CN110684640A - Microfluidic device and gene sequencer - Google Patents
Microfluidic device and gene sequencer Download PDFInfo
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- CN110684640A CN110684640A CN201910976112.3A CN201910976112A CN110684640A CN 110684640 A CN110684640 A CN 110684640A CN 201910976112 A CN201910976112 A CN 201910976112A CN 110684640 A CN110684640 A CN 110684640A
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
- B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
- B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
- B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6869—Methods for sequencing
Abstract
The invention relates to the technical field of microfluidics, and discloses a microfluid device and a gene sequencer. The micro fluid channel comprises a liquid inlet channel and a liquid outlet channel, and the liquid inlet channel and the liquid outlet channel respectively comprise a transverse channel extending along the transverse direction and a longitudinal channel extending along the longitudinal direction. The liquid inlet flow channel and the liquid outlet flow channel of the micro fluid channel in the micro fluid device are communicated with the solution cavity for testing, so that liquid can be fed through the liquid inlet flow channel of the micro fluid channel and discharged through the liquid outlet flow channel. The combination of the transverse flow channel and the longitudinal flow channel can form a three-dimensional micro flow channel, and compared with a two-dimensional micro flow channel, the liquid inlet efficiency can be improved, so that the liquid inlet device is suitable for the test requirement of high flux and high efficiency. The gene sequencer with the microfluid device can meet the requirement of high-throughput and high-performance gene sequencing.
Description
Technical Field
The invention relates to the technical field of microfluidics, in particular to a microfluid device and a gene sequencer.
Background
Microfluidic devices can integrate a series of operations such as sample pretreatment, separation, reaction, detection, and data analysis on a single substrate. Microfluidic technology has been rapidly developed because it greatly reduces the cost of microfluidic analysis and shortens the time of microfluidic analysis, and has been widely used in the fields of DNA sequencing, protein analysis, single cell analysis, drug detection, food safety, and the like.
The structure of the current microfluidic device is usually simple, and usually, the microfluidic channel is realized only on a two-dimensional plane, so that the current high-throughput analysis instrument has many limitations in use, and the high-throughput and high-efficiency test requirements are difficult to meet.
Disclosure of Invention
The present invention is directed to solving at least one of the problems of the prior art. Therefore, the invention provides a microfluidic device which is provided with three-dimensional microfluidic channels and can meet the testing requirements of high throughput and high efficiency.
The invention also provides a gene sequencer with the microfluid device.
In a first aspect, one embodiment of the present invention provides a microfluidic device comprising a substrate having disposed thereon:
the micro fluid channel comprises a liquid inlet flow channel and a liquid outlet flow channel, and the liquid inlet flow channel and the liquid outlet flow channel respectively comprise a transverse flow channel extending along the transverse direction and a longitudinal flow channel extending along the longitudinal direction;
the first inlet is communicated with the liquid inlet flow channel;
the second inlet is communicated with the liquid inlet flow channel;
and the liquid outlet is communicated with the liquid outlet flow passage and is used for discharging liquid.
The microfluidic device of the embodiment of the invention has at least the following beneficial effects: the transverse flow channel and the longitudinal flow channel form a three-dimensional micro-fluid channel, and compared with the currently adopted two-dimensional micro-fluid channel, the three-dimensional micro-fluid channel can meet the testing requirement of high flux and high efficiency.
According to other embodiments of the present invention, the fluid outlet channel includes a buffer channel and an outlet channel, and the outlet channel is connected to the fluid outlet; one end of the buffer flow passage is used for communicating the solution cavity, and the other end of the buffer flow passage is communicated with the outlet channel.
According to other embodiments of the microfluidic device of the present invention, the outlet channel is connected to a third inlet, the third inlet is connected to the buffer channel, and the outlet channel is connected to a third valve hole.
According to other embodiments of the present invention, the first inlet includes a plurality of liquid inlets, the liquid inlet channel includes inlet channels respectively connected to the liquid inlets, and the inlet channels are independent of each other and respectively connected to a first valve hole.
According to other embodiments of the present invention, a second valve opening is in communication between the second inlet and the inlet channel.
According to other embodiments of the present invention, the microfluidic device further includes a cleaning flow channel, the cleaning flow channel is respectively communicated with the second inlet and the liquid outlet, and the cleaning flow channel is further provided with a fourth valve hole.
According to further embodiments of the present invention, the substrate includes a first base layer, a second base layer, a third base layer, and a fourth base layer, which are sequentially stacked and connected, wherein: the surface of the second base layer facing the first base layer is provided with a transverse groove, and the surface of the first base layer facing the second base layer and the transverse groove form a transverse flow channel; the second base layer is provided with the longitudinal flow channel, and the longitudinal flow channel extends along the stacking direction; the third base layer is provided with a slotted hole extending along the transverse direction, and the slotted hole is correspondingly communicated with the longitudinal flow channel; a through hole is formed in the fourth base layer and communicated with the slotted hole of the third base layer; the surfaces of the second base layer and the fourth base layer facing the third base layer and the slotted holes jointly form the transverse flow channel.
