles (7) fixed in the trapezoidal micro-channel (6) and the wall surface of the channel to obtain the nano-pores (8).
6. The production method according to claim 5, characterized in that: the buffer solution is phosphate buffer salt solution.
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US6942771B1 (en) * 1999-04-21 2005-09-13 Clinical Micro Sensors, Inc. Microfluidic systems in the electrochemical detection of target analytes
WO2018148501A1 (en) * 2017-02-10 2018-08-16 The Rockefeller University Methods for cell-type specific profiling to identify drug targets
CN109622078A (en) * 2018-12-11 2019-04-16 西安交通大学 A kind of micro-fluidic chip for the single position enrichment of particle in non-newtonian fluid
CN109590038A (en) * 2018-12-29 2019-04-09 天津大学 A kind of sub-micron runner micro-fluidic chip and preparation method thereof

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