Cell manipulation device and method based on quartz lined tube array

Abstract

The present invention provides a cell manipulation device and method based on a quartz lined tube array. The present invention includes an array well plate placement slot, a quartz sleeve, a cell wall breaking device and a cell liquid transfer mechanism. The array well plate placement slot is a cell culture plate, which is provided with uniform openings. The quartz sleeve can be sleeved on the openings. The cell wall breaking device is used to break down the cells with high voltage electricity after approaching the cells in the solution, so that the cells can break the wall. The cell liquid transfer mechanism is used to extract the cell solution after the wall is broken and transfer it. The present invention can realize multi-mode operation of cells under less solution, from the pre-treatment of cells to ensure the rupture of cell walls, the automated sampling process of cells, and the detection process of cells, to realize multi-process operation.

Description

Technical Field

[0001] The present invention relates to the technical field of cell sampling, and in particular to a cell operation device and method based on a quartz lined tube array. Background Art

[0002] In recent years, with the gradual deepening of cell physiological research, single-cell analysis has become an important means to understand and decipher many key life processes, and single-cell separation technology is the basis for single-cell level research. How to obtain a sufficient number of single cells is the key to research. The "Single-cell separation microfluidic chip" with application number 201610454360 uses a microfluidic chip to arrange single cells and then realize the separation process of single cells. Similar methods are disclosed in the "Microfluidic cell culture chip with uniform cell distribution and cell sampling method" with application number 201710667551. The operation of cells in the prior art mainly includes physical processing processes such as using microfluidic cells and capillaries with a large aperture to operate cells. It also includes biological operation processes such as culturing and proliferating cells in an environment such as a culture solution. The current existing patent technology mainly emphasizes the operation of a certain process of cells. The existing technology cannot realize the multi-process operation flow of previous processing, sampling, and detection of a small number of cells. Summary of the invention

[0003] In response to the above technical problems, a cell manipulation device and method based on a quartz lined tube array is provided. The technical means adopted by the present invention are as follows:

[0004] A cell manipulation device based on a quartz lined tube array comprises an array well plate placement slot, a quartz sleeve, a cell wall breaking device and a cell fluid transfer mechanism. The array well plate placement slot is a cell culture plate with uniform openings, the quartz sleeve can be sleeved on the openings, the cell wall breaking device is used to break down the cells with high voltage electricity after approaching the cells in the solution so that the cells can be broken, and the cell fluid transfer mechanism is used to extract the cell solution after the wall is broken and transfer it.

[0005] Furthermore, the openings on the cell culture plate are arrayed openings, and positioning holes for positioning the cell fluid transfer mechanism are arranged around each opening.

[0006] Furthermore, the cell wall disruption device includes positive and negative probes, a power source connected to each probe, and a support assembly for placing a needle tip.

[0007] The present invention also discloses a specific operation method of the cell manipulation device, including continuously or independently completing one of the following steps:

[0008] Cell pretreatment steps: pre-inject the cell solution into the quartz sleeve, and place the quartz sleeve on the cell culture plate; place the cell wall breaking device in the cell solution to be broken; turn on the power switch of the cell wall breaking device for a preset time;

[0009] Cell sampling steps: the robot arm is positioned with the positioning hole of the array well plate placement slot, and moves in the XOY plane to the top of the quartz sleeve; then it descends to a preset height to perform cell sampling;

[0010] Cell injection and detection steps: The robotic arm rises to a preset height, translates in the XOY plane, and moves to the injection port of the detector.

[0011] The present invention has the following advantages:

[0012] 1. The present invention provides a method for operating the entire cell process, which realizes multi-mode operation of cells without solution, from cell pretreatment to ensure the rupture of cell walls, cell automated sampling process, and cell detection process, to realize multi-process operation.

[0013] 2. The present invention utilizes a double tungsten needle pulse high voltage electric breakdown method to achieve cell wall rupture. The cell pretreatment method is used to effectively extract intracellular compounds, which is more conducive to detecting the types of intracellular compounds. At the same time, only two electrode wires are close to or in slight contact with the cell solution, which minimizes the contact process with the cell solution and reduces the probability of cell contamination. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative labor.

[0015] Figure 1 Schematic diagram of the overall structure of the cell pretreatment step of the present invention.

[0016] Figure 2 for Figure 1 Front view of .

[0017] Figure 3 It is a quartz lined tube and a cell wall breaking monomer structure.

[0018] Figure 4 Isometric view of cell wall disruption in a quartz-lined tube.

[0019] Figure 5 Schematic diagram of a quartz lined tube.

[0020] Figure 6 Isometric view of the array plate fixture.

[0021] Figure 7 for Figure 6 main view.

[0022] Figure 8 Schematic diagram of each operation method.

[0023] In the figure: 1. Array well plate placement slot; 2. Quartz liner tube and cell wall breaking mechanism; 3. Discharge needle tip assembly; 4. Quartz sleeve; 5. Positive electrode discharge tungsten needle; 6. Negative electrode discharge tungsten needle; 7. Robotic arm; 8. Sampling needle; 9. Detector. DETAILED DESCRIPTION

[0024] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0025] like Figures 1 to 7 As shown, an embodiment of the present invention discloses a cell manipulation device based on a quartz lined tube array, including an array well plate placement slot 1, a quartz sleeve, a cell wall breaking device 2 and a cell fluid transfer mechanism, wherein the array well plate placement slot is a cell culture plate having uniform openings thereon, the quartz sleeve 4 can be sleeved on the openings, the cell wall breaking device is used to utilize high voltage electricity to break down the cells after approaching the cells in the solution so that the cells can be broken, and the cell fluid transfer mechanism is used to extract the cell solution after the wall is broken and transfer it.

[0026] The openings on the cell culture plate are array-type openings, and each opening is surrounded by positioning holes for positioning the cell solution transfer mechanism. Specifically, the cell culture plate can use a 96-well plate. For cell pretreatment and cell sampling, the cell solution in the process is all in a disposable quartz sleeve. Each quartz tube is used alone and does not interfere with each other. The 96-well plate can place the quartz sleeve well and ensure that the quartz sleeve does not slide down and fall.

[0027] The cell wall breaking device includes a discharge needle tip assembly 3, specifically including a positive electrode discharge tungsten needle 5, a negative electrode discharge tungsten needle 6, a power supply connected to each probe, and a support assembly for placing the needle tip, two tungsten needