CN113862140B - Portable liquid drop digital PCR device - Google Patents
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- CN113862140B CN113862140B CN202111136369.1A CN202111136369A CN113862140B CN 113862140 B CN113862140 B CN 113862140B CN 202111136369 A CN202111136369 A CN 202111136369A CN 113862140 B CN113862140 B CN 113862140B
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- 239000007788 liquid Substances 0.000 title claims abstract description 44
- 238000007847 digital PCR Methods 0.000 title claims abstract description 10
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- 239000008346 aqueous phase Substances 0.000 claims abstract description 24
- 239000000758 substrate Substances 0.000 claims abstract description 15
- 238000003752 polymerase chain reaction Methods 0.000 claims abstract description 5
- 238000012408 PCR amplification Methods 0.000 claims description 55
- 238000001917 fluorescence detection Methods 0.000 claims description 10
- 238000000034 method Methods 0.000 claims description 10
- 238000011304 droplet digital PCR Methods 0.000 claims description 9
- 238000005485 electric heating Methods 0.000 claims description 9
- 230000003321 amplification Effects 0.000 claims description 8
- 239000002184 metal Substances 0.000 claims description 8
- 238000003199 nucleic acid amplification method Methods 0.000 claims description 8
- 239000007850 fluorescent dye Substances 0.000 claims description 6
- 239000003292 glue Substances 0.000 claims description 5
- 239000004812 Fluorinated ethylene propylene Substances 0.000 claims description 4
- 239000004696 Poly ether ether ketone Substances 0.000 claims description 4
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 4
- HQQADJVZYDDRJT-UHFFFAOYSA-N ethene;prop-1-ene Chemical group C=C.CC=C HQQADJVZYDDRJT-UHFFFAOYSA-N 0.000 claims description 4
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- 239000011888 foil Substances 0.000 claims description 4
- 229920009441 perflouroethylene propylene Polymers 0.000 claims description 4
- 229920002530 polyetherether ketone Polymers 0.000 claims description 4
- 229920001721 polyimide Polymers 0.000 claims description 4
- -1 polytetrafluoroethylene Polymers 0.000 claims description 4
- 229920001343 polytetrafluoroethylene Polymers 0.000 claims description 4
- 239000004810 polytetrafluoroethylene Substances 0.000 claims description 4
- 230000008569 process Effects 0.000 claims description 4
- 239000000523 sample Substances 0.000 claims description 4
- 239000000741 silica gel Substances 0.000 claims description 4
- 229910002027 silica gel Inorganic materials 0.000 claims description 4
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- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 19
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- 229920000642 polymer Polymers 0.000 description 8
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- 238000006467 substitution reaction Methods 0.000 description 3
- 238000003491 array Methods 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6844—Nucleic acid amplification reactions
- C12Q1/6851—Quantitative amplification
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Abstract
The invention discloses a portable liquid drop digital PCR device which comprises a substrate, a movable slide block, an oil phase injector, an aqueous phase injector, a constant force spring, an injector fixing bracket, a connecting capillary, a liquid drop generating module, a PCR amplifying capillary, a first electrothermal film and a second electrothermal film, wherein the injector fixing bracket is fixed at one end of the substrate, the movable slide block is arranged at the other end of the substrate, the injector fixing bracket is respectively connected with one end of the oil phase injector, one end of the aqueous phase injector and one end of the constant force spring, the movable slide block is respectively connected with the other ends of the oil phase injector, the aqueous phase injector and the constant force spring, one end of the connecting capillary is respectively connected with the oil phase injector and the aqueous phase injector, the other end of the connecting capillary is connected with the liquid drop generating module, the output end of the liquid drop generating module is the PCR amplifying capillary, and the first electrothermal film and the second electrothermal film respectively wrap the PCR amplifying capillary. The invention can be widely applied to the technical field of polymerase chain reaction devices.
Description
Technical Field
The invention relates to the technical field of polymerase chain reaction devices, in particular to a portable liquid drop digital PCR device.
