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CN116899639A - Microfluidic device and application thereof - Google Patents

Microfluidic device and application thereof Download PDF

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Publication number
CN116899639A
CN116899639A CN202310794805.7A CN202310794805A CN116899639A CN 116899639 A CN116899639 A CN 116899639A CN 202310794805 A CN202310794805 A CN 202310794805A CN 116899639 A CN116899639 A CN 116899639A
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Prior art keywords
microfluidic device
microfluidic
detection
area
quality control
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CN202310794805.7A
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CN116899639B (en
Inventor
余伟明
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Shanghai Jinxin Biotechnology Co ltd
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Shanghai Jinxin Biotechnology Co ltd
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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
    • 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
    • B01L3/50273—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 means or forces applied to move the fluids
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
    • G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
    • G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/5302—Apparatus specially adapted for immunological test procedures
    • G01N33/5304—Reaction vessels, e.g. agglutination plates
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
    • G01N33/56983—Viruses
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/58—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
    • G01N33/585—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances with a particulate label, e.g. coloured latex
    • G01N33/587—Nanoparticles
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/10—Integrating sample preparation and analysis in single entity, e.g. lab-on-a-chip concept
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0809—Geometry, shape and general structure rectangular shaped
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/12—Specific details about materials
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0403—Moving fluids with specific forces or mechanical means specific forces
    • B01L2400/0406—Moving fluids with specific forces or mechanical means specific forces capillary forces
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/005—Assays involving biological materials from specific organisms or of a specific nature from viruses
    • G01N2333/08—RNA viruses
    • G01N2333/165—Coronaviridae, e.g. avian infectious bronchitis virus
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2469/00—Immunoassays for the detection of microorganisms
    • G01N2469/20—Detection of antibodies in sample from host which are directed against antigens from microorganisms

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  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Immunology (AREA)
  • Hematology (AREA)
  • Biomedical Technology (AREA)
  • Urology & Nephrology (AREA)
  • General Health & Medical Sciences (AREA)
  • Molecular Biology (AREA)
  • Analytical Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Chemical Kinetics & Catalysis (AREA)
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  • Biochemistry (AREA)
  • Cell Biology (AREA)
  • Microbiology (AREA)
  • Biotechnology (AREA)
  • Food Science & Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Virology (AREA)
  • Dispersion Chemistry (AREA)
  • Clinical Laboratory Science (AREA)
  • Plasma & Fusion (AREA)
  • Nanotechnology (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Automatic Analysis And Handling Materials Therefor (AREA)

Abstract

The invention relates to the field of biological detection, in particular to a microfluidic device and application thereof, wherein the microfluidic device comprises a microfluidic element formed by microsphere sintering or bonding, the microfluidic element sequentially comprises a sample adding area, a mixing area, an observation area and a waste liquid area, a reagent capable of reacting with a detected substance in liquid to be detected and a reagent capable of reacting with a quality control substance in the liquid to be detected are arranged in the mixing area so as to respectively form a detected substance conjugate and a quality control substance conjugate, the observation area comprises a detection partition and a quality control partition, the detection partition is internally provided with a reagent capable of reacting with the detected substance conjugate, and the quality control partition is internally provided with a substance capable of reacting with the quality control substance conjugate. The microfluidic device can flow without the need of closed packaging of a flow channel, has simple manufacturing process, can realize semi-quantitative detection by adjusting the flow speed of fluid in the microfluidic device, can reduce background signals and improve sensitivity, and is used for detection with high sensitivity requirements.

Description

Microfluidic device and application thereof
Technical Field
The invention relates to the field of biological detection, in particular to a microfluidic device and application thereof.
