CN114609388B - Microfluidic immunodetection method and device - Google Patents
Microfluidic immunodetection method and device Download PDFInfo
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- CN114609388B CN114609388B CN202210508816.XA CN202210508816A CN114609388B CN 114609388 B CN114609388 B CN 114609388B CN 202210508816 A CN202210508816 A CN 202210508816A CN 114609388 B CN114609388 B CN 114609388B
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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
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56966—Animal cells
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
- B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
- B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
- B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
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- 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
- B01L3/502707—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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- 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
Abstract
The invention relates to the technical field of cell detection, in particular to a microfluidic immunodetection method and a microfluidic immunodetection device, which comprise the following steps: receiving a structural schematic diagram of a microfluidic chip, cutting a chip material plate by using a laser cutting machine to obtain a microfluidic solid chip according to the structural schematic diagram of the microfluidic chip, setting compression parameters of a microfluidic vacuum hot press, inputting the microfluidic solid chip into the microfluidic vacuum hot press, performing compression to obtain a microfluidic compression chip, receiving an immunodetection reagent and blood to be detected, introducing the blood to be detected into an immunodetection layer, injecting the immunodetection reagent into the immunodetection layer containing the blood to be detected, uniformly mixing, standing in a dark place for a specified time at a preset temperature to obtain a mixed solution, centrifuging the mixed solution to obtain a layered solution, and determining an immunodetection result of the blood to be detected according to the layered solution. The invention can solve the problem of low efficiency of immunodetection.
Description
Technical Field
The invention relates to the technical field of cell detection, in particular to a microfluidic immunodetection method and a microfluidic immunodetection device.
Background
With the rapid development of medical level, various medical problems have been overcome one by one, but from the perspective of medical prevention, effective early discovery of physical abnormalities is an extremely important step, and particularly, the immune system, as a core system of a human body, is liable to cause cold, fever, tumor and even canceration when the immunity is low. Therefore, how to effectively detect the number and the proportion of immune cells of a human body so as to determine the immunocompetence of the human body is an extremely important health early warning measure.
At present, most of the immunoassay methods mainly adopt manual detection, for example, blood to be detected of a human body is collected and placed in a test tube, immunoassay reagent is injected into the test tube, and the immunoassay reagent is mixed, stood and shaken to separate the proportions of different immune systems. Although the manual detection method can achieve the detection purpose, the detection efficiency is low, and human resources are wasted.
Disclosure of Invention
The invention provides a microfluidic immunoassay method, a microfluidic immunoassay device and a computer readable storage medium, and mainly aims to solve the problem of low immunoassay efficiency.
In order to achieve the above object, the present invention provides a microfluidic immunoassay method, comprising:
receiving an immunodetection instruction, and receiving a microfluidic chip structure schematic diagram which is stored in a database in advance according to the immunodetection instruction, wherein the microfluidic chip structure schematic diagram shows 5 layers of microfluidic chips which are respectively a first protective layer, a second protective layer, a bottom embedded layer, an immunodetection layer and a top embedded layer;
the structural schematic diagram of the microfluidic chip is led into a laser cutting machine, wherein a chip material plate is stored in the laser cutting machine in advance;
according to the schematic structural diagram of the microfluidic chip, cutting the chip material plate by using the laser cutting machine to obtain a microfluidic solid chip, wherein the microfluidic solid chip comprises 5 layers, and the first protective layer, the second protective layer, the bottom embedded layer, the immunodetection layer and the top embedded layer are sequentially arranged from the bottom layer to the top layer;
starting a microfluidic vacuum hot press, setting compression parameters of the microfluidic vacuum hot press, inputting the microfluidic entity chip into the microfluidic vacuum hot press to perform compression, and obtaining a microfluidic compression chip;
receiving an immunodetection reagent and blood to be detected, introducing the blood to be detected into the immunodetection layer, injecting the immunodetection reagent into the immunodetection layer containing the blood to be detected, uniformly mixing, and standing at a preset temperature in a dark place for a specified time to obtain a mixed solution;
and centrifuging the mixed solution to obtain a layering solution, and determining the immunoassay result of the blood to be detected according to the layering solution.
Optionally, the introducing the schematic structural diagram of the microfluidic chip into a laser cutting machine, where a chip material plate is stored in the laser cutting machine in advance, includes:
starting the laser cutting machine, and checking whether the area, the quality and the thickness of a chip material plate stored in the laser cutting machine reach expected values;
and if the area, the quality and the thickness of the chip material plate do not reach the expected values, replacing a new chip material plate until the area, the quality and the thickness of the chip material plate reach the expected values, and leading the structural schematic diagram of the microfluidic chip into a laser cutting machine.
Optionally, the expected values include an area expected value of 25 square centimeters, a mass expected value of 0.35kg, and a thickness expected value of 3 centimeters.
Optionally, the cutting, by the laser cutting machine, the chip material plate according to the schematic structural diagram of the microfluidic chip to obtain a microfluidic entity chip includes:
analyzing the size of each layer of the first protective layer, the second protective layer, the bottom embedded layer, the immunodetection layer and the top embedded layer in the structural schematic diagram of the microfluidic chip, wherein the size comprises the thickness of the layer and the area of the layer;
cutting the chip material plate into five layers by using the laser cutting machine to obtain an original first protective layer, an original second protective layer, an original bottom embedded layer, an original immunodetection layer and an original top embedded layer, wherein the cutting size of each layer is sequentially the same as the layer thickness and the layer area in the schematic view of the microfluidic chip structure;
analyzing the surface texture of each layer according to the structural schematic diagram of the microfluidic chip;
sequentially engraving the original first protective layer, the original second protective layer, the original bottom end embedding layer, the original immunodetection layer and the original top end embedding layer according to the surface texture of each layer to obtain an engraved first protective layer, an engraved second protective layer, an engraved bottom end embedding layer, an engraved immunodetection layer and an engraved top end embedding layer;
and sequentially combining the carved first protective layer, the carved second protective layer, the carved bottom end embedding layer, the carved immunodetection layer and the carved top end embedding layer from the bottom layer to the top layer to obtain the microfluidic solid chip.
