CN117074655A - Microfluidic chip, chip box and tumor marker detection method - Google Patents
Microfluidic chip, chip box and tumor marker detection method Download PDFInfo
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- 238000001514 detection method Methods 0.000 title claims abstract description 93
- 239000000439 tumor marker Substances 0.000 title claims abstract description 22
- 238000006243 chemical reaction Methods 0.000 claims abstract description 73
- 239000007788 liquid Substances 0.000 claims abstract description 53
- 238000004806 packaging method and process Methods 0.000 claims abstract description 35
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 29
- 230000000087 stabilizing effect Effects 0.000 claims abstract description 13
- 238000007789 sealing Methods 0.000 claims description 23
- 238000004140 cleaning Methods 0.000 claims description 16
- 239000007850 fluorescent dye Substances 0.000 claims description 13
- UPSFMJHZUCSEHU-JYGUBCOQSA-N n-[(2s,3r,4r,5s,6r)-2-[(2r,3s,4r,5r,6s)-5-acetamido-4-hydroxy-2-(hydroxymethyl)-6-(4-methyl-2-oxochromen-7-yl)oxyoxan-3-yl]oxy-4,5-dihydroxy-6-(hydroxymethyl)oxan-3-yl]acetamide Chemical compound CC(=O)N[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@H]1[C@H](O)[C@@H](NC(C)=O)[C@H](OC=2C=C3OC(=O)C=C(C)C3=CC=2)O[C@@H]1CO UPSFMJHZUCSEHU-JYGUBCOQSA-N 0.000 claims description 13
- 239000002699 waste material Substances 0.000 claims description 9
- 238000011010 flushing procedure Methods 0.000 claims description 8
- 238000000034 method Methods 0.000 claims description 8
- 230000000712 assembly Effects 0.000 claims description 6
- 238000000429 assembly Methods 0.000 claims description 6
- 229940127121 immunoconjugate Drugs 0.000 claims description 5
- 239000000427 antigen Substances 0.000 claims description 4
- 102000036639 antigens Human genes 0.000 claims description 4
- 108091007433 antigens Proteins 0.000 claims description 4
- 239000011248 coating agent Substances 0.000 claims description 3
- 238000000576 coating method Methods 0.000 claims description 3
- 239000000243 solution Substances 0.000 description 8
- 206010028980 Neoplasm Diseases 0.000 description 6
- 230000009286 beneficial effect Effects 0.000 description 5
- 210000002966 serum Anatomy 0.000 description 5
- 201000010099 disease Diseases 0.000 description 4
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 4
- 238000010586 diagram Methods 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012544 monitoring process Methods 0.000 description 2
- 239000000758 substrate Substances 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 206010061902 Pancreatic neoplasm Diseases 0.000 description 1
- 238000006757 chemical reactions by type Methods 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 238000003018 immunoassay Methods 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 208000015486 malignant pancreatic neoplasm Diseases 0.000 description 1
- 201000002528 pancreatic cancer Diseases 0.000 description 1
- 208000008443 pancreatic carcinoma Diseases 0.000 description 1
- 239000000376 reactant Substances 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
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- G—PHYSICS
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- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
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- 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
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- 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
- G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
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Abstract
The invention discloses a microfluidic chip, which comprises a packaging module and a reaction module, wherein the packaging module is connected with the reaction module through a microfluidic pipeline, the packaging module is used for storing a reagent and a sample to be tested, and the reagent and the sample to be tested flow into the reaction module through the microfluidic pipeline for reaction; the microfluidic chip further comprises a pneumatic driving module, wherein the pneumatic driving module is communicated with the microfluidic pipeline and used for injecting gas into the microfluidic pipeline, the microfluidic pipeline is communicated with a pressure stabilizing pipeline, and the pressure stabilizing pipeline is communicated with an external environment and used for maintaining pressure stability in the microfluidic pipeline. The micro-fluidic chip fully injects reagents and samples required by the reaction into the reaction cavity through the pneumatic driving module, so that liquid residues in a micro-fluidic pipeline are reduced, and the detection precision is improved. The invention further discloses a chip box suitable for the microfluidic chip and a tumor marker detection method adopting the microfluidic chip.
Description
Technical Field
The invention belongs to the technical field of biomedical diagnosis, and particularly relates to a microfluidic chip, a chip box and a tumor marker detection method.
