Two-dimensional liquid chromatography micro-fluidic chip and manufacturing method thereof

Abstract

The invention relates to the technical field of substance separation and detection, and relates to a two-dimensional liquid chromatography micro-fluidic chip and a manufacturing method thereof. The two-dimensional liquid chromatography micro-fluidic chip comprises a chip body, wherein a channel structure is formed in the chip body; the sample channel is connected with a first end of the one-dimensional cross-shaped sample introduction channel, the one-dimensional mobile phase channel is connected with a second end of the one-dimensional cross-shaped sample introduction channel, a first end of the one-dimensional separation channel is connected with a third end of the one-dimensional cross-shaped sample introduction channel, and the plunger channel is connected with the third end of the one-dimensional cross-shaped sample introduction channel. The two-dimensional mobile phase channel is connected with a first end of the two-dimensional cross-shaped sample introduction channel, a second end of the one-dimensional separation channel is connected with a second end of the two-dimensional cross-shaped sample introduction channel, the two-dimensional separation channel is connected with a third end of the two-dimensional cross-shaped sample introduction channel, and the post-column channel is connected with the two-dimensional separation channel. According to the two-dimensional liquid chromatography micro-fluidic chip and the manufacturing method thereof provided by the invention, the defect that a channel of a quartz chip is difficult to be filled with a chromatographic column in the prior art is overcome.

Description

Technical Field

[0001] The present invention relates to the technical field of substance separation and detection, and in particular to a two-dimensional liquid chromatography microfluidic chip and a manufacturing method thereof. Background Art

[0002] For the separation and detection of certain complex mixtures, very strong separation capabilities are usually required, and traditional one-dimensional separation methods are difficult to meet the separation requirements of such compounds (such as large molecular proteins and peptides). At present, in order to achieve larger peak capacity and higher separation efficiency, multidimensional separation modes are widely used. Among them, two-dimensional liquid chromatography has become an effective method for separating complex samples due to its excellent separation ability and peak capacity. However, traditional two-dimensional liquid chromatography systems usually rely on valve switching to achieve this, which not only makes the system structure complicated, but also introduces a large dead volume, thus failing to meet the separation requirements for precious small-volume samples.

[0003] In comparison, microfluidic chip chromatography analysis technology has shown significant advantages. It can greatly reduce the consumption of precious samples, significantly shorten the analysis time, and has the characteristics of flexible design, adaptability to small-volume rare samples, efficient and rapid separation, and easy miniaturization and portability. Quartz microfluidic chips have become a common preparation material for liquid chromatography chips due to their excellent pressure resistance and good repeatability. However, due to the high rigidity of quartz chips, it is difficult to fill chromatographic columns in the channels of quartz chips in the prior art. Summary of the invention

[0004] The invention provides a two-dimensional liquid chromatography microfluidic chip and a manufacturing method thereof, which are used to solve the defect in the prior art that it is difficult to fill a chromatographic column in a channel of a quartz chip.

[0005] On the one hand, the present invention provides a two-dimensional liquid chromatography microfluidic chip, comprising: a chip body, wherein a channel structure is formed in the chip body, and the channel structure comprises a sample channel, a one-dimensional mobile phase channel, a two-dimensional mobile phase channel, a one-dimensional cross injection channel, a two-dimensional cross injection channel, a one-dimensional separation channel, a two-dimensional separation channel, a plunger channel and a post-column channel.

[0006] The sample channel is connected to the first end of the one-dimensional cross injection channel, the one-dimensional mobile phase channel is connected to the second end of the one-dimensional cross injection channel, the first end of the one-dimensional separation channel is connected to the third end of the one-dimensional cross injection channel, the plunger channel is connected to the third end of the one-dimensional cross injection channel, the two-dimensional mobile phase channel is connected to the first end of the two-dimensional cross injection channel, the second end of the one-dimensional separation channel is connected to the second end of the two-dimensional cross injection channel, the first end of the two-dimensional separation channel is connected to the third end of the two-dimensional cross injection channel, and one end of the post-column channel is connected to the second end of the two-dimensional separation channel.

[0007] According to the two-dimensional liquid chromatography microfluidic chip provided by the present invention, a first plunger is provided at the connection between the plunger channel and the first end of the one-dimensional separation channel, and a second plunger is provided at the connection between the two-dimensional mobile phase channel and the first end of the two-dimensional separation channel; a one-dimensional chromatography filling column is provided in the one-dimensional separation channel, and a two-dimensional chromatography filling column is provided in the two-dimensional separation channel.

