CN110614126B - Micro-fluidic chip for realizing optical detection and manufacturing method thereof - Google Patents
Micro-fluidic chip for realizing optical detection and manufacturing method thereof Download PDFInfo
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- 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
- B01L3/502707—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 characterised by the manufacture of the container or its components
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- 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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Abstract
The invention discloses a micro-fluidic chip for realizing optical detection and a manufacturing method thereof. The chip is composed of an inner functional area and an outer functional area. The chip design is simplified, a complex structure and a micro valve are not needed, and the cost and the processing difficulty are reduced. The chip is made of organic materials with good optical performance, so that optical detection is facilitated, and meanwhile, the chip can integrate nucleic acid extraction and amplification detection functions, so that automatic nucleic acid extraction, amplification and detection are realized, labor is saved, and time is shortened. The detection reagent can be pre-packaged in the chip, and the chip is stored at normal temperature and operated in a closed mode, so that cross contamination of samples is avoided.
Description
Technical Field
The invention relates to the field of microfluidic chips, in particular to a microfluidic chip for realizing optical detection and a manufacturing method of the microfluidic chip.
Background
The micro-fluidic chip realizes a multi-step biochemical reaction process by utilizing highly integrated functional units in a micro-scale space, reduces manual operation, realizes Sample in-result out (Sample in-Answer out), and provides an automatic solution for the fields of biomedical diagnosis, analytical chemistry, life science and the like. The microfluidic chip has the advantages of reducing the consumption of a reaction system and required reagents, greatly shortening the sample processing time, avoiding cross contamination of the totally-enclosed chip-reagent system, reducing the manual operation process through automatic control and improving the operation consistency. The integration of processes such as nucleic acid sample extraction and detection analysis by using a microfluidic chip is a hot point of research in recent years. The existing micro-fluidic chip research mainly focuses on the scientific research field, the integration level is high, a large number of micro valves or precise structures are designed on the chip, the complexity of the chip manufacturing process is increased, the batch production is difficult, the cost is high, and an external control system is complex.
The nucleic acid amplification reaction is a molecular biology technology for copying and amplifying specific DNA fragments by using polymerase, after the amplification reaction, the number of the specific DNA fragments reaches more than 100 ten thousand, and a high-sensitivity molecular identification result is obtained by developing with a special dye/probe. The nucleic acid amplification technology mainly comprises temperature-variable amplification and isothermal amplification. Temperature-variable amplification, also known as Polymerase Chain Reaction (PCR), is the earliest technique for nucleic acid amplification, and obtains exponentially amplified DNA fragments by temperature cycling to achieve denaturation-renaturation between specific primer probes and template DNA. The isothermal amplification technology can realize exponential amplification of DNA molecules at a constant temperature, gets rid of the dependence on excellent equipment, and shows good application prospects in clinical and on-site rapid diagnosis. Both of these two nucleic acid amplification technologies require an optical detection element to read signals, so the requirement for the quality of the reaction tube is relatively high, and amplification detection devices commonly used in the market, such as 7500 of ABI corporation and myccycler of BioRad corporation, need to be used in cooperation with the optical amplification reaction tube. At present, the processing technology of transparent organic materials in the field of microfluidic chip manufacturing is more and more mature, glass and silicon-based chips are gradually replaced, the chip cost is greatly reduced, and the optical characteristics of the organic materials are kept.
Disclosure of Invention
In order to solve the defects in the prior art, the invention provides a micro-fluidic chip for realizing optical detection, which integrates the extraction, amplification and detection of nucleic acid on the micro-fluidic chip. The chip design is simplified, a complex structure and a micro valve are not needed, and the manufacturing difficulty is reduced.
Meanwhile, the invention also provides a manufacturing method of the micro-fluidic chip for realizing the optical detection.
The invention provides a micro-fluidic chip for realizing optical detection.
A micro-fluidic chip for realizing optical detection comprises an outer functional area and an inner functional area; wherein, outer functional area comprises cover plate and storehouse layer, and outer functional area has 3 constant head tanks. The bin body layer of the outer functional zone comprises a purified reagent storage unit, a waste liquid unit and an amplification detection unit. The inner functional region comprises a nucleic acid extraction unit and a linking unit, wherein the nucleic acid extraction unit comprises at least one inlet, at least one reaction chamber and at least one outlet; the connecting unit comprises an elastic buckle structure.
There is sealed the pad between outer functional area and the interior functional area, and interior functional area passes through the elasticity buckle and fixes with outer functional area.
The outer functional area and the inner functional area can move relatively under the action of external force.
Preferably, the purification reagent storage unit comprises at least one reagent chamber.
Preferably, the amplification and detection unit comprises a freeze-drying reagent chamber, a drying agent chamber, an amplification reaction chamber and a communication pipeline.
Preferably, the inner functional region is of circular configuration.
Preferably, the optical performance of the chip is good, and the manufacturing material is one or a combination of more of PMMA, epoxy resin, PTFE, PS, PC, COC, COP or PDMS.
Preferably, the processing method of the structural component of the chip comprises high-precision mold injection molding, micro-machining, 3D printing, deposition and bonding.
Preferably, the inner surface of the chip has a functional modification group or is subjected to surface treatment, and has hydrophilicity.
Preferably, the sealing gasket is made of silica gel or rubber.
Preferably, the off-chip functional regions are bonded separately, the on-chip functional regions are bonded separately, and the bonding method includes low-temperature bonding, thermal compression bonding, solvent bonding, anodic bonding, plasma bonding, ultrasonic bonding, and adhesive bonding.
Preferably, the cover plate and the bin layer of the outer functional area need a special fixture for precise positioning in the bonding process; the nucleic acid extraction unit and the connection unit of the inner functional region are aligned by using a limiting hole in the bonding process.
The principle of the micro-fluidic chip for realizing optical detection in the first aspect of the invention is that the inner functional area penetrates through the center of the outer functional area by using an elastic buckle structure and is fixed on the outer functional area, a certain elastic sealing gasket is fixed between the inner functional area and the outer functional area, and small holes are arranged on the elastic sealing gasket and are communicated with a chamber of a purifying reagent storage unit, a chamber of an amplification detection unit and a chamber of a waste liquid unit. Meanwhile, the purification chamber of the nucleic acid extraction unit on the inner functional region is provided with an outlet and an inlet, the position of the purification chamber is 180 ℃, when the inner functional region and the outer functional region move relatively, if the outlet or the inlet is located at the same position as the small hole in the sealing gasket, the communication between the outer functional region and the inner functional region is realized, and if the outlet or the inlet is not located at the same position as the small hole in the sealing gasket, the communication between the outer functional region and the inner.
The appearance of the micro-fluidic chip for realizing the optical detection is designed to be circular. Wherein, the inner functional area must be a circular structure, and the communication or the sealing of different chambers/pipelines of the outer functional area and the inner functional area is realized through rotation.
The second aspect of the present invention provides a method for manufacturing a microfluidic chip for performing optical detection, including the steps of:
1) selecting materials of all structural components of the chip;
2) selecting a processing method according to the material characteristics of each part of the chip for processing;
3) sequentially placing the cover plate and the bin body layer of the outer functional area into a special fixture for bonding;
4) aligning the nucleic acid extraction unit of the internal functional region with the limiting hole of the connecting unit, inserting a limiting sheath, and bonding;
5) fixing the sealing gasket at the corresponding position of the back of the bin body layer of the outer functional area;
6) and the elastic buckle of the inner functional area after bonding penetrates through the sealing gasket and is fixed at the center of the outer functional area after bonding.
Preferably, before step 3), necessary pre-treatments are performed according to the characteristics of the selected chip fabrication material, including surface hydrophilic treatment, dehydration treatment of organic materials, and dust removal of organic materials.
The invention has the beneficial effects that:
1) the chip design is simplified, a complex structure and a micro valve are not needed, and the cost and the processing difficulty are reduced;
2) the chip is made of organic materials with good optical performance, so that optical detection is facilitated, and meanwhile, the chip can integrate nucleic acid extraction and amplification detection functions, so that automatic nucleic acid extraction, amplification and detection are realized, labor is saved, and time is shortened;
3) the detection reagent can be pre-packaged in the chip, and the chip is stored at normal temperature and operated in a closed mode, so that cross contamination of samples is avoided.
Drawings
Fig. 1 is a top view of an exploded view of a microfluidic chip structure for implementing optical detection according to the present invention.
Fig. 2 is a bottom view of an exploded view of a microfluidic chip structure for performing optical detection according to the present invention.
Wherein the reference numerals are: 1. the kit comprises a cover plate, 2, a cover plate positioning groove, 3, a bin body layer, 4, a sealing gasket, 5, a connecting unit, 6, a nucleic acid extraction unit, 7, a connecting unit limiting hole, 8, a nucleic acid extraction unit limiting hole, 9, an elastic buckle, 10, a waste liquid unit, 11, an amplification reaction chamber, 12, a bin body layer positioning groove, 13, a purification reagent storage unit, 14, a freeze-drying reagent chamber, 15 and a drying agent chamber.
Detailed Description
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 exemplary and in no way limit the present invention and its applications or uses.
Meanwhile, the sizes of the respective portions shown in the drawings are not drawn in an actual proportional relationship for the convenience of description.
Example 1.
As shown in fig. 1 and fig. 2, the microfluidic chip for implementing optical detection disclosed by the present invention is composed of two independent parts, namely an outer functional region and an inner functional region. Wherein, the outer functional area is round and consists of a cover plate (1) and a bin body layer (3); the outer functional area is provided with 3 positioning grooves which are respectively positioned at the corresponding positions of the cover plate (2) and the bin body layer (12) and can be fixed in position; the storehouse layer (3) of the external functional area comprises a purification reagent storage unit (13), a waste liquid unit (10) and amplification detection units (11, 14, 15).
The inner functional area comprises a nucleic acid extraction unit (6) and a connection unit (5), wherein the nucleic acid extraction unit (6) comprises an inlet, a serpentine reaction chamber and an outlet; the connection unit (5) comprises an elastic buckle (9).
A sealing gasket (4) is arranged between the outer functional area and the inner functional area, and the inner functional area passes through the center of the circle of the center of the outer functional area through an elastic buckle (9) and is fixed on the outer functional area.
The outer functional area and the inner functional area can move relatively under the action of external force, so that the switching of different reagent chambers of the purification reagent storage unit (3) is realized, and the communication or the sealing of the inner functional area and the outer functional area is realized.
The purification reagent storage unit (3) contains 6 reagent chambers and corresponding tubing.
The amplification detection unit comprises a freeze-dried reagent chamber (14), a drying agent chamber (15) and an amplification reaction chamber (11).
The working principle of the microfluidic chip is as follows: reagents required for nucleic acid purification are preloaded in a reagent chamber of a purification reagent storage unit (13) of the chip, and a special modification group or preset magnetic beads exist in a snake-shaped reaction chamber of a nucleic acid extraction unit (6). The rotation of the inner functional area is controlled to switch between different reagent chambers. When the sample flows through the serpentine reaction chamber, nucleic acid is adsorbed, and impurities and waste liquid directly enter a waste liquid unit (10); washing the nucleic acid as the wash solution flows through the serpentine reaction chamber; when the eluent flows through the serpentine reaction chamber, the nucleic acid is eluted and then is sequentially injected into a freeze-drying reagent chamber (14) and an amplification reaction chamber (11) of the amplification detection unit to carry out amplification reaction and detect a fluorescent signal.
The inner functional area is of a circular structure, the chip material is selected from PC, and all structural parts of the chip are formed by injection molding through a high-precision mold.