Optionally, the first protective layer has a layer thickness of 0.3 cm and a layer area of 16 cm; the layer thickness of the second protective layer is 0.2 cm, and the layer area is 16 square cm; the layer thickness of the bottom embedded layer is 0.5 cm, and the layer area is 12.25 square cm; the thickness of the immunodetection layer is 1 cm, and the area of the layer is 10.24 square centimeters; the layer thickness of the tip embedding layer was 0.5 cm and the layer area was 12.25 cm square.
Optionally, the immunoassay layer comprises a blood storage region, a fluid channel region, a flexible valve region, and a detection reaction region.
Optionally, after the blood to be detected is introduced into the immunoassay layer, the immunoassay reagent is injected into the immunoassay layer containing the blood to be detected and mixed uniformly, and the mixture is kept standing for a specified time at a preset temperature in a dark place, so as to obtain a mixed solution, including:
closing the flexible valve area, and introducing the blood to be detected into the blood storage area;
injecting the immunodetection reagent into the detection reaction area, opening the flexible valve area, and draining the blood to be detected in the blood storage area to the detection reaction area;
and after vibrating the micro-fluidic solid chip, standing the vibrated micro-fluidic solid chip for 3min to obtain the mixed solution.
Optionally, the centrifuging the mixed solution to obtain a stratified solution, includes:
placing the micro-fluidic chip comprising the mixed liquid in a centrifuge;
adding hemolysin into the mixed solution and standing for 3 minutes at room temperature in a dark environment;
setting the centrifugal frequency of the centrifugal machine, starting the centrifugal machine, and centrifuging the mixed solution to obtain the layering solution.
Optionally, the compression parameters include a back pressure, a positive pressure, a compression temperature and a compression time, wherein the back pressure is between 0.7MPa and 0.8MPa, the positive pressure is between 0.35MPa and 0.45MPa, the compression temperature is between 52 ° and 62 °, and the compression time is between 5min and 6 min.
In order to solve the above problems, the present invention also provides a microfluidic immunoassay device, comprising:
the system comprises an immunodetection starting module, a database, a micro-fluidic chip and a data processing module, wherein the immunodetection starting module is used for receiving an immunodetection instruction and receiving a micro-fluidic chip structural schematic diagram which is stored in the database in advance according to the immunodetection instruction, and the micro-fluidic chip structural schematic diagram comprises 5 layers which are respectively a first protective layer, a second protective layer, a bottom end embedded layer, an immunodetection layer and a top end embedded layer;
the chip cutting module is used for guiding the micro-fluidic chip structure schematic diagram into a laser cutting machine, wherein a chip material plate is stored in the laser cutting machine in advance, and the chip material plate is cut by the laser cutting machine according to the micro-fluidic chip structure schematic diagram to obtain a micro-fluidic entity chip, wherein the micro-fluidic entity chip comprises 5 layers, and the first protective layer, the second protective layer, the bottom end embedding layer, the immunodetection layer and the top end embedding layer are sequentially arranged from the bottom layer to the top layer;
the compression module is used for starting the microfluidic vacuum hot press, setting compression parameters of the microfluidic vacuum hot press, and inputting the microfluidic entity chip into the microfluidic vacuum hot press to perform compression to obtain a microfluidic compression chip;
the mixing module is used for receiving an immunodetection reagent and blood to be detected, introducing the blood to be detected into the immunodetection layer, injecting the immunodetection reagent into the immunodetection layer containing the blood to be detected, uniformly mixing the immunodetection reagent and the immunodetection layer, and standing the mixture for a specified time in a dark place at a preset temperature to obtain a mixed solution.
And the centrifugal detection module is used for centrifuging the mixed solution to obtain a layering solution, and determining the immunodetection result of the blood to be detected according to the layering solution.
In order to solve the above problem, the present invention also provides an electronic device, including:
a memory storing at least one instruction; and
and the processor executes the instructions stored in the memory to realize the microfluidic immunoassay method.
In order to solve the above problem, the present invention further provides a computer-readable storage medium having at least one instruction stored therein, where the at least one instruction is executed by a processor in an electronic device to implement the microfluidic immunoassay method as described above.