[0008] According to the two-dimensional liquid chromatography microfluidic chip provided by the present invention, the third end outlet of the one-dimensional cross injection channel is provided with a first limiting channel for limiting the first plunger, and the fourth end inlet of the two-dimensional cross injection channel is provided with a second limiting channel for limiting the second plunger.

[0009] According to the two-dimensional liquid chromatography microfluidic chip provided by the present invention, the cross-sections of the first limiting channel and the second limiting channel are both semicircular.

[0010] According to the two-dimensional liquid chromatography microfluidic chip provided by the present invention, the diameters of the first limiting channel and the second limiting channel are both 80 μm to 120 μm.

[0011] According to the two-dimensional liquid chromatography microfluidic chip provided by the present invention, the first plunger and the second plunger are both porous silica microspheres.

[0012] According to the two-dimensional liquid chromatography microfluidic chip provided by the present invention, the diameters of the one-dimensional separation channel and the two-dimensional separation channel are both 80 μm to 120 μm.

[0013] According to the two-dimensional liquid chromatography microfluidic chip provided by the present invention, the channel structure also includes a first liquid discharge channel and a second liquid discharge channel, the first liquid discharge channel is connected to the fourth end of the one-dimensional cross injection channel, and the second liquid discharge channel is connected to the fourth end of the two-dimensional cross injection channel.

[0014] According to the two-dimensional liquid chromatography microfluidic chip provided by the present invention, the chip body includes two quartz single chips, the quartz single chips are provided with grooves for forming the channel structure, and the two quartz single chips are arranged opposite to each other and bonded to each other so that the grooves on the two quartz single chips form the channel structure.

[0015] Another aspect of the present invention provides a two-dimensional liquid chromatography microfluidic core and a manufacturing method thereof based on any one of the above items, comprising the following steps.

[0016] Grooves corresponding to the channel structure are respectively made on two quartz monoliths.

[0017] The two quartz monoliths with the grooves are aligned and thermally bonded to form the chip body, so that the grooves of the two quartz monoliths are combined to form the channel structure.

[0018] A first plunger is introduced into the connection between the plunger channel and the first end of the one-dimensional separation channel through the plunger channel, and then a chromatographic filler is filled into the one-dimensional separation channel through the post-column channel to form a one-dimensional chromatographic filling column in the one-dimensional separation channel; a second plunger is introduced into the connection between the two-dimensional mobile phase channel and the first end of the two-dimensional separation channel through the two-dimensional mobile phase channel, and then a chromatographic filler is filled into the two-dimensional separation channel through the post-column channel to form a two-dimensional chromatographic filling column in the two-dimensional separation channel.

[0019] The two-dimensional liquid chromatography microfluidic chip provided by the present invention provides a channel structure in a chip body. When a one-dimensional separation channel is filled with chromatographic fillers to form a one-dimensional chromatographic filling column, a first plunger is first introduced into the connection between the plunger channel and the first end of the one-dimensional separation channel through the plunger channel, and then the chromatographic fillers are filled into the one-dimensional separation channel through the post-column channel to form a one-dimensional chromatographic filling column in the one-dimensional separation channel. When a two-dimensional separation channel is filled with chromatographic fillers to form a two-dimensional chromatographic filling column, a second plunger is introduced into the connection between the two-dimensional mobile phase channel and the first end of the two-dimensional separation channel through the two-dimensional mobile phase channel, and then the chromatographic fillers are filled into the two-dimensional separation channel through the post-column channel to form a two-dimensional chromatographic filling column in the two-dimensional separation channel. That is, the two-dimensional liquid chromatography microfluidic chip provided by the present invention can quickly and easily realize the preparation of one-dimensional chromatographic filling columns and two-dimensional chromatographic filling columns in the chip body, thereby solving the defect that it is difficult to fill chromatographic columns in channels of quartz chips in the prior art.

[0020] Additional aspects and advantages of the present invention will be given in part in the following description and in part will be obvious from the following description, or will be learned through practice of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] In order to more clearly illustrate the technical solutions in the present invention or the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0022] Figure 1 This is one of the schematic diagrams of the two-dimensional liquid chromatography microfluidic chip provided in an embodiment of the present invention.

[0023] Figure 2 It is a schematic diagram of thermal bonding of two quartz monoliths with channel structures in a two-dimensional liquid chromatography microfluidic chip provided by an embodiment of the present invention.

[0024] Figure 3 It is a schematic diagram of a circular cross-section channel in a two-dimensional liquid chromatography microfluidic chip provided in an embodiment of the present invention.

[0025] Figure 4 It is a schematic diagram of a semicircular cross-section channel in a two-dimensional liquid chromatography microfluidic chip provided in an embodiment of the present invention.

[0026] Figure 5 It is a schematic diagram of the process of manufacturing a microfluidic chip provided by an embodiment of the present invention.

[0027] Reference numerals: 100. Chip body; 101. Sample channel; 102. One-dimensional mobile phase channel; 103. Two-dimensional mobile phase channel; 104. One-dimensional cross injection channel; 105. Two-dimensional cross injection channel; 106. One-dimensional separation channel; 107. Two-dimensional separation channel; 108. Plunger channel; 109. Post-column channel; 110. First limiting channel; 111. Second limiting channel; 112. First liquid discharge channel; 113. Second liquid discharge channel. DETAILED DESCRIPTION

[0028] In order to make the purpose, technical solution and advantages of the present invention clearer, the technical solution of the present invention will be clearly and completely described below in conjunction with the drawings of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0029] In the description of the embodiments of the present invention, it should be noted that the terms "center", "longitudinal", "lateral", "up", "down", "front", "back", "left", "right", "vertical", "horizontal", "top", "bottom", "inside", "outside" and the like indicate positions or positional relationships based on the positions or positional relationships shown in the accompanying drawings, and are only for the convenience of describing the embodiments of the present invention and simplifying the description, and do not indicate or imply that the devices or elements referred to must have a specific orientation, be constructed and operate in a specific orientation, and therefore cannot be understood as limitations on the embodiments of the present invention. In addition, the terms "first", "second", and "third" are used for descriptive purposes only and cannot be understood as indicating or implying relative importance.

[0030] In the description of the embodiments of the present invention, it should be noted that, unless otherwise clearly specified and limited, the terms "connected" and "connection" should be understood in a broad sense, for example, it can be a fixed connection, a detachable connection, or an integral connection; it can be a mechanical connection or an electrical connection; it can be a direct connection or an indirect connection through an intermediate medium. For ordinary technicians in this field, the specific meanings of the above terms in the embodiments of the present invention can be understood according to specific circumstances.

[0031] In the embodiments of the present invention, unless otherwise clearly specified and limited, the first feature being "above" or "below" the second feature may mean that the first and second features are in direct contact, or the first and second features are in indirect contact through an intermediate medium. Moreover, the first feature being "above", "above" and "above" the second feature may mean that the first feature is directly above or obliquely above the second feature, or simply means that the first feature is higher in level than the second feature. The first feature being "below", "below" and "below" the second feature may mean that the first feature is directly below or obliquely below the second feature, or simply means that the first feature is lower in level than the second feature.

[0032] In the description of this specification, the description with reference to the terms "one embodiment", "some embodiments", "example", "specific example", or "some examples" etc. means that the specific features, structures, materials or characteristics described in conjunction with the embodiment or example are included in at least one embodiment or example of the embodiment of the present invention. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described may be combined in any one or more embodiments or examples in a suitable manner. In addition, those skilled in the art may combine and combine the different embodiments or examples described in this specification and the features of the different embodiments or examples, without contradiction.

[0033] Combine the following Figures 1 to 5 The invention describes a two-dimensional liquid chromatography microfluidic chip and a method for making the same.

[0034] See also Figure 1 As shown, the two-dimensional liquid chromatography microfluidic chip provided in an embodiment of the present invention includes: a chip body 100, a channel structure is formed in the chip body 100, and the channel structure includes a sample channel 101, a one-dimensional mobile phase channel 102, a two-dimensional mobile phase channel 103, a one-dimensional cross injection channel 104, a two-dimensional cross injection channel 105, a one-dimensional separation channel 106, a two-dimensional separation channel 107, a plunger channel 108 and a post-column channel 109.