According to another embodiment of the present invention, the first base layer, the second base layer, the third base layer, and the fourth base layer are bonded by an adhesive layer, and the adhesive layer has a relief notch corresponding to the groove, the slot, and the through hole.
According to other embodiments of the present invention, the microfluidic device further includes a fifth base layer, a sixth base layer and a chip sub-plate, the fifth base layer is connected to the bottom of the fourth base layer, the sixth base layer and the fifth base layer are connected in a stacked manner, and the chip sub-plate has a test chip disposed thereon, wherein: a solution cavity is formed between the sixth base layer and the test chip, and the fifth base layer and the sixth base layer are provided with openings for communicating the through holes of the fourth base layer; the opening is communicated with the solution cavity; the fourth base layer and the sixth base layer are provided with printed circuits and are connected and conducted through a conductive component, and one end of the conductive component is located at the upper part of the solution cavity.
According to other embodiments of the microfluidic device of the present invention, the test chip is packaged with the circuit board by a COB packaging technique to integrally form the chip sub-board.
According to other embodiments of the present invention, the conductive assembly includes a first conductive pillar and a second conductive pillar, the first conductive pillar connects the printed circuit on the sixth base layer and the printed circuit on the fourth base layer, and the second conductive pillar connects the printed circuit on the fourth base layer and the chip daughter board, so as to form a conductive path capable of transmitting the electrical signal in the solution chamber to the chip daughter board.
In a second aspect, an embodiment of the present invention provides a gene sequencer comprising a microfluidic device according to any one of the preceding claims.
The gene sequencer of the embodiment of the invention at least has the following beneficial effects: the gene sequencer with the microfluid device can meet the gene sequencing requirement of high flux and high efficiency through the arrangement of the three-dimensional microfluid channel.
Drawings
FIG. 1 is a top view (in perspective) of the first embodiment;
FIG. 2 is a perspective view of the first embodiment;
FIG. 3 is a schematic view of another angle of FIG. 2;
FIG. 4 is an exploded schematic view of the first embodiment;
FIG. 5 is a schematic view of the structure of a second base layer in the first embodiment;
FIG. 6 is a perspective view of FIG. 5;
FIG. 7 is a schematic structural view of a third base layer in the first embodiment;
FIG. 8 is a top view (in perspective) of the second embodiment;
FIG. 9 is a perspective view of the second embodiment;
FIG. 10 is an exploded view of the second embodiment;
fig. 11 is a schematic view (perspective) of a combined structure of the chip sub-board, the fifth base layer, and the sixth base layer in the second embodiment;
fig. 12 is a schematic perspective view of the second embodiment.
Detailed Description
The concept and technical effects of the present invention will be clearly and completely described below in conjunction with the embodiments to fully understand the objects, features and effects of the present invention. It is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments, and those skilled in the art can obtain other embodiments without inventive effort based on the embodiments of the present invention, and all embodiments are within the protection scope of the present invention.
In the description of the embodiments of the present invention, if an orientation description is referred to, for example, an orientation or positional relationship indicated by "up", "down", etc. is based on an orientation or positional relationship shown in the drawings, it is only for convenience of description and simplification of description, and does not indicate or imply that the referred device or element must have a specific orientation, be constructed in a specific orientation, and be operated, and thus, it is not to be understood as a limitation of the present invention.
In the description of the embodiments of the present invention, if a certain feature is referred to as being "disposed" or "connected" to another feature, it may be directly disposed or connected to the other feature or indirectly disposed or connected to the other feature. In the description of the embodiments of the present invention, if "a number" is referred to, it means more than one, if "a plurality" is referred to, it is understood that the number is not included, and if "first", "second" … … "sixth" and "seventh" are referred to, it is understood that the technical features are distinguished, and the relative importance or the number of the indicated technical features is implicitly indicated, or the precedence of the indicated technical features is implicitly indicated.
