CN110819507B - Micro-droplet preparation chip for intestinal microorganism detection - Google Patents
Micro-droplet preparation chip for intestinal microorganism detection Download PDFInfo
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- CN110819507B CN110819507B CN201911118190.6A CN201911118190A CN110819507B CN 110819507 B CN110819507 B CN 110819507B CN 201911118190 A CN201911118190 A CN 201911118190A CN 110819507 B CN110819507 B CN 110819507B
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- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
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- 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
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Abstract
The invention relates to a liquid drop preparation chip for intestinal microorganism detection, which can accurately control and regulate the size of multiple packaged liquid drops by arranging a regulating channel; according to the change of the preparation requirements of the liquid drops, multiple wrapped liquid drops with different sizes can be obtained by adjusting the change of the dynamic parameters of the channels; meanwhile, the adjustment of the invention is not limited to the overall size of the multi-wrapped liquid drops, and the inner core size, the thickness of the wrapping layer and the like of the multi-wrapped liquid drops can be independently adjusted; the liquid drop preparation chip provided by the invention allows the preparation of the uniform double emulsion liquid drops with different specifications on the same chip, has universality within a certain liquid drop size range, and can be used as an independent liquid drop preparation unit to be combined with liquid drop using units with various specifications, so that the detection cost is greatly reduced.
Description
Technical Field
The invention relates to the technical field of microfluidics, in particular to a micro-droplet preparation chip for intestinal microorganism detection.
Background
Intestinal microorganisms refer to the general term of microbial communities residing in the human intestinal tract, including bacteria, archaebacteria, unicellular eukaryotes, and the like, and together with the intestinal environment constitute a huge and complex ecological system. The human intestinal tract is a nutrient-rich microenvironment, the number of carried bacteria is up to 100 trillion, the number of genes of human intestinal microorganisms is up to 500 ten thousand, and the number of genes is 150 times of the basic factors of human. The intestinal microorganisms have certain relativity with metabolic diseases, cardiovascular diseases, digestive system diseases, cancers, immune system diseases and central nervous system diseases, and the research on the structure, the function and the pathogenic mechanism of the intestinal microorganisms is beneficial to playing a role in the treatment of diseases and developing new treatment modes.
With the development of microfluidic technology, the importance of the technology in the field of microorganism/nucleic acid detection is gradually highlighted. The digital PCR method is a mainstream method for detecting microorganisms/nucleic acids based on a microfluidic means at present, has extremely high sensitivity and accuracy, and is suitable for accurately and quantitatively analyzing specific nucleic acids in a trace sample. The digital PCR technology is a detection means based on a statistical principle, and the main means is that a sample solution or a diluent thereof is distributed into a large number of micro-droplets, the droplets are used as a micro-reactor for amplification reaction, fluorescence detection is carried out on the reaction result, and then the initial copy number of target DNA in a sample is obtained according to a Poisson distribution principle.
At present, micro-droplets used for digital PCR detection are generally water-in-oil type single-package droplet systems. Chinese patent (CN 109746061 a) describes a controlled preparation method of such droplets in its disclosure, which describes that for three common droplet generation modes, i.e. co-axial flow, cross flow and flow focusing modes, the size of the generated droplets can be controlled by adjusting the flow rate of the continuous phase fluid, wherein in a compression mode with a lower flow rate the droplets have a larger size, and in a jet mode with a higher flow rate the droplet size is relatively minimal.
Chinese patent (CN 107429426 a) proposes in its disclosure that since the emulsion carrier phase of the single-coated droplet system used for digital PCR detection is an inert oil phase, this prevents the droplet reactor from being detected, quantified and sorted using available methods such as fluorescence activated cell sorting that is incompatible with the oil carrier phase. The patent also proposes to overcome the above problems by performing digital PCR detection using double emulsions.
However, the preparation of multiple emulsions, particularly for the purpose of digital PCR detection, presents challenges in terms of uniformity of droplet size, droplet weight, number of inner core droplets, etc. dimensions.