In order to solve the problems in the background art, an immunodetection instruction is received firstly, and a schematic structure diagram of a microfluidic chip stored in a database in advance is received according to the immunodetection instruction, wherein the microfluidic chip shown in the schematic structure diagram of the microfluidic chip comprises 5 layers, namely a first protective layer, a second protective layer, a bottom embedded layer, an immunodetection layer and a top embedded layer; further, the structural schematic diagram of the microfluidic chip is led into a laser cutting machine, wherein a chip material plate is stored in the laser cutting machine in advance, the chip material plate is cut by the laser cutting machine according to the structural schematic diagram of the microfluidic chip to obtain a microfluidic solid chip, wherein the microfluidic solid chip comprises 5 layers, the first protective layer, the second protective layer, the bottom embedding layer, the immunodetection layer and the top embedding layer are sequentially arranged from the bottom layer to the top layer, a microfluidic vacuum hot press is started, after the compression parameters of the microfluidic vacuum hot press are set, the microfluidic solid chip is input into the microfluidic vacuum hot press to be compressed to obtain a microfluidic compression chip, the manufacturing process of the microfluidic chip is sequentially cutting and compressing, and the microfluidic compression chip is subjected to high-precision operation by the laser cutting machine and the microfluidic vacuum hot press, the chip manufacturing of higher specification can be realized, subsequent detection efficiency is improved, finally, the immunoassay reagent is received and the blood to be detected is received, the blood to be detected is led into to be detected after the immunoassay layer, the immunoassay reagent is injected into the immunoassay layer including the blood to be detected and is uniformly mixed, and is kept stand for the appointed time in a dark place at the preset temperature, mixed liquid is obtained, the mixed liquid is centrifuged, the mixed liquid is obtained layered liquid, the immunoassay result of the blood to be detected is determined according to the layered liquid, it is seen that the immunoassay is realized in the chip without artificial excessive test tube passing operation, and the automation degree is higher. Therefore, the microfluidic immunoassay method, the microfluidic immunoassay device, the electronic equipment and the computer readable storage medium provided by the invention can solve the problem of low immunoassay efficiency.
Drawings
FIG. 1 is a schematic flow chart of a microfluidic immunoassay method according to an embodiment of the present invention;
FIG. 2 is an exemplary graph of the psychrometric chart of FIG. 1;
FIG. 3 is a diagram of a design of one of the microfluidic chips of the embodiment of FIG. 1;
FIG. 4 is a functional block diagram of a microfluidic immunoassay device according to an embodiment of the present invention;
fig. 5 is a schematic structural diagram of an electronic device for implementing the microfluidic immunoassay method according to an embodiment of the present invention.
The implementation, functional features and advantages of the present invention will be further described with reference to the accompanying drawings.
Detailed Description
It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.
The embodiment of the application provides a microfluidic immunoassay method. The execution subject of the microfluidic immunoassay method includes, but is not limited to, at least one of the electronic devices that can be configured to execute the method provided by the embodiments of the present application, such as a server, a terminal, and the like. In other words, the microfluidic immunoassay method may be performed by software or hardware installed in a terminal device or a server device. The server includes but is not limited to: a single server, a server cluster, a cloud server or a cloud server cluster, and the like.
Fig. 1 is a schematic flow chart of a microfluidic immunoassay method according to an embodiment of the present invention. In this embodiment, the microfluidic immunoassay method comprises:
and S1, receiving an immunodetection instruction, and receiving a microfluidic chip structure schematic diagram pre-stored in a database according to the immunodetection instruction, wherein the microfluidic chip structure schematic diagram shows 5 layers, namely a first protective layer, a second protective layer, a bottom end embedding layer, an immunodetection layer and a top end embedding layer.
It is explained that people with low immunity are susceptible to common cold, fever, tumor and even canceration. The human immune system consists of immune organs, immune cells, immune molecules and a network of lymphatic circulation. Immune cells reach immune functions by circulating blood to various organs and tissues of the whole body, wherein the main immune cells comprise T cells. How to effectively detect T cells of a human body so as to determine the immunocompetence of the human body is an extremely important health early warning measure.
It can be understood that after the immunodetection instruction is received, a testing environment for immunodetection needs to be constructed first, and the first step of the testing environment is to construct a microfluidic chip, so that the microfluidic chip can effectively provide a good immunodetection environment.
In the embodiment of the invention, the structural schematic diagram of the microfluidic chip is designed in advance by research personnel and stored in a database, and when an immunoassay needs to be executed, the structural schematic diagram of the microfluidic chip in the database can be directly called.
Further, in order to provide a superior immunoassay environment for the microfluidic chip, the microfluidic chip designed in the embodiment of the present invention has 5 layers, which are a first protection layer, a second protection layer, a bottom embedded layer, an immunoassay layer, and a top embedded layer. The first protective layer is generally arranged at the lowest end, the second protective layer is arranged on the first protective layer, the bottom embedded layer is connected with the second protective layer and the immunodetection layer, and the other surface of the immunodetection layer is connected with the top embedded layer, so that the complete microfluidic chip is constructed.
And S2, introducing the structural schematic diagram of the microfluidic chip into a laser cutting machine, wherein a chip material plate is stored in the laser cutting machine in advance.
It can be understood that the microfluidic chip of the entity class needs to be generated by combining the structural schematic diagram of the microfluidic chip, and in detail, the structural schematic diagram of the microfluidic chip is introduced into a laser cutting machine, wherein a chip material plate is stored in the laser cutting machine in advance, and the method comprises the following steps:
starting the laser cutting machine, and checking whether the area, the quality and the thickness of a chip material plate stored in the laser cutting machine reach expected values;
and if the area, the quality and the thickness of the chip material plate do not reach the expected values, replacing a new chip material plate until the area, the quality and the thickness of the chip material plate reach the expected values, and leading the structural schematic diagram of the microfluidic chip into a laser cutting machine.
In the embodiment of the invention, the model of the laser cutting machine is VLS3.50 Universal, and the material of the chip material plate is polymethyl methacrylate. The expected values comprise an area expected value, a mass expected value and a thickness expected value, wherein the area expected value is 25 square centimeters, the mass expected value is 0.35kg, and the thickness is 3 centimeters, namely when the area of the chip material plate does not reach 25 square centimeters, or the mass does not reach 0.35kg, or the thickness does not reach 3 centimeters, the area and the mass of the chip material plate do not reach the expected values. And when the area, the mass and the thickness of the chip material plate are all larger than expected values, guiding the structural schematic diagram of the micro-fluidic chip into a VLS3.50 Universal laser cutting machine to perform cutting operation.