The microfluid device comprises a substrate, wherein a microfluid channel, a first inlet, a second inlet and a liquid outlet are arranged on the substrate. The micro fluid channel comprises a liquid inlet channel and a liquid outlet channel, and the liquid inlet channel and the liquid outlet channel respectively comprise a transverse channel extending along the transverse direction and a longitudinal channel extending along the longitudinal direction. The liquid inlet flow channel and the liquid outlet flow channel of the micro fluid channel in the micro fluid device are communicated with the solution cavity for testing, so that liquid can be fed through the liquid inlet flow channel of the micro fluid channel and discharged through the liquid outlet flow channel. The combination of the transverse flow channel and the longitudinal flow channel can form a three-dimensional micro flow channel, and compared with a two-dimensional micro flow channel, the liquid inlet efficiency can be improved, so that the liquid inlet device is suitable for the test requirement of high flux and high efficiency. In addition, in terms of overall structure design, on the premise of meeting the requirement of high-throughput and high-efficiency test, the three-dimensional microfluidic channel is arranged, so that the structural space can be fully utilized, the overall structure of the microfluidic device is smaller than that of a conventional two-dimensional microfluidic device, and the miniaturization development of the microfluidic device in the technical field of microfluidic is facilitated.
The first inlet and the second inlet are both communicated with the liquid inlet flow channel and used for feeding liquid. The first inlet comprises a plurality of liquid inlets for manually adding a sample and a reagent, and the liquid inlet flow channel comprises inlet channels respectively communicated with the liquid inlets, so that different reagents can be respectively fed into the liquid inlet flow channel for mixing detection. The liquid outlet is communicated with the liquid outlet flow passage and is used for discharging waste liquid, and after the test is finished, the waste liquid can be discharged through the liquid outlet flow passage and the liquid outlet.
First embodiment
Referring to fig. 1 to 3, a top view (perspective) and a perspective view, respectively, of a first embodiment of a microfluidic device is shown. The microfluidic device of this embodiment has a microfluidic channel, a first inlet, a second inlet, and a liquid outlet disposed on a substrate. Referring to a coordinate system in the figure, a direction z perpendicular to the surface of the substrate is defined as a longitudinal direction, directions x and y perpendicular to the longitudinal direction are defined as transverse directions, the flow channels extending in the transverse direction are transverse flow channels, and the flow channels extending in the longitudinal direction are longitudinal flow channels. The base body is placed on a horizontal plane, when the upper surface of the base body is parallel to the horizontal plane, the transverse flow channel can convey the solution along the horizontal direction, and the longitudinal flow channel can convey the solution along the vertical direction. The liquid inlet flow channel and the liquid outlet flow channel respectively comprise a transverse flow channel extending along the transverse direction and a longitudinal flow channel extending along the longitudinal direction, and the transverse direction and the longitudinal direction are relative to the substrate, so that the transverse flow channel and the longitudinal flow channel form a three-dimensional micro-fluid channel which can convey the solution along the transverse direction and the longitudinal direction.
The micro fluid channel comprises a liquid inlet channel 100 and a liquid outlet channel 200, and the first inlet and the second inlet are both communicated with the liquid inlet channel and used for feeding liquid. The first inlet of the present embodiment includes three inlet ports 31, 32, 33, which are provided at the side of the substrate and communicate with the inlet flow channel 100 through one first inlet passage 51, 52, 53, respectively. According to specific detection requirements, a sample or a reagent can be added from the three liquid inlets 31, 32, 33, and enters a liquid containing cavity for detection (in this embodiment, no solution cavity is provided) through the liquid inlet channel 100 for mixing and corresponding detection. The first inlet channels 51, 52 and 53 are independent of each other and are respectively communicated with the first valve holes 61, 62 and 63 through the first longitudinal valve channels 34, 35 and 36, correspondingly, the liquid inlet channel 100 comprises three liquid inlet transverse channels 47, 48 and 49 and a liquid inlet longitudinal channel 44, one ends of the three liquid inlet transverse channels 47, 48 and 49 are respectively communicated with the first valve holes 61, 62 and 63, and the other ends are respectively communicated with the liquid inlet longitudinal channel 44. The solution fed from each inlet port 31, 32, 33 passes through each first inlet channel 51, 52, 53, first longitudinal valve channel 34, 35, 36, first valve hole 61, 62, 63 to each inlet transverse flow channel 47, 48, 49, and then merges into the inlet longitudinal flow channel 44, thereby passing into the solution chamber for detection to be mixed and tested. The opening and closing of the first valve holes 61, 62, 63 are controlled to realize the amount of the sample or reagent in the liquid inlets 31, 32, 33 entering the liquid inlet flow channel 100, thereby realizing the adjustment of the liquid inlet amount of various solutions.
A second valve hole 69 is communicated between the second inlet 68 and the liquid inlet channel 100 for automatic liquid inlet, and a second liquid inlet transverse channel 50 communicated with the second valve hole 69 and the liquid inlet longitudinal channel 44 is also arranged in the liquid inlet channel. In actual use, the second inlet 68 is connected to the solution bottle, and the liquid inlet control is performed by opening and closing the second valve hole 69. In this embodiment, the second inlet 68 is disposed on the upper surface of the substrate, the second inlet 68 is connected to the second inlet channel 54 through the second longitudinal channel 41, the second inlet channel 54 is connected to the second valve hole 69 through the second longitudinal valve channel 37, the second valve hole 69 is communicated with the liquid inlet transverse flow channel 50, and the second liquid inlet transverse flow channel 50 is communicated with the liquid inlet longitudinal flow channel 44. The solution enters the solution cavity for detection through the second inlet 68, the second longitudinal channel 41, the second inlet channel 54, the second longitudinal valve channel 37, the second valve hole 69, the second transverse liquid inlet channel 50 and the longitudinal flow channel 44 in sequence.