Background
Droplet digital polymerase chain reaction (ddPCR) is a method of separating PCR samples into water-in-oil droplets using microfluidic droplet technology, and can be used for absolute quantification of nucleic acids in samples. The injection module in the existing PCR device drives the water phase and the oil phase in a mechanical pump or pneumatic pump mode, the module volume is large, and the cost is high; one side of the amplification pipeline is heated by using a heating table, a heat conduction module or a heating plate in the amplification module, and the temperature of one side close to a heat source is higher than that of the other side, so that the pipeline is heated unevenly. In addition, one side of the pipeline in the low temperature area at the top is a heat conducting module, the other side is exposed to the air, and a large amount of heat can be taken away by air convection, so that the stability in the low temperature area is poor, and the ambient temperature obviously influences the amplification efficiency. In the conventional devices, a heating table or a heating plate with a large volume is required in a single heat source or multiple heat sources, and portability and light weight cannot be realized.
Disclosure of Invention
In view of the above, the embodiment of the invention provides a simple and practical portable droplet digital PCR device to realize portability of the device.
In one aspect, the invention provides a portable liquid drop digital PCR device, which comprises a substrate, a movable sliding block, an oil phase injector, an aqueous phase injector, a constant force spring, an injector fixing bracket, a connecting capillary, a liquid drop generating module, a first electrothermal film and a second electrothermal film, wherein the liquid drop generating module comprises a PCR amplification capillary, the injector fixing bracket is fixed at one end of the substrate, the movable sliding block is arranged at the other end of the substrate, the injector fixing bracket is respectively connected with one end of the oil phase injector, one end of the aqueous phase injector and one end of the constant force spring, the movable sliding block is respectively connected with the other ends of the oil phase injector, the aqueous phase injector and the constant force spring, one end of the connecting capillary is respectively connected with the oil phase injector and the aqueous phase injector, the other end of the connecting capillary is connected with the liquid drop generating module, the output end of the liquid drop generating module is the PCR amplification capillary, and the first electrothermal film and the second electrothermal film respectively wrap the PCR amplification capillary.
Optionally, the device further comprises a digital display temperature control console, wherein the digital display temperature control console is respectively connected with the first electrothermal film and the second electrothermal film and is used for controlling the temperatures of the first electrothermal film and the second electrothermal film.
Optionally, the device further comprises a fluorescence detection module, the fluorescence detection module comprises a microscope camera, an optical filter and a computer, wherein the microscope camera is used for shooting and recording the state of the liquid drops flowing through the observation area, the optical filter is used for filtering light except the fluorescent probe in the sample, and the computer is used for detecting the fluorescence intensity of each liquid drop passing through the observation area and counting the number of the liquid drops passing through the observation area.
Optionally, the temperature of the first electrothermal film is set to ninety-five degrees celsius.
Optionally, the temperature of the second electrothermal film is set to sixty degrees celsius.
Alternatively, the PCR amplification capillary includes a silica gel capillary, a fluorinated ethylene propylene copolymer capillary, a polytetrafluoroethylene capillary, and a polyether ether ketone capillary.
Alternatively, the PCR amplification capillary is processed by an extrusion or hot stretching process.
Optionally, the first electrothermal film and the second electrothermal film use metal foil and metal wire as inner electric heating body, polyimide film as outer insulating heat conducting layer, and back glue is arranged on the surface of the outer insulating heat conducting layer and used for fixing PCR amplification capillary tube.
Optionally, the first electrothermal film and the second electrothermal film wrap the PCR amplification capillary with a width ratio of three to one.
