Application device for sorting microorganisms on micro-fluidic chip

Abstract

The utility model relates to the technical field of microorganism sorting, in particular to an application device for sorting microorganisms on a micro-fluidic chip. Re-suspending the cell or microorganism sample by using the photocuring hydrogel solution; injecting the sample into a micro-fluidic sorting chip; after standing, the sample falls into a large number of micro-pits which are uniformly distributed on the bottom surface of the chamber of the microfluidic sorting chip and are equivalent to a single sample in size; the signal detection module is used for carrying out fluorescence or Raman spectrum detection on a sample in the micro-pit array; the curing light source irradiates the position of the target sample; according to the method and the structure, the gel lysis solution is injected into a microfluidic sorting chip, a liquid-phase target sample is obtained, the detection sensitivity of microorganisms of about 1 micron is remarkably improved, meanwhile, potential damage in the process is prevented, the sorted microorganisms can be continuously cultured and amplified to be used for bacterial strain screening of synthetic biology, operation is convenient, and the method and the structure are easy to implement. And the separation efficiency is higher.

Description

Technical Field

[0001] The utility model relates to the technical field of microorganism sorting, in particular to an application device for microorganism sorting on a microfluidic chip. Background Art

[0002] Existing technologies primarily include fluorescence flow cytometry sorting and optical tweezers extraction. Fluorescence flow cytometry is primarily targeted at samples such as cells larger than 7 microns, but its sensitivity for detecting microorganisms around 1 micron often falls short of practical requirements. Furthermore, antibody dyes and excessive fluid pressure can potentially damage sample activity. Even with sorting, further culture and amplification are impossible, making it unsuitable for strain screening in synthetic biology. Optical tweezers also introduce significant damage during sample separation, complicate equipment operation, and result in low separation efficiency, significantly limiting their application in synthetic biology. Utility Model Content

[0003] The purpose of this utility model is to provide an application device for microbial sorting on a microfluidic chip to address the defects in the existing technology, thereby significantly improving the detection sensitivity of microorganisms of about 1 micron, while preventing potential damage during the process. The sorted microorganisms can be further cultured and amplified for strain screening in synthetic biology, and the operation is convenient and the separation efficiency is higher.

[0004] In order to achieve the above-mentioned object, the technical solution adopted by the present invention is: an application device for microorganism sorting on a microfluidic chip, comprising a sorting control module, a fluid driving module, a signal detection module, a solidified light source and a microfluidic sorting chip;

[0005] A large number of micro-pits with the same size as a single sample are evenly distributed on the bottom surface of the microfluidic sorting chip chamber;

[0006] Resuspend the cell or microbial sample in the light-curable hydrogel solution;

[0007] Injecting the sample into the microfluidic sorting chip, maintaining a certain flow rate so that the sample evenly fills the chamber of the microfluidic sorting chip;

[0008] After the sample is left to stand, it falls into a large number of micro-pits evenly distributed on the bottom surface of the chamber of the microfluidic sorting chip, which are of the same size as a single sample;

[0009] The signal detection module performs fluorescence or Raman spectroscopy detection on the sample in the micro-pit array, and screens and records the location of the target sample according to different specific detection methods;

[0010] The microfluidic sorting chip is moved to the sorting control module, and the curing light source is irradiated on the position of the target sample, so that the hydrogel around the target sample is cured and wraps the target sample;

[0011] Injecting a cleaning solution into the microfluidic sorting chip for cleaning to remove any uncured sample residue;

[0012] The gel lysis solution is injected into the microfluidic sorting chip to lyse the solidified hydrogel to obtain the target sample in liquid phase.

[0013] Furthermore, a liquid storage chamber and a control liquid circuit are provided in the fluid driving module, and the control liquid circuit includes a vacuum pump, a clamping valve, and a connecting hose; the chip liquid inlet of the microfluidic sorting chip is connected to the output pipeline of the fluid driving module and the liquid storage chamber, and the chip liquid outlet of the microfluidic sorting chip is connected to the waste liquid pipeline of the fluid driving module.

