CN211111987U - Device for generating and fusing liquid drops - Google Patents
Device for generating and fusing liquid drops Download PDFInfo
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- CN211111987U CN211111987U CN201921380574.0U CN201921380574U CN211111987U CN 211111987 U CN211111987 U CN 211111987U CN 201921380574 U CN201921380574 U CN 201921380574U CN 211111987 U CN211111987 U CN 211111987U
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Abstract
The utility model relates to a device that droplet generated and fused, it includes: the two ends of the first pipeline respectively comprise a first connecting port and a second connecting port, one end of the second pipeline comprises a third connecting port, the other end of the second pipeline is communicated with the first pipeline, one end of the third pipeline comprises a fourth connecting port, the other end of the third pipeline is communicated with the first pipeline, one end of the fourth pipeline comprises a pipeline with 3 or more upstream branches and a fifth connecting port, the other end of the fourth pipeline is communicated with the first pipeline through a common pipeline, and the common pipeline comprises a continuous bending pipeline. The utility model discloses a little volume of microorganism, multifactor many condition are cultivateed and are detected.
Description
Technical Field
The utility model belongs to the technical field of the micro-fluidic, a little liquid drop generates and the technique of fusing is a simple and practical's realization multi-condition multifactor little liquid drop generates and wraps up microorganism or unicellular device.
Background
Conventional microbial strain screening and culture condition optimization are usually carried out in shake flasks, and the whole process is low in efficiency, high in cost, long in time and large in equipment and space. The micro-fluidic technology has extremely high efficiency, and as the structure is tiny, hundreds of microorganism culture units are easy to integrate on the chip at one time, so that a large amount of culture media can be saved; the integrated operation of software is adopted, the whole experiment process operation can be simulated on the chip, and the micro-fluidic chip is applied to culture and screening microorganisms, so that the problems can be effectively avoided.
Although the above patent documents all relate to microfluidic chips, since the breeding of microorganisms is a special biological process, different culture factors may be changed for culturing, and then the target microorganisms are screened out after detection and evaluation. Therefore, how to realize the culture and detection of the microorganism with the micro-volume and the multi-factor and multi-condition by the micro-fluidic technology is one of the key problems of microorganism breeding.
Documents of the prior art
Patent document
Patent document 1: chinese patent application publication CN 104099247A;
patent document 2: chinese patent application publication CN 104007091A;
patent document 3: chinese patent application publication CN 105944775A.
Disclosure of Invention
An object of the utility model is to overcome prior art's not enough, on micro-fluidic technology's basis, provide a little liquid drop and generate and device that fuses.
The purpose of the utility model is realized by the following technical scheme.
A micro-droplet generation and fusion device, comprising:
a first pipeline, two ends of which respectively comprise a first connecting port and a second connecting port,
a second pipeline, one end of which comprises a third connecting port and the other end of which is communicated with the first pipeline,
one end of the third pipeline comprises a fourth connecting port, the other end of the third pipeline is communicated with the first pipeline,
a fourth pipeline, one end of which comprises a pipeline with 3 or more branches at the upstream and a fifth connecting port, the other end of which is communicated with the first pipeline through a section of common pipeline, the section of common pipeline comprises a continuous bending pipeline,
wherein the first communication part of the second pipeline and the first pipeline is positioned between the second communication part of the third pipeline and the first pipeline and the third communication part of the fourth pipeline and the first pipeline, the distance between the first communication part and the second communication part and the distance between the first communication part and the third communication part are 500-2000 mu m respectively,
the first pipeline is communicated with the first power source through the first connecting port and the capillary pipeline, the first pipeline is communicated with the first valve through the second connecting port and the capillary pipeline, the second pipeline is communicated with the second power source and the second valve through the third connecting port and the capillary pipeline respectively, the third pipeline is communicated with the third valve through the fourth connecting port and the capillary pipeline, and the fourth pipeline is communicated with the power source connected with the branch and the corresponding control valve through the fifth connecting port and the capillary pipeline respectively.
The upstream branches of the fourth pipeline are 3, and the upstream branches are respectively a fourth pipeline a, a fourth pipeline b and a fourth pipeline c.
The upstream branches of the fourth pipelines are 4, and the four branches are respectively a fourth pipeline a, a fourth pipeline b, a fourth pipeline c and a fourth pipeline d.
The upstream branches of the fourth pipelines are 5, and the upstream branches are respectively a fourth pipeline a, a fourth pipeline b, a fourth pipeline c, a fourth pipeline d and a fourth pipeline e.
The continuous bent pipeline communicated with the first pipeline through the upstream branch of the fourth pipeline is formed by one or more structures of a P type, an S type, a U type, a broken line type and a wave type.
The total volume of the liquid contained in the continuously bent pipeline is 4-100 mu L, preferably 4-50 mu L, preferably 4-20 mu L, preferably 6-12 mu L, and further preferably 8-10 mu L.
The distance between the first communication part and the second communication part and the distance between the first communication part and the third communication part are 750-1800 mu m, and 1000-1500 mu m is further preferable.
The first communicating part is respectively the same as the second communicating part and the third communicating part in distance.