The storehouse layer of the outer functional area on the chip and the storehouse and the pipeline included in the nucleic acid extracting unit of the inner functional area are both subjected to hydrophilic treatment by using a plasma technology.
The sealing gasket is made of silica gel, is formed by injection molding of a high-precision mold and has elasticity.
And the outer functional area and the inner functional area of the chip are independently bonded, and a hot-press bonding technology is selected. Firstly, put into special design's fixture after cover plate (1) and storehouse body layer (3) with outer functional area through cover plate constant head tank (2) and storehouse body layer constant head tank (12) alignment, have 3 archs on the fixture, can accurately match the three constant head tank of outer functional area. The PC hot pressing conditions were as follows: and cooling to room temperature after lasting for 15 minutes at a temperature of 160 ℃, a vacuum degree of <0.1bar and a pressure of 1 MPa.
Secondly, after aligning the limiting hole (8) of the nucleic acid extraction unit (6) in the inner functional region with the limiting hole (7) of the connection unit (5), inserting a limiting sheath, and placing the limiting sheath into a specially designed fixture for hot-press bonding under the same conditions.
Example 2.
Interior functional area and outer functional area outward appearance are circular, and main structural design is with embodiment 1, and interior functional area aligns through spacing aperture, and outer functional area aligns through the constant head tank. The manufacturing process comprises the following steps:
1) selecting a PDMS material as a chip material;
2) each structural part of the chip is injection molded by adopting a high-precision mold;
3) and (3) carrying out surface treatment on the PDMS structural member by using an oxygen plasma technology, aligning and laminating the cover plate (1) and the bin body layer (3) of the treated outer functional region through a positioning groove, and then putting the cover plate and the bin body layer into a fixture, and keeping the temperature for 100 ℃ for one hour to realize permanent sealing. After the connecting unit (5) and the nucleic acid extracting unit (6) of the inner functional area are aligned through the limiting hole and fixed by the limiting sheath, the other processing methods are the same as those of the outer functional area;
4) adhering a sealing gasket (4) formed by injection molding to the corresponding position of the back surface of the bin body layer (3) of the outer functional area;
5) and (3) fixing the elastic buckle (9) of the inner functional area after bonding on the outer functional area after bonding by penetrating the center of the outer functional area.
It is within the scope of the invention to cover such minor variations within the spirit and scope of the invention as defined by the appended claims. Such as the material, shape and size of the microfluidic chip, the shape and size of the chamber, the shape and size of various functional and connective channels, etc.
Claims (9)
1. A micro-fluidic chip for realizing optical detection is characterized in that the chip consists of an outer functional area and an inner functional area; the outer functional area consists of a cover plate and a bin body layer; the outer functional area is provided with 3 positioning grooves; the bin body layer of the outer functional zone comprises a purified reagent storage unit, a waste liquid unit and an amplification detection unit; the internal functional region comprises a nucleic acid extraction unit and a connection unit; the nucleic acid extraction unit comprises at least one inlet, at least one reaction chamber, and at least one outlet; the connecting unit comprises an elastic buckle structure; a sealing gasket is arranged between the outer functional area and the inner functional area; the inner functional area is fixed with the outer functional area through an elastic buckle; the outer functional area and the inner functional area can rotate relatively under the action of external force;
the purification reagent storage unit comprises at least one reagent chamber;
the amplification detection unit comprises a freeze-drying reagent chamber, a drying agent chamber, an amplification reaction chamber and a communication pipeline;
the inner functional area is of a circular structure.
2. The microfluidic chip for realizing optical detection according to claim 1, wherein the chip has good optical properties, and the material is selected from one or more of PMMA, epoxy resin, PTFE, PS, PC, COC, COP and PDMS.
3. The microfluidic chip for realizing optical detection according to claim 1, wherein the chip structure part processing method comprises high precision mold injection, micro-machining, 3D printing, deposition and bonding.