[0014] Furthermore, the sample suspension is added to the liquid storage chamber, and the output pipeline is closed by the pinch valve;

[0015] The vacuum pump is connected to the waste liquid pipeline, and the vacuum pump operates to form a certain vacuum degree in the sorting chip;

[0016] Turn off the vacuum pump, open the pinch valve, and use negative pressure to draw the sample into the liquid inlet of the microfluidic sorting chip;

[0017] The sample liquid spreads out after entering the microfluidic sorting chip;

[0018] After standing for a certain period of time, the sample is randomly distributed into a plurality of the micro-pits.

[0019] Furthermore, the sorting control module includes a motion control mechanism, which moves the microfluidic sorting chip, aligns the signal detection module and the microfluidic sorting chip to perform signal detection and find the micro-pit position where the desired target sample is located.

[0020] Furthermore, the sorting control module records the positions of the micro-pits;

[0021] The motion control mechanism moves the microfluidic sorting chip so that the corresponding micro-pit position is aligned with the curing light source;

[0022] The curing light source cures the target sample at the corresponding micro-pit position;

[0023] The remaining unsolidified liquid is sucked away through the liquid outlet of the chip via the waste liquid pipeline by the fluid driving module;

[0024] The remaining target sample is reserved for subsequent use.

[0025] Furthermore, the curing light source is blue or purple light, and the curing light source uses a laser or a monochromatic LED light source in combination with the microholes to achieve a light spot equivalent to the micropit.

[0026] Furthermore, the signal detection module selects a fluorescence microscopy imaging module or a Raman spectroscopy detection module to meet different requirements for screening target samples.

[0027] Furthermore, the photocurable hydrogel solution uses a GelMA solution to resuspend the cell or microbial sample.

[0028] The system comprises a sorting control module, a fluid driving module, a signal detection module, a curing light source and a microfluidic sorting chip; a large number of micropits of the same size as a single sample are evenly distributed on the bottom of the chamber of the microfluidic sorting chip; the cell or microbial sample is resuspended with a light-curing hydrogel solution; the sample is injected into the microfluidic sorting chip, and a certain flow rate is maintained so that the sample is evenly spread over the chamber of the microfluidic sorting chip; after the sample is allowed to stand, it falls into a large number of micropits of the same size as a single sample that are evenly distributed on the bottom of the chamber of the microfluidic sorting chip; the signal detection module performs fluorescence or Raman spectroscopy detection on the sample in the micropit array, and screens and records the target sample according to different specific detection methods. at the position; moving the microfluidic sorting chip to the sorting control module, the solidified light source irradiating the position of the target sample, and the hydrogel around the target sample solidifying and wrapping the target sample; injecting a cleaning solution into the microfluidic sorting chip for cleaning, and cleaning the unsolidified sample residue; injecting a gel lysis solution into the microfluidic sorting chip, lysing the solidified hydrogel, and obtaining a liquid phase of the target sample. The method and structure achieve significantly improved detection sensitivity of microorganisms of about 1 micron, while preventing potential damage in the process. The sorted microorganisms can be continued to be cultured and amplified for strain screening in synthetic biology, and the operation is convenient and the separation efficiency is higher. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are only some embodiments recorded in the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0030] Figure 1 This is a schematic diagram of the overall structure of the microbial sorting application device on a microfluidic chip;

[0031] Figure 2 This is a top view of the overall structure of the microbial sorting application device on a microfluidic chip;

[0032] Figure 3 This is a front view of the overall structure of the microfluidic chip microorganism sorting application device without the pipeline;

[0033] Figure 4 This is a side view of the overall structure of the microfluidic chip microorganism sorting application device without pipelines;

[0034] Figure 5 A perspective view of a microfluidic sorting chip for microorganism sorting application device on a microfluidic chip;

[0035] Figure 6 This is an exploded view of the overall structure of the microfluidic chip microorganism sorting application device;

[0036] Reference numerals:

[0037] Sorting control module 1, fluid driving module 2, output pipeline 21, waste liquid pipeline 22, signal detection module 3, curing light source 4, microfluidic sorting chip 5, micro pit 51, chip liquid inlet 52, chip liquid outlet 53. DETAILED DESCRIPTION

[0038] The technical solutions in the embodiments of the present invention will be described clearly and completely below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, rather than all the embodiments.

[0039] In the description of the present invention, it should be noted that the orientations or positional relationships indicated by “center”, “up”, “down”, “left”, “right”, “vertical”, “horizontal”, “inside” and “outside” are based on the orientations or positional relationships shown in the accompanying drawings, and are only for the convenience of describing the present invention and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, be constructed and operated in a specific orientation, and therefore cannot be understood as a limitation on the present invention.

[0040] An application device for microbial sorting on a microfluidic chip, such as Figure 1 、 56, comprising a sorting control module 1, a fluid driving module 2, a signal detection module 3, a curing light source 4 and a microfluidic sorting chip 5; a large number of micro-pits equivalent to the size of a single sample are evenly distributed on the bottom surface of the chamber of the microfluidic sorting chip 5; the cell or microbial sample is resuspended with a light-curing hydrogel solution; the sample is injected into the microfluidic sorting chip 5, and a certain flow rate is maintained so that the sample is evenly spread across the chamber of the microfluidic sorting chip 5; after the sample is allowed to stand, it falls into a large number of micro-pits 51 equivalent to the size of a single sample evenly distributed on the bottom surface of the chamber of the microfluidic sorting chip 5; the signal detection module 3. Perform fluorescence or Raman spectroscopy detection on the samples in the micropit array, and screen and record the location of the target sample according to different specific detection methods; move the microfluidic sorting chip 5 to the sorting control module 1, and irradiate the location of the target sample with the solidified light source 4. The hydrogel around the target sample solidifies and wraps the target sample; inject cleaning fluid into the microfluidic sorting chip 5 for cleaning to clean the unsolidified sample residue; inject gel lysis fluid into the microfluidic sorting chip 5 to lyse the solidified hydrogel to obtain the liquid phase of the target sample.

[0041] Specifically, a large number of micro-pits 51 of the same size as a single microbial sample are evenly distributed on the bottom surface of the microfluidic sorting chip, which greatly improves the detection and capture capabilities of tiny microorganisms, such as bacteria of about 1 micron, and achieves high-sensitivity sorting. The signal detection module 3 performs fluorescence or Raman spectroscopy detection on the sample in the micro-pit 51, which can accurately identify and record the position of the target sample, providing precise guidance for subsequent operations. The entire sorting process avoids sample damage factors that may be caused by traditional methods, such as the use of antibody dyes and excessive fluid pressure, thereby effectively protecting the biological activity of the sample. The target sample is wrapped with light-cured hydrogel, which not only achieves sample fixation, but also reduces the potential damage to the sample by physical and chemical treatments, which is beneficial to subsequent cultivation. and amplification experiments, the selection control module 1, the fluid drive module 2, the signal detection module 3, the solidification light source 4 and the microfluidic sorting chip 5 are highly integrated, making the entire sorting process more compact, efficient, and easy to automate. The target sample is quickly fixed and separated in the micropit 51 through irradiation by the solidification light source 4, and then the target sample is quickly cleaned and recovered using the cleaning liquid and gel lysis liquid, which improves the sorting efficiency and recovery rate. It is further suitable for strain screening in synthetic biology, and can efficiently and accurately separate microorganisms with specific functions, providing strong support for synthetic biology research. In the biomedical field, the device also has broad application potential, such as disease diagnosis, drug screening, etc., providing efficient and reliable microbial sorting solutions for related research.

[0042] As a preferred embodiment of the above, Figure 5As shown, a liquid storage chamber and a control liquid circuit are provided in the fluid driving module 2, and the control liquid circuit includes a vacuum pump, a clamping valve, and a connecting hose; the chip liquid inlet 52 of the microfluidic sorting chip 5 is connected to the output pipeline 21 of the fluid driving module 2 and the liquid storage chamber, and the chip liquid outlet 53 of the microfluidic sorting chip 5 is connected to the waste liquid pipeline 22 of the fluid driving module 2.