The first, second, third and fourth pipes are disposed on a substrate, and the substrate and the first, second, third and fourth pipes are formed of glass, Polymethylmethacrylate (PMMA), Polycarbonate (PC), Polystyrene (PS), acrylonitrile-butadiene-styrene copolymer (ABS), preferably Polymethylmethacrylate (PMMA).
The cross-sectional area of the first, second, third and fourth conduits ranges from 2.5 × 10 to 3mm2~4mm2，Preferably 0.01 to 3mm2More preferably 0.1 to 2.5mm2More preferably 0.25 to 1mm in thickness2It is further preferred that the cross-sectional areas of the first, second, third and fourth conduits are the same.
Drawings
Fig. 1 is a schematic structural diagram of a device for generating and fusing micro droplets according to the present invention.
Fig. 2 is a schematic diagram of a chip for generating and fusing micro droplets according to the present invention.
Description of the symbols:
1a first channel, 2 a second channel, 3 a third channel, 4a fourth channel (4a, 4b, 4c, 4d, 4e), 5a substrate, 7 holes, 8 holes, 9 a detection window I, 10 a detection window II, 11 a first connection port, 12 a second connection port, 21 a third connection port, 31 a fourth connection port, 41 '' '' a fifth connection port, 13 a first communication portion, 14 a second communication portion, 15 a third communication portion, 16 a fourth communication portion, 17 a fifth communication portion, 18 a sixth communication portion, 19 a seventh communication portion, 201 an electrode, 202 an electrode, 22 a first power source, 23 a second power source, 24 a third power source, 25 a fourth power source, 26 a fifth power source, 27 a sixth power source, 28 a seventh power source, 29 first valve, 30 second valve, 32 third valve, 33 fourth valve, 34 fifth valve, 35 sixth valve, 36 seventh valve, 37 eighth valve.
Detailed Description
Specific embodiments of the present invention will be described in more detail below with reference to the accompanying drawings. While specific embodiments of the invention are shown in the drawings, it will be understood that the invention may be embodied in various forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the invention to those skilled in the art.
It should be noted that certain terms are used throughout the description and claims to refer to particular components. As one skilled in the art will appreciate, various names may be used to refer to a component. This specification and claims do not intend to distinguish between components that differ in name but not function. In the following description and in the claims, the terms "include" and "comprise" are used in an open-ended fashion, and thus should be interpreted to mean "include, but not limited to. The following description is of the preferred embodiment of the invention, and is made for the purpose of illustrating the general principles of the invention and not for the purpose of limiting the invention. The protection scope of the present invention is subject to the limitations defined by the appended claims.
In order to facilitate understanding of embodiments of the present invention, the following description will be given by way of example of several embodiments with reference to the accompanying drawings, which do not limit the present invention.
A little liquid drop generate and device that fuses, as shown in figure 1, it includes:
a first pipeline, two ends of which respectively comprise a first connecting port and a second connecting port,
a second pipeline, one end of which comprises a third connecting port and the other end of which is communicated with the first pipeline,
one end of the third pipeline comprises a fourth connecting port, the other end of the third pipeline is communicated with the first pipeline,
a fourth pipeline, one end of which comprises a pipeline with 3 or more branches at the upstream and a fifth connecting port, the other end of which is communicated with the first pipeline through a section of common pipeline, the section of common pipeline comprises a continuous bending pipeline,
wherein the first communicating portion of the second duct and the first duct is located between the second communicating portion of the third duct and the first duct and the third communicating portion of the fourth duct and the first duct,
the first pipeline is communicated with a first power source through a first connecting port and a capillary pipeline, and is communicated with a first valve through a second connecting port and the capillary pipeline; the second pipeline is respectively communicated with a second power source and a second valve through a third connecting port and a capillary tube; the third pipeline is communicated with a third valve through a fourth connecting port and a capillary tube; and the fourth pipeline is respectively communicated with the power source connected with the branch and the corresponding control valve through a fifth connecting port and a capillary tube.
In a specific embodiment, the first, second, third and fourth channels are disposed on a substrate, the substrate is a microfluidic chip substrate formed of glass, polymethyl methacrylate (PMMA), Polycarbonate (PC), Polystyrene (PS) and acrylonitrile-butadiene-styrene (ABS), and the first, second, third and fourth channels are formed inside the substrate. The utility model discloses an among the micro-fluidic chip, first pipeline, second pipeline, third pipeline, fourth pipeline can the base plate inside through photoetching, hot pressing, sculpture, mould plastics etc. technology form, also can be independent fashioned pipeline.
The material of the pipeline is selected from any one of glass, polymethyl methacrylate (PMMA), Polycarbonate (PC), Polystyrene (PS) and acrylonitrile-butadiene-styrene copolymer (ABS), and the preferable forming material is polymethyl methacrylate (PMMA).