For single-coated droplets, the droplet size can be controlled by adjusting the continuous phase flow rate; however, in the preparation of multiple emulsions, where there are multiple continuous phases and where the injection rate of the subsequent continuous phase is generally not allowed to vary at will, for example, in the preparation of double emulsions where the first continuous phase (oil) shears the aqueous core fluid to produce water-in-oil droplets and moves forward along the microchannel and the second continuous phase (aqueous phase) shears the first continuous phase containing the water-in-oil droplets to produce water-in-oil droplets, the injection rate of the second continuous phase is substantially controlled so that the first continuous phase breaks down just in the middle of the two water-in-oil droplets under the shear of the second continuous phase due to the spacing of the water-in-oil droplets in the first continuous phase; otherwise, as the process continues, the fracture position of the first continuous phase will be changed and uncontrollable under the shearing of the second continuous phase, and as a result, double droplets generated are uncontrollable, for example, water-in-oil-in-water double droplets may be generated under the shearing of the second continuous phase, oil-in-water single package droplets may be generated, and water-in-oil droplets generated during the shearing of the first continuous phase are included; thereby causing confusion in the droplet system.
Therefore, the current multiple emulsion preparation chips are usually dedicated chips. The multiple emulsion preparation chip often requires different hydrophilic and hydrophobic modification on different parts of the micro-channel, so that the manufacturing cost and difficulty of the multiple emulsion preparation chip are high, and if the multiple emulsion preparation chip has the general property of preparing liquid drops with different sizes, the cost of preparing the liquid drops is reduced to a great extent.
In other words, the resulting water-in-oil-in-water droplets are essentially fragmented
It is difficult to produce droplets having uniform dimensions. As mentioned above, in the preparation of droplets of different sizes, there is a difference in the flow rate of the continuous phase fluid, which results in the distance between two adjacent droplets within the continuous phase possibly differing due to their size variations (e.g. in the preparation of droplets of different batches); this may result in both water-in-oil-in-water double droplets and oil-in-water single-wrapped droplets being obtained when the droplets are secondarily wrapped, the reason for the single-wrapped droplets occurring during the secondary wrapping being that the distance between two adjacent droplets generated in the previous stage is too large, resulting in that the partially wrapped oil continuous phase does not contain aqueous droplets when the secondary wrapping is performed. While such problems can be avoided by adjusting the flow rate of the continuous phase at the time of secondary encapsulation to match the rate of droplet generation to exactly the droplet pitch of the preceding stage to avoid single encapsulation droplets at the time of secondary encapsulation, such adjustment is clearly difficult; more importantly, as previously mentioned, a change in continuous phase flow rate will result in a change in the size of the droplets produced, e.g., a decrease in continuous phase flow rate will result in an increase in droplet size, which is detrimental to digital PCR detection.
Disclosure of Invention
In order to solve the problems in the prior art, the invention provides a micro-droplet preparation chip for intestinal microorganism detection. The micro-droplet preparation chip provided by the invention allows the precise control and adjustment of the size of double-emulsion droplets, can prepare uniform double-emulsion droplets with different specifications on the basis of the same chip, has universality within a certain droplet size range, and can be used as an independent droplet preparation unit to be combined with droplet use units (such as digital PCR chips) with various specifications, so that the detection cost is greatly reduced.
Although the size of the singly-packed droplets can be flexibly controlled by adjusting the flow rate of the continuous phase; the final size of the multipack droplets is limited by a number of factors and adjustment is not generally achieved. For example, in the preparation of a double emulsion, referring to fig. 1-3, single encapsulated droplets generated after a core fluid (typically an aqueous phase) is sheared by a first continuous phase (typically an oil phase) are arranged in microchannels at a substantially constant distance from each other, the first continuous phase entraining the single encapsulated droplets is then sheared as a new core fluid by a second continuous phase (typically an aqueous phase), thereby forming water-in-oil-in-water double emulsion droplets.
Theoretically, the sum of the volumes of each single-wrapped droplet in the microchannel and the first continuous phase dispensed by the single-wrapped droplet in the microchannel (such as V1 shown in fig. 1-3 and 5-7 and V1' in fig. 5-7, where V1' is the adjusted V1, the continuous phase between two adjacent micro-droplets is equally divided, the sum of the volumes of the continuous phases dispensed by two sides of the micro-droplet and the volume of the micro-droplet itself are V1 or V1', hereinafter referred to as the dispensing volume) should be equal to the volume of the finally prepared double-wrapped droplet (V2 in fig. 1-3), so as to ensure continuous and stable formation of the homogenized double-wrapped droplet.