Background
The rapid diagnosis test strip is a novel in-vitro diagnosis technology developed on the basis of monoclonal antibody technology, colloidal gold immunochromatography technology and new material technology in the 90 th century, has the advantages of rapidness, simplicity, single person detection and economy (figure 1), and is widely applied to the fields of medical detection, food quality monitoring, environmental monitoring, agriculture, animal husbandry, entrance and exit inspection and quarantine, forensic proposal and the like. The colloidal gold immunochromatography technology uses a microporous filter membrane (NC membrane, nitrocellulose membrane) with a large aperture as a carrier, fixes a specific antigen or antibody on the NC membrane in a strip shape, moves forward through capillary action after a sample to be detected is added to a sample pad at one end of a test strip, generates a specific immune reaction with a colloidal gold marking reagent on a binding pad, moves to the NC membrane, is captured by the antigen or antibody fixed on the NC membrane surface, is gathered on a detection belt, and obtains an intuitive color development result through visual observation of the density of light reflection signals of a nitrocellulose surface marker (colloidal gold or latex particles). The other unbound labels pass through the detection belt and flow into the water absorption pad, so as to achieve the purpose of automatic separation. However, the existing rapid diagnosis test strip cannot realize quantitative or semi-quantitative detection, has low detection speed, complex manufacturing process, low sensitivity and accuracy, and cannot be used for detection with high sensitivity requirements.
Disclosure of Invention
In view of the above-mentioned drawbacks of the prior art, an object of the present invention is to provide a microfluidic device and use thereof for solving the problems of the prior art.
To achieve the above and other related objects, the present invention provides a microfluidic device, which includes a microfluidic element formed by sintering or bonding microspheres, the microfluidic element sequentially includes a sample adding area, a mixing area, an observation area, and a waste liquid area, a reagent capable of reacting with a substance to be tested in a liquid to be tested and a reagent capable of reacting with a substance to be tested in the liquid to be tested are disposed in the mixing area, so as to form a substance to be tested conjugate and a substance to be tested conjugate respectively, the observation area includes a detection area and a substance to be tested, a reagent capable of reacting with the substance to be tested is disposed in the detection area, and a substance capable of reacting with the substance to be tested conjugate is disposed in the substance to be tested.
The microsphere is a polymer microsphere. The polymer microsphere is selected from polystyrene microsphere or organic glass microsphere. The polymer microsphere is a monodisperse microsphere.
The polymer microsphere is a polymer microsphere with a modified surface. The polymer microsphere is a polymer microsphere with the surface chemically modified. The surface modification is a hydrophilic modification. The hydrophilic modification refers to grafting hydrophilic groups on the surfaces of the polymer microspheres. The hydrophilic groups are selected from any one or more of the following: carboxyl, hydroxyl and amino.
The microfluidic element is a plate-shaped microfluidic element or a rod-shaped microfluidic element.
The surface of the sample adding area is provided with a impurity removing part. The impurity removing part is arranged at one end of the near mixing zone. The impurity removing part is in a zigzag shape. In the impurity removing part, the angle alpha formed by the front edge (namely the front edge of the saw tooth) forming the saw tooth and the surface (namely the basal plane) of the microfluidic element is larger than the angle beta formed by the rear edge (namely the rear edge of the saw tooth) and the surface of the microfluidic element.
The mixing zone comprises a plurality of flow passages which are mutually intersected and mutually communicated.
In the plate-shaped microfluidic element, folds are further arranged on the surfaces of the mixing area and the observation area.
In the rod-shaped microfluidic element, the microfluidic element is further provided with a reinforcing rib, and the reinforcing rib is arranged in the axial direction of the microfluidic element and penetrates through the sample adding area, the mixing area, the observation area and the waste liquid area.
The invention also provides application of the microfluidic device in preparing detection products.
The invention also provides a detection method, which comprises the following steps: and adding the liquid to be detected into a sample adding area of the microfluidic device, and checking or detecting an observation area after the liquid to be detected flows to a waste liquid area to obtain a detection result.
As described above, the microfluidic device and the use thereof of the present invention have the following advantageous effects: the microfluidic device can flow through the capillary suction tube phenomenon without the need of closed encapsulation of a runner, and the manufacturing process is simple. The flow rate of the fluid within the microfluidic device is adjustable. Quantitative or semi-quantitative can be achieved. The microfluidic device can be cleaned, so that background signals can be reduced, the sensitivity can be improved, and the microfluidic device can be used for detection with high sensitivity requirements.
Drawings
Fig. 1 shows a schematic front perspective view of a plate-like microfluidic element according to the invention.
Fig. 2 shows a schematic rear perspective view of the plate-like microfluidic element of fig. 1 according to the present invention.
Fig. 3 shows a schematic front view of a plate-like microfluidic element according to the invention as shown in fig. 1.
Fig. 4 shows a schematic front perspective view of a plate-like microfluidic element according to the invention.
Fig. 5 shows a schematic rear perspective view of the plate-like microfluidic element of fig. 4 according to the present invention.
Fig. 6 is a side view of the plate-shaped microfluidic element shown in fig. 4 and a partially enlarged view of the impurity removing unit according to the present invention.
Fig. 7 shows a schematic front perspective view of a plate-like microfluidic element according to the invention.
Fig. 8 is a schematic rear perspective view of the plate-like microfluidic element of fig. 7 according to the present invention.
Fig. 9 shows a side view of the plate-like microfluidic element of fig. 7 according to the invention.
Fig. 10 shows a front view of a rod-like microfluidic element according to the invention.
Fig. 11 shows a left side view of the rod-shaped microfluidic element of fig. 10 according to the present invention.
Fig. 12 shows a perspective view of the rod-shaped microfluidic element of fig. 10 according to the present invention.
Fig. 13 shows a top view of the rod-shaped microfluidic element of fig. 10 according to the present invention.
Fig. 14 shows a front view of a rod-like microfluidic element according to the invention.
Fig. 15 shows a left side view of the rod-shaped microfluidic element of fig. 14 according to the present invention.
Fig. 16 shows one of the perspective views of the rod-shaped microfluidic element of fig. 14 according to the present invention.
Fig. 17 shows one of the perspective views of the rod-like microfluidic element of fig. 14 according to the present invention.
Fig. 18 shows a top view of the rod-shaped microfluidic element of fig. 14 according to the present invention.
Fig. 19 shows a bottom view of the rod-shaped microfluidic element of fig. 14 according to the present invention.
Fig. 20 shows a front view of a rod-like microfluidic element according to the invention.
Fig. 21 shows a left side view of the rod-shaped microfluidic element of fig. 20 according to the present invention.
Fig. 22 shows one of the perspective views of the rod-shaped microfluidic element shown in fig. 20 according to the present invention.
Fig. 23 shows one of the perspective views of the rod-shaped microfluidic element shown in fig. 20 according to the present invention.
Fig. 24 shows a top view of the rod-shaped microfluidic element of fig. 20 according to the present invention.
Fig. 25 shows one of the perspective views of a plate-like microfluidic element according to the present invention.
Fig. 26 shows one of the perspective views of the plate-like microfluidic element of fig. 25 according to the present invention.
Fig. 27 shows one of the perspective views of a plate-like microfluidic element of the present invention.
Fig. 28 is a schematic perspective view of a plate-like microfluidic element according to the present invention.
Fig. 29 shows one of the schematic side views of the plate-like microfluidic element of fig. 28 according to the present invention.
Description of element reference numerals
1. Sample application area
11. Impurity removing part
2. Mixing zone
21. Fold
22. Annular component
3. Viewing area
31. Detecting partitions
32. Quality control partition
4. Waste liquid area
41. Waste liquid absorbing piece mounting groove
5. Reinforcing rib
Detailed Description
Further advantages and effects of the present invention will become apparent to those skilled in the art from the disclosure of the present invention, which is described by the following specific examples.
Please refer to fig. 1 to 24. It should be understood that the structures, proportions, sizes, etc. shown in the drawings are for illustration purposes only and should not be construed as limiting the invention to the extent that it can be practiced, since modifications, changes in the proportions, or otherwise, used in the practice of the invention, are not intended to be critical to the essential characteristics of the invention, but are otherwise, required to achieve the objective and effect taught by the invention. Also, the terms such as "upper," "lower," "left," "right," "middle," and "a" and the like recited in the present specification are merely for descriptive purposes and are not intended to limit the scope of the invention, but are intended to provide relative positional changes or modifications without materially altering the technical context in which the invention may be practiced.