In the present embodiment, the cross-sectional shapes of the first duct, the second duct, the third duct, and the fourth duct are not limited, and may be any shapes that are convenient for forming and facilitating the flow of droplets, such as a circle, a rectangle, and an ellipse, and the cross-sectional area thereof is in the range of 2.5 × 10-3mm2~4mm2Preferably 0.01 to 3mm in thickness2More preferably 0.1 to 2.5mm2More preferably 0.25 to 1mm in thickness2。
In a specific embodiment, the cross-sectional areas of the first, second, third and fourth conduits may be the same or different from each other. The first, second, third and fourth conduits may be tapered. In the present invention, the cross-sectional area of the pipe can be limited as long as it satisfies the above-mentioned limitation. The shape and cross-sectional area of the conduit can be designed by one skilled in the art depending on the size of the droplets contained in the conduit and the intended use of the droplets. In some embodiments, it is further preferable that the first conduit, the second conduit, the third conduit and the fourth conduit have the same shape and cross-sectional area, and under the same condition, the pressure of the liquid in the conduits can be ensured to be consistent, so as to facilitate the movement operation of the liquid drops.
In a specific embodiment, the distance between the first communicating part and the second communicating part and the distance between the first communicating part and the third communicating part are 500 to 2000 μm, preferably 750 to 1800 μm, and preferably 1000 to 1500 μm. Specifically, the distance may be 600 μm, 700 μm, 800 μm, 900 μm, 1000 μm, 1100 μm, 1200 μm, 1300 μm, 1400 μm, 1500 μm, 1600 μm, 1700 μm, 1800 μm, 1900 μm.
In one embodiment, the distances between the first communication portion and the second and third communication portions may be different or the same. In one embodiment, the first communication site is the same distance from the second communication site to the third communication site.
In an embodiment, the second duct, the third duct, and the fourth duct upstream branch duct are arranged in parallel with each other. The distance between the upstream branch pipelines of the fourth pipeline is 500-2000 mu m, preferably 750-1800 mu m, and preferably 1000-1500 mu m. Specifically, the upstream branch pipes of the fourth pipe are arranged in parallel with each other, and the distance between the upstream branch pipes is related to the size of liquid drops in the pipe and the cross-sectional area of the pipe. Specifically, the particles may be 600 μm, 700 μm, 800 μm, 900 μm, 1000 μm, 1100 μm, 1200 μm, 1300 μm, 1400 μm, 1500 μm, 1600 μm, 1700 μm, 1800 μm, 1900 μm. The spacing of the upstream branch conduits of the fourth conduit may be the same or different.
In a specific embodiment, the upstream branch of the fourth pipeline is composed of a nonlinear pipeline, preferably a continuously curved pipeline, and further preferably one or more of a P-type, an S-type, a U-type, a broken line type and a wave type, which is in communication with the first pipeline. That is, the structure may be a single P-type, S-type, U-type, zigzag-type, or wave-type repeating structure, or a combination of a plurality of structures in the shape, and may be regularly alternated or randomly combined.
No matter what shape is the nonlinear pipeline or the bent pipeline, the volume of the nonlinear pipeline or the bent pipeline is not smaller than the total volume of liquid contained in the nonlinear pipeline or the bent pipeline, preferably 4-100 mu L, preferably 4-50 mu L, preferably 4-20 mu L, preferably 6-12 mu L, further preferably 8-10 mu L, further, the total volume of the liquid contained in the nonlinear pipeline is 4-100 mu L, preferably 4-50 mu L, preferably 4-20 mu L, preferably 6-12 mu L, further preferably 8-10 mu L.
The utility model discloses in can realize the high accuracy through using the power supply, steady no pulsatory liquid transmission. In a specific embodiment of the present invention, the first power source, the second power source, and the fourth pipe-connected power source are independently selected from any one of an injection pump, a pressure pump, a peristaltic pump, a diaphragm pump, and/or a plunger pump, respectively, and preferably, the first power source, the second power source, and the fourth pipe-connected power source are injection pumps. In the present invention, the size of the range of the power source is not limited, and those skilled in the art can appropriately select the injection pump, the pressure pump, the peristaltic pump, the diaphragm pump and/or the plunger pump with a suitable range according to the amount of the sample to be injected.
The utility model discloses in through using the valve to and change the pressure in each airtight pipeline through opening and closing the valve, control the flow direction of liquid drop in each pipeline through the change of pressure. The utility model discloses in, first valve, second valve, third valve to and the control valve of the fifth interface connection of fourth pipeline branch independently select any kind in solenoid valve, rotary valve, rocking arm valve, the pinch-off valve respectively. Preferably, the control valves connected to the first valve, the second valve, the third valve, and the fifth connecting port of the fourth pipe branch are rotary valves.
In some embodiments, it will be understood by those skilled in the art that the valve may be replaced by other mechanisms or components, such as a syringe pump used as a power source to serve as the valve, as long as the pressure in the closed pipeline can be changed by opening and closing.
The utility model discloses in, first power supply, second power supply are only the power supply that is used for showing the performance difference and acts on to do not intend to inject the quantity of power supply, first power supply can be the first power supply of several, and the second power supply can be several second power supply.
The present invention is directed to a valve having different functions, and is not intended to limit the number of valves, and the first valve may be a plurality of first valves, the second valve may be a plurality of second valves, and the third valve may be a plurality of third valves.