4. The microfluidic chip for realizing optical detection according to claim 1, wherein the inner surface of the chip has a functional modification group or is surface-treated to have hydrophilicity.
5. The microfluidic chip for realizing optical detection according to claim 1, wherein the gasket is made of silicone or rubber.
6. The microfluidic chip for realizing optical detection according to claim 1, wherein the off-chip functional regions are bonded separately, and the on-chip functional regions are bonded separately, and the bonding method includes low temperature bonding, thermal compression bonding, solvent bonding, anodic bonding, plasma bonding, ultrasonic bonding, and adhesive bonding.
7. The microfluidic chip for realizing optical detection according to claim 6, wherein the cover plate and the chamber layer of the outer functional region require a special fixture for precise positioning during bonding; the nucleic acid extraction unit and the connection unit of the inner functional region are aligned by using a limiting hole in the bonding process.
8. The method for manufacturing the microfluidic chip for realizing optical detection according to claim 1, comprising the following steps:
1) selecting materials of all structural components of the chip;
2) selecting a processing method according to the material characteristics of each part of the chip for processing;
3) placing the cover plate and the bin body layer of the outer functional area into a special fixture in sequence for bonding;
4) aligning the nucleic acid extraction unit of the internal functional region with the limiting hole of the connecting unit, inserting a limiting sheath, and bonding;
5) fixing the sealing gasket at the corresponding position of the back of the bin body layer of the outer functional area;
6) and the elastic buckle of the inner functional area after bonding penetrates through the sealing gasket and is fixed at the center of the outer functional area after bonding.
9. The method of claim 8, wherein prior to step 3), the steps of pre-treating the chip are performed according to the characteristics of the selected chip-making material, such as surface hydrophilic treatment, dehydration treatment of organic material, and dust removal of organic material.
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| CN201810684326.9A CN110614126B (en) | 2018-06-28 | 2018-06-28 | Micro-fluidic chip for realizing optical detection and manufacturing method thereof |
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| CN201810684326.9A CN110614126B (en) | 2018-06-28 | 2018-06-28 | Micro-fluidic chip for realizing optical detection and manufacturing method thereof |
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| CN110614126B true CN110614126B (en) | 2021-04-09 |
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| CN101517413A (en) * | 2006-09-27 | 2009-08-26 | 霍夫曼-拉罗奇有限公司 | Rotatable test element |
| US9101935B2 (en) * | 2011-09-14 | 2015-08-11 | Samsung Electronics Co., Ltd. | Microfluidic apparatus and control method thereof |
| CN105316224A (en) * | 2015-12-07 | 2016-02-10 | 中国科学院苏州生物医学工程技术研究所 | Full-automatic nucleic acid extraction and PCR amplification micro-fluidic chip and application method thereof |
| CN106552683A (en) * | 2016-12-06 | 2017-04-05 | 天津岳达科技有限公司 | Efficient microsphere biological detection chip |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101517413A (en) * | 2006-09-27 | 2009-08-26 | 霍夫曼-拉罗奇有限公司 | Rotatable test element |
| US9101935B2 (en) * | 2011-09-14 | 2015-08-11 | Samsung Electronics Co., Ltd. | Microfluidic apparatus and control method thereof |
| CN105316224A (en) * | 2015-12-07 | 2016-02-10 | 中国科学院苏州生物医学工程技术研究所 | Full-automatic nucleic acid extraction and PCR amplification micro-fluidic chip and application method thereof |
| CN106552683A (en) * | 2016-12-06 | 2017-04-05 | 天津岳达科技有限公司 | Efficient microsphere biological detection chip |
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Application publication date: 20191227 Assignee: Beijing Linke Biotechnology Co.,Ltd. Assignor: BEIJING INTEGRATED BIOSYSTEMS Co.,Ltd. Contract record no.: X2022990000932 Denomination of invention: A microfluidic chip for optical detection and its fabrication method Granted publication date: 20210409 License type: Common License Record date: 20221114 |