Fig. 1 shows the double emulsion preparation process in v1=v2, extrusion mode. In the extrusion mode, the first continuous phase shears the core fluid at a lower flow rate to form single-package droplets with larger particle sizes, and the distribution volume V1 is small due to the lower flow rate of the first continuous phase and the small distance between two adjacent droplets. Since v2=v1, the shear force required to be provided by the second continuous phase is predetermined, corresponding to its predetermined flow rate.
Fig. 2 shows the double emulsion preparation process in v1=v2, jet mode. In the jet mode, the first continuous phase shears the core fluid at a higher flow rate to form a single-coated emulsion with smaller particle size, but the distribution volume V1 is large due to the higher flow rate of the first continuous phase and the large spacing between two adjacent droplets. Since v2=v1, the shear force required to be provided by the second continuous phase is also predetermined, corresponding to its predetermined flow rate.
Both cases shown in fig. 1 and 2 show that while a smaller size single-encapsulated emulsion can be prepared by increasing the flow rate of the first continuous phase, an increase in flow rate introduces more of the first continuous phase fluid, resulting in an increase in the spacing of the single-encapsulated droplets such that the volume of the resulting double-encapsulated droplets in the first continuous phase jet-stream mode (fig. 2) is instead greater than the volume of the resulting double-encapsulated droplets in the first continuous phase-stream mode (fig. 1). This makes it very difficult to prepare double-wrapped droplets of smaller size.
Fig. 3 shows the double emulsion preparation process in v1+.v2, jet mode. V1+.v2 means that the flow rate of the second continuous phase is adjusted at will, irrespective of other factors. In this case, although it is possible to reduce the size of the droplets sheared by the second continuous phase, this shearing mode does not guarantee that the first continuous phase comprising the single-coated droplets breaks down at exactly the intermediate position of the two single-coated droplets under the shearing of the second continuous phase, and therefore the droplets obtained are those comprising: a chaotic system of water-in-oil-in-water double-coated droplets, oil-in-water single-coated droplets and water-in-oil single-coated droplets. Such drop systems cannot meet downstream application requirements and should therefore be avoided.
Based on such recognition and analysis, the present invention provides a micro-droplet preparation chip for intestinal microorganism detection, the droplet preparation chip is used for preparing at least two-layer wrapped multiple emulsion droplets with controllable particle size, and comprises a core fluid channel, a first continuous phase channel, a single wrapped droplet channel, a second continuous phase channel, a double wrapped droplet channel …, and so on, and can have an n+1th continuous phase channel and an N-th wrapped droplet channel according to the wrapping weight of the prepared droplets; the inner core fluid channel is intersected with the first continuous phase channel, and downstream of the intersection is in fluid communication with the single-package liquid drop channel; similarly, the N heavily wrapped drop channel intersects the n+1 th continuous communication channel, downstream of which junction is in fluid communication with the n+1 heavily wrapped drop channel; the microdroplet preparation chip also includes a conditioning channel that is in fluid communication with the N-fold wrapped channel, thus having a progression, such as an nth-order conditioning channel, that allows for the size of the dispensing volume V1 of the N-fold wrapped drop within the corresponding drop channel to be adjusted.
Preferably, the conditioning channels are present only between two adjacent continuous phase channels.
Preferably, the conditioning channel is used to withdraw the continuous phase fluid from the droplet channel, thereby reducing the dispense volume V1; the regulating channels are connected to separate power units.
Preferably, the micro-droplet preparation chip comprises a cover plate layer and a micro-channel layer, wherein the regulating channel, the core fluid channel, the first continuous phase channel, the single-package droplet channel, the second continuous phase channel and the double-package droplet channel are all positioned on the same side of the micro-channel layer. All of the micro-channels on the micro-channel layer preferably have the same depth to allow the micro-channel layer to be processed by a single photolithographic process.