In a particular embodiment of the invention, the capillary channel has a cross-sectional area in the range of 2.5 × 10- 3mm2~4mm2Preferably 0.01 to 3mm in thickness2More preferably 0.1 to 2.5mm2More preferably 0.25 to 1mm in thickness2。
The pipeline connecting port is connected with the capillary, the connecting position is sealed, and then the pipeline connecting port is communicated with the power source and/or the valve through the capillary. In order to ensure the stable conduction of the power source pressure, the capillary tube is a hard tube, and the pipeline has no flexible change. Further preferably, the capillary tube is any one of a polytetrafluoroethylene tube (PTFE tube), a copolymer tube of perfluoropropyl perfluorovinyl ether and polytetrafluoroethylene (PFA tube), a polyetheretherketone tube (PEEK tube), a polycarbonate tube (PC tube), and a polystyrene tube (PS tube).
In an embodiment of the present invention, the first pipeline, the second pipeline, the third pipeline, and the fourth pipeline are disposed on a substrate to form a micro-fluidic chip, i.e. a chip for generating and fusing micro-droplets, the substrate is a micro-fluidic chip substrate, and there are 2 holes 7 and 8 on the chip substrate, the holes 7 and 8 are located near the first communication portion 13 of the first pipeline 1 and the second pipeline 2, and the positions of the holes are not fixed, and can be located on both sides of the second pipeline 2, or on both sides of the first pipeline 1, respectively, the distance between the holes 7 and 8 and the first communication portion 13 of the first pipeline 1 and the first communication portion 13 of the second pipeline 2 is 0.1mm-1cm, preferably 0.3mm-5mm, and further preferably 0.5mm-2mm, the holes 7 and 8 are used for accommodating the plus and minus poles 201 and 202 of the fusion electrode that are additionally disposed, and those skilled in the art can set the holes 7 and 201, 201 according to the chip design requirements in the above ranges, And 8 position. The frequency of the voltage loaded on the fusion electrode is 0-20000Hz, preferably 1000-10000Hz, and the voltage of the electrode is 1-5000V, preferably 500-1000V. When the fusion electrode is connected with a power supply, an electric field is generated to act on the liquid drops at the first communication part, and the purpose is to promote the liquid drop fusion. The electric field can be any of an alternating electric field or a constant electric field, and the voltage applied by the electrodes is 1-5000V, preferably 500-1000V.
Of course, in the present invention, the specific shape of the holes 7 and 8 and the size of the holes are not limited as long as the fusion electrodes can be placed. Wherein the fusion electrode may refer to the fusion electrode described above for the device of the present invention.
In a specific embodiment, the chip substrate further includes a first detection window 9 and a second detection window 10 formed on the first pipeline, the first detection window 9 and the second detection window 10 are only required to be used for monitoring and detecting micro-droplets in the pipeline, and there is no limitation on the specific shape and form thereof, if the basic material of the chip is formed by a transparent material, the first detection window 9 and the second detection window 10 are two detection points of the first pipeline, and if the chip substrate and the pipeline material are opaque, the first detection window 9 and the second detection window 10 are located on the first pipeline itself to form two transparent portions, that is, both the pipeline and the substrate material are transparent here. The position of the second detection window 10 is not limited, preferably, the first detection window is arranged on the first pipeline and is positioned between the third communication part and the first connection port, and the distance between the first detection window and the third communication part is 1mm-1cm, preferably 1.5-3mm, so that the micro-droplets moving in the pipeline can be monitored and detected more conveniently and more accurately.
In an embodiment of the present invention, the first pipe accommodates a droplet to be fused entering from the fourth pipe. The first power source is started, the first valve is opened, the fused liquid drops are pushed to stay under the first communication part under the driving of the first power source, the first power source is closed, the driving is stopped, the second power source is started to provide driving force for the pipeline through the third connecting port, and therefore the liquid of the third connecting port is pushed to enter the liquid drops quantitatively, and the liquid drop fusion is completed. During the liquid drop fusion process, the fusion electrode is connected with a power supply, and an electric field is generated to act on the first connection part to promote the liquid drop fusion.
Further, a plurality of droplets to be fused, for example, a droplet a1 to be fused, a2 to be fused, a3, … to be fused, an to be fused, may be present in the chip. The droplet fusion was performed sequentially by the above method.
The utility model discloses in, fourth pipeline one end includes that its upper reaches branch is 3 and above pipeline and the fifth connector that is parallel to each other. The upstream branch pipeline can realize automatic configuration of culture solution with different concentration and different factors.
In a specific embodiment, a microfluidic chip includes a substrate, and a first channel, a second channel, a third channel, and a fourth channel formed in the substrate. Wherein, first pipeline both ends include first connector and second connector respectively. One end of the second pipeline comprises a third connecting port, and the other end of the second pipeline is communicated with the first pipeline; one end of the third pipeline comprises a fourth connecting port, and the other end of the third pipeline is communicated with the first pipeline; the upstream of the fourth pipeline is branched into 3, which are respectively a fourth pipeline a, a fourth pipeline b and a fourth pipeline c, and one end of the fourth pipeline is respectively connected with fifth connecting ports 41, 41' and 41 ″.