And S3, cutting the chip material plate by using the laser cutting machine according to the schematic structural diagram of the microfluidic chip to obtain the microfluidic solid chip, wherein the microfluidic solid chip comprises 5 layers, and the first protective layer, the second protective layer, the bottom end embedding layer, the immunodetection layer and the top end embedding layer are sequentially arranged from the bottom layer to the top layer.
In detail, referring to fig. 2, the obtaining of the microfluidic solid chip by cutting the chip material plate with the laser cutting machine according to the schematic structural diagram of the microfluidic chip includes:
s31, analyzing the size of each layer of the first protection layer, the second protection layer, the bottom embedded layer, the immunodetection layer and the top embedded layer in the structural schematic diagram of the microfluidic chip, wherein the size comprises the layer thickness and the layer area;
s32, cutting the chip material plate into five layers by using the laser cutting machine to obtain an original first protective layer, an original second protective layer, an original bottom end embedded layer, an original immunodetection layer and an original top end embedded layer, wherein the cutting size of each layer is the same as the layer thickness and the layer area in the schematic view of the microfluidic chip structure;
s33, analyzing the surface texture of each layer according to the schematic structural diagram of the microfluidic chip;
s34, sequentially engraving the original first protection layer, the original second protection layer, the original bottom end embedding layer, the original immunodetection layer and the original top end embedding layer according to the surface texture of each layer to obtain an engraved first protection layer, an engraved second protection layer, an engraved bottom end embedding layer, an engraved immunodetection layer and an engraved top end embedding layer;
and S35, sequentially combining the carved first protective layer, the carved second protective layer, the carved bottom end embedding layer, the carved immunodetection layer and the carved top end embedding layer from the bottom layer to the top layer to obtain the microfluidic solid chip.
It should be explained that the microfluidic chip constructed in the embodiment of the present invention includes 5 layers, which are the first protection layer, the second protection layer, the bottom embedded layer, the immunodetection layer, and the top embedded layer, respectively, and in order for the laser cutting machine to accurately cut the microfluidic chip with 5 layers of structures, the size of each layer needs to be shown in the schematic structural diagram of the microfluidic chip, where the size includes the layer thickness and the layer area.
In detail, the layer thickness of the first protective layer is 0.3 cm, and the layer area is 16 cm; the layer thickness of the second protective layer is 0.2 cm, the layer area is 16 square centimeters; the thickness of the bottom embedded layer is 0.5 cm, and the layer area is 12.25 square cm; the thickness of the immunodetection layer is 1 cm, and the area of the layer is 10.24 square cm; the top embedded layer had a layer thickness of 0.5 cm and a layer area of 12.25 cm.
Therefore, after the layer thickness and the area of each layer are sequentially mastered in the laser cutting machine, the chip material plate can be divided into five, and the original first protection layer, the original second protection layer, the original bottom end embedding layer, the original immunodetection layer and the original top end embedding layer with the same layer thickness and area are obtained.
Referring to fig. 3, in an example of a microfluidic solid chip according to an embodiment of the present invention, since functions of each layer are different in a five-layer structure of the microfluidic chip, a first protection layer is used to protect safety of the whole microfluidic chip placed on a device stage, and the first protection layer needs to be designed according to structure adaptability of the device stage, if the device stage includes two convex clamping grooves, surface texture in the first protection layer should include two concave clamping grooves with the same size. The second protective layer is used for placing a prevention layer for the first protective layer to break to cause the abnormity of the whole microfluidic chip. The bottom embedded layer and the top embedded layer are both used for protecting the safety of the immunodetection layer, and the immunodetection layer is used as a core layer for carrying out immunodetection. Therefore, the surface lines of each layer are different because the functions of each layer are different.
In detail, the immunodetection layer comprises a blood storage region, a fluid channel region, a flexible valve region and a detection reaction region, wherein the blood storage region is used for storing blood to be detected, the fluid channel region is used for introducing the blood to be detected into the detection reaction region, the flexible valve region is used for controlling the opening and closing of a channel of the fluid channel region, and the detection reaction region is used for detecting the concentration of immune cells of the blood to be detected.
And S4, starting the microfluidic vacuum hot press, setting the compression parameters of the microfluidic vacuum hot press, and inputting the microfluidic solid chip into the microfluidic vacuum hot press to perform compression to obtain a microfluidic compression chip.
It can be explained that, in order to combine the engraved first protection layer, the engraved second protection layer, the engraved bottom end embedding layer, the engraved immunodetection layer and the engraved top end embedding layer firmly together to obtain the microfluidic solid chip, the engraved first protection layer, the engraved second protection layer, the engraved bottom end embedding layer, the engraved immunodetection layer and the engraved top end embedding layer need to be compressed.
In the embodiment of the invention, the compression parameters comprise backpressure pressure, positive pressure, compression temperature and compression time, and specifically, the backpressure pressure is between 0.7MPa and 0.8MPa, the positive pressure is between 0.35MPa and 0.45MPa, the compression temperature is between 52 degrees and 62 degrees, and the compression time is between 5min and 6 min. Wherein the positive pressure is the side of the engraving tip embedding layer and the back pressure is the side of the engraving first protection layer.
S5, receiving an immunodetection reagent and blood to be detected, introducing the blood to be detected into the immunodetection layer, injecting the immunodetection reagent into the immunodetection layer containing the blood to be detected, mixing uniformly, and standing in a dark place at a preset temperature for a specified time to obtain a mixed solution.