Background
Cancer is a disease that severely threatens human health, with a high incidence and mortality rate. The microfluidic chip has wide application prospect, and can realize high-sensitivity, high-flux and rapid detection analysis by flexibly designing the structure of the chip so as to be applied to high-efficiency tumor analysis.
The early detection of tumor markers has important significance for predicting the disease state, but the current products capable of realizing the instant detection and the home detection are less, so that the patient can usually discover the disease state only when the hospital is in medical treatment, and the disease state is discovered too late, thereby delaying treatment. In the existing microfluidic chip, the reaction reagent and the sample need to flow into the reaction area through the microfluidic pipeline, and due to the smaller diameter of the microfluidic pipeline, a part of residues are unavoidable to remain in the pipeline, so that the detection accuracy is affected.
Disclosure of Invention
In view of the above-described deficiencies of the prior art, the present invention aims to:
(1) The microfluidic chip comprises a pneumatic driving module, and can fully inject reagents and samples required by reaction into a reaction cavity, so that liquid residues in a microfluidic pipeline are reduced, and the detection precision is improved;
(2) Providing a chip card box matched with the microfluidic chip;
(3) The tumor marker detection method adopts the pneumatic driving module to inject the reagent and the sample, can draw a fluorescence standard curve, and has higher precision.
In order to achieve all or part of the above objects, the present invention provides the following technical solutions:
the invention provides a microfluidic chip, which comprises a packaging module and a reaction module, wherein the packaging module is connected with the reaction module through a microfluidic pipeline, the packaging module is used for storing a reagent and a sample to be tested, and the reagent and the sample to be tested flow into the reaction module through the microfluidic pipeline for reaction; the microfluidic chip further comprises a pneumatic driving module, wherein the pneumatic driving module is communicated with the microfluidic pipeline and used for injecting gas into the microfluidic pipeline, the microfluidic pipeline is communicated with a pressure stabilizing pipeline, and the pressure stabilizing pipeline is communicated with an external environment and used for maintaining pressure stability in the microfluidic pipeline. The technical scheme has the beneficial effects that the pneumatic driving module drives the reagent and the sample in the microfluidic pipeline to be fully injected into the reaction module by injecting gas into the microfluidic pipeline, so that the detection accuracy is prevented from being reduced due to the fact that the sample to be detected remains in the microfluidic pipeline; the pneumatic driving module can clean the pipeline by injecting gas to remove residues in the microfluidic pipeline.
The pneumatic driving module is arranged on one side of the packaging module, and gas is injected into the microfluidic pipeline along the flowing direction of the reagent and the sample to be tested in the microfluidic pipeline.
The pneumatic driving module comprises an air storage cavity, a first sealing element, a first passage and a second passage, wherein the first sealing element is movably arranged in the air storage cavity, two ends of the first passage are respectively connected with the air storage cavity and an external air source, and a first one-way valve is arranged on the first passage to control air in the external air source to enter the air storage cavity; the two ends of the second passage are respectively connected with the gas storage cavity and the microfluidic pipeline, and a second one-way valve is arranged on the second passage to control the gas in the gas storage cavity to be injected into the microfluidic pipeline. The technical scheme has the advantages that the first one-way valve controls the gas in the external gas source to enter the gas storage cavity through the first passage, and the second one-way valve controls the gas in the gas storage cavity to enter the micro-flow pipeline through the second passage, so that the injection of the gas into the micro-flow pipeline is realized.
The reaction module comprises at least one sample detection cavity, wherein the sample detection cavity is used for reacting a sample to be detected with a reagent.
The reaction module comprises a comparison module, the comparison module is connected with the sample detection cavity through the micro-flow pipeline, and the packaging module, the sample detection cavity and the comparison module are sequentially connected. The technical scheme has the beneficial effects that the contrast module is used for drawing the fluorescence standard curve, so that the signal data for drawing the standard curve and the signal for detecting the sample to be detected are obtained under the same reaction condition and detection condition, and the accuracy of the detection result is further ensured.
The control module comprises at least three control reaction chambers so as to ensure the accuracy of a standard curve.
The packaging module comprises a plurality of packaging components, each packaging component comprises a liquid storage cavity, a second sealing piece and a third one-way valve, a reagent or a sample to be tested is stored in each liquid storage cavity, the second sealing piece is detachably arranged at the liquid adding end of each liquid storage cavity, and the third one-way valve is arranged at the liquid outlet end of each liquid storage cavity and connected with the corresponding microfluidic pipeline.
The microfluidic chip further comprises a waste liquid collecting module, and the waste liquid collecting module is connected with the reaction module.
The invention also provides a chip box for placing the microfluidic chip in the technical scheme, which comprises a box body, wherein a first through hole, a second through hole and a liquid adding hole are formed in the box body, the first through hole is communicated with the pneumatic driving module, and gas enters the pneumatic driving module through the first through hole; the second through hole is communicated with the pressure stabilizing pipeline; the liquid adding holes are arranged corresponding to the packaging modules of the microfluidic chip and are used for adding reagents and adding samples.
The invention further provides a tumor marker detection method, the microfluidic chip in the technical scheme is adopted, a detection sample, a secondary antibody, a fluorescent substrate and a cleaning solution are arranged in the packaging module, and a coating antibody is arranged in the reaction module.
The detection method of the tumor marker comprises the following steps:
s1: enabling a detection sample to enter the sample detection cavity through the micro-flow pipeline, and injecting a cleaning liquid for flushing after the reaction is finished;
s2: enabling the secondary antibody to enter the sample detection cavity through the micro-flow pipeline, and injecting cleaning liquid for flushing after the reaction is finished;
s3: enabling a fluorogenic substrate to enter the sample detection cavity through the microfluidic pipeline to be reacted;
s4, detecting the fluorescence intensity of the sample detection cavity by adopting a chemiluminescent instrument, and calculating the concentration of the tumor marker in the sample according to the fluorescence intensity;
wherein, in at least one step of S1, S2, S3, the method further comprises the following steps: and starting a pneumatic driving module, and injecting gas into the microfluidic pipeline to inject at least one of a sample to be detected, a secondary antibody, a fluorogenic substrate and a cleaning solution in the microfluidic pipeline into the reaction module.
The detection method has the beneficial effects that the liquid in the microfluidic pipeline is discharged by adopting a method of injecting gas, so that the detection precision is improved.
Further, at least three groups of antigen-coated antibody conjugates with different concentrations are pre-embedded in the control module;
in the step S2, allowing a secondary antibody to enter the control module for reaction;
in said S3, allowing fluorogenic substrate to enter said control module for reaction;
in the step S4, detecting the fluorescence degree of the antigen-coated antibody conjugate of at least three groups of different concentrations, drawing a standard curve according to the fluorescence degree value, and calculating the tumor marker concentration corresponding to the fluorescence degree of the sample detection cavity according to the standard curve.
The detection method has the beneficial effects that the standard curve is drawn through the fluorescence, so that the signal data drawn in the standard curve and the signal for detecting the sample to be detected are obtained under the same reaction condition and detection condition, and the accuracy of the detection result is further ensured.
Compared with the prior art, the invention has at least the following beneficial effects:
(1) The microfluidic chip provided by the invention is provided with the pneumatic driving module, and the gas is injected into the microfluidic pipeline to drive the reagent and the sample in the microfluidic pipeline to be fully injected into the reaction module, so that the detection accuracy is prevented from being reduced due to the fact that the sample to be detected remains in the microfluidic pipeline; the pneumatic driving module can clean residues in the microfluidic pipeline by injecting gas so as to achieve the purpose of cleaning the pipeline;
(2) The microfluidic chip provided by the invention is provided with the contrast module for drawing the standard curve, so that the signal data for drawing the standard curve and the signal for detecting the sample to be detected are obtained under the same reaction condition and detection condition, and the accuracy of the detection result is further ensured;
(3) The tumor marker detection method provided by the invention adopts the microfluidic chip, can detect tumor markers of various cancers in time and at home, and provides an early detection method and an effective monitoring means which can be realized at home for cancer detection.
Drawings
In order to more clearly illustrate the technical solutions of specific embodiments of the present invention, the drawings that are needed in the description of the embodiments will be briefly described below, and it is obvious that the drawings described below are only some embodiments of the present invention, and other drawings may be obtained according to these drawings without inventive effort to a person of ordinary skill in the art.