Compared with the prior art, the technical scheme provided by the invention has the following technical effects: the invention relates to a portable liquid drop digital PCR device which comprises a substrate, a movable slide block, an oil phase injector, an aqueous phase injector, a constant force spring, an injector fixing bracket, a connecting capillary, a liquid drop generating module, a PCR amplification capillary, a first electrothermal film and a second electrothermal film, wherein the injector fixing bracket is fixed at one end of the substrate, the movable slide block is arranged at the other end of the substrate, the injector fixing bracket is respectively connected with one end of the oil phase injector, one end of the aqueous phase injector and one end of the constant force spring, the movable slide block is respectively connected with the other ends of the oil phase injector, the aqueous phase injector and the constant force spring, one end of the connecting capillary is respectively connected with the oil phase injector and the aqueous phase injector, the other end of the connecting capillary is connected with the liquid drop generating module, the output end of the liquid drop generating module is the PCR amplification capillary, and the first electrothermal film and the second electrothermal film respectively wrap the PCR amplification capillary. According to the invention, the injector is driven by the constant force spring, so that the system structure can be simplified, the liquid drops in the PCR amplification capillary are heated by the first electrothermal film and the second electrothermal film, the heated uniformity and stability of the liquid drops are improved, and the portability of the device can be realized.
Drawings
In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings that are needed in the description of the embodiments will be briefly introduced below, and it is obvious that the drawings in the following description are only some embodiments of the present application, and that other drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a schematic diagram of a portable droplet digital PCR device according to an embodiment of the present invention;
fig. 2 is a schematic diagram of a droplet generation module of a portable droplet digital PCR device according to an embodiment of the present invention.
Detailed Description
In order to make the objects, technical solutions and advantages of the present application more apparent, the present application will be further described in detail with reference to the accompanying drawings and examples. It should be understood that the specific embodiments described herein are for purposes of illustration only and are not intended to limit the present application.
Referring to fig. 1, an embodiment of the present invention provides a portable droplet digital PCR device, including a substrate 1, a moving slide block 2, an oil phase injector 3, an aqueous phase injector 4, a constant force spring 5, an injector fixing support 6, a connecting capillary 7, a droplet generation module 8, a first electrothermal film 11 and a second electrothermal film 12, wherein the droplet generation module 8 includes a PCR amplification capillary 9, the injector fixing support 6 is fixed at one end of the substrate 1, the moving slide block 2 is disposed at the other end of the substrate 1, the injector fixing support 6 is respectively connected with one ends of the oil phase injector 3, the aqueous phase injector 4 and the constant force spring 5, the moving slide block 2 is respectively connected with the other ends of the oil phase injector 3, the aqueous phase injector 4 and the constant force spring 5, one end of the connecting capillary 7 is respectively connected with the oil phase injector 3 and the aqueous phase injector 4, the other end of the connecting capillary 7 is connected with the droplet generation module 8, the other end of the droplet generation module 8 is connected with the PCR amplification capillary 9, and the first electrothermal film 11 and the second electrothermal film 12 are respectively wrapped around the PCR amplification capillary 9.
The oil phase injector and the water phase injector are fixed on the injector fixing bracket, and tail pistons of the oil phase injector and the water phase injector are in contact with the movable sliding block. When the constant force spring is in a stretching state after the setting is finished, constant pulling force is provided for the movable sliding block, and the movable sliding block drives the oil phase injector and the water phase injector, so that the oil phase and the water phase inside the injector enter the liquid drop generating module together through the connecting capillary. The tension can be adjusted by the elastic coefficient of the constant force spring. Referring to fig. 2, the droplet generation module further includes an aqueous phase capillary 15, an oil phase capillary 16, a PCR amplification capillary 9, and a sealant 17, and generates the micro droplet 13 through the droplet generation module, wherein the aqueous phase capillary 15, the oil phase capillary 16, and the PCR amplification capillary 9 are all polymer capillaries, one end of each polymer capillary is an output end, the other end of each polymer capillary is an input end, an inner diameter of the output end is smaller than an inner diameter of the input end, the output end of the aqueous phase capillary 15 is inserted from the input end of the PCR amplification capillary 9 to the output end of the PCR amplification capillary 9, the output end of the oil phase capillary 16 is connected with the input end of the PCR amplification capillary 9, and the sealant 17 is used for sealing the input end of the PCR amplification capillary 9. The PCR amplification capillary 9 is an ultra-long pipeline for conveying micro-droplets, the input end of the PCR amplification capillary 99 is a pipeline with a thicker inner diameter, and the output end of the PCR amplification capillary 9 is a pipeline with a thinner inner diameter. The input ends of the water phase capillary 15 and the oil phase capillary 16 are connected with the connecting capillary, and the oil phase and the water phase inside the oil phase injector and the water phase injector are driven by the moving slide block to enter the oil phase capillary 16 and the water phase capillary 15 respectively. The output end of the oil phase capillary 16 is inserted into the input end of the PCR amplification capillary 9, and after the oil phase enters the oil phase capillary 16, the oil phase flows into the input end of the PCR amplification capillary 9 from the output end of the oil phase capillary 16 and flows to the output end of the PCR amplification capillary 9. The output end of the water phase capillary 15 is inserted into the output end of the PCR amplification capillary 9 from the input end of the PCR amplification capillary 9, the water phase directly flows into the output end of the PCR amplification capillary 9, and the water-in-oil liquid drops are formed under the action of the oil phase cutting force with high peripheral flow velocity. The sealant is sealed at the input end of the PCR amplification capillary 9 and is used for preventing leakage and the entry of external air.
Further as a preferred embodiment, referring to fig. 1, the apparatus further includes a digital display temperature control console 10, and the digital display temperature control console 10 is respectively connected to the first electrothermal film 11 and the second electrothermal film 12, and is used for controlling the temperatures of the first electrothermal film 11 and the second electrothermal film 12.
The temperature of the first electric heating film is controlled to be 95 ℃ through the digital display temperature control console, and the temperature of the second electric heating film is controlled to be 60 ℃ through the digital display temperature control console. The real-time temperature of the first electrothermal film and the second electrothermal film can be observed through the digital display temperature control console, and the first electrothermal film and the second electrothermal film can be accurately controlled.
Further as a preferred embodiment, referring to fig. 1, the apparatus further includes a fluorescence detection module 14, where the fluorescence detection module 14 includes a microscope camera, an optical filter, and a computer, where the microscope camera is used to capture and record a state of a droplet flowing through the observation area, the optical filter is used to filter light other than the fluorescent probe in the sample, and the computer is used to detect a fluorescence intensity of each droplet passing through the observation area and count a number of droplets passing through the observation area.
The output end of the PCR amplification capillary tube is an observation area, after the PCR amplification of the liquid drops is completed through 40 high-low temperature cycles, the fluorescent intensity detection and the statistics of the number of the liquid drops are carried out on the liquid drops in the observation area through a fluorescent detection module, a microscope camera is used for shooting and recording the state of the liquid drops flowing through the observation area, a light filter is further arranged in front of a lens of the microscope camera and used for filtering light except for a fluorescent probe in a sample, the microscope camera uploads recorded data to a computer, the computer analyzes the uploaded data, the fluorescent intensity of each liquid drop passing through the observation area and the number of the liquid drops passing through the observation area are detected, and finally a chart is automatically drawn and the data is stored.
Further as a preferred embodiment, the temperature of the first electrothermal film is set to ninety-five degrees celsius.
The temperature of the first electrothermal film is set to be ninety five ℃, and a PCR amplified capillary region wrapped by the first electrothermal film is a high temperature region and is used for denaturing target DNA in the high temperature region and unwinding the target DNA into two RNAs.
Further as a preferred embodiment, the temperature of the second electrothermal film is set to sixty degrees celsius.
The temperature of the second electrothermal film is set to be sixty ℃, and the PCR amplification capillary region wrapped by the second electrothermal film is a low-temperature region and is used for assembling RNA into 2 new DNA by utilizing bases and fluorescent probes in the water phase, so that the amplification of the DNA is completed.
Further preferred embodiments, the PCR amplification capillaries include silica gel capillaries, fluorinated ethylene propylene copolymer capillaries, polytetrafluoroethylene capillaries, and polyether ether ketone capillaries.
The oil phase capillary, the water phase capillary and the PCR amplification capillary are all polymer capillaries, and the polymer capillaries comprise silica gel capillaries, fluorinated ethylene propylene copolymer capillaries, polytetrafluoroethylene capillaries and polyether-ether-ketone capillaries, have the advantage of good hydrophobicity, and can be better applied to the generation of micro-droplets.