In detail, after the blood to be detected is introduced into the immunodetection layer, the immunodetection reagent is injected into the immunodetection layer containing the blood to be detected and is uniformly mixed, and the mixture is kept stand for a specified time at a preset temperature in a dark place to obtain a mixed solution, which comprises:
closing the flexible valve area, and introducing the blood to be detected into the blood storage area;
injecting the immunodetection reagent into the detection reaction area, opening the flexible valve area, and draining the blood to be detected in the blood storage area to the detection reaction area;
and after vibrating the micro-fluidic solid chip, standing the vibrated micro-fluidic solid chip for 3min to obtain the mixed solution.
In the embodiment of the invention, the cell number of immune cells, particularly T cells, in blood to be detected is mainly detected, so that the used immunoassay reagent mainly comprises CD4-FITC \ CD8-PE \ CD3-APC and the like. The immunoassay reagent can perform chemical reaction with different immunocytes in blood, so that chemical substances with different qualities are generated, and a layering phenomenon can be generated in a mixed solution.
S6, centrifuging the mixed solution to obtain a layering solution, and determining the immunodetection result of the blood to be detected according to the layering solution.
In detail, the centrifuging the mixed solution to obtain a stratified solution includes:
placing the microfluidic chip comprising the mixed solution in a centrifuge;
adding hemolysin into the mixed solution and standing for 3 minutes at room temperature in a dark environment;
setting the centrifugal frequency of the centrifugal machine, starting the centrifugal machine, and centrifuging the mixed solution to obtain the layering solution.
In the embodiment of the invention, the hemolysin can be prepared by FACSLYSING, and has the main effects of quickly dissolving red blood cells and releasing hemoglobin, thereby improving the accuracy of immunoassay. In addition, chemical substances with different qualities are generated under the action of different centrifugal forces through the chemical reaction of the immunoassay reagent and the blood to be detected, so that the original mixed solution is converted into a stratified solution, different layers in the stratified solution represent the proportion of different immune cells, and an immunoassay result is deduced according to the proportion of different immune cells.
In order to solve the problems in the background art, the embodiment of the invention firstly receives an immunodetection instruction, and receives a microfluidic chip structural schematic diagram which is stored in a database in advance according to the immunodetection instruction, wherein the microfluidic chip structural schematic diagram shows 5 layers, namely a first protective layer, a second protective layer, a bottom embedded layer, an immunodetection layer and a top embedded layer, so that the microfluidic chip is used for replacing a test tube to execute immunodetection, and the microfluidic chip comprises 5 layers of structures, so that the design is more reasonable and safer, and the problem of low immunodetection efficiency caused by manual operation is prevented; further, the structural schematic diagram of the microfluidic chip is led into a laser cutting machine, wherein a chip material plate is stored in the laser cutting machine in advance, the chip material plate is cut by the laser cutting machine according to the structural schematic diagram of the microfluidic chip to obtain a microfluidic solid chip, wherein the microfluidic solid chip comprises 5 layers, the first protective layer, the second protective layer, the bottom embedding layer, the immunodetection layer and the top embedding layer are sequentially arranged from the bottom layer to the top layer, a microfluidic vacuum hot press is started, after the compression parameters of the microfluidic vacuum hot press are set, the microfluidic solid chip is input into the microfluidic vacuum hot press to be compressed to obtain a microfluidic compression chip, the manufacturing process of the microfluidic chip is sequentially cutting and compressing, and the microfluidic compression chip is subjected to high-precision operation by the laser cutting machine and the microfluidic vacuum hot press, the chip manufacturing of higher specification can be realized, subsequent detection efficiency is improved, finally, the immunoassay reagent is received and the blood to be detected is received, the blood to be detected is led into to be detected after the immunoassay layer, the immunoassay reagent is injected into the immunoassay layer including the blood to be detected and is uniformly mixed, and is kept stand for the appointed time in a dark place at the preset temperature, mixed liquid is obtained, the mixed liquid is centrifuged, the mixed liquid is obtained layered liquid, the immunoassay result of the blood to be detected is determined according to the layered liquid, it is seen that the immunoassay is realized in the chip without artificial excessive test tube passing operation, and the automation degree is higher. Therefore, the microfluidic immunoassay method, the microfluidic immunoassay device, the electronic equipment and the computer readable storage medium provided by the invention can solve the problem of low immunoassay efficiency.
Fig. 4 is a functional block diagram of a microfluidic immunoassay device according to an embodiment of the present invention.
The microfluidic immunoassay device 100 of the present invention may be installed in an electronic device. According to the realized functions, the microfluidic immunoassay device 100 may include an immunoassay starting module 101, a chip cutting module 102, a compression module 103, a mixing module 104, and a centrifugal detection module 105. The module of the present invention, which may also be referred to as a unit, refers to a series of computer program segments that can be executed by a processor of an electronic device and that can perform a fixed function, and that are stored in a memory of the electronic device.