[0035] The sample channel 101 is connected to a first end of a one-dimensional cross injection channel 104, the one-dimensional mobile phase channel 102 is connected to a second end of the one-dimensional cross injection channel 104, the first end of a one-dimensional separation channel 106 is connected to a third end of the one-dimensional cross injection channel 104, the plunger channel 108 is connected to the third end of the one-dimensional cross injection channel 104, the two-dimensional mobile phase channel 103 is connected to a first end of a two-dimensional cross injection channel 105, the second end of the one-dimensional separation channel 106 is connected to a second end of the two-dimensional cross injection channel 105, the first end of a two-dimensional separation channel 107 is connected to a third end of the two-dimensional cross injection channel 105, and one end of a post-column channel 109 is connected to a second end of the two-dimensional separation channel 107.

[0036] It should be noted that the first end to the fourth end of the one-dimensional cross injection channel 104 described in this embodiment are Figure 1 Similarly, the first end to the fourth end of the two-dimensional cross injection channel 105 described in this embodiment are Figure 1 The ① to ④ ends shown in the figure.

[0037] The two-dimensional liquid chromatography microfluidic chip provided by the present invention is provided with a channel structure in the chip body 100. When the one-dimensional separation channel 106 is filled with chromatographic fillers to form a one-dimensional chromatographic filling column, the first plunger is first introduced into the connection between the plunger channel 108 and the first end of the one-dimensional separation channel 106 through the plunger channel 108, and then the chromatographic fillers are filled into the one-dimensional separation channel 106 through the post-column channel 109 to form a one-dimensional chromatographic filling column in the one-dimensional separation channel 106, and the chromatographic fillers are filled into the two-dimensional separation channel 107 to form a two-dimensional chromatographic filling column. When preparing the column, a second plunger is introduced into the connection between the two-dimensional mobile phase channel 103 and the first end of the two-dimensional separation channel 107 through the two-dimensional mobile phase channel 103, and then the chromatographic filler is filled into the two-dimensional separation channel 107 through the post-column channel 109 to form a two-dimensional chromatographic filling column in the two-dimensional separation channel 107. That is, the two-dimensional liquid chromatography microfluidic chip provided by the present invention can quickly and easily realize the preparation of one-dimensional chromatographic filling columns and two-dimensional chromatographic filling columns in the chip body 100, which solves the defect that it is difficult to fill chromatographic columns in the channels of quartz chips in the prior art.

[0038] Specifically, in this embodiment, the channel structure includes a sample channel 101, a one-dimensional mobile phase channel 102, a two-dimensional mobile phase channel 103, a one-dimensional cross injection channel 104, a two-dimensional cross injection channel 105, a one-dimensional separation channel 106, a two-dimensional separation channel 107, a plunger channel 108 and a post-column channel 109.

[0039] Among them, the sample channel 101 is used to input the sample solution; the one-dimensional mobile phase channel 102 is used for the one-dimensional mobile phase to flow in, the two-dimensional mobile phase channel 103 is used for the two-dimensional mobile phase to flow in, and is used to introduce a second plunger into the connection between the two-dimensional mobile phase channel 103 and the first end of the two-dimensional separation channel 107, so as to prepare a one-dimensional chromatographic filling column and a two-dimensional chromatographic filling column in the one-dimensional separation channel 106 and the two-dimensional separation channel 107, respectively.

[0040] In the one-dimensional cross injection channel 104 , the solvent and the sample form a stable laminar flow and enter the one-dimensional separation channel 106 to achieve small volume injection. In the two-dimensional cross injection channel 105 , the fraction flowing out of the one-dimensional separation channel 106 and the two-dimensional mobile phase solvent enter the two-dimensional separation channel 107 .

[0041] The one-dimensional separation channel 106 is used to load one-dimensional chromatographic filler for one-dimensional separation of samples, and the two-dimensional separation channel 107 is used to load two-dimensional chromatographic filler for two-dimensional separation of samples.

[0042] The plunger channel 108 is used to introduce a first plunger into the connection between the plunger channel 108 and the first end of the one-dimensional separation channel 106 .

[0043] The post-column channel 109 is used to fill the one-dimensional separation channel 106 and the two-dimensional separation channel 107 with chromatographic fillers and to allow the sample solution to flow out.

[0044] When in use, in the channel structure, the sample channel 101, the one-dimensional mobile phase channel 102, the two-dimensional mobile phase channel 103, the post-column channel 109 and the first liquid discharge channel 112 and the second liquid discharge channel 113 described below are all connected with capillaries (not shown in the figure). The capillaries are used to communicate with corresponding external containers to facilitate the introduction of corresponding liquid media into the channel structure, and valves can be set on the capillaries to open or close the corresponding channels.