FIG. 1 is a schematic diagram of a microfluidic chip in an embodiment of the present invention;
FIG. 2 is a schematic diagram of a pneumatic driving module of a microfluidic chip according to an embodiment of the present invention;
FIG. 3 is a schematic diagram of a chip case according to a second embodiment of the present invention;
FIG. 4 is a schematic view of a portion of a chip case according to a second embodiment of the present invention;
FIG. 5 is a standard curve obtained by the test in the third embodiment of the present invention;
reference numerals: 1-packaging module, 101-packaging assembly, 1011-liquid storage cavity, 1012-second sealing member, 1013-third one-way valve, 2-reaction module, 201-sample detection cavity, 202-control reaction cavity, 3-pneumatic driving module, 301-gas storage cavity, 302-first sealing member, 303-first one-way valve, 304-second one-way valve, 305-first passage, 306-second passage, 4-pressure stabilizing pipeline, 5-waste liquid collecting module, 6-chip box, 601-first through hole, 602-second through hole, 603-liquid adding hole.
Detailed Description
The following description of the embodiments of the present invention will be made clearly and fully, and it is apparent that the embodiments described are only some, but not all, of the embodiments of the present invention. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Example 1
The invention provides a microfluidic chip, as shown in figure 1, which comprises a packaging module 1 and a reaction module 2, wherein the packaging module 1 and the reaction module 2 are connected through a microfluidic pipeline, the packaging module 1 is used for storing a reagent and a sample to be tested, and the reagent and the sample flow into the reaction module 2 through the microfluidic pipeline for reaction; the micro-fluidic chip further comprises a pneumatic driving module 3, wherein the pneumatic driving module 3 is communicated with the micro-flow pipeline and used for injecting gas into the micro-flow pipeline, the micro-flow pipeline is communicated with a pressure stabilizing pipeline 4, and the pressure stabilizing pipeline is communicated with the external environment and used for maintaining pressure stability in the micro-flow pipeline. According to the invention, the pneumatic driving module 3 is arranged in the microfluidic chip, so that the pneumatic driving module 3 injects gas into the microfluidic pipeline, and reagents and samples in the microfluidic pipeline are fully injected into the reaction module 2 through the driving of the gas, so that the detection accuracy is prevented from being reduced due to the fact that samples to be detected remain in the microfluidic pipeline.
The pneumatic driving module 3 is arranged at one side of the packaging module 1, and gas is injected into the microfluidic pipeline along the flowing direction of the reagent and the sample to be tested.
Referring to fig. 1 and 2 in combination, the pneumatic driving module 3 includes a gas storage cavity 301, a first sealing member 302, a first check valve 303, a first passage 305, a second check valve 304 and a second passage 306, where the first sealing member 302 is movably installed in the gas storage cavity 301 and can reciprocate along the gas storage cavity 301 under the driving of gas; the two ends of the first passage 305 are respectively connected with the gas storage cavity 301 and an external gas source, the first one-way valve 303 is arranged on the first passage 305, the first one-way valve 303 controls gas in the external gas source to enter the gas storage cavity 302 through the first passage 305, and when the gas enters the gas storage cavity 301, the first sealing element 302 moves upwards along the gas storage cavity 301 under the driving of the gas; the two ends of the second passage 306 are respectively connected with the gas storage cavity and the microfluidic pipeline, the second one-way valve 304 is installed on the second passage 306, the second one-way valve 304 controls the gas in the gas storage cavity 301 to enter the microfluidic pipeline through the second passage 306 so as to drive the liquid in the microfluidic pipeline to be injected into the reaction module 2, and at the moment, the first sealing member 302 moves downwards along the gas storage cavity 302. The first seal 302 may be, for example, a sealing element or a sealing assembly such as a piston.
In one embodiment, the external air source is the environment, i.e., the pneumatic drive module 3 draws air from the environment into the air storage cavity 301. In other embodiments, the pneumatic driving module 3 may be externally connected with an air supply device such as an air tank.