The immunoassay starting module 101 is configured to receive an immunoassay instruction, and receive a schematic structure diagram of a microfluidic chip pre-stored in a database according to the immunoassay instruction, where the schematic structure diagram of the microfluidic chip includes 5 microfluidic chips, which are a first protective layer, a second protective layer, a bottom embedded layer, an immunoassay layer, and a top embedded layer;
the chip cutting module 102 is configured to introduce the schematic structural diagram of the microfluidic chip into a laser cutting machine, where a chip material plate is pre-stored in the laser cutting machine, and the chip material plate is cut by the laser cutting machine according to the schematic structural diagram of the microfluidic chip to obtain a microfluidic solid chip, where the microfluidic solid chip has 5 layers, and the first protective layer, the second protective layer, the bottom end embedding layer, the immunodetection layer, and the top end embedding layer are sequentially arranged from the bottom layer to the top layer;
the compression module 103 is used for starting the microfluidic vacuum hot press, setting compression parameters of the microfluidic vacuum hot press, and inputting the microfluidic solid chip into the microfluidic vacuum hot press to perform compression to obtain a microfluidic compression chip;
the mixing module 104 is configured to receive an immunoassay reagent and blood to be detected, introduce the blood to be detected into the immunoassay layer, inject the immunoassay reagent into the immunoassay layer including the blood to be detected, mix the reagents uniformly, and stand the mixture for a specified time at a preset temperature in a dark place to obtain a mixed solution;
the centrifugation detection module 105 is configured to centrifuge the mixed solution to obtain a stratified solution, and determine an immunoassay result of the blood to be detected according to the stratified solution.
In detail, the specific implementation manner of using each module in the microfluidic immunoassay device 100 according to the embodiment of the present invention is the same as that in embodiment 1, and is not repeated herein.
Fig. 5 is a schematic structural diagram of an electronic device for implementing a microfluidic immunoassay method according to an embodiment of the present invention.
The electronic device 1 may comprise a processor 10, a memory 11 and a bus 12, and may further comprise a computer program, such as a microfluidic immunoassay method program, stored in the memory 11 and executable on the processor 10.
The memory 11 includes at least one type of readable storage medium, which includes flash memory, removable hard disk, multimedia card, card-type memory (e.g., SD or DX memory, etc.), magnetic memory, magnetic disk, optical disk, etc. The memory 11 may in some embodiments be an internal storage unit of the electronic device 1, e.g. a removable hard disk of the electronic device 1. The memory 11 may also be an external storage device of the electronic device 1 in other embodiments, such as a plug-in mobile hard disk, a Smart Media Card (SMC), a Secure Digital (SD) Card, a Flash memory Card (Flash Card), and the like, which are provided on the electronic device 1. Further, the memory 11 may also include both an internal storage unit and an external storage device of the electronic device 1. The memory 11 may be used not only to store application software installed in the electronic device 1 and various types of data, such as codes of programs of the microfluidic immunoassay method, but also to temporarily store data that has been output or will be output.
The processor 10 may be composed of an integrated circuit in some embodiments, for example, a single packaged integrated circuit, or may be composed of a plurality of integrated circuits packaged with the same or different functions, including one or more Central Processing Units (CPUs), microprocessors, digital Processing chips, graphics processors, and combinations of various control chips. The processor 10 is a Control Unit (Control Unit) of the electronic device, connects various components of the electronic device by using various interfaces and lines, and executes various functions and processes data of the electronic device 1 by operating or executing programs or modules (such as microfluidic immunoassay method programs) stored in the memory 11 and calling data stored in the memory 11.
The bus 12 may be a Peripheral Component Interconnect (PCI) bus, an Extended Industry Standard Architecture (EISA) bus, or the like. The bus 12 may be divided into an address bus, a data bus, a control bus, etc. The bus 12 is arranged to enable connection communication between the memory 11 and at least one processor 10 or the like.
Fig. 5 only shows an electronic device with components, and it will be understood by a person skilled in the art that the structure shown in fig. 5 does not constitute a limitation of the electronic device 1, and may comprise fewer or more components than shown, or a combination of certain components, or a different arrangement of components.
For example, although not shown, the electronic device 1 may further include a power supply (such as a battery) for supplying power to each component, and preferably, the power supply may be logically connected to the at least one processor 10 through a power management device, so as to implement functions of charge management, discharge management, power consumption management, and the like through the power management device. The power supply may also include any component of one or more dc or ac power sources, recharging devices, power failure detection circuitry, power converters or inverters, power status indicators, and the like. The electronic device 1 may further include various sensors, a bluetooth module, a Wi-Fi module, and the like, which are not described herein again.
Further, the electronic device 1 may further include a network interface, and optionally, the network interface may include a wired interface and/or a wireless interface (such as a WI-FI interface, a bluetooth interface, etc.), which are generally used for establishing a communication connection between the electronic device 1 and other electronic devices.
Optionally, the electronic device 1 may further comprise a user interface, which may be a Display (Display), an input unit (such as a Keyboard), and optionally a standard wired interface, a wireless interface. Alternatively, in some embodiments, the display may be an LED display, a liquid crystal display, a touch-sensitive liquid crystal display, an OLED (Organic Light-Emitting Diode) touch device, or the like. The display, which may also be referred to as a display screen or display unit, is suitable for displaying information processed in the electronic device 1 and for displaying a visualized user interface, among other things.
It is to be understood that the described embodiments are for purposes of illustration only and that the scope of the appended claims is not limited to such structures.