[0045] According to some embodiments of the present invention, a first plunger (not shown in the figure) is provided at the connection between the plunger channel 108 and the first end of the one-dimensional separation channel 106, and a second plunger (not shown in the figure) is provided at the connection between the two-dimensional mobile phase channel 103 and the first end of the two-dimensional separation channel 107; a one-dimensional chromatography filling column (not shown in the figure) is provided in the one-dimensional separation channel 106, and a two-dimensional chromatography filling column (not shown in the figure) is provided in the two-dimensional separation channel 107.

[0046] By arranging the first plunger and the second plunger in the channel structure, the plungers can be used to block the response position of the channel structure, so as to facilitate filling the one-dimensional chromatography filling column and the two-dimensional chromatography filling column in the one-dimensional separation channel 106 and the two-dimensional separation channel 107 respectively. After the one-dimensional chromatography filling column and the two-dimensional chromatography filling column are formed in the channel structure, the one-dimensional chromatography filling column and the two-dimensional chromatography filling column can be limited to ensure the stability of the one-dimensional chromatography filling column and the two-dimensional chromatography filling column.

[0047] It should be noted that the first plunger is used as the inlet end plunger of the one-dimensional chromatographic filling column, and the second plunger is used as both the end plunger of the one-dimensional chromatographic filling column and the inlet end plunger of the two-dimensional chromatographic filling column. In addition, the first plunger and the second plunger in this embodiment are both porous plunger structures, which can allow liquid media to flow.

[0048] See also Figure 1 As shown, according to some embodiments of the present invention, the third end outlet of the one-dimensional cross injection channel 104 is provided with a first limiting channel 110 for limiting the first plunger, and the fourth end inlet of the two-dimensional cross injection channel 105 is provided with a second limiting channel 111 for limiting the second plunger.

[0049] By providing the first limiting channel 110 and the second limiting channel 111, the first plunger and the second plunger in the channel structure can be limited respectively to prevent the first plunger and the second plunger from moving in the channel structure while ensuring that the liquid medium can flow smoothly.

[0050] See also Figure 3 and Figure 4 As shown, according to some embodiments of the present invention, the cross-sections of the first limiting channel 110 and the second limiting channel 111 are both semicircular.

[0051] By setting the diameters of the first limiting channel 110 and the second limiting channel 111 to 80 μm to 120 μm, and setting the diameters of the one-dimensional separation channel 106 and the two-dimensional separation channel 107 to 80 μm to 120 μm, the first plunger and the second plunger can be limited by the solid structure at the bottom of the semicircular channel, and the liquid medium can also flow smoothly through the semicircular channel.

[0052] Specifically, the cross-sections of the first limiting channel 110 and the second limiting channel 111 are both semicircular, the cross-sections of the one-dimensional separation channel 106 and the two-dimensional separation channel 107 are both circular, the diameters of the first limiting channel 110 and the second limiting channel 111 can be 80μm, 90μm, 100μm, 110μm or 120μm, etc., and the diameters of the one-dimensional separation channel 106 and the two-dimensional separation channel 107 can be 80μm, 90μm, 100μm, 110μm or 120μm, etc.

[0053] According to some embodiments of the present invention, the first plunger and the second plunger are both porous silica microspheres.

[0054] By setting the first plunger and the second plunger as porous silica microspheres, the porous silica microspheres can be used to limit the chromatographic column, and the pore structure on the porous silica microspheres can be used to provide a flow channel for the liquid medium. In addition, the porous silica microspheres have a simple structure and are easy to be introduced into the corresponding position of the channel structure through the plunger channel. They are stable in nature, are not easy to react with the liquid medium or the chromatographic filler, and have good stability and durability.

[0055] See also Figure 1 As shown, according to some embodiments of the present invention, the channel structure also includes a first liquid discharge channel 112 and a second liquid discharge channel 113, the first liquid discharge channel 112 is connected to the fourth end of the one-dimensional cross injection channel 104, and the second liquid discharge channel 113 is connected to the fourth end of the two-dimensional cross injection channel 105.

[0056] By providing the first liquid discharge channel 112 and the second liquid discharge channel 113 , the liquid medium at the corresponding position in the channel structure can be discharged.