Further as a preferred embodiment, the PCR amplification capillary is processed by an extrusion or hot stretching process.
The oil phase capillary, the water phase capillary and the PCR amplification capillary are all polymer capillaries, and the polymer capillaries are obtained by extrusion or hot stretching process, so that the inner diameter and the outer diameter of the polymer capillaries can be better controlled, and the method is better applied to the generation of micro liquid drops.
Further as a preferred embodiment, the first electrothermal film and the second electrothermal film use metal foil and metal wire as inner electric heating body, polyimide film as outer insulating heat conducting layer, and the surface of the outer insulating heat conducting layer is provided with back glue for fixing PCR amplification capillary.
After the liquid drops are generated by the liquid drop generating module, the liquid drops continuously flow in the PCR amplification capillary tube, and pass through a high temperature region (95 ℃) wrapped by the first electrothermal film and a low temperature region (60 ℃) wrapped by the second electrothermal film in sequence, wherein the first electrothermal film and the second electrothermal film take metal foils and metal wires as inner conductive heating bodies, polyimide films as outer insulating heat conducting layers, the surface of the outer insulating heat conducting layers is provided with back glue, the back glue is used for fixing the PCR amplification capillary tube, and the first electrothermal film and the second electrothermal film are folded after the PCR amplification capillary tube is fixed, so that the upper surface and the lower surface of the PCR amplification capillary tube are heated uniformly.
Further as a preferred embodiment, the first electrothermal film and the second electrothermal film wrap the PCR amplification capillary with a width ratio of three to one.
Wherein, the time ratio required for the high temperature region and the low temperature region is 3:1, the width ratio of the PCR amplification capillary covered in the first electrothermal film and the second electrothermal film is 3:1, i.e. the time for the droplet to pass through the high temperature zone is 20 seconds and the time for the droplet to pass through the low temperature zone is 60 seconds. The target DNA is denatured in a high temperature region, and is unwound into two RNAs, and the RNAs pass through a low temperature region, and the RNAs are assembled into 2 new DNAs by utilizing bases and fluorescent probes in an aqueous phase, so that one round of amplification is completed. By analogy, after 40 cycles of amplification, the fluorescence intensity of the droplets containing DNA will be significantly enhanced, and finally pass through the fluorescence detection zone at the end of the PCR amplification capillary.
The invention relates to a realization flow of a portable liquid drop digital PCR device, which comprises the following steps: the water phase injector and the oil phase injector are driven by the movable slide block and the constant force spring, and the pulling force is controlled by the constant force spring, so that the water phase and the oil phase in the water phase injector and the oil phase injector enter the liquid drop generating module through the connecting capillary, and the water phase and the oil phase generate liquid drops through the liquid drop generating module and enter the PCR amplification capillary. The temperature of the first electrothermal film and the temperature of the second electrothermal film are controlled through a digital display temperature control console, a high-temperature area and a low-temperature area are formed in the PCR amplification capillary, the liquid drops pass through the high-temperature area and the low-temperature area to carry out PCR amplification reaction, and finally the liquid drops pass through a fluorescence detection area, namely an observation area, at the tail end of the PCR amplification capillary, and are subjected to fluorescence detection through a fluorescence detection module.
In the related art, the injection module drives the water phase and the oil phase in a mechanical pump or a pneumatic pump mode, so that the injection module is large in volume and high in cost; the PCR amplification module uses a heating table, a heat conduction module or a heating plate to heat one side of the amplification pipeline, so that the temperature of one side close to a heat source is higher than that of the other side, and the pipeline is heated unevenly. In addition, in both the single heat source and the multiple heat sources, a heating table or a heating plate having a large volume is required, and portability and weight saving cannot be achieved.