Furthermore, the fifth connection ports 41, 41', 41 ″ are respectively connected with different power sources and valves for realizing the sample injection of the oil phase and the water phase. For example, the fifth connection port 41 is connected to the third power source and the fourth valve through a capillary tube, the fifth connection port 41' is connected to the fourth power source and the fifth valve through a capillary tube, and the fifth connection port 41 ″ is connected to the fifth power source and the sixth valve through a capillary tube. Before use, all the pipes are filled with oil phase, preferably mineral oil as filling medium, so that the pipes connected to the fifth connection ports 41, 41', 41 ″ are filled with mineral oil. In use, the fifth connection ports 41', 41 ″ are connected to a fresh basic culture solution, a basic culture solution containing a chemical factor, a substrate reaction solution, and the like, via the corresponding connection capillaries and the power source. For example, the fifth connection port 41' is connected to a fresh basic culture solution, and the fifth connection port 41 ″ is connected to a basic culture solution containing the culture factor x. And opening the first valve, and respectively starting the fourth power source and the fifth power source to fill the fourth pipeline b and the fourth pipeline c with corresponding culture solution. Starting a fourth power source, opening a first valve at the same time, closing the fourth power source when the basal culture solution in the fourth pipeline b quantitatively enters the main pipeline of the fourth pipeline through a fourth communication part, starting the third power source, and forming a liquid drop p of the water-in-oil basal culture solution at the fourth communication part; the liquid drop p further reaches the position right below the fifth communication part under the drive of the third power source, the third power source is closed, the fifth power source is started to inject the basic culture solution containing the culture factor x into the liquid drop p, and a culture medium liquid drop p' containing the culture factor x is formed. The culture factor x is used as a single factor or a single factor, the flow velocity and the flow of the injected liquid are controlled by a corresponding power source, namely a fifth power source, the entering amount of the culture factor is changed, so that the concentration of the culture factor x is controlled, different concentration gradients are formed, and the formation of the culture solution with the single factor and different concentrations is realized.
The utility model discloses in, accept the above-mentioned embodiment that forms culture medium liquid drop p 'of different concentration culture factors, culture medium liquid drop p' flows continuous crooked pipeline department, through the special shape and the structure of bend, effectively promotes different buffer and the mixture of culturing the factor.
In one embodiment, the upstream of the fourth pipeline is branched into 4, which are respectively a fourth pipeline a, a fourth pipeline b, a fourth pipeline c and a fourth pipeline d, and one end of each of the four pipelines is connected to the fifth connection ports 41, 41 ', 41 ″ and 41 ″', respectively. In addition to the structure in which the fourth channel is branched at 3 upstream sides, a fourth channel d and a fifth connection port 41 '' 'thereof are added, the fifth connection port 41' '' is connected to a sixth power source and a seventh valve through a capillary tube, and a culture medium containing a culture factor y is connected to the fifth connection port 41 '' 'through a capillary tube, and when a culture medium droplet p' containing the culture factor x reaches a sixth connection portion, the sixth power source connected to the fifth connection port 41 '' 'is driven to quantitatively control the culture medium containing the culture factor y to enter the culture medium droplet p' to form a culture medium droplet p '' containing the culture factor x and the culture factor y. In this embodiment, the control of the culture medium with different concentration by a single factor may be realized by individually controlling the amount of the culture factor x or the culture factor y, or the formation of the culture medium with different concentration by two factors may be realized by separately controlling the amount of the culture factor x and the culture factor y.
In one embodiment, the upstream of the fourth pipeline is branched into 5, which are respectively a fourth pipeline a, a fourth pipeline b, a fourth pipeline c, a fourth pipeline d and a fourth pipeline e, and one end of each of the four pipelines is connected with the fifth connection ports 41, 41 ', 41 "' and 41" ', respectively. Further, the first pipeline, the second pipeline, the third pipeline and the fourth pipeline are arranged on the substrate to form the microfluidic chip, as shown in fig. 2. Specifically, the fifth connection ports 41, 41 ', 41 "', and 41" 'are connected to different power sources and valves, respectively, for injecting the oil phase and the water phase, and the upstream of the fourth line is branched into 4 structures, and the fifth connection port 41 "' is connected to a seventh power source and an eighth valve, respectively, through a capillary tube. The using method is the same as the formation of the culture solution with single factor and different concentration and the formation of the culture solution with two factors and different concentration, just before the chip is used, an oil phase sample injection system is connected with the pipeline connecting ports, the pipeline is filled with oil phase medium, and the fifth connecting ports 41 ', 41 ' ' ' ' can be connected with fresh culture solution, chemical factors, substrate reaction solution and the like when in use. Further, for example, a fresh medium is connected to the fifth connection port 41 ', a water-in-oil culture medium droplet p is formed at the fourth communication site, and the fifth connection ports 41 ' ', 41 ' ' ', and 41 ' ' ' ' are connected to a culture factor x, a culture factor y, and a culture factor z, respectively, which are fused with the water-in-oil culture medium droplet p in the pipe at the fifth communication site, the sixth communication site, and the seventh communication site, respectively, to finally form a new culture medium droplet p ' ' ' or a droplet a containing a culture factor of a different culture factor and/or a culture factor of a different concentration. Different culture factors or culture factors with different concentrations are realized by controlling the entering amount of the corresponding power source. The culture medium drops p ' ' ' or drops a containing different culture factors and/or culture factors with different concentrations are arranged at the continuous bent pipeline, and effectively promote the buffering and mixing of the different culture factors. In the present embodiment, the automatic preparation of the micro-system of the three-factor multi-level microbial culture solution is realized by the power source connected to the fifth connection ports 41, 41 '' ''. Of course, those skilled in the art can realize the preparation of the multi-factor and multi-level microorganism culture solution according to more branches and connectors of the fourth pipeline, thereby realizing the culture of the multi-factor and multi-level microorganisms.