The liquid outlet 67 is communicated with the liquid outlet flow passage 200 for discharging liquid. After the detection or the cleaning is finished, the solution can be discharged through the liquid outlet flow passage 200 and the liquid outlet 67. The outlet channel 200 of this embodiment includes the buffer channel 58, the buffer area channel 57, the first outlet transverse channel 45 and the first transverse outlet channel 56. One end of the buffer flow channel 58 is communicated with the solution cavity for testing through the first liquid outlet transverse channel 45, the other end is communicated with the buffer area channel 57, and the buffer area channel 57 can be communicated with the first transverse outlet channel 56 through a plurality of sections of micro fluid channels which are communicated transversely and longitudinally. This buffering runner 58, buffer zone passageway 57 can keep in solution, and its effect lies in, there is the solution that awaits measuring at test appearance liquid intracavity solution to under the condition that needs continue to add solution, can send into this buffering runner with the solution in the appearance liquid chamber when adding solution and keep in, thereby avoid the solution in the former solution intracavity directly to discharge from the liquid outlet. After the solution is added, the solution in the buffer flow channel is only needed to be conveyed back to the solution containing cavity, so that the test is not needed to be interrupted due to the fact that the solution is added again, and the complexity of the test step is avoided being increased. Air may be pumped by a peristaltic pump into the third inlet 66, through the third inlet longitudinal channel 43 and the third inlet channel 46, to re-pump the mixed liquid temporarily stored in the buffer flow channel 58 and the buffer zone channel 57 into the liquid-containing chamber.
The liquid outlet channel 200 is connected to the third inlet 66, the third inlet 66 is connected to the buffer channel 58, and a peristaltic pump may be used to introduce air from the third inlet 66, so as to return the solution temporarily stored in the buffer channel 58 to the liquid accommodating chamber. A third valve hole 65 is communicated with the flow passage between the outlet passage and the third inlet 66, and the opening and closing of the buffer flow passage 58 and the liquid outlet 42 are realized through the opening and closing of the third valve hole 65. The solution in the solution cavity is discharged through the liquid outlet longitudinal flow passage 70, the first liquid outlet transverse flow passage 45, the buffer flow passage 58, the third inlet passage 46, the buffer area passage 57, the third valve hole 65, the third valve longitudinal passage 39, the second transverse outlet passage 73, the outlet valve pipeline 40, the first transverse outlet passage 56, the liquid outlet passage 42 and the liquid outlet 67 in sequence. The first liquid outlet transverse channel 45, the buffer area channel 57 and the second transverse outlet channel 73 are flow channels arranged above, the buffer flow channel 58, the third inlet channel 46 and the first transverse outlet channel 56 are flow channels arranged below, and the communication parts of the flow channels arranged above and below are communicated through the longitudinally extending flow channels, so that the liquid outlet flow channel 200 with the micro fluid channels which are communicated in a longitudinal and transverse manner is formed.
The microfluidic device further includes a cleaning flow channel 300, the cleaning flow channel 300 is respectively communicated with the second inlet 68 and the liquid outlet 42, and the cleaning flow channel 200 is further provided with a fourth valve hole 64. The on-off of the cleaning flow channel and the liquid outlet is realized through the opening and closing of the fourth valve hole 64. The cleaning liquid is discharged through the second inlet 68, the second longitudinal channel 41, the second inlet channel 54, the third inlet channel 55, the fourth valve hole channel 38, the fourth valve hole 64, the third liquid outlet transverse channel 71, the fourth transverse outlet channel 72, the first transverse outlet channel 56, the liquid outlet channel 42 and the liquid outlet 67.
Referring to fig. 4 to 7, there are respectively shown an exploded view of the first embodiment, a schematic structural view of the second base layer, a perspective view of the second base layer, and a schematic structural view of the third base layer; in the embodiment, the substrate comprises a first base layer 5, a second base layer 7, a third base layer 9 and a fourth base layer 11 which are sequentially connected in a stacked manner, and the base layers are provided with grooves, holes and other structures and then stacked to form the micro fluid channel, so that the substrate is simple and convenient to process and assemble.