The invention provides a microfluidic device, which comprises a microfluidic element formed by microsphere sintering or bonding, wherein the microfluidic element sequentially comprises a sample adding area 1, a mixing area 2, an observation area 3 and a waste liquid area 4, a reagent capable of reacting with a detected substance in a liquid to be detected and a reagent capable of reacting with a quality control substance in the liquid to be detected are arranged in the mixing area 2 so as to respectively form a detected substance conjugate and a quality control substance conjugate, the observation area 3 comprises a detection partition 31 and a quality control partition 32, the detection partition 31 is internally provided with a reagent capable of reacting with the detected substance conjugate, and the quality control partition 32 is internally provided with a substance capable of reacting with the quality control substance conjugate.
In certain embodiments of the invention, the microspheres are polymeric microspheres. In certain embodiments of the invention, the polymeric microspheres are selected from Polystyrene (PS) microspheres or Plexiglas (PMMA) microspheres.
The microfluidic element formed by microsphere sintering or bonding has certain porosity, and the porosity can be used as a runner to enable the liquid to be tested to flow based on the capillary suction tube phenomenon.
The porosity of the microfluidic element is 40-60%. The porosity refers to the volume fraction of the void space.
The polymer microsphere has the advantages of small particle size, large specific surface area, strong adsorption capacity, good dispersibility, easy modification and the like.
The diameter of the polymer microsphere is 30-120 microns. For example 30 to 50 microns, 50 to 70 microns, 70 to 90 microns, 90 to 110 microns, 110 to 130 microns, 130 to 150 microns. The polymeric microspheres used in different microfluidic elements may be different or the same.
The polymer microsphere is a monodisperse microsphere. By monodisperse microspheres is meant that at least 90% of the microspheres have an average particle size distribution within 5%, the distribution of microspheres can be considered monodisperse. By monodisperse microspheres is meant that the microspheres are monodisperse within each microfluidic element.
In certain embodiments of the invention, the monodisperse microspheres are solid microspheres, mesoporous microspheres, or sphere-to-sphere (SOS) microspheres having hollow cavities and a hierarchical structure.
The flow rates of the liquid to be measured in the microfluidic elements prepared from the microspheres with different diameters are different. The diameter of the microspheres can thus be selected according to the requirements of the assay. In general, the test solution can flow from the sample application region 1 to the waste solution region 4 within 15s to 3 min.
In certain embodiments of the invention, the sintering temperature is 120 to 220 ℃. The sintering temperature may be selected from any of the following ranges: 120-140 ℃, 140-160 ℃, 160-180 ℃, 180-200 ℃ or 200-220 ℃.
In some embodiments of the invention, the bonding may be bonding with a photo-curing agent. The photocuring agent may be a named radical photoinitiator, such as 2-hydroxy-2-methyl-1-phenyl-1-propanone or 1-hydroxycyclohexyl phenyl ketone.
The polymer microsphere is a polymer microsphere with a modified surface. The polymer microsphere is a polymer microsphere with the surface chemically modified. In certain embodiments of the invention, the surface modification is a hydrophilic modification. Specifically, the hydrophilic modification refers to grafting hydrophilic groups to the surface of the polymer microsphere. The hydrophilic groups are selected from any one or more of the following: carboxyl, hydroxyl and amino.
The hydrophilic modified polymer microsphere can be obtained through commercial purchase or self-preparation. The self-made method can refer to the patent with publication numbers of CN 107936167A, CN 114736414A and CN 103601854B.
The microfluidic element prepared from the microsphere subjected to hydrophilic modification can flow through the capillary suction tube phenomenon without the need of closed encapsulation of a runner, so that the preparation method of the microfluidic element is simple and easy to industrialize.
In the embodiments of the present invention shown in fig. 1 to 9, the microfluidic element is plate-shaped, which is called a plate-shaped microfluidic element.
In the embodiments of the present invention shown in fig. 10 to 24, the microfluidic element is in the form of a rod, which is called a rod-shaped microfluidic element.
The sample adding area 1 is columnar or plate-shaped. Specifically, in the plate-shaped microfluidic element, the sample application region 1 is plate-shaped. In the rod-shaped microfluidic element, the sample application region 1 is rod-shaped.