[0057] See also Figure 2 As shown, according to some embodiments of the present invention, the chip body 100 includes two quartz monoliths, on which grooves for forming a channel structure are provided. The two quartz monoliths are arranged opposite to each other and bonded to each other so that the grooves on the two quartz monoliths form a channel structure.

[0058] By configuring the chip body 100 as two quartz monoliths facing each other and bonded to each other, it is possible to easily fabricate a channel structure between the two quartz monoliths.

[0059] The microfluidic chip manufacturing method provided by the present invention is described below. The microfluidic chip manufacturing method described below and the two-dimensional liquid chromatography microfluidic chip described above can be referenced to each other.

[0060] See also Figure 5 As shown, the microfluidic chip manufacturing method provided in the embodiment of the present invention is used to manufacture the two-dimensional liquid chromatography microfluidic chip as described in any of the above embodiments, and includes the following steps.

[0061] S510, making grooves corresponding to the channel structures on two quartz monoliths respectively.

[0062] S520, aligning and thermally bonding two quartz monoliths with grooves to form a chip body 100, so that the grooves on the two quartz monoliths are combined to form a channel structure.

[0063] S530, introducing a first plunger through the plunger channel 108 into the connection between the plunger channel 108 and the first end of the one-dimensional separation channel 106, and then filling the one-dimensional separation channel 106 with chromatographic filler through the post-column channel 109 to form a one-dimensional chromatographic filling column in the one-dimensional separation channel 106, introducing a second plunger through the two-dimensional mobile phase channel 103 into the connection between the two-dimensional mobile phase channel 103 and the first end of the two-dimensional separation channel 107, and then filling the two-dimensional separation channel 107 with chromatographic filler through the post-column channel 109 to form a two-dimensional chromatographic filling column in the two-dimensional separation channel 107.

[0064] The microfluidic chip manufacturing method provided by the present invention can facilitate filling one-dimensional chromatographic filling columns and two-dimensional chromatographic filling columns in the one-dimensional separation channel 106 and the two-dimensional separation channel 107 in the chip body 100, thereby solving the defect that it is difficult to fill chromatographic columns in the channels of quartz chips in the prior art.

[0065] Specifically, in step S510, a quartz chrome plate (quartz monolith) coated with a 500nm thick AZ-1500 positive photoresist and a 100nm thick chromium layer is fixed in a high-resolution contact exposure machine through standard photolithography and wet etching technology, the position of the film mask is adjusted to align it with the quartz chrome plate, and then exposed by ultraviolet light to transfer the designed mask pattern to the quartz chrome plate. The chrome plate after exposure, development and de-chroming is placed on a shaking table containing hydrofluoric acid for etching, and the etching depth is detected by a profilometer. After reaching the required depth, secondary etching is performed to prepare chip channels of different depths. The secondary etching step after the glue filling is the same as the primary etching step. After complete glue removal and cutting, a single-piece quartz microfluidic chip (quartz monolith) is obtained.

[0066] In step S520, the two quartz plates are bonded by plasma-assisted bonding technology and thermal bonding technology to obtain a quartz microfluidic two-dimensional liquid chromatography chip, namely, the chip body 100. For details on the stacking layers of the upper and lower quartz monolithic sheets when bonding, see the attached Figure 2 shown.

[0067] In step S530, a tool (a thin wire with a certain strength, such as a metal wire) is used to introduce porous silica microspheres as a first plunger through the plunger channel 108 to the connection between the plunger channel 108 and the first end of the one-dimensional separation channel 106, and then the plunger channel 108 is blocked. Tetrahydrofuran is used as a dispersant to prepare a chromatographic filler slurry solution (such as a 3 mg / mL to 5 mg / mL C18 chromatographic filler slurry solution), which is ultrasonically mixed evenly. The one-dimensional chromatographic filling column is filled by using a method of driving the slurry solution with air pressure. When the column bed length of the filling column reaches a set value (such as 1 cm), the filling is stopped and the one-dimensional chromatographic filling column is compacted at a set flow rate (such as 0.01 mL / min) and a pressure change of 0-1000 psi. After the one-dimensional chromatographic filling column is filled, a capillary foam method is used to introduce porous silica microspheres as a second plunger through the two-dimensional mobile phase channel 103 to the connection between the two-dimensional mobile phase channel 103 and the first end of the two-dimensional separation channel 107. The two-dimensional chromatographic filling column is prepared by the same method, and finally a quartz microfluidic two-dimensional liquid chromatography chip containing a one-dimensional chromatographic filling column and a two-dimensional chromatographic filling column is prepared.