Referring to fig. 4, 5 and 6, the surface of the second base layer 7 facing the first base layer 5 is provided with a transverse groove 7-1, and the surface of the first base layer 5 facing the second base layer 7 and the transverse groove 7-1 enclose a transverse flow channel; the second base layer 7 is provided with a longitudinal through hole 7-2, and the longitudinal through hole 7-2 extends along the stacking direction to form a longitudinal flow channel.
Referring to fig. 4 and 7, the third base layer 9 is provided with a slot 9-1 extending along the transverse direction and a through hole 9-2 penetrating through the third base layer, wherein the slot 9-1 and the through hole 9-2 are respectively communicated with the longitudinal through hole 7-2 of the second base layer 7; the fourth base layer 11 is provided with a through hole 11-1, and the through hole 11-1 is communicated with a slotted hole 9-1 of the third base layer 9; the surfaces of the second base layer 7 and the fourth base layer 11 facing the third base layer 9 and the slotted holes jointly form a transverse flow channel. The longitudinal through hole 7-2, the through hole 9-2 and the through hole 11-1 are correspondingly communicated to form a longitudinal flow channel.
The first base layer 5, the second base layer 7, the third base layer 9 and the fourth base layer 11 are respectively bonded through bonding layers, the bonding layers of the embodiment are double-sided adhesive tapes 6, 8 and 10, and abdicating notches are formed in the positions, corresponding to the hole and groove structures, of the base layers, for forming the micro fluid channel, the inlet, the outlet and the valve hole, so that the solution can flow.
The double-sided adhesive tape 6 between the first base layer 5 and the second base layer 7 is provided with abdicating gaps with the same position and the same shape corresponding to the hole and groove structure of the surface of the second base layer 7 facing the first base layer 5; the double-sided adhesive tape 8 between the second base layer 7 and the third base layer 9 is provided with abdicating gaps with the same position and the same shape corresponding to the hole and groove structure on the third base layer 9; the double-sided adhesive tape 10 between the third base layer 9 and the fourth base layer 11 is provided with abdicating gaps with the same position and the same shape corresponding to the hole and groove structure on the third base layer 9.
The microfluidic device of this embodiment forms a three-dimensional microfluidic channel by the combination of the holes and the grooves on the first base layer 5, the second base layer 7, the third base layer 9, and the fourth base layer 11, and the notches on the double-sided tapes 6, 8, and 10, and is suitable for the detection of liquid phase materials. The materials of the first base layer 5, the second base layer 7, the third base layer 9 and the fourth base layer 11 can be selected from PC materials, and the PC materials have wide material sources, excellent processing performance and stability and lower cost. The thickness of each basic unit can be rationally set up according to actual demand.
In other embodiments, the substrate may be integrally formed by means of 3D printing or the like.
Second embodiment
This embodiment is further optimized based on the first embodiment, and referring to fig. 8 to 12, a top view, a perspective view, an exploded schematic view, a partial structural perspective view and a three-dimensional structural schematic view of the microfluidic device of this embodiment are respectively shown.
Referring to fig. 10 and 12, in this embodiment, the base body includes a first base layer 5, a second base layer 7, a third base layer 9, a fourth base layer 11, a fifth base layer 14, a sixth base layer 16, a seventh base layer 20, a chip daughter board, a double-sided adhesive tape, a first conductive pillar 12, and a second conductive pillar 18, which are sequentially stacked and connected, and the base layers are bonded by the double-sided adhesive tape. The first base layer 5, the second base layer 7, the third base layer 9, and the fourth base layer 11 may be configured as the same structure as the first embodiment, and a silicone valve, a sealing ring, and a pressing sheet may be further disposed at each opening on the upper surface of the first base layer 5. Wherein:
the silica gel valve 1 can open or close a corresponding inlet or outlet and is used for controlling the inlet and outlet of a sample, a reagent, air or waste liquid;
the sealing ring 2 is used for sealing the gap between the microfluid device and the liquid inlet needle A, the air inlet needle B or the outlet needle C;
the laminating sheet 3 is used for fixing and laminating the double-sided adhesive tape 4 used for the silica gel valve 1 so as to prevent the silica gel valve from leaking liquid or air and cover a horizontal micro fluid channel formed by the first base layer 5;
the first base layer 5 is used for fixing the positions of the silica gel valve 1 and the sealing ring 2 on the base body, and leakage caused by displacement is prevented;
the second substrate 7 is used for forming a first transverse micro-fluid channel (transverse flow channel) and a micro-fluid channel (longitudinal flow channel) along the longitudinal direction;
the third substrate 9 is used to form a second microfluidic channel (lateral flow channel) in the lateral direction;
the fourth base layer 11 is used for covering the transverse flow channel formed by the third base layer 9 and carrying a printed conducting circuit;
the fifth base layer 14 is used for forming a liquid containing cavity together with the sixth base layer 16 and the chip to be tested;
the sixth base layer 16 is used for forming a liquid containing cavity together with the fifth base layer 14 and the chip to be tested and carrying a printed conducting circuit;
the seventh base layer 20 is supported between the microfluidic device fourth base layer 11 and the chip daughter board to form a gap;
the first conductive pillar 12 and the second conductive pillar 18 are used for connecting printed conductive circuits of different layers;
the chip daughter board 21 is used for bearing a chip to be tested;
the double-sided adhesive tapes 3, 6, 8, 10, 13, 15 and 19 are used for adhering a silica gel valve, a sealing ring, each layer of base layer and a chip daughter board.