Preferably, the adjusting channels are symmetrically arranged at two sides of the liquid drop channel, and the sizes (including width, height and diagonal size) of the adjusting channels are smaller than the liquid drop diameter in the corresponding liquid drop channel; more preferably smaller than the radius of the droplet.
Preferably, the adjustment channel on each side is made up of a number of sub-channels arranged side by side, which sub-channels distribute the connection openings of the adjustment channel and the droplet channel over a larger length, allowing each sub-channel to be connected with the droplet channel using smaller connection openings and achieving equal or even larger adjustment capabilities, the smaller connection opening dimensions allowing smaller turbulence to the droplets in the droplet channel during adjustment, facilitating the small-sized droplets to maintain their position and shape in the droplet channel.
Preferably, the regulating channel comprises a collecting channel gradually expanding along the liquid flow direction and a drainage grating arranged at the joint of the collecting channel and the liquid drop channel, the drainage grating consists of a plurality of microcolumns which are arranged at intervals, and a gap between two adjacent microcolumns forms a flow path of a continuous phase between the liquid drop channel and the collecting channel; the collecting channel is connected with a liquid collecting channel at the maximum flaring along the liquid flow direction; the microcolumn may be a body having any cross-sectional shape, preferably having a square or circular cross-section.
The drainage grating has the same function as that of the plurality of parallel sub-channels, so that the size of a single opening connected with the liquid drop channel can be reduced, and the interference on liquid drops is reduced; the gaps of the drainage grating are finally collected in the liquid collecting channel through the collecting channel, so that the liquid collecting channel can be allowed to have a size exceeding that of liquid drops, the distribution volume V1 can be adjusted at a higher speed, and meanwhile, the position and the form of the liquid drops in the channel are not obviously disturbed; and, because the collecting channel has a diverging structure along the direction of the liquid flow, the continuous phase entraining the liquid droplets can enter the collecting channel through the gap at an increased speed along with the increase of the flaring of the collecting channel in the process of flowing through the drainage grating; thereby further improving the adjusting speed of the dispensing volume V1, being beneficial to the high speed of the whole process of preparing the liquid drops, and being capable of better meeting the requirements of digital PCR detection and the like for rapidly preparing a large number of uniform liquid drops in a short time.
In the foregoing embodiment, all the micro-channels preferably have the same depth, in other words, the micro-channels in the foregoing embodiment are all planarized channel structures; for example, a non-penetrating microchannel may be formed by partial lithography or machining on a single side of the microchannel layer, or a microchannel structure may be formed by penetrating the microchannel layer and then closing the microchannel layer by a cover sheet and a base sheet.
When the microdroplet preparation chip is used to prepare a double-encapsulated droplet, it may also have a three-dimensional microchannel structure. The three-dimensional micro-channel structure has the following characteristics: the upstream side of the core fluid channel, the first continuous phase channel and the single-package liquid drop channel is a non-through equal-depth micro-channel, and the non-through equal-depth micro-channel is arranged on the upper side of the micro-channel layer; the downstream side of the single-wrapped droplet channel, the second continuous phase channel, and the double-wrapped droplet channel are also non-through equal-depth microchannels, but are disposed on the underside of the microchannel layer; the middle part of the single-package liquid drop channel penetrates through the micro-channel layer and is respectively in fluid communication with the upstream side and the downstream side of the single-package liquid drop channel; the head end and the tail end of the middle part of the single-package liquid drop channel are respectively provided with a non-through adjusting channel, wherein the adjusting channel at the head end is arranged at the lower side of the micro-channel layer, and the adjusting channel at the tail end is arranged at the upper side of the micro-channel layer; the depth of the regulating channel is not greater than the radius of the single wrapped droplet.
The single-coated droplet flows in such a microchannel structure, and when moving from the upstream side of the single-coated droplet channel located at the upper side to the middle of the through single-coated droplet channel, the droplet will sink in a curved path in the through channel due to the higher specific gravity of the aqueous droplet than the oily continuous phase, and then enter the downstream side of the single-coated droplet channel; the regulating channels at the head and tail ends can respectively suck the continuous phase at the bottom of the channel near the head end where the liquid drop has not yet completely sunk and suck the continuous phase at the top of the channel near the tail end where the liquid drop has basically sunk to the bottom, so as to regulate the distribution volume V1.
Preferably, the conditioning channel is a flat channel having a depth of no more than 1/4 of the diameter of the individual wrapped drop.
Preferably, the width of the adjusting channel is half of the length of the middle part of the single-package liquid drop channel, so that when the single-package liquid drop moves in the middle part of the penetrating single-package liquid drop channel, the vertical acting force on the single-package liquid drop generated by the adjusting channel due to the continuous phase suction is weak, and the adjusting channel is basically stable on the whole path, so that the interference on the moving process and the form of the single-package liquid drop is weakened.
Preferably, the two adjusting channels located at the upper and lower layers of the microchannel layer may be communicated through one through hole, thereby allowing the same power to be supplied to the adjusting channels at different positions through the same power unit.
Preferably, the junction of the core fluid channel and the first continuous phase channel is in fluid communication with the single wrap droplet channel through a reduced diameter section.
In the foregoing scheme, the preparation process of double-wrapping liquid drops is mainly exemplified, but the device of the invention can be equally applicable to the controllable preparation of liquid drops with more wrapping weights. When more preparation of the packed heavy liquid drops is needed, a regulating channel can be arranged on one side or two sides of the liquid drop channel at the downstream side of each stage of continuous phase channel except the downstream of the last stage of continuous phase channel, so that the distribution volume in the corresponding liquid drop channel is regulated, and further, the controllable preparation of the multiple packed liquid drops is realized.
The chip of the invention can adopt the existing method to carry out hydrophilic-hydrophobic modification on each part of the micro-channel, wherein the surface property of the regulating channel is the same as the surface property of the liquid drop channel communicated with the regulating channel.
The invention also provides a liquid drop preparation method, which is carried out based on the micro liquid drop preparation chip provided by the invention, and the distribution volume of liquid drops in the corresponding liquid drop channels is regulated through the regulating channels, so that the volume of liquid drops obtained by the next stage of encapsulation is controllable.
Compared with the prior art, the invention has the following beneficial effects: the invention discovers key factors influencing the size of the multi-packed liquid drops from the formation mechanism analysis of the multi-packed liquid drops, and designs a liquid drop preparation chip and a method capable of adjusting the distribution volume of the liquid drops in a liquid drop channel so as to enable the final volume of the multi-packed liquid drops to be controllable; the droplet preparation chip can allow the size of the multiple packaged droplets to be accurately controlled and adjusted; according to the change of the preparation requirements of the liquid drops, multiple wrapped liquid drops with different sizes can be obtained by adjusting the change of the dynamic parameters of the channels; meanwhile, the adjustment of the invention is not limited to the overall size of the multi-wrapped liquid drops, and the inner core size, the thickness of the wrapping layer and the like of the multi-wrapped liquid drops can be independently adjusted; the liquid drop preparation chip provided by the invention allows the preparation of the uniform double emulsion liquid drops with different specifications on the same chip, has universality within a certain liquid drop size range, and can be used as an independent liquid drop preparation unit to be combined with liquid drop using units with various specifications, so that the detection cost is greatly reduced.
Drawings
Fig. 1 is a schematic illustration of a double-wrapped droplet preparation process with v1=v2 in an extrusion mode;
fig. 2 is a schematic illustration of a double-wrapped droplet preparation process in jet mode, v1=v2;
FIG. 3 is a schematic illustration of a reported droplet preparation process for V1+.V2 in a jet mode;
FIG. 4 is a top view of the chip structure of the present invention;
FIG. 5 is a schematic representation of a modulation channel modulating a dispense volume V1 to V1';
FIG. 6 is a schematic representation of a modulation channel having sub-channels;
FIG. 7 is a schematic representation of a conditioning channel with drainage grille;
FIG. 8 is an enlarged partial illustration of a conditioning channel with a drainage grille;
FIG. 9 is a schematic cross-section of a chip with stereoscopic microchannels;
FIG. 10 is a schematic three-dimensional structure of a chip with stereoscopic microchannels;
FIG. 11 is a top view illustration of the chip of FIG. 10;
FIG. 12 is another schematic three-dimensional structure of the chip of FIG. 10;
fig. 13 is another top view illustration of a chip with stereoscopic microchannels.
In the figure: 1 is a core fluid channel, 2 is a first continuous phase channel, 3 is a single-package liquid drop channel, 4 is a reduced diameter section, 5 is a second continuous phase channel, 6 is a double-package liquid drop channel, 7 is an adjusting channel, 71 is a sub-channel, 72 is a collecting channel, 73 is a drainage grating, 731 is a micro-column, 732 is a micro-column gap, 74 is a liquid collecting channel, and 75 is a through hole.
Detailed Description
To further clarify the idea of the invention, the invention provides the following specific embodiments.
Example 1
As shown in fig. 4-5, a micro-droplet preparation chip for intestinal microorganism detection is provided, wherein the micro-droplet preparation chip is used for preparing double-wrapping droplets and comprises a cover plate layer and a micro-channel layer; the upper surface of the microchannel layer is provided with a non-penetrating microchannel structure, the cover plate layer covers the upper surface of the microchannel layer, and one side of the cover plate layer is provided with four through holes which are in fluid communication with the microchannel layer; the micro-channel structure comprises an inner core fluid channel 1, a first continuous phase channel 2, a single-package liquid drop channel 3, a second continuous phase channel 5 and a double-package liquid drop channel 6; the inner core fluid channel 1 is intersected with the first continuous phase channel 2 in a cross way, the downstream of the intersection is in fluid communication with the single-package liquid drop channel 3 through a reducing section 4, the single-package liquid drop channel 3 is intersected with the second continuous phase channel 5 in a cross way, and the downstream of the intersection is in fluid communication with the double-package liquid drop channel 6; two regulating channels 7 are symmetrically arranged on two sides of the downstream section of the single-package liquid drop channel 3, and the two symmetrically arranged regulating channels 7 are in fluid communication with the single-package liquid drop channel 3 and connected with the same injection pump, so that the first continuous phase is sucked from the single-package liquid drop channel 3 or the first continuous phase is supplemented into the single-package liquid drop channel 3 through the regulating channels 7, and the distribution volume of the single-package liquid drop is adjusted from V1 to V1'. The width of the conditioning channel 7 is smaller than the diameter of the individual droplets and it has the same surface properties as the individual droplet channels 3.
Example 2
As shown in fig. 4 and 7, a micro-droplet preparation chip for intestinal microorganism detection is provided, which has a main body structure similar to that of the micro-droplet preparation chip described in embodiment 1, except that the adjustment channels 7 located on each side of the single-package droplet channel 3 are each composed of four sub-channels 71 arranged side by side, and the interval between any two adjacent sub-channels 71 is 0.5 to 3 times the width of the sub-channel 71 itself.
Example 3
As shown in fig. 4 and 7-8, a micro-droplet preparation chip for intestinal microorganism detection is provided, unlike embodiments 1 and 2, the adjustment channels 7 located at each side of the single-coated droplet channel 3 each include a collection channel 72 gradually expanding along the flow direction and a drainage grating 73 disposed at the connection between the collection channel and the droplet channel, and the connection lines formed by the inner edges of the drainage gratings 73 are coincident with or located outside the extension lines of the side walls of the single-coated droplet channel 3, so that the single-coated droplet is not blocked during the flowing process in the single-coated droplet channel 3; the drainage grating 73 is composed of a plurality of micro-columns 731 which are arranged at intervals, and a gap 732 between every two adjacent micro-columns 731 forms a flow path of the first continuous phase between the single-package liquid drop channel 3 and the collecting channel 72; the collecting channel 72 is connected with a liquid collecting channel 74 at the maximum flaring along the liquid flow direction; the microcolumn 731 has a square cross section.