In summary, the embodiment of the invention has the following advantages:
(1) According to the embodiment of the invention, the injector is driven by the constant force spring, so that the system structure is simplified, the system volume and weight are reduced, the equipment cost is reduced, and the portability of the device is realized;
(2) According to the embodiment of the invention, the folded electrothermal film is used for carrying out wrapped heating on the micro-droplets flowing in the capillary, so that the uniformity and stability of heated ground are improved, the problem of influence of environmental temperature is solved, and the heated time ratio of different temperature areas is accurately controlled by controlling the heated areas of the high temperature area and the low temperature area, so that the target DNA is ensured to be amplified smoothly;
(3) Compared with the heating table, the electrothermal film in the embodiment of the invention greatly reduces the volume and weight of the system, and further realizes the portability of the device.
In some alternative embodiments, the functions/acts noted in the block diagrams may occur out of the order noted in the operational illustrations. For example, two blocks shown in succession may in fact be executed substantially concurrently or the blocks may sometimes be executed in the reverse order, depending upon the functionality/acts involved. Furthermore, the embodiments presented and described in the flowcharts of the present invention are provided by way of example in order to provide a more thorough understanding of the technology. The disclosed methods are not limited to the operations and logic flows presented herein. Alternative embodiments are contemplated in which the order of various operations is changed, and in which sub-operations described as part of a larger operation are performed independently.
Furthermore, while the invention is described in the context of functional modules, it should be appreciated that, unless otherwise indicated, one or more of the functions and/or features may be integrated in a single physical device and/or software module or may be implemented in separate physical devices or software modules. It will also be appreciated that a detailed discussion of the actual implementation of each module is not necessary to an understanding of the present invention. Rather, the actual implementation of the various functional modules in the apparatus disclosed herein will be apparent to those skilled in the art from consideration of their attributes, functions and internal relationships. Accordingly, one of ordinary skill in the art can implement the invention as set forth in the claims without undue experimentation. It is also to be understood that the specific concepts disclosed are merely illustrative and are not intended to be limiting upon the scope of the invention, which is to be defined in the appended claims and their full scope of equivalents.
The functions, if implemented in the form of software functional units and sold or used as a stand-alone product, may be stored in a computer-readable storage medium. Based on this understanding, the technical solution of the present invention may be embodied essentially or in a part contributing to the prior art or in a part of the technical solution in the form of a software product stored in a storage medium, comprising several instructions for causing a computer device (which may be a personal computer, a server, a network device, etc.) to perform all or part of the steps of the method of the embodiments of the present invention. And the aforementioned storage medium includes: a usb disk, a removable hard disk, a Read-only Memory (ROM), a random access Memory (RAM, random Access Memory), a magnetic disk, or an optical disk, or other various media capable of storing program codes.
Logic and/or steps represented in the flowcharts or otherwise described herein, e.g., a ordered listing of executable instructions for implementing logical functions, can be embodied in any computer-readable medium for use by or in connection with an instruction execution system, apparatus, or device, such as a computer-based system, processor-containing system, or other system that can fetch the instructions from the instruction execution system, apparatus, or device and execute the instructions. For the purposes of this description, a "computer-readable medium" can be any means that can contain, store, communicate, propagate, or transport the program for use by or in connection with the instruction execution system, apparatus, or device.
More specific examples (a non-exhaustive list) of the computer-readable medium would include the following: an electrical connection (electronic device) having one or more wires, a portable computer diskette (magnetic device), a Random Access Memory (RAM), a read-only memory (ROM), an erasable programmable read-only memory (EPROM or flash memory), an optical fiber device, and a portable compact disc read-only memory (CDROM). Additionally, the computer-readable medium may even be paper or other suitable medium upon which the program is printed, as the program may be electronically captured, via, for instance, optical scanning of the paper or other medium, then compiled, interpreted or otherwise processed in a suitable manner, if necessary, and then stored in a computer memory.
It is to be understood that portions of the present invention may be implemented in hardware, software, firmware, or a combination thereof. In the above-described embodiments, the various steps or methods may be implemented in software or firmware stored in a memory and executed by a suitable instruction execution system. For example, if implemented in hardware, as in another embodiment, may be implemented using any one or combination of the following techniques, as is well known in the art: discrete logic circuits having logic gates for implementing logic functions on data signals, application specific integrated circuits having suitable combinational logic gates, programmable Gate Arrays (PGAs), field Programmable Gate Arrays (FPGAs), and the like.