According to the above, the three-dimensional micro fluid channel is mainly constituted by the first base layer 5, the second base layer 7, the third base layer 9, and the fourth base layer 11. The liquid containing cavity is formed by the fifth base layer 14, the sixth base layer 16 and the tested chip on the chip daughter board 21 and is used for mixing the sample of the detected body and various reagents; and conductive paths between the liquid containing cavity and the chip daughter board are formed among the fifth base layer 14, the sixth base layer 16 and the chip daughter board so as to facilitate the transmission of detection signals. The hollow part in the middle of the seventh base layer 20 corresponds to the position of the chip to be tested on the chip daughter board, and is used for forming a gap between the fourth base layer and the chip daughter board.
Fig. 11 shows a schematic structure of the fifth base layer 14 and a schematic structure of the electronic daughter board, the fifth base layer 14 and the sixth base layer 16, and referring to fig. 8, 9 and 11, the fifth base layer 14 is connected to the bottom of the fourth base layer 11, the sixth base layer 16 and the fifth base layer 14 are connected in a stacked manner, and the chip daughter board is provided with a tested chip. The tested chip can be packaged with the circuit board by the COB packaging technology to form an integrated chip daughter board, and then glued with the sixth base layer 16 by the double-sided adhesive tape. A solution cavity 24 is formed between the sixth base layer 16 and the tested chip, and the fifth base layer 14 and the sixth base layer 16 are provided with openings for communicating with the through holes of the fourth base layer 11, and the openings are communicated with the solution cavity 24, so that the longitudinal flow channel can be communicated with the liquid containing cavity.
The fourth substrate 11 and the sixth substrate 16 are provided with printed circuits and are electrically connected through a conductive member, one end of which is located at the upper portion of the solution chamber 24. In this embodiment, the conductive assembly includes a first conductive pillar 12 and a second conductive pillar 18, and the first conductive pillar 12 and the second conductive pillar 18 may be copper pillars. The first conductive pillar 12 connects the sixth base layer 16 and the printed circuits 22 and 23 on the fourth base layer 11, and the second conductive pillar 18 connects the printed circuit 22 on the fourth base layer 11 and the chip daughter board 21, so as to form a conductive path, which can transmit the electrical signal at the top in the solution cavity 24 to the chip daughter board 21.
In the field of microfluidics, a series of operations such as sample pretreatment, separation, reaction, detection, data analysis, and the like can be integrated on one substrate by using the microfluidic device. Microfluidic technology has been rapidly developed because it greatly reduces the cost of microfluidic analysis and shortens the time of microfluidic analysis, and has been widely used in the fields of DNA sequencing, protein analysis, single cell analysis, drug detection, food safety, and the like.
With the rapid development of science and technology, the application of medical engineering has become more and more popular, and the demand for medical devices has become more and more great, wherein the development of preventive medicine is most concerned, because Chinese emphasizes on preventive treatment, how to find diseases early through early screening and intervene early, and even promoted to the national level, the importance of strategic position is conceivable.
The development of gene detection is the most fierce in preventive medical engineering, and the application of non-invasive prenatal gene detection serves tens of millions of pregnant women within a few years, and other applications such as early screening of cancers are not mentioned. However, with the widespread use of gene detection, the operation and detection mode in the laboratory by manual methods has been far from meeting the demand, so the demand of gene sequencers with automatic detection function is also high, and the key point of the development of gene sequencing technology is that the design of microfluidic channels and gene sequencing chips is a necessary condition of automatic gene sequencers, and in order to ensure the sample, the reagents and the chips have sufficient mixing and reaction time, and also in order to reduce the volume of the gene sequencers, efficient microfluidic devices are very important.
The structural design of the current microfluidic devices is usually simple, and the microfluidic channels are usually realized only on a two-dimensional plane, which obviously has many limitations in use for high-throughput gene testing applications. In combination with the foregoing description of the structure of the microfluidic device, the embodiment of the invention provides the microfluidic device with three-dimensional microfluidic channels, which can solve the problems in production and use of the multidimensional microfluidic, and has very important practical significance in practical application.