The microfluidic immunoassay method program stored in the memory 11 of the electronic device 1 is a combination of instructions that, when executed in the processor 10, can implement:
receiving an immunodetection instruction, and receiving a microfluidic chip structure schematic diagram which is stored in a database in advance according to the immunodetection instruction, wherein the microfluidic chip structure schematic diagram shows 5 layers of microfluidic chips which are respectively a first protective layer, a second protective layer, a bottom embedded layer, an immunodetection layer and a top embedded layer;
the structural schematic diagram of the microfluidic chip is led into a laser cutting machine, wherein a chip material plate is stored in the laser cutting machine in advance, and the chip material plate is cut by the laser cutting machine according to the structural schematic diagram of the microfluidic chip to obtain a microfluidic solid chip, wherein the microfluidic solid chip comprises 5 layers, and the first protective layer, the second protective layer, the bottom end embedding layer, the immunodetection layer and the top end embedding layer are sequentially arranged from the bottom layer to the top layer;
starting a microfluidic vacuum hot press, setting compression parameters of the microfluidic vacuum hot press, inputting the microfluidic entity chip into the microfluidic vacuum hot press to perform compression, and obtaining a microfluidic compression chip;
receiving an immunodetection reagent and blood to be detected, introducing the blood to be detected into the immunodetection layer, injecting the immunodetection reagent into the immunodetection layer containing the blood to be detected, uniformly mixing, and standing at a preset temperature in a dark place for a specified time to obtain a mixed solution;
and centrifuging the mixed solution to obtain a layering solution, and determining the immunoassay result of the blood to be detected according to the layering solution.
Specifically, the specific implementation method of the processor 10 for the instruction may refer to the description of the relevant steps in the embodiments corresponding to fig. 1 to fig. 5, which is not repeated herein.
Further, the integrated modules/units of the electronic device 1, if implemented in the form of software functional units and sold or used as separate products, may be stored in a computer readable storage medium. The computer readable storage medium may be volatile or non-volatile. For example, the computer-readable medium may include: any entity or device capable of carrying said computer program code, recording medium, U-disk, removable hard disk, magnetic disk, optical disk, computer Memory, Read-Only Memory (ROM).
The present invention also provides a computer-readable storage medium, storing a computer program which, when executed by a processor of an electronic device, may implement:
receiving an immunodetection instruction, and receiving a microfluidic chip structure schematic diagram which is stored in a database in advance according to the immunodetection instruction, wherein the microfluidic chip structure schematic diagram shows 5 layers of microfluidic chips which are respectively a first protective layer, a second protective layer, a bottom embedded layer, an immunodetection layer and a top embedded layer;
the structural schematic diagram of the microfluidic chip is led into a laser cutting machine, wherein a chip material plate is stored in the laser cutting machine in advance, and the chip material plate is cut by the laser cutting machine according to the structural schematic diagram of the microfluidic chip to obtain a microfluidic solid chip, wherein the microfluidic solid chip comprises 5 layers, and the first protective layer, the second protective layer, the bottom end embedding layer, the immunodetection layer and the top end embedding layer are sequentially arranged from the bottom layer to the top layer;
starting a microfluidic vacuum hot press, setting compression parameters of the microfluidic vacuum hot press, inputting the microfluidic entity chip into the microfluidic vacuum hot press to perform compression, and obtaining a microfluidic compression chip;
receiving an immunodetection reagent and blood to be detected, introducing the blood to be detected into the immunodetection layer, injecting the immunodetection reagent into the immunodetection layer containing the blood to be detected, uniformly mixing, and standing at a preset temperature in a dark place for a specified time to obtain a mixed solution;
and centrifuging the mixed solution to obtain a layering solution, and determining the immunoassay result of the blood to be detected according to the layering solution.
In the embodiments provided in the present invention, it should be understood that the disclosed apparatus, device and method can be implemented in other ways. For example, the above-described apparatus embodiments are merely illustrative, and for example, the division of the modules is only one logical functional division, and other divisions may be realized in practice.
The modules described as separate parts may or may not be physically separate, and parts displayed as modules may or may not be physical units, may be located in one position, or may be distributed on multiple network units. Some or all of the modules may be selected according to actual needs to achieve the purpose of the solution of the present embodiment.
In addition, functional modules in the embodiments of the present invention may be integrated into one processing unit, or each unit may exist alone physically, or two or more units are integrated into one unit. The integrated unit can be realized in a form of hardware, or in a form of hardware plus a software functional module.
It will be evident to those skilled in the art that the invention is not limited to the details of the foregoing illustrative embodiments, and that the present invention may be embodied in other specific forms without departing from the spirit or essential attributes thereof.
The present embodiments are therefore to be considered in all respects as illustrative and not restrictive, the scope of the invention being indicated by the appended claims rather than by the foregoing description, and all changes which come within the meaning and range of equivalency of the claims are therefore intended to be embraced therein. Any reference signs in the claims shall not be construed as limiting the claim concerned.
Furthermore, it will be obvious that the term "comprising" does not exclude other elements or steps, and the singular does not exclude the plural. A plurality of units or means recited in the system claims may also be implemented by one unit or means in software or hardware. The terms second, etc. are used to denote names, but not to denote any particular order.
Finally, it should be noted that the above embodiments are only intended to illustrate the technical solutions of the present invention and not to limit the same, and although the present invention is described in detail with reference to the preferred embodiments, it should be understood by those skilled in the art that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention.
Claims (8)
1. A microfluidic immunoassay method, the method comprising:
receiving an immunodetection instruction, and receiving a microfluidic chip structural schematic diagram which is pre-stored in a database according to the immunodetection instruction, wherein the microfluidic chip structural schematic diagram comprises 5 layers, namely a first protective layer, a second protective layer, a bottom embedded layer, an immunodetection layer and a top embedded layer;
introducing the structural schematic diagram of the microfluidic chip into a laser cutting machine, wherein a chip material plate is stored in the laser cutting machine in advance;
according to the schematic structural diagram of the microfluidic chip, cutting the chip material plate by using the laser cutting machine to obtain a microfluidic solid chip, wherein the microfluidic solid chip comprises 5 layers, and the first protective layer, the second protective layer, the bottom embedded layer, the immunodetection layer and the top embedded layer are sequentially arranged from the bottom layer to the top layer;
starting a microfluidic vacuum hot press, setting compression parameters of the microfluidic vacuum hot press, inputting the microfluidic entity chip into the microfluidic vacuum hot press to perform compression, and obtaining a microfluidic compression chip;
receiving an immunodetection reagent and blood to be detected, introducing the blood to be detected into the immunodetection layer, injecting the immunodetection reagent into the immunodetection layer containing the blood to be detected, uniformly mixing, and standing at a preset temperature in a dark place for a specified time to obtain a mixed solution;
and centrifuging the mixed solution to obtain a layering solution, and determining the immunodetection result of the blood to be detected according to the layering solution.