In the embodiment of the invention shown in fig. 1 to 13, the application area 1 is provided with a bifurcation. The contact area of the sample adding area 1 and the liquid to be detected can be increased by the bifurcation of the sample adding area 1, so that more liquid to be detected is absorbed, and the bifurcation is also used for coarse filtration and diversion of the liquid to be detected, so that blockage is prevented when the microfluidic device is used.
In the plate-like microfluidic element of the present invention, the surface of the sample application region 1 is provided with a removing portion 11. The impurity removing unit 11 is disposed at the rear end of the sample adding region (in the present invention, "front" and "rear" refer to the flowing direction of the liquid to be measured, the direction in which the liquid to be measured passes first is the front and the direction in which the liquid to be measured passes second), that is, near one end of the mixing region 2.
In some embodiments of the present invention, the impurity removing unit 11 is serrated. As shown in fig. 6, the zigzag shape is that the surface of the microfluidic element is taken as a base surface, and the surface is firstly and then is gradually raised and slowly lowered, and the circulation is performed so that the angle alpha formed by the front edge (i.e. the front edge of the zigzag) forming the zigzag and the base surface is larger than the angle beta formed by the rear edge (i.e. the rear edge of the zigzag) and the base surface.
The impurity removing part can increase the filtering area and the sample adding resistance, so that substances affecting detection in the liquid to be detected are removed. For example, blood cells in whole blood can be removed, and the serum remaining after the blood cells are filtered can continue to flow along the microfluidic element to be detected.
The microfluidic device of the present invention is suitable for use with a variety of samples. Such as saliva, urine, amniotic fluid, blood or blood products, umbilical cord blood, villus, cerebrospinal fluid, spinal fluid.
The mixing region 2 may be formed as a single channel, may be formed as a plurality of channels, or may be formed as a combination of a single channel and a plurality of channels.
In the plate-like microfluidic element of the present invention as shown in fig. 1, the mixing zone 2 comprises a single channel. In the plate-like microfluidic element of the present invention as shown in fig. 4, 8, 27, the mixing zone 2 comprises a plurality of flow channels intersecting each other and communicating with each other.
Specifically, in the plate-like microfluidic element shown in fig. 4, the flow channels of the mixing region 2 are formed in series, that is, by a plurality of annular members 22, and two flow channels of each annular member 22 in the mixing region 2 are separated and then joined, and then separated again, thus circulating. In this embodiment, the mixing zone 2 is formed by three annular members 22 in series. In one embodiment, the annular member proximal to the loading zone 1 may also act directly as a zone for the addition of wash or eluent.
Specifically, in the plate-shaped microfluidic element shown in fig. 8, the flow channel distribution of the mixing region 2 is in a scale shape.
In the plate-like microfluidic element shown in fig. 25, the mixing zone 2 is formed by a combination of an annular member 22 and a single channel connection. In the plate-like microfluidic element shown in fig. 28, the mixing region 2 is formed by a combination of an annular member 22 and a plurality of channels distributed in a fish scale shape.
In the embodiment shown in fig. 25 and 28, the annular member 22 may be directly used as a washing or eluting solution addition area.
The fish scale and/or serial mixing zone 2 can uniformly mix the liquid to be tested and then react with substances in the liquid to be tested. In the two embodiments, after flowing to the mixing area 2 from the sample adding area 1, the liquid to be measured flows forward through a plurality of flow channels, is mixed and reacted at the crossing points of the flow channels, and then continuously flows to the next crossing point along different flow channels after being mixed, and flows, mixes and reacts for a plurality of times.
In the plate-like microfluidic element, the surfaces of the mixing zone 2 and the observation zone 3 are also provided with folds 21. The arrangement of the folds 21 can increase the flow resistance, so that the liquid to be measured can be mixed more uniformly, and the contact area of the reagent can be increased, so that the reaction is more sufficient.
The folds 21 may be saw-tooth-shaped, circular protrusions, or depressions. The shape of the wrinkles 21 is not particularly limited as long as the aforementioned function can be achieved.
As shown in fig. 2 and 5, the folds 21 on the surfaces of the mixing zone 2 and the observation zone 3 are arranged on the same side of the microfluidic element.