The reaction module 2 comprises a sample detection cavity 201 and a control module, the control module is connected with the sample detection cavity 201 through a micro-flow pipeline, and the sample detection cavity 201 is positioned between the packaging module 1 and the control module. The control module comprises at least three control reaction chambers 202, and the control reaction chambers 202 are sequentially connected through a micro-flow pipeline. The control module is used for providing a control signal for drawing a standard curve and realizing drawing of a fluorescence standard curve. In the prior art, the detection environment for drawing the standard curve is different from the detection environment of the sample to be detected (for example, the concentration of the reagent used for drawing the standard curve is different from that of the reagent used for detecting the sample to be detected, the activity of the antibody is different, etc.), which can cause that the different errors between the two detection results cannot be eliminated. The contrast module in the microfluidic chip provided by the invention is used for drawing a standard curve, so that signal data for drawing the standard curve and a signal for detecting a sample to be detected are obtained under the same reaction condition and detection condition, and the accuracy of a detection result is further ensured. In this embodiment, the number of the reference reaction chambers 202 is four, and the number of the reference reaction chambers 202 may be set according to practical situations, such as chip size, detection environment, and required accuracy, but in order to ensure accuracy, the number of the reference reaction chambers 202 is preferably not less than 3.
The package module 1 comprises a plurality of package assemblies 101, wherein one package assembly 101 is used for storing a sample to be tested, and the other package assemblies 101 are used for storing different reagents. The packaging assembly 101 comprises a liquid storage cavity 1011, a second sealing element 1012, a third one-way valve 1013, wherein the second sealing element 1012 is detachably arranged at the liquid adding end of the liquid storage cavity, and the third one-way valve 1013 is arranged at the liquid outlet end of the liquid storage cavity and connected with a microfluidic pipeline for controlling the liquid in the liquid storage cavity 1011 to flow into the microfluidic pipeline. In this embodiment, the number of the package assemblies 101 is 4, and the number of the package assemblies 101 can be set according to the specific reaction type and the requirement. The second seal 1012 may be, for example, a sealing element or a sealing assembly such as a piston.
The microfluidic chip further comprises a waste liquid collecting module 5, and the waste liquid collecting module 5 is connected with the reaction module 2 through a microfluidic pipeline and is used for collecting waste liquid generated in the reaction process. In this embodiment, the pressure stabilizing pipeline 4 is connected to the microfluidic pipeline through the waste liquid collecting module 5, so as to maintain the pressure in the microfluidic pipeline stable.
Example two
Referring to fig. 3 and 4 in combination, the present embodiment provides a chip box for placing the microfluidic chip in the above technical solution, which includes a box body 6, and a first through hole 601, a second through hole 602, and a filling hole 603 are formed in the box body 6. The first through hole 601 is communicated with the pneumatic driving module 3, specifically, the first through hole 601 is communicated with the first passage 305 of the pneumatic driving module 601, and gas enters the first passage 305 of the pneumatic driving module 3 through the first through hole 601; the second through hole 602 is communicated with the pressure stabilizing pipeline 4; the liquid adding hole 603 is arranged corresponding to the packaging component 101 of the microfluidic chip, when reagents and samples are added, the second sealing piece 1012 in the packaging component 101 is taken out through the liquid adding hole 603, and after the reagents are added, the second sealing piece 101 is installed at the liquid adding end of the liquid storage cavity 1011 in a sealing mode.
Example III
The embodiment provides a detection method of a tumor marker, and the microfluidic chip in the technical scheme is adopted. The tumor marker detection method adopts a chemiluminescent immunoassay method, the packaging module 1 of the microfluidic chip comprises four packaging components 101 which are respectively used for storing a detection sample, a secondary antibody, a fluorescent substrate and a cleaning solution, and a coated antibody capable of being specifically combined with the tumor marker in the detection sample is laid in the sample detection cavity 201 of the reaction module 2.
The tumor marker detection method comprises the following steps:
s1: allowing a detection sample to enter a sample detection cavity 201 through a micro-flow pipeline, and injecting a cleaning solution for flushing after the reaction is finished;
s2: allowing the secondary antibody to enter a sample detection cavity 201 through a micro-flow pipeline, and injecting a cleaning liquid for flushing after the reaction is finished;
s3: allowing the fluorogenic substrate to enter the sample detection cavity 201 through a microfluidic pipeline to be reacted;
s4: detecting fluorescence of the sample detection cavity 201 by adopting a chemiluminescent instrument, and calculating the concentration of the tumor marker in the sample according to the fluorescence;
wherein, in at least one step of S1, S2, S3, the method further comprises the following steps: the pneumatic driving module 3 is started to inject gas into the microfluidic circuit so as to inject the sample to be detected, the secondary antibody, the fluorogenic substrate and/or the cleaning liquid in the microfluidic circuit into the sample detection cavity 201.