[0068] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or make equivalent replacements for some of the technical features therein. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A two-dimensional liquid chromatography microfluidic chip, characterized in that: include: A chip body, wherein a channel structure is formed in the chip body, and the channel structure includes a sample channel, a one-dimensional mobile phase channel, a two-dimensional mobile phase channel, a one-dimensional cross injection channel, a two-dimensional cross injection channel, a one-dimensional separation channel, a two-dimensional separation channel, a plunger channel, and a post-column channel; The sample channel is connected to the first end of the one-dimensional cross injection channel, the one-dimensional mobile phase channel is connected to the second end of the one-dimensional cross injection channel, the first end of the one-dimensional separation channel is connected to the third end of the one-dimensional cross injection channel, the plunger channel is connected to the third end of the one-dimensional cross injection channel, the two-dimensional mobile phase channel is connected to the first end of the two-dimensional cross injection channel, the second end of the one-dimensional separation channel is connected to the second end of the two-dimensional cross injection channel, the first end of the two-dimensional separation channel is connected to the third end of the two-dimensional cross injection channel, and one end of the post-column channel is connected to the second end of the two-dimensional separation channel.

2. The two-dimensional liquid chromatography microfluidic chip according to claim 1, characterized in that: A first plunger is provided at the connection between the plunger channel and the first end of the one-dimensional separation channel, and a second plunger is provided at the connection between the two-dimensional mobile phase channel and the first end of the two-dimensional separation channel; A one-dimensional chromatographic filling column is arranged in the one-dimensional separation channel, and a two-dimensional chromatographic filling column is arranged in the two-dimensional separation channel.

3. The two-dimensional liquid chromatography microfluidic chip according to claim 2, characterized in that: The third end outlet of the one-dimensional cross injection channel is provided with a first limiting channel for limiting the first plunger, and the fourth end inlet of the two-dimensional cross injection channel is provided with a second limiting channel for limiting the second plunger.

4. The two-dimensional liquid chromatography microfluidic chip according to claim 3, characterized in that: The cross-sections of the first limiting channel and the second limiting channel are both semicircular.

5. The two-dimensional liquid chromatography microfluidic chip according to claim 4, characterized in that: The diameters of the first limiting channel and the second limiting channel are both 80 μm to 120 μm.

6. The two-dimensional liquid chromatography microfluidic chip according to claim 2, characterized in that: The first plunger and the second plunger are both porous silica microspheres.

7. The two-dimensional liquid chromatography microfluidic chip according to any one of claims 1 to 6, characterized in that: The diameters of the one-dimensional separation channel and the two-dimensional separation channel are both 80 μm to 120 μm.

8. The two-dimensional liquid chromatography microfluidic chip according to any one of claims 1 to 6, characterized in that: The channel structure further includes a first liquid discharge channel and a second liquid discharge channel, wherein the first liquid discharge channel is connected to the fourth end of the one-dimensional cross injection channel, and the second liquid discharge channel is connected to the fourth end of the two-dimensional cross injection channel.

9. The two-dimensional liquid chromatography microfluidic chip according to any one of claims 1 to 6, characterized in that: The chip body includes two quartz monoliths, on which grooves for forming the channel structure are arranged, and the two quartz monoliths are arranged opposite to each other and bonded to each other so that the grooves on the two quartz monoliths form the channel structure.

10. A method for manufacturing a microfluidic chip based on the two-dimensional liquid chromatography microfluidic chip according to any one of claims 1 to 9, characterized in that: include: Making grooves corresponding to the channel structure on two quartz monoliths respectively; Aligning and thermally bonding the two quartz monoliths with the grooves to form the chip body, so that the grooves of the two quartz monoliths are combined to form the channel structure; A first plunger is introduced into the connection between the plunger channel and the first end of the one-dimensional separation channel through the plunger channel, and then a chromatographic filler is filled into the one-dimensional separation channel through the post-column channel to form a one-dimensional chromatographic filling column in the one-dimensional separation channel; a second plunger is introduced into the connection between the two-dimensional mobile phase channel and the first end of the two-dimensional separation channel through the two-dimensional mobile phase channel, and then a chromatographic filler is filled into the two-dimensional separation channel through the post-column channel to form a two-dimensional chromatographic filling column in the two-dimensional separation channel.

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