The microfluidic device of the present embodiment can be used in conjunction with an existing gene sequencer for gene sequencing. The gene sequencer can be provided with a pipeline for introducing a reagent, a pipeline for discharging solution or air, a valve control device for opening or closing the silica gel valves at the first valve port, the second valve port, the third valve port and the fourth valve port, a pump for feeding liquid or discharging liquid, a pump for introducing air, a gene sequencing program, an automatic tightness testing program and a detection device for detecting the tightness of the joint of the liquid feeding device of the gene sequencer and the microfluidic device, wherein the gene sequencing program is pre-installed for automatically controlling the opening or closing of the silica gel valves and the pump. Gene sequencing can be performed by the following steps:
1) placing the device on a gene sequencer, respectively connecting a second inlet, a third inlet and a liquid outlet to corresponding pipelines of the gene sequencer, starting an automatic tightness testing program (the gene sequencer is preassembled), and automatically closing all valve ports after testing;
2) manually adding the specimen sample, the first reagent, and the second reagent to the first inlets 31, 32, and 33, respectively;
3) starting a gene sequencing program (preassembling a gene sequencer);
5) the gene sequencing program automatically controls the opening of the fourth valve port 64 (the straight-through valve);
5) the gene sequencing program automatically controls the flow of the cleaning solution from the reagent bottle into the second inlet 68;
6) the cleaning liquid is discharged from the second inlet 68 through the second longitudinal channel 41, the second inlet channel 54, the third inlet channel 55, the fourth valve hole channel 38, the fourth valve hole 64, the third liquid outlet transverse channel 71, the fourth transverse outlet channel 72, the first transverse outlet channel 56, the liquid outlet channel 42 and the liquid outlet 67 to a waste liquid bottle communicated with the liquid outlet 67.
7) The gene sequencing program automatically controls the fourth valve port 64 (the straight-through valve) to be closed;
8) the gene sequencing program automatically controls to open the first valve hole 61 (sample valve) and the third valve hole 65 (outlet valve), and opens the vacuum pump to suck the sample from the manual sample and the first inlet 31, and the sample enters the liquid containing cavity 24 through the first inlet channel 51, the first longitudinal valve channel 34, the liquid inlet transverse flow channel 47 and the liquid inlet longitudinal flow channel 44;
9) the gene sequencing program automatically controls the closing of the first valve hole 61 (sample valve) and the vacuum pump;
10) the gene sequencing program automatically controls to open the first valve hole 62 (first reagent valve), and opens the vacuum pump to suck the first reagent from the first inlet 32, and the first reagent enters the liquid containing cavity 24 through the first inlet channel 52, the first longitudinal valve channel 35, the liquid inlet transverse flow channel 48 and the liquid inlet longitudinal flow channel 44;
11) the gene sequencing program automatically controls the closing of the first valve port 62 (first reagent valve) and the vacuum pump;
12) the gene sequencing program automatically controls to open a first valve hole 63 (a second reagent valve), and opens a vacuum pump to suck a second reagent from the first inlet 33, and the second reagent enters the liquid containing cavity 24 through the first inlet channel 53, the first longitudinal valve channel 36, the liquid inlet transverse flow channel 49 and the liquid inlet longitudinal flow channel 44;
13) the gene sequencing program automatically controls the closing of the first valve port 63 (second reagent valve), the third valve port 65 (outlet valve) and the vacuum pump;
14) the gene sequencing program automatically executes the coverage rate test of the mixed liquid in the liquid containing cavity 24 and the tested chip;
15) if the coverage test is passed (i.e., the solution completely covers the chip under test), then go to step 18;
16) if the coverage test is not passed (i.e. the solution does not completely cover the chip to be tested), the gene sequencing program automatically opens the third valve hole 65 (outlet valve) and the vacuum pump, sucks the mixed liquid out of the liquid chamber 24, then enters the buffer flow channel 58 and the buffer area channel 57 for temporary storage through the first liquid outlet transverse channel 45, then closes the third valve hole 65 (outlet valve) and the vacuum pump, opens the peristaltic pump to pump air into the third inlet 66, pumps the air into the liquid chamber 24 again through the third inlet longitudinal channel 43 and the third inlet channel 46, pumps the mixed liquid temporarily stored in the buffer flow channel 58 and the buffer area channel 57 into the liquid chamber 24, and then closes the peristaltic pump;
17) repeating steps 14) -17);
18) the gene sequencing program automatically executes the gene sequencing work, and after the gene sequencing work is finished, the third valve hole 65 (outlet valve) and the vacuum pump are opened, the mixed liquid is sucked out from the liquid cavity 24 and is discharged into the waste liquid bottle through the first liquid outlet transverse channel 45, the buffer flow channel 58, the buffer area channel 57, the third inlet channel 46, the third valve longitudinal channel 39, the outlet valve pipeline 40, the first transverse outlet channel 56, the liquid outlet channel 42 and the liquid outlet 67 in sequence.