In one embodiment, the detection method can be used for pancreatic cancer monitoring and detection, CA19-9 is selected as a tumor marker, a sample is human serum, and the detection principle is as follows: in step S1, CA19-9 in serum is specifically combined with a coating antibody pre-embedded in a reaction detection cavity 201, and other substances which are not combined are washed by a washing liquid; in the step S2, the secondary antibody is combined with CA19-9, the secondary antibody can be combined with a fluorogenic substrate to emit light, and the excessive secondary antibody is washed by a washing liquid; in step S3, the fluorogenic substrate is sufficiently labeled with CA19-9 reactant; the content of CA19-9 in serum directly influences the content of a fluorogenic substrate marked on the secondary antibody, and the concentration of CA19-9 in serum can be obtained by detecting fluorescence by a chemiluminescent instrument. The pneumatic driving module 3 can enable liquid in the microfluidic pipeline to be injected into the sample detection cavity 201, and enable cleaning liquid to wash the sample detection cavity 201, so that the reaction accuracy is improved.
In one embodiment, a microfluidic chip with a control module comprising four control reaction chambers 202, CA 19-9-coated antibody conjugates of different concentrations are pre-embedded in the four control reaction chambers 202 is used. The tumor marker detection method adopting the microfluidic chip comprises the following steps:
s1: allowing a detection sample to enter a sample detection cavity 201 through a micro-flow pipeline, and injecting a cleaning solution for flushing after the reaction is finished;
s2: enabling the secondary antibody to enter a sample detection cavity 201 and four control reaction cavities 202 through a micro-flow pipeline, and injecting cleaning liquid for flushing after the reaction is finished;
s3: enabling a fluorogenic substrate to enter a sample detection cavity 201 and four control reaction cavities 202 through a microfluidic pipeline to be reacted;
s4, detecting the fluorescence of the sample detection cavity 201 and the fluorescence of the four control reaction cavities 202 by adopting a chemiluminescent instrument, drawing a standard curve of fluorescence-CA 19-9 concentration according to the fluorescence of the four control reaction cavities 202, and calculating the CA19-9 concentration in serum according to the standard curve and the fluorescence of the sample detection cavity 201.
By adopting the method, the fluorescence standard curve can be drawn, and the signal drawn by the standard curve and the sample detection signal are obtained under the same detection condition and the same reaction condition, so that the influence of different errors on the detection precision is avoided. As shown in fig. 5, the standard curve drawn by the method has good linearity, and the correlation coefficient (R2) is greater than 0.99.
The above description of the embodiments is only intended to assist in understanding the method and core idea of the invention. It should be noted that it will be apparent to those skilled in the art that various improvements and modifications can be made to the present invention without departing from the principles of the invention, and such improvements and modifications fall within the scope of the appended claims.
Claims (12)
1. A microfluidic chip, characterized in that: the device comprises a packaging module (1) and a reaction module (2), wherein the packaging module (1) and the reaction module (2) are connected through a micro-flow pipeline, the packaging module (1) is used for storing a reagent and a sample to be tested, and the reagent and the sample to be tested flow into the reaction module (2) through the micro-flow pipeline for reaction; the micro-fluidic chip further comprises a pneumatic driving module (3), the pneumatic driving module (3) is communicated with the micro-flow pipeline and is used for injecting gas into the micro-flow pipeline, the micro-flow pipeline is communicated with a pressure stabilizing pipeline (4), and the pressure stabilizing pipeline (4) is communicated with the external environment and is used for maintaining pressure stability in the micro-flow pipeline.
2. The microfluidic chip of claim 1, wherein: the pneumatic driving module (3) is arranged at one side of the packaging module (1), and gas is injected into the microfluidic pipeline along the flowing direction of the reagent and the sample to be tested in the microfluidic pipeline.
3. The microfluidic chip according to claim 1 or 2, wherein: the pneumatic driving module (3) comprises a gas storage cavity (301), a first sealing element (302), a first passage (305) and a second passage (306), wherein the first sealing element (302) is movably installed in the gas storage cavity (301), two ends of the first passage (305) are respectively connected with the gas storage cavity (301) and an external gas source, and a first one-way valve (303) is installed on the first passage (305) to control gas in the external gas source to enter the gas storage cavity (301); two ends of the second passage (306) are respectively connected with the gas storage cavity (301) and the microfluidic pipeline, and a second one-way valve (304) is arranged on the second passage (306) to control gas in the gas storage cavity (301) to be injected into the microfluidic pipeline.