After the configuration of different horizontal culture mediums with different factors is completed, the formed liquid drop p ' ' ' or the liquid drop a directly enters the first pipeline through the driving of a third power source, or a plurality of parallel liquid drops enter the first pipeline through the liquid drop cutting at a third communication position. In the present invention, the formed liquid drop p' ″ or a liquid drop a can be one liquid drop or several liquid drops, such as liquid drop a1, liquid drop a2, …, liquid drop an, and the liquid drops and the number thereof are prepared according to the culture requirement. When all the liquid drops enter the first pipeline, the liquid drops are to-be-fused liquid drops, under the driving of the first power source, the liquid drops sequentially reach the position right below the first communication part, the first power source is closed, and the second power source connected with the second pipeline drives the bacteria liquid connected with the second power source to sequentially enter the liquid drops a1, the liquid drops a2, … and the liquid drops an respectively and complete the fusion of the liquid drops. In the process of liquid drop fusion, the fusion electrode is communicated with a power supply to generate an electric field effect to promote fusion. In the operation process, the first detection window 9 identifies the liquid drops, and fusion operation control is facilitated. The utility model discloses in, the liquid drop passes through drive power supply and moves continuous crooked pipeline department, through the special shape and the structure of bend, can effectively promote the buffering and the mixture of different cultivation factors.
In one embodiment, the parallel droplets a1, a2, … and an are droplets to be fused, and sequentially reach the position right below the first communication part under the driving of the first power source, the first power source is closed, and the second power source connected with the second pipeline drives the bacteria liquid connected with the second power source to sequentially enter the droplets a1, a2, … and an and complete the fusion of the droplets. In the process of liquid drop fusion, the fusion electrode is communicated with a power supply to generate an electric field effect to promote fusion. In the operation process, the first detection window 9 identifies the liquid drops, and fusion operation control is facilitated.
Examples
The first pipeline, the second pipeline, the third pipeline and the fourth pipeline are pipelines formed in a chip substrate, the size of the chip substrate is 3cm x 5cm x 4mm (length x width x thickness), the material used by the chip substrate is PMMA, the first pipeline, the second pipeline, the third pipeline and the fourth pipeline formed in the chip substrate are pipelines with square cross sections, the cross section area is 1mm2 (the side length is 1mm), the first pipeline is respectively communicated with the second pipeline, the third pipeline and the fourth pipeline, the first communication parts of the second pipeline and the first pipeline are positioned at the second communication parts of the third pipeline and the first pipeline, and the distance between the first communication parts of the fourth pipeline and the first pipeline is 1.5mm respectively from the second communication part to the third communication part. The pipeline is filled with oily medium. The upstream branches of the fourth pipelines are 5, namely a fourth pipeline a, a fourth pipeline b, a fourth pipeline c, a fourth pipeline d and a fourth pipeline e which are arranged in parallel, and the distance between the fourth pipeline a, the fourth pipeline b, the fourth pipeline c, the fourth pipeline d and the fourth pipeline e is 1.5 mm.
A first connecting port of the first pipeline is connected with an injection pump A, and a second connecting port is connected with a rotary first valve; the third connecting port of the second pipeline is connected with a syringe pump B and a rotary second valve, the fourth connecting port of the third pipeline is connected with a rotary third valve, and the fourth pipeline a, the fourth pipeline B, the fourth pipeline C, the fourth pipeline D and the fourth pipeline E are respectively connected with a syringe pump C, a rotary fourth valve, a syringe pump D, a rotary fifth valve, a syringe pump E, a rotary sixth valve, a syringe pump F, a rotary seventh valve, a syringe pump G and a rotary eighth valve. The pipeline is connected with the injection pump and the rotary valve through a capillary, the inner diameter of the capillary is 1.0 mm, and the material is polytetrafluoroethylene.
In this embodiment, the syringe pumps a, B, C, D, E, F and G used are all industrial syringe pumps, and the valve heads of the syringe pumps are three-way valves, and are commercially available from MSP1-C2 of north Heiben Baoding Lange constant flow pump, Inc.
The first valve, the second valve, the third valve, the fourth valve, the fifth valve, the sixth valve, the seventh valve and the eighth valve are all Mrv-01 high-pressure two-way valves purchased from Nanjing wetting fluid control devices, Inc.
The utility model discloses the chip is before using, the pipeline is the well oil phase that is full of earlier, then the oil phase has been connected respectively to the fifth connector, the culture medium, various culture factor solution, and be full of corresponding liquid in making fourth pipeline upper reaches lateral conduit, wherein, the medium oil phase is mineral oil, syringe pump C is used for driving the oil phase, syringe pump D is used for the drive to be full of 10G/L tryptone, the culture solution of 10G/L yeast extract (being called for short "initial culture solution"), syringe pump E, syringe pump F, syringe pump G are used for driving culture factor X respectively, culture factor Y, culture factor Z's initial culture solution, in this embodiment, culture factor X, culture factor Y, culture factor Z is 50G/L NaCl, 25 mol/L NaOH, 250 mg/L ampicillin respectively.