Further, as shown in fig. 1 to 2, the impurity removing unit 11, the mixing section 2, and the wrinkles 21 of the observation section 3 are provided on the opposite surfaces of the microfluidic element.
As shown in fig. 12, in the rod-shaped microfluidic element, the microfluidic element is further provided with a reinforcing rib 5, and the reinforcing rib 5 is arranged in the axial direction of the microfluidic element and penetrates through the sample adding region 1, the mixing region 2, the observation region 3 and the waste liquid region 4. Since the rod-shaped microfluidic element is elongated in shape, the microspheres are fragile after sintering or bonding, and thus the reinforcing ribs 5 can increase the strength of the rod-shaped microfluidic element. The reinforcing bars 5 may be Polycarbonate (PC) bars.
In a rod-like microfluidic element, the mixing zone 2 extends in a ring-shape and/or square along the ribs 5 to the viewing zone 3.
The length of the mixing zone 2 may be varied according to the kind and amount of the liquid to be measured. The reagent in the mixing zone 2 may be an antibody, antigen or other reagent. The reagent in the mixing zone 2 may be a labeled reagent, such as a fluorescein label, a colloidal gold label, or the like.
In the rod-like microfluidic element, a plurality of reagents, such as antibodies, antigens or other reagents, capable of capturing different substances to be tested are provided in the mixing zone 2. Correspondingly, reagents capable of reacting with a plurality of different test substance combinations are also provided in the detection zone 31. I.e. the microfluidic element can detect a plurality of substances to be detected simultaneously.
For example, in the case of detecting whether or not a new coronavirus exists in the body by using a blood sample, a colloidal gold-labeled new coronavirus antigen and a colloidal gold-labeled mouse IgG may be provided in the mixing region 2, a mouse anti-human IgG antibody is provided in the detection region 31, and a goat anti-mouse IgG antibody is provided in the quality control region 32. After the blood sample is added to the sample adding area 1, the sample flows under capillary action, if the sample contains new coronavirus antibody IgG, the sample is combined with new coronavirus antigen marked by colloidal gold at the mixing area 2, and then the sample continues to flow forwards, and the combined IgG at the detection area 3 is combined with mouse anti-human IgG antibody to form a sandwich compound. Since the mouse anti-human IgG antibody is adsorbed on the detection zone 31, the colloidal gold is aggregated and precipitated therein to develop color. At the quality control partition 32, the colloidal gold labeled mouse IgG is combined with the goat anti-mouse IgG antibody, and the colloidal gold can be aggregated and precipitated to develop color because the goat anti-mouse IgG antibody is adsorbed on the quality control partition 32, so that the detection process is successfully completed, and the microfluidic element works normally. If the sample does not contain the novel coronavirus antibody IgG, the detection zone 31 does not have the color due to the interception of colloidal gold, but the color at the quality control zone 32 still exists.
In the case of detecting an antigen, nucleic acid or other substance, the principle is the same as that of detecting an antibody, except that the reagents provided in the mixing zone 2, the detection zone 31 and the quality control zone 32 are different. The reagents to be set may be specifically selected according to common general knowledge in the art, and the present invention is not particularly limited.
The detection zone 31 and the quality control zone 32 are both block-shaped areas, not linear areas. The length of the detection partition 31 and the quality control partition 32 is 3 mm-15 mm. For example, the detecting section 31 and the quality control section 32 have lengths of 3 to 5mm, 5 to 10mm, and 10 to 15mm. The length refers to the length of the region through which the liquid to be measured flows. Longer detection zones 31 may enable semi-quantification: when the liquid to be measured flows through the detection partition 31, if the content of the target substance to be measured in the liquid to be measured is high, the color of the whole area will be deep when the liquid to be measured flows through all the detection partition 31, and if the content of the target substance to be measured in the liquid to be measured is low, the color of the area through which the liquid to be measured flows first will be deep when the liquid to be measured flows through the detection partition 31, and the color of the area through which the liquid to be measured flows later will be light, so that the target substance to be measured in the liquid to be measured can be semi-quantified.