[0043] Specifically, a liquid storage chamber is provided inside the fluid driving module 2 for storing solutions to be injected into the microfluidic sorting chip 5, such as light-cured hydrogel solution, cleaning solution and gel lysis solution, etc. The liquid storage chamber ensures a stable supply of liquids required during the sorting process, reducing operational delays or failures caused by insufficient or interrupted liquids. The control liquid path is that the fluid driving module 2 includes a vacuum pump, a pinch valve and a connecting hose. These components work together to achieve precise control of liquid flow, wherein the vacuum pump is used to generate negative pressure to drive the liquid from the liquid storage chamber through the connecting hose into the microfluidic sorting chip 5. By adjusting the power or working time of the vacuum pump, the flow rate and flow of the liquid can be precisely controlled to ensure uniform distribution and effective sorting of the sample within the chip. The pinch valve is located on the control liquid path and is used to control the on-off of the liquid. By opening or closing the pinch valve, switching and sequential injection of different types of liquids can be achieved, such as first injecting a light-cured hydrogel solution to resuspend the sample, then injecting a cleaning solution to clean non-target samples, and finally injecting a gel lysis solution to crack the solidified hydrogel to release the target sample. The quick response and precise control of the pinch valve improve the automation level and operational efficiency of the sorting process. The connecting hose is used to connect the liquid storage chamber, the control liquid circuit and the microfluidic sorting chip 5 to ensure the smooth flow of liquid in the system. The hose material must have good corrosion resistance and high pressure resistance to meet the transportation needs of different types of liquids. The chip liquid inlet 52 of the microfluidic sorting chip 5 is connected to the output pipeline 21 of the fluid drive module 2 through the connecting hose to receive the liquid from the liquid storage chamber; the chip liquid outlet 53 is connected to the waste liquid pipeline 22 of the fluid drive module 2 to discharge unused liquid or waste liquid out of the system. This design realizes the circulation of liquid in the sorting chip and the timely discharge of waste liquid, ensuring the continuity and efficiency of the sorting process.

[0044] As a preferred embodiment of the above, Figure 1 As shown, the sample suspension is added to the liquid storage chamber, and the output pipeline 21 is closed by the pinch valve; the vacuum pump is connected to the waste liquid pipeline 22, and the vacuum pump works to form a certain vacuum degree in the sorting chip; the vacuum pump is turned off, and the pinch valve is opened, and the negative pressure causes the sample to be sucked into the chip liquid inlet 52 of the microfluidic sorting chip 5; the sample liquid spreads out after entering the microfluidic sorting chip 5; after standing for a certain period of time, the sample is randomly distributed to the plurality of micro-pits 51.

[0045] Specifically, first, the microbial sample suspension to be sorted, including the target microorganisms and possible other non-target microorganisms, is added to the liquid storage chamber of the fluid driving module 2. This step ensures that the sample suspension is in a ready state and can be injected into the microfluidic sorting chip 5 at any time. After adding the sample suspension, the output pipeline 21 of the fluid driving module 2 is closed by controlling the clamping valve. This step is to prevent the sample suspension from being accidentally sucked into the output pipeline when the vacuum pump is working, but to ensure that the sample suspension can enter the microfluidic sorting chip 5 according to the predetermined path. Next, the vacuum pump is connected to the waste liquid pipeline 22 and the vacuum pump is started. The operation of the vacuum pump will form a certain vacuum degree inside the microfluidic sorting chip 5. This negative pressure environment is the key factor in driving the sample suspension into the chip. After the vacuum pump has been operating for a period of time, the vacuum pump is turned off and the pinch valve is quickly opened to reconnect the output line 21 to the liquid storage chamber. At this time, due to the negative pressure already formed in the microfluidic sorting chip 5, the sample suspension is attracted by the negative pressure and is sucked into the microfluidic sorting chip 5 through the chip liquid inlet 52. After the sample suspension enters the microfluidic sorting chip 5, it will spread out under the guidance of the internal structure of the chip. As the liquid flows and spreads, the microorganisms in the sample will be randomly distributed into the multiple micropits 51 evenly distributed on the bottom surface of the chip. This process utilizes the microchannel and micropit structure of the microfluidic chip to achieve uniform dispersion and random distribution of the sample. To ensure that the sample can be stably distributed in each micropit 51, it needs to be left to stand for a certain period of time. During this period, the microorganisms in the sample will gradually settle to the bottom of the micropit due to factors such as gravity and surface tension, and interact with the micropit surface.