The method comprises the steps of starting an injection pump C, simultaneously opening a first valve, driving an oil phase in a fourth pipeline a to enter a pipeline, closing the injection pump C, simultaneously starting the injection pump D, starting the injection pump C, enabling 4 mu L 'initial culture solution' to form a droplet p of culture solution in oil under the fourth communication position when the 'initial culture solution' in the fourth pipeline b is injected below the fourth communication position, immediately closing the injection pump D, starting the injection pump C, enabling the 4 mu L 'initial culture solution' to form the droplet p of the culture solution in oil, enabling the droplet p to move under the fifth communication position under the driving of the injection pump C, simultaneously starting the injection pump C, driving 50G/L NaCl 'initial culture solution' 2 mu L 'in the fourth pipeline C to enter the droplet p in the pipeline, closing the injection pump E, starting the injection pump C, forming the droplet p' at the moment, driving the injection pump C to move under the sixth communication position to reach the 'initial culture solution' 2 mu L p ', closing the injection pump F, driving the injection pump F to reach the fourth pipeline b 5 mu p', enabling the injection pump C '2 mu p' to move, enabling the injection pump C '2 mu p' 2 p 'to form the droplet p', and enabling the ampicillin < 2 > 2 < p 'to move under the injection pump C', and starting the injection pump C < 2 > 2 < 2 > ampicillin < 2 > and the injection pump C < 2 > 2 < p > infusion pump < 2 > and starting the injection pump C < 2 > infusion pump C < 2 < p < 2 > infusion pump < 2 < p > infusion pump < 2 <.
And further pushing the liquid drop a to pass through a continuously bent pipeline by using a syringe pump C to reach a third communication part, and cutting the liquid drop, wherein 3 liquid drops a1, a2 and a3 which form middle parallel liquid drops and have the volume of 2 mu L are formed, the size of the liquid drops a3 which are pinched and removed is about 2 mu L, and the liquid drops are discharged through the third pipeline, in the process of forming the liquid drops a1, the liquid drops a2 and the liquid drops a3, a first detection window 9 is arranged on the first pipeline and is positioned between the third communication part and the first connection port, the first detection window 9 is 2mm away from the third communication part, the operation control is carried out, when the liquid drops a1 reach the first detection window 9, the cutting operation of the liquid drops a2 is carried out, and when the liquid drops a2 reach the first detection window 9, the cutting operation of the liquid drops a3 is carried out.
The first rotary valve is opened, the syringe pump A is driven continuously, and the liquid drops a1, a2 and a3 are pushed to move towards the second connecting port. When the liquid drop a1 reaches the position right below the first communication part, the injection pump A stops driving, the injection pump B is started to provide driving force, so that the bacterium liquid B connected with the second pipeline is pushed to enter the liquid drop a1, the fusion electrode is communicated with a 500V power supply while the injection pump B provides the driving force, and an electric field is generated to promote the fusion of the liquid drops. In the examples, the bacterial suspension b is Escherichia coli. The steps are circularly repeated, and the fusion operation of the liquid drop a2 and the liquid drop a3 is completed, so that the inoculation culture of 3 colibacillus with parallel concentration is realized. The first detection window 9 serves as a droplet identification point, identifying the position of the droplet; and the second detection window 10 is used for detecting the liquid drop detection point and the bacteria concentration OD.
Industrial applicability
The utility model discloses a micro-fluidic chip for microbial cultivation and detection can make and use in the micro-fluidic chip field. The utility model is suitable for a satisfy the microbial cultivation growth condition through multifactor multilevel, be suitable for adaptive evolution, resistant mechanism research and cultivate optimization and microbial detection.
Although the embodiments of the present invention have been described above with reference to the accompanying drawings, the present invention is not limited to the above-described embodiments and application fields, and the above-described embodiments are illustrative, instructive, and not restrictive. Those skilled in the art, having the benefit of this disclosure, may effect numerous modifications thereto without departing from the scope of the invention as defined by the appended claims.
The present application is susceptible to various modifications and alternative forms, specific embodiments have been shown by way of example in the drawings and have been described in detail herein. However, the application is not intended to be limited to the particular forms disclosed. On the contrary, the intention is to cover all modifications, equivalents, and alternatives falling within the scope of the application, which is defined by the appended claims and their legal equivalents.
The numerical ranges recited in the present invention all include the data of the two endpoints of the numerical range, and also include each specific numerical value in the numerical range, and the numerical values can be combined with the endpoints at will to form a new small range.