In the description of the present specification, a description referring to terms "one embodiment," "some embodiments," "examples," "specific examples," or "some examples," etc., means that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the present invention. In this specification, schematic representations of the above terms do not necessarily refer to the same embodiments or examples. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.
While embodiments of the present invention have been shown and described, it will be understood by those of ordinary skill in the art that: many changes, modifications, substitutions and variations may be made to the embodiments without departing from the spirit and principles of the invention, the scope of which is defined by the claims and their equivalents.
While the preferred embodiment of the present invention has been described in detail, the present invention is not limited to the embodiments, and those skilled in the art can make various equivalent modifications or substitutions without departing from the spirit of the present invention, and these equivalent modifications or substitutions are included in the scope of the present invention as defined in the appended claims.
Claims (8)
1. The portable liquid drop digital PCR device is characterized by comprising a substrate (1), a movable sliding block (2), an oil phase injector (3), an aqueous phase injector (4), a constant force spring (5), an injector fixing bracket (6), a connecting capillary tube (7), a liquid drop generating module (8), a first electrothermal film (11) and a second electrothermal film (12), wherein the liquid drop generating module (8) comprises a PCR (polymerase chain reaction) amplification capillary tube (9), the injector fixing bracket (6) is fixed at one end of the substrate (1), the movable sliding block (2) is arranged at the other end of the substrate (1), the injector fixing bracket (6) is respectively connected with one end of the oil phase injector (3), one end of the aqueous phase injector (4) and one end of the constant force spring (5), the movable sliding block (2) is respectively connected with the other end of the oil phase injector (3), one end of the aqueous phase injector (4) and the other end of the constant force spring (5), one end of the connecting capillary tube (7) is respectively connected with the oil phase injector (3) and the aqueous phase injector (4), the other end of the connecting capillary tube (7) is connected with the other end of the liquid drop generating module (8), the first electrothermal film (11) and the second electrothermal film (12) respectively wrap the PCR amplification capillary (9);
the device also comprises a digital display temperature control table (10), wherein the digital display temperature control table (10) is respectively connected with the first electric heating film (11) and the second electric heating film (12) and is used for controlling the temperature of the first electric heating film (11) and the temperature of the second electric heating film (12).
2. The portable digital PCR device of claim 1 further comprising a fluorescence detection module (14), the fluorescence detection module (14) including a microscope camera for capturing and recording the status of droplets flowing within the observation area, a light filter for filtering light outside of the fluorescent probe in the sample, and a computer for detecting the fluorescence intensity of each droplet passing through the observation area and counting the number of droplets passing through the observation area.
3. A portable droplet digital PCR device according to claim 1, characterized in that the temperature of the first electrothermal film (11) is set to ninety-five degrees celsius.
4. A portable droplet digital PCR device according to claim 1, characterized in that the temperature of the second electrothermal film (12) is set to sixty degrees celsius.
5. A portable droplet digital PCR device according to claim 3, characterized in that the PCR amplification capillary (9) comprises a silica gel capillary, a fluorinated ethylene propylene copolymer capillary, a polytetrafluoroethylene capillary and a polyetheretherketone capillary.
6. A portable droplet digital PCR device according to claim 5, characterized in that the PCR amplification capillary (9) is manufactured by extrusion or hot stretching processes.
7. The portable liquid drop digital PCR device according to claim 1, wherein the first electrothermal film (11) and the second electrothermal film (12) use metal foil and metal wire as inner electric heating elements, polyimide film as outer insulating heat conducting layer, and back glue is arranged on the surface of the outer insulating heat conducting layer and used for fixing the PCR amplification capillary (9).
8. A portable droplet digital PCR device according to claim 1, characterized in that the width ratio of the first electrothermal film (11) and the second electrothermal film (12) surrounding the PCR amplification capillary (9) is three to one.
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