In the embodiment shown in fig. 2 to 3 and 8, the observation area 3 is U-shaped, and the detection area 31 and the quality control area 32 are respectively disposed on two parallel sides of the U-shape.
In the embodiments shown in fig. 5, 12, 16, and 20, the detection zone 31 and the quality control zone 32 are disposed adjacent to each other.
The waste liquid zone 4 is used for collecting residual waste liquid after detection.
The shape of the waste liquid area 4 is not particularly limited, and may be, for example, an approximate square (as shown in fig. 1 to 9), a U-shape, or a double-layer (as shown in fig. 25 to 26) or a multi-layer structure formed by square and U-shape.
In the plate-like microfluidic element, the waste liquid region 4 is provided with a waste liquid absorber housing groove 41 (as shown in fig. 9). The waste liquid absorbing member may be absorbent cotton or the like.
The microfluidic device may further comprise detection aids such as a housing.
The invention also provides application of the microfluidic device in preparing detection products.
The detection product can be biological detection product or chemical detection product. In one embodiment, the test product is a diagnostic product. Such as nucleic acid detection products, antigen detection products, antibody detection products, biomarker detection products, and the like.
The invention also provides a detection method, which comprises the following steps: and adding the liquid to be detected into a sample adding area 1 of the microfluidic device, and checking or detecting an observation area 3 after the liquid to be detected flows into a waste liquid area 4 to obtain a detection result.
The detection method is a qualitative detection method or a semi-quantitative detection method.
The use method of the microfluidic device provided by the invention comprises the following steps:
and adding the liquid to be measured into the sample adding area 1, enabling the liquid to be measured to flow into the mixing area 2 through the sample adding area 1, reacting with the reagent in the mixing area 2, enabling the liquid to be measured to continuously flow into the detection area 31 and the quality control area 32 after the reaction is finished, and absorbing the residual waste liquid after the reaction is finished by the waste liquid area 4. The micro-fluidic element directly observes the color change of the detection partition 31 and the quality control partition 32 after washing or without washing, if the color of the quality control partition 32 changes, the micro-fluidic device is indicated to work normally, and if the quality control partition does not change, the micro-fluidic device is indicated to work abnormally and needs to be detected again. On the premise that the microfluidic device works normally, if the color of the detection partition 31 does not change, the liquid to be detected does not contain the detected target substance, and if the color of the detection partition 31 changes, the liquid to be detected contains the detected target substance. Further, if the content of the measured target substance in the liquid to be measured is high, the color of the entire area of the detection partition 31 will be deep, and if the content of the measured target substance in the liquid to be measured is low, the color of the area of the detection partition 31 through which the liquid to be measured flows first will be deep, and the color of the area through which the liquid to be measured flows later will be light.
The regular and sequential arrangement of the sintered or bonded microspheres can promote the liquid to be tested to swim in sequence to form a regular liquid layer, thereby improving the combination efficiency and resolution. Meanwhile, the microfluidic device can swim without sealing and packaging the flow channels, and the multi-layer flow channels can improve the combination capacity and are not easy to block, so that the microfluidic device can be cleaned, the resolution is improved, and the false positive or false negative rate is reduced. The special flow channel design of the invention can lead the reactant to uniformly pass through a narrow channel to perform the concentration function, thereby further improving the detection sensitivity.
In summary, the present invention effectively overcomes the disadvantages of the prior art and has high industrial utility value.
The above embodiments are merely illustrative of the principles of the present invention and its effectiveness, and are not intended to limit the invention. Modifications and variations may be made to the above-described embodiments by those skilled in the art without departing from the spirit and scope of the invention. Accordingly, it is intended that all equivalent modifications and variations of the invention be covered by the claims, which are within the ordinary skill of the art, be within the spirit and scope of the present disclosure.