[0046] As a preferred embodiment of the above, Figure 1 As shown, the sorting control module 1 includes a motion control mechanism, which moves the microfluidic sorting chip 5, and aligns the signal detection module 3 and the microfluidic sorting chip 5 to perform signal detection and find the position of the micro-pit 51 where the desired target sample is located.

[0047] Specifically, by integrating a precise motion control mechanism in the sorting control module 1, the microfluidic sorting chip 5 can be accurately moved so that it can be precisely aligned with the signal detection module 3, ensuring the precise movement and positioning of the microfluidic chip in two-dimensional or three-dimensional space. After the sample suspension is loaded into the microfluidic sorting chip 5 and left to stand for a period of time, the signal detection module 3 starts to perform signal detection on the sample in the chip. This step usually involves fluorescence spectroscopy, Raman spectroscopy or other types of signal detection technology, which is used to identify and distinguish target samples from non-target samples. Through the detection of the signal detection module 3, the position of the micropit 51 where the target sample is located can be identified. At this time, the motion control mechanism in the sorting control module 1 will accurately move the microfluidic sorting chip 5 according to the position information provided by the signal detection module 3, so that the micropit 51 where the target sample is located is precisely aligned with the detection window of the signal detection module 3. In order to achieve high-precision alignment and detection, the motion control mechanism needs to have a high degree of accuracy and stability.

[0048] As a preferred embodiment of the above, Figure 5 As shown, the sorting control module 1 records the position of the micro-pit 51; the motion control mechanism moves the microfluidic sorting chip 5 so that the corresponding position of the micro-pit 51 is aligned with the curing light source 4; the curing light source 4 solidifies the target sample at the corresponding position of the micro-pit 51; the remaining unsolidified liquid is sucked away through the chip outlet 53 via the waste liquid pipeline 22 by the fluid driving module 2; the remaining target sample is retained for subsequent use.

[0049] Specifically, the sorting control module 1 first records the position of the micropit 51 where the target sample is located, ensuring that the curing light source 4 can accurately illuminate the micropit where the target sample is located. Then, the motion control mechanism accurately moves the microfluidic sorting chip 5 based on the micropit position information provided by the sorting control module 1. This step ensures that the micropit 51 where the target sample is located can be accurately aligned with the irradiation area of the curing light source 4. When the position of the micropit 51 is aligned with the curing light source 4, the curing light source 4 is activated and emits light of a specific wavelength. This light irradiates the photocurable hydrogel in the micropit 51, triggering its polymerization reaction, thereby firmly fixing the target sample in the micropit. This step achieves rapid and efficient curing of the target sample. At the same time as the target sample is curing, the fluid drive module 2 begins to work. It uses negative or positive pressure to aspirate the remaining uncured liquid in the microfluidic sorting chip 5, including non-target samples and photocurable hydrogel that has not participated in curing, through the chip outlet 53, and discharges it out of the system through the waste liquid pipeline 22. This step effectively removes non-target samples and waste liquid, facilitating subsequent sample recovery and processing. After the above steps, only the target samples immobilized by the photocurable hydrogel remain in the microfluidic sorting chip 5. These samples can be retained for subsequent use, such as further culture, analysis, or amplification experiments.