4. The microfluidic chip of claim 1, wherein: the reaction module (2) comprises at least one sample detection cavity (201), wherein the sample detection cavity (201) is used for reacting a sample to be detected with a reagent.
5. The microfluidic chip according to claim 4, wherein: the reaction module (2) comprises a comparison module, the comparison module is connected with the sample detection cavity (201) through the microfluidic pipeline, and the packaging module (1), the sample detection cavity (201) and the comparison module are sequentially connected.
6. The microfluidic chip of claim 5, wherein: the control module includes at least three control reaction chambers (202).
7. The microfluidic chip of claim 1, wherein: the packaging module (1) comprises a plurality of packaging assemblies (101), the packaging assemblies (101) comprise a liquid storage cavity (1011), a second sealing element (1012) and a third one-way valve (1013), reagents or samples to be tested are stored in the liquid storage cavity (1011), the second sealing element (1012) is detachably arranged at the liquid adding end of the liquid storage cavity (1011), and the third one-way valve (1013) is arranged at the liquid outlet end of the liquid storage cavity (1011) and is connected with the microfluidic pipeline.
8. The microfluidic chip of claim 1, wherein: the microfluidic chip further comprises a waste liquid collecting module (5), and the waste liquid collecting module (5) is connected with the reaction module (2).
9. A cartridge for placing the microfluidic chip of any one of claims 1-8, characterized in that: the pneumatic device comprises a box body (6), wherein a first through hole (601), a second through hole (602) and a liquid adding hole (603) are formed in the box body (6), the first through hole (601) is communicated with the pneumatic driving module (3), and gas enters the pneumatic driving module (3) through the first through hole (601); the second through hole (602) is communicated with the pressure stabilizing pipeline (4); the liquid adding hole (603) is arranged corresponding to the packaging module (1) of the microfluidic chip and is used for adding reagents and adding samples.
10. A tumor marker detection method, which is characterized in that: the microfluidic chip according to claim 5 or 6 is adopted, a detection sample, a secondary antibody, a fluorogenic substrate and a cleaning solution are filled in the packaging module (1), and a coating antibody is pre-buried in the sample detection cavity (201).
11. The tumor marker detection method according to claim 10, wherein: the method comprises the following steps:
s1: enabling a detection sample to enter the sample detection cavity (201) through the micro-flow pipeline, and injecting a cleaning liquid for flushing after the reaction is finished;
s2: enabling the secondary antibody to enter the sample detection cavity (201) through the micro-flow pipeline, and injecting cleaning liquid for flushing after the reaction is finished;
s3: allowing a fluorogenic substrate to enter the sample detection cavity (201) through the microfluidic pipeline to be reacted;
s4, detecting the fluorescence degree of the sample detection cavity (201) by adopting a chemiluminescent instrument, and calculating the concentration of the tumor marker in the sample according to the fluorescence degree;
wherein, in at least one step of S1, S2, S3, the method further comprises the following steps: and starting a pneumatic driving module (3), and injecting gas into the microfluidic pipeline to inject at least one of a sample to be detected, a secondary antibody, a fluorogenic substrate and a cleaning solution in the microfluidic pipeline into the reaction module (2).
12. The tumor marker detection method according to claim 11, wherein: at least three groups of antigen-coated antibody conjugates with different concentrations are pre-embedded in the control module;
in the step S2, allowing a secondary antibody to enter the control module for reaction;
in said S3, allowing fluorogenic substrate to enter said control module for reaction;
in the step S4, the fluorescence degree of the antigen-coated antibody conjugate of at least three groups of different concentrations is detected, a standard curve is drawn according to the fluorescence degree value, and the tumor marker concentration corresponding to the fluorescence degree of the sample detection cavity (201) is calculated according to the standard curve.
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| CN202310051446.6A CN117074655A (en) | 2023-02-02 | 2023-02-02 | Microfluidic chip, chip box and tumor marker detection method |
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| CN202310051446.6A CN117074655A (en) | 2023-02-02 | 2023-02-02 | Microfluidic chip, chip box and tumor marker detection method |
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