Compared with the existing microfluidic device with two-dimensional microfluidic channels, the microfluidic device of the embodiment can be more suitable for the high-throughput and high-efficiency test requirements of gene detection, realizes reasonable utilization of space, and is beneficial to miniaturization development of the microfluidic device.
The embodiment of the invention also provides a gene sequencer which comprises the microfluidic device in any one of the above embodiments, so that the requirement of high-throughput and high-performance gene sequencing is met, and the further development of a gene sequencing technology is facilitated.
The embodiments of the present invention have been described in detail with reference to the accompanying drawings, but the present invention is not limited to the above embodiments, and various changes can be made within the knowledge of those skilled in the art without departing from the gist of the present invention. Furthermore, the embodiments of the present invention and the features of the embodiments may be combined with each other without conflict.
Claims (10)
1. A microfluidic device comprising a substrate having disposed thereon:
the micro fluid channel comprises a liquid inlet flow channel and a liquid outlet flow channel, and the liquid inlet flow channel and the liquid outlet flow channel respectively comprise a transverse flow channel extending along the transverse direction and a longitudinal flow channel extending along the longitudinal direction;
the first inlet is communicated with the liquid inlet flow channel;
the second inlet is communicated with the liquid inlet flow channel;
and the liquid outlet is communicated with the liquid outlet flow passage and is used for discharging liquid.
2. The microfluidic device according to claim 1, wherein the outlet flow channel comprises a buffer flow channel and an outlet channel, and the outlet channel is communicated with the outlet; one end of the buffer flow passage is used for communicating the solution cavity, and the other end of the buffer flow passage is communicated with the outlet channel.
3. The microfluidic device according to claim 2, wherein the outlet channel is connected to a third inlet, the third inlet is connected to the buffer channel, and the outlet channel is connected to a third valve hole.
4. The microfluidic device according to claim 1, wherein the first inlet comprises a plurality of liquid inlets, the liquid inlet channel comprises inlet channels respectively connected to the liquid inlets, and each of the inlet channels is independent of each other and respectively connected to a first valve hole.
5. The microfluidic device according to claim 1, further comprising a purge flow channel, wherein the purge flow channel is connected to the second inlet and the liquid outlet, and a fourth valve hole is disposed on the purge flow channel.
6. The microfluidic device according to any one of claims 1 to 5, wherein the substrate comprises a first base layer, a second base layer, a third base layer, and a fourth base layer, which are sequentially connected in a stack, wherein:
the surface of the second base layer facing the first base layer is provided with a transverse groove, and the surface of the first base layer facing the second base layer and the transverse groove form a transverse flow channel;
the second base layer is provided with the longitudinal flow channel, and the longitudinal flow channel extends along the stacking direction;
the third base layer is provided with a slotted hole extending along the transverse direction, and the slotted hole is correspondingly communicated with the longitudinal flow channel;
a through hole is formed in the fourth base layer and communicated with the slotted hole of the third base layer;
the surfaces of the second base layer and the fourth base layer facing the third base layer and the slotted holes jointly form the transverse flow channel.
7. The microfluidic device according to claim 6, wherein the first substrate, the second substrate, the third substrate and the fourth substrate are bonded by bonding layers, and the bonding layers have notches corresponding to the grooves, the slots and the through holes.
8. The microfluidic device according to claim 6, further comprising a fifth base layer, a sixth base layer and a chip daughter board, wherein the fifth base layer is connected to the bottom of the fourth base layer, the sixth base layer and the fifth base layer are connected in a stacked manner, and the chip daughter board has a test chip disposed thereon, wherein:
a solution cavity is formed between the sixth base layer and the test chip, and the fifth base layer and the sixth base layer are provided with openings for communicating the through holes of the fourth base layer; the opening is communicated with the solution cavity;
the fourth base layer and the sixth base layer are provided with printed circuits and are connected and conducted through a conductive component, and one end of the conductive component is located at the upper part of the solution cavity.
9. The microfluidic device according to claim 8, wherein the conductive assembly comprises a first conductive pillar and a second conductive pillar, the first conductive pillar connects the printed circuit on the sixth base layer and the printed circuit on the fourth base layer, and the second conductive pillar connects the printed circuit on the fourth base layer and the chip daughter board, forming a conductive path capable of transmitting the electrical signal in the solution chamber to the chip daughter board.
10. A gene sequencer comprising the microfluidic device of any one of claims 1 to 9.
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| CN114405563A (en) * | 2022-01-13 | 2022-04-29 | 深圳清华大学研究院 | Micro-fluidic chip and gene sequencing system |
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