Claims (20)
1. An apparatus for droplet generation and fusion, comprising:
a first pipeline, two ends of which respectively comprise a first connecting port and a second connecting port,
a second pipeline, one end of which comprises a third connecting port and the other end of which is communicated with the first pipeline,
one end of the third pipeline comprises a fourth connecting port, the other end of the third pipeline is communicated with the first pipeline,
a fourth pipeline, one end of which comprises a pipeline with 3 or more branches at the upstream and a fifth connecting port, the other end of which is communicated with the first pipeline through a section of common pipeline, the section of common pipeline comprises a continuous bending pipeline,
wherein the first communicating parts of the second pipeline and the first pipeline are positioned between the second communicating parts of the third pipeline and the first pipeline and between the fourth pipeline and the third communicating parts of the first pipeline, the distance between the first communicating parts and the second communicating parts and the distance between the first communicating parts and the third communicating parts are 500-2000 mu m respectively,
the first pipeline is communicated with the first power source through the first connecting port and the capillary pipeline, the first pipeline is communicated with the first valve through the second connecting port and the capillary pipeline, the second pipeline is communicated with the second power source and the second valve through the third connecting port and the capillary pipeline respectively, the third pipeline is communicated with the third valve through the fourth connecting port and the capillary pipeline, and the fourth pipeline is communicated with the power source connected with the branch and the corresponding control valve through the fifth connecting port and the capillary pipeline respectively.
2. The device for droplet generation and fusion of claim 1, wherein the upstream branch of the fourth pipeline is 3, namely a fourth pipeline a, a fourth pipeline b and a fourth pipeline c.
3. The device for droplet generation and fusion of claim 1, wherein the number of the fourth pipeline upstream branches into 4, namely a fourth pipeline a, a fourth pipeline b, a fourth pipeline c and a fourth pipeline d.
4. The device for droplet generation and fusion of claim 1, wherein the number of the fourth pipeline upstream branches into 5, namely a fourth pipeline a, a fourth pipeline b, a fourth pipeline c, a fourth pipeline d and a fourth pipeline e.
5. The device for generating and fusing the liquid drops as claimed in any one of claims 1 to 4, wherein the continuous curved pipeline of which the upstream branch of the fourth pipeline is communicated with the first pipeline is composed of one or more structures selected from P type, S type, U type, broken line type and wave type.
6. The device for droplet generation and fusion of claim 1, wherein the continuous curved conduit contains a total volume of liquid of 4-100 μ L.
7. The device for droplet generation and fusion of claim 6, wherein the continuous curved conduit contains a total volume of liquid of 4-50 μ L.
8. The device for droplet generation and fusion of claim 7, wherein the continuous curved conduit contains a total volume of liquid of 4-20 μ L.
9. The device for droplet generation and fusion of claim 8, wherein the continuous curved conduit contains a total volume of liquid of 6-12 μ L.
10. The device of claim 9, wherein the continuous curved conduit contains a total volume of liquid of 8-10 μ L.
11. The apparatus for droplet generation and fusion of claim 1, wherein the distance between the first communicating portion and the second and third communicating portions is 750 μm to 1800 μm.
12. The device for droplet generation and fusion of claim 11, wherein the distance between the first communication site and the second and third communication sites is 1000 μm to 1500 μm.
13. The droplet generation and fusion device of claim 1, wherein the first communication site is the same distance from the second and third communication sites, respectively.
14. The device of claim 1, wherein the first, second, third and fourth conduits are disposed on a substrate, and the substrate and the first, second, third and fourth conduits are formed from any one of glass, Polymethylmethacrylate (PMMA), Polycarbonate (PC), Polystyrene (PS), Acrylonitrile Butadiene Styrene (ABS).
15. The device of claim 14, wherein the first, second, third, and fourth conduits are disposed on a substrate, and the substrate and the first, second, third, and fourth conduits are formed from Polymethylmethacrylate (PMMA).
16. The droplet generation and fusion apparatus of claim 1, wherein the first, second, third, and fourth conduits have cross-sectional areas in the range of 2.5 × 10-3mm2～4mm2。
17. The droplet generation and fusion apparatus of claim 16, wherein the first, second, third and fourth conduits have cross-sectional areas in the range of 0.01 to 3mm2。
18. The droplet generation and fusion apparatus of claim 17, wherein the first, second, third and fourth conduits have cross-sectional areas in the range of 0.1-2.5 mm2。
19. The device of claim 18, wherein the first droplet is generated and mergedThe cross-sectional areas of the pipeline, the second pipeline, the third pipeline and the fourth pipeline are in the range of 0.25-1 mm2。
20. The droplet generation and fusion apparatus of claim 19, wherein the first, second, third, and fourth conduits have the same cross-sectional area.
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| CN201921380574.0U CN211111987U (en) | 2019-08-23 | 2019-08-23 | Device for generating and fusing liquid drops |
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| CN201921380574.0U CN211111987U (en) | 2019-08-23 | 2019-08-23 | Device for generating and fusing liquid drops |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112403538A (en) * | 2019-08-23 | 2021-02-26 | 无锡源清天木生物科技有限公司 | Device and method for generating and fusing liquid drops |
| CN114054105A (en) * | 2020-08-07 | 2022-02-18 | 无锡源清天木生物科技有限公司 | Micro-droplet quick sorting system |
-
2019
- 2019-08-23 CN CN201921380574.0U patent/CN211111987U/en active Active
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112403538A (en) * | 2019-08-23 | 2021-02-26 | 无锡源清天木生物科技有限公司 | Device and method for generating and fusing liquid drops |
| CN114054105A (en) * | 2020-08-07 | 2022-02-18 | 无锡源清天木生物科技有限公司 | Micro-droplet quick sorting system |
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