2. The microfluidic immunoassay method of claim 1, wherein the introducing the microfluidic chip structure diagram into a laser cutting machine, wherein the laser cutting machine stores a chip material plate in advance, comprises: starting the laser cutting machine, and checking whether the area, the quality and the thickness of a chip material plate stored in the laser cutting machine reach expected values;
and if the area, the quality and the thickness of the chip material plate do not reach the expected values, replacing a new chip material plate until the area, the quality and the thickness of the chip material plate reach the expected values, and introducing the structural schematic diagram of the microfluidic chip into a laser cutting machine.
3. The microfluidic immunoassay method of claim 2, wherein the expected values comprise an area expected value of 25 square centimeters, a mass expected value of 0.35kg, and a thickness expected value of 3 centimeters.
4. The microfluidic immunodetection method according to claim 1, wherein the obtaining the microfluidic solid chip by cutting the chip material plate by the laser cutting machine according to the schematic structural diagram of the microfluidic chip comprises: analyzing the size of each layer of the first protective layer, the second protective layer, the bottom embedded layer, the immunodetection layer and the top embedded layer in the structural schematic diagram of the microfluidic chip, wherein the size comprises the thickness of the layer and the area of the layer;
cutting the chip material plate into five layers by using the laser cutting machine to obtain an original first protective layer, an original second protective layer, an original bottom embedded layer, an original immunodetection layer and an original top embedded layer, wherein the cutting size of each layer is sequentially the same as the layer thickness and the layer area in the schematic view of the microfluidic chip structure;
analyzing the surface texture of each layer according to the structural schematic diagram of the microfluidic chip;
sequentially engraving the original first protection layer, the original second protection layer, the original bottom end embedding layer, the original immunodetection layer and the original top end embedding layer according to the surface texture of each layer to obtain an engraved first protection layer, an engraved second protection layer, an engraved bottom end embedding layer, an engraved immunodetection layer and an engraved top end embedding layer;
and sequentially combining the carved first protective layer, the carved second protective layer, the carved bottom end embedded layer, the carved immunodetection layer and the carved top end embedded layer from the bottom layer to the top layer to obtain the microfluidic solid chip.
5. The microfluidic immunoassay method of claim 4, wherein the first protective layer has a layer thickness of 0.3 cm and a layer area of 16 cm; the layer thickness of the second protective layer is 0.2 cm, and the layer area is 16 square cm; the layer thickness of the bottom embedded layer is 0.5 cm, and the layer area is 12.25 square cm; the thickness of the immunodetection layer is 1 cm, and the layer area is 10.24 square cm; the layer thickness of the top embedded layer was 0.5 cm, and the layer area was 12.25 square cm.
6. The microfluidic immunoassay method of claim 1, wherein the immunoassay layer comprises a blood storage region, a fluid channel region, a flexible valve region, and a detection reaction region.
7. The microfluidic immunoassay method of claim 1, wherein the compression parameters comprise a back pressure between 0.7MPa and 0.8MPa, a positive pressure between 0.35MPa and 0.45MPa, a compression temperature between 52 ℃ and 62 ℃, and a compression time between 5min and 6 min.
8. A microfluidic immunoassay device, the device comprising:
the system comprises an immunodetection starting module, a database and a database, wherein the immunodetection starting module is used for receiving an immunodetection instruction and receiving a microfluidic chip structural schematic diagram which is stored in the database in advance according to the immunodetection instruction, and the microfluidic chip structural schematic diagram shows 5 layers, namely a first protective layer, a second protective layer, a bottom end embedding layer, an immunodetection layer and a top end embedding layer;
the chip cutting module is used for guiding the micro-fluidic chip structure schematic diagram into a laser cutting machine, wherein a chip material plate is stored in the laser cutting machine in advance, and the chip material plate is cut by the laser cutting machine according to the micro-fluidic chip structure schematic diagram to obtain a micro-fluidic entity chip, wherein the micro-fluidic entity chip comprises 5 layers, and the micro-fluidic entity chip comprises a first protective layer, a second protective layer, a bottom end embedding layer, an immunodetection layer and a top end embedding layer in sequence from a bottom layer to a top layer;
the compression module is used for starting the microfluidic vacuum hot press, setting compression parameters of the microfluidic vacuum hot press, and inputting the microfluidic entity chip into the microfluidic vacuum hot press to perform compression to obtain a microfluidic compression chip;
the mixing module is used for receiving an immunodetection reagent and blood to be detected, introducing the blood to be detected into the immunodetection layer, injecting the immunodetection reagent into the immunodetection layer containing the blood to be detected, uniformly mixing, and standing for a specified time in a dark place at a preset temperature to obtain a mixed solution;
and the centrifugal detection module is used for centrifuging the mixed solution to obtain a layering solution, and determining the immunodetection result of the blood to be detected according to the layering solution.
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