Claims (18)

1. The microfluidic device is characterized by comprising a microfluidic element formed by microsphere sintering or bonding, the microfluidic element sequentially comprises a sample adding area (1), a mixing area (2), an observation area (3) and a waste liquid area (4), a reagent capable of reacting with a detected substance in a liquid to be detected and a reagent capable of reacting with a quality control substance in the liquid to be detected are arranged in the mixing area (2) so as to respectively form a detected substance conjugate and a quality control substance conjugate, the observation area (3) comprises a detection partition (31) and a quality control partition (32), the detection partition (31) is internally provided with a reagent capable of reacting with the detected substance conjugate, and the quality control partition (32) is internally provided with a substance capable of reacting with the quality control substance conjugate.
2. The microfluidic device of claim 1, wherein the microspheres are polymeric microspheres; preferably, the polymeric microspheres are selected from polystyrene microspheres or organic glass microspheres.
3. The microfluidic device of claim 2, wherein the polymeric microspheres have a diameter of 30-120 microns and/or the polymeric microspheres are monodisperse microspheres; preferably, the monodisperse microsphere is a solid microsphere, a mesoporous microsphere or an SOS microsphere.
4. The microfluidic device of claim 2, wherein the polymeric microspheres are surface modified polymeric microspheres; preferably, the polymer microsphere is a polymer microsphere with a chemically modified surface; more preferably, the surface modification is a hydrophilic modification; more preferably, the hydrophilic modification is grafting of carboxyl, hydroxyl and/or amino groups to the surface of the polymeric microspheres.
5. The microfluidic device according to claim 1, wherein the microfluidic element has a porosity of 40-60% and/or a sintering temperature of 120-200 ℃.
6. The microfluidic device according to claim 1, wherein the microfluidic element is a plate-like microfluidic element or a rod-like microfluidic element.
7. Microfluidic device according to claim 1, characterized in that the sample application zone (1) is provided with a bifurcation for straining or guiding the liquid to be measured.
8. The microfluidic device according to claim 6, wherein a sample addition section (1) of the plate-like microfluidic device is formed with a impurity removal section (11) on the surface thereof; preferably, the impurity removing part (11) is arranged at one end of the near mixing zone (2); more preferably, the impurity removing unit (11) has a zigzag protrusion; more preferably, in the impurity removing unit (11), an included angle α between the front edge of the sawtooth and the base surface is larger than an included angle β between the rear edge of the sawtooth and the base surface.
9. The microfluidic device according to claim 6, wherein the mixing zone (2) of the plate-like microfluidic element comprises a plurality of flow channels intersecting and communicating with each other.
10. The microfluidic device according to claim 8, wherein in the plate-like microfluidic element, the mixing zone (2) and the viewing zone (3) surfaces are provided with corrugations (21); preferably, the folds (21) on the surfaces of the mixing zone (2) and the observation zone (3) are arranged on the same surface of the microfluidic element.
11. The microfluidic device according to claim 10, wherein the folds (21) of the impurity removal section (11) and the mixing zone (2) and the observation zone (3) are provided on opposite sides of the microfluidic device.
12. The microfluidic device according to claim 6, wherein the rod-like microfluidic element is provided with ribs (5) in an axial direction, the ribs (5) penetrating the sample application region (1), the mixing region (2), the observation region (3) and the waste region (4).
13. The microfluidic device according to claim 12, wherein in the rod-like microfluidic element the mixing zone (2) extends in a ring-shape and/or square-shape along the ribs (5) to the observation zone (3).
14. The microfluidic device according to claim 6, wherein in the rod-like microfluidic element, a plurality of reagents capable of capturing different substances to be tested are provided in the mixing zone (2); and/or the detection partition (31) is internally provided with a reagent which can react with a plurality of different detected substance combinations.
15. The microfluidic device according to claim 1, wherein the detection zone (31) and the quality control zone (32) are both block-shaped zones; preferably, the length of the detection partition (31) and/or the quality control partition (32) is 3-15 mm.
16. Microfluidic device according to claim 1, characterized in that the waste region (4) is provided with a waste absorber receiving slot (41).
17. Use of a microfluidic device according to any one of claims 1 to 16 for the preparation of a detection product.
18. A method of detection, the method comprising the steps of: adding a liquid to be detected into a sample adding area (1) of the microfluidic device according to any one of claims 1-16, and checking or detecting an observation area (3) after the liquid to be detected flows into a waste liquid area (4) to obtain a detection result.
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