Example 4
The scheme of this example is not shown in the drawings, but differs from examples 1-3 in that the microdroplet preparation chip is used to prepare triple encapsulated droplets. The microchannel layer also comprises a third continuous phase channel which is crossed with the double-wrapping liquid channel 6 and a triple-wrapping liquid drop channel which is in fluid connection with the intersection; second adjusting channels are symmetrically arranged on two downstream sides of the double-wrapping liquid drop channel 6, and the second adjusting channels on two sides are commonly connected to another injection pump for adjusting the distribution volume of double-wrapping liquid drops in a second continuous phase channel, and the second adjusting channels have the same surface properties as the double-wrapping liquid drop channel 6.
Example 5
As shown in fig. 4 and 9-11, a micro-droplet preparation chip for intestinal microorganism detection is provided, which is different from embodiments 1-3 in that the micro-droplet preparation chip further includes a substrate layer, and the micro-channel layer has a three-dimensional micro-channel structure, specifically, the three-dimensional micro-channel structure has the following characteristics: the upstream sides of the core fluid channel 1, the first continuous phase channel 2 and the single-package liquid drop channel 3 are non-through equal-depth micro channels, and the micro channels are arranged on the upper side of the micro channel layer; the downstream side of the single-wrapped droplet channel 3, the second continuous phase channel 5, and the double-wrapped droplet channel 6 are also non-through equal-depth micro-channels, which are arranged on the lower side of the micro-channel layer; the middle part of the single-package liquid drop channel 3 penetrates through the micro-channel layer and is respectively in fluid communication with the upstream side and the downstream side of the single-package liquid drop channel 3; the head end and the tail end of the middle part of the single-package liquid drop channel 3 are respectively provided with a non-through adjusting channel 7, wherein the adjusting channel 7 at the head end is arranged at the lower side of the micro-channel layer, and the adjusting channel 7 at the tail end is arranged at the upper side of the micro-channel layer; the depth of the regulating channel 7 is not greater than the radius of the individual wrapped drops. The regulating channel 7 is a flat channel, and the width of the regulating channel is half of the length of the through middle part of the single-package liquid drop channel 3. The regulating channels 7 on the upper and lower sides of the microchannel layer are connected to different syringe pumps.
Example 6
As shown in fig. 4, 9, 12-13, a micro-droplet preparation chip for intestinal microorganism detection is provided. Different from the embodiment 5, two adjusting channels 7 respectively located on the upper and lower surfaces of the micro-channel layer on the same side of the single-package droplet channel 3 are respectively communicated through a through hole 75; the two through holes 75 are commonly connected to the same syringe pump.
Example 7
There is provided a multi-pack droplet preparation method based on the micro droplet chip of any one of the foregoing embodiments, wherein the size of the N-th pack droplet is controlled by adjusting the flow rate of the N-th continuous phase, and then the distribution volume of the N-th pack droplet in the corresponding droplet channel is adjusted by adjusting the channel 7 connected to the N-th pack droplet channel, so that the flow rate of the n+1-th continuous phase can be changed along with the change of the distribution volume, thereby adjusting the thickness of the pack layer of the next stage. And N is a positive integer.
The above examples are merely illustrative of preferred embodiments of the present invention and should not be construed as limiting all possible embodiments of the present invention, which may be obtained by conventional substitution or modification of the present invention by those skilled in the art without inventive effort on the basis of the technical idea of the present invention. The actual scope of the invention is defined in the following claims.
Claims (7)
1. The utility model provides a little liquid droplet preparation chip for intestinal microorganism detects, its is used for preparing the controllable little liquid droplet of at least two-layer parcel of particle diameter, including cover plate layer and microchannel layer, its characterized in that: the micro-channel layer comprises a core fluid channel, a first continuous phase channel, an N-fold wrapped droplet channel, an (n+1) -th continuous phase channel and an N+1-fold wrapped droplet channel; the inner core fluid channel is intersected with the first continuous phase channel, and downstream of the intersection is in fluid communication with the single-package liquid drop channel; the N heavily wrapped liquid drop channel is intersected with the N+1 continuous communication channel, and the downstream of the intersection is in fluid communication with the N+1 heavily wrapped liquid drop channel; the micro-droplet preparation chip further comprises an N-th stage adjusting channel, wherein the N-th stage adjusting channel is in fluid communication with the N-heavy-package droplet channel, and each stage of adjusting channel is connected with an independent power unit, wherein N is a positive integer; the N-th-stage regulating channels are symmetrically arranged on two sides of the N-fold wrapped liquid drop channel, and are used for pumping continuous phase fluid from the liquid drop channel so as to reduce the distribution volume V1; the N-th-stage adjusting channel consists of a plurality of sub-channels which are arranged in parallel, and the distance between the sub-channels is 0.5-3 times of the width of the sub-channels; the N-th-stage adjusting channel comprises a collecting channel gradually expanding along the liquid flow direction and a drainage grating arranged at the joint of the collecting channel and the N-heavy-package liquid drop channel, wherein the drainage grating consists of a plurality of microcolumns which are arranged at intervals, and a gap between two adjacent microcolumns forms a flow path of an N-th continuous phase between the N-heavy-package liquid drop channel and the collecting channel; the collecting channel is connected with a liquid collecting channel at the maximum flaring along the liquid flow direction; the micro-droplet preparation chip is used for preparing double-coated droplets and further comprises a substrate layer; the upstream side of the core fluid channel, the first continuous phase channel and the single-package liquid drop channel is a non-through equal-depth micro-channel, and the non-through equal-depth micro-channel is arranged on the upper side of the micro-channel layer; the downstream side of the single-wrapped droplet channel, the second continuous phase channel, and the double-wrapped droplet channel are also non-through equal-depth microchannels, which are arranged on the underside of the microchannel layer; the middle part of the single-package liquid drop channel penetrates through the micro-channel layer and is respectively in fluid communication with the upstream side and the downstream side of the single-package liquid drop channel; the head end and the tail end of the middle part of the single-package liquid drop channel are respectively provided with a non-through adjusting channel.
2. The micro-droplet preparation chip for intestinal microorganism detection of claim 1, wherein: the nth stage modulation channel has the same surface properties as the N heavily wrapped drop channel.
3. The micro-droplet preparation chip for intestinal microorganism detection of claim 1, wherein: the microcolumns have square or circular cross sections.
4. The micro-droplet preparation chip for intestinal microorganism detection of claim 1, wherein: the regulating channel at the head end of the middle part of the single-package liquid drop channel is arranged at the lower side of the micro-channel layer, and the regulating channel at the tail end is arranged at the upper side of the micro-channel layer; the depth of the regulating channel is not greater than the radius of the single wrapped droplet.
5. The micro-droplet preparation chip for intestinal microorganism detection according to claim 4, wherein: the regulating channel is a flat channel, and the depth of the regulating channel is not more than 1/4 of the diameter of the single-package liquid drop; and has a width that is half the length of the middle of the single wrap drop channel.
6. The micro-droplet preparation chip for intestinal microbiota detection according to any one of claims 1-5, wherein: two adjusting channels on the same side of the single-package liquid drop channel are respectively communicated through a through hole; the two through holes are commonly connected to the same syringe pump.
7. A method for preparing multiple coated droplets, characterized in that the method is performed based on the micro droplet preparation chip according to any one of the preceding claims 1 to 6, wherein the size of the N-th coated droplet is controlled by adjusting the flow rate of the N-th continuous phase, and then the distribution volume of the N-th coated droplet in the corresponding droplet channel is adjusted by adjusting the channel connected to the N-th coated droplet channel, so that the flow rate of the n+1-th continuous phase can be changed along with the change of the distribution volume, thereby adjusting the thickness of the coating layer of the next stage.
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| CN114247485B (en) * | 2020-09-25 | 2022-12-30 | 中国科学院青岛生物能源与过程研究所 | Micro-fluidic chip for particle screening and separation |
| CN114210378B (en) * | 2021-11-22 | 2023-04-28 | 广东省科学院健康医学研究所 | Liquid drop generating device based on microporous capillary and preparation method thereof |
| CN114260035B (en) * | 2021-12-23 | 2023-04-07 | 广东工业大学 | Multilayer wrapped micro-fluidic chip and cell particle generator |
| CN117007800B (en) * | 2023-08-09 | 2024-05-10 | 祥符实验室 | Dual-droplet microfluidic chip for escherichia coli detection and application thereof |
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