[0050] As a preferred embodiment of the above, Figure 1 As shown, the curing light source 4 is blue or purple light. The curing light source 4 uses a laser or a monochromatic LED light source in combination with microholes to achieve a light spot equivalent to the micropit 51 .

[0051] Specifically, the curing light source 4 is designed to be blue or purple light. These two colors of light are widely used in the field of photocuring chemistry because they can effectively excite the photoinitiator in the photocurable material, thereby initiating a polymerization reaction. Blue and purple light have higher energy and can penetrate a certain thickness of photocurable hydrogel to ensure that the target sample is fully and evenly cured. The curing light source 4 can be a laser or a monochromatic LED light source. The laser light source has a high degree of directionality and focus, and can form a very small light spot, which is suitable for precise curing of small areas. The monochromatic LED light source has lower cost and is simpler to operate.

[0052] As a preferred embodiment of the above, Figure 1 As shown, the signal detection module 3 selects a fluorescence microscopy imaging module or a Raman spectrum detection module to meet different requirements for screening target samples.

[0053] Specifically, in a microbial sorting system, depending on the specific needs of the target sample being screened, either a fluorescence microscopy module or a Raman spectroscopy module can be selected as the signal detection module. Fluorescence microscopy is suitable for scenarios requiring high sensitivity and specificity, such as cell biology and molecular biology research; while Raman spectroscopy is suitable for scenarios requiring non-destructive testing and a wide detection range, such as substance identification and structural analysis. By properly selecting the signal detection module, precise identification and screening of different target samples can be achieved.

[0054] As a preferred embodiment of the above, Figure 1 As shown, the photocurable hydrogel solution uses GelMA solution to resuspend cells or microbial samples.

[0055] Specifically, by choosing GelMA solution as the photocurable hydrogel solution to resuspend cell or microbial samples, it is based on its excellent biocompatibility, photocrosslinking properties, degradability and porous structure, which helps to improve the accuracy and reliability of the microfluidic chip sorting system and provide strong support for subsequent experimental research.

[0056] The above shows and describes the basic principles, main features, and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The above embodiments and descriptions are merely illustrative of the principles of the present invention. Various changes and improvements may be made to the present invention without departing from the spirit and scope of the present invention. Such changes and improvements are intended to fall within the scope of the present invention. The scope of protection claimed in this invention is defined by the appended claims and their equivalents.

Claims

1. An application device for microorganism sorting on a microfluidic chip, characterized by: It comprises a sorting control module (1), a fluid driving module (2), a signal detection module (3), a curing light source (4) and a microfluidic sorting chip (5); A large number of micro-pits with a size comparable to that of a single sample are evenly distributed on the bottom surface of the chamber of the microfluidic sorting chip (5).

2. The device for microorganism sorting on a microfluidic chip according to claim 1, characterized in that: A liquid storage chamber and a control liquid circuit are provided in the fluid driving module (2), and the control liquid circuit comprises a vacuum pump, a pinch valve, and a connecting hose; the chip liquid inlet (52) of the microfluidic sorting chip (5) is connected to the output pipeline (21) of the fluid driving module (2) and the liquid storage chamber, and the chip liquid outlet (53) of the microfluidic sorting chip (5) is connected to the waste liquid pipeline (22) of the fluid driving module (2).

3. The device for microorganism sorting on a microfluidic chip according to claim 2, characterized in that: The sorting control module (1) includes a motion control mechanism, which moves the microfluidic sorting chip (5), aligns the signal detection module (3) and the microfluidic sorting chip (5), performs signal detection, and finds the position of the micropit (51) where the desired target sample is located.

4. The device for microorganism sorting on a microfluidic chip according to claim 3, characterized in that: The curing light source (4) is blue or purple light, and the curing light source (4) uses a laser or a monochromatic LED light source in conjunction with microholes to achieve a light spot equivalent to the micropit (51).

5. The device for microorganism sorting on a microfluidic chip according to claim 1, characterized in that: The signal detection module (3) selects a fluorescence microscopy imaging module or a Raman spectrum detection module to meet different requirements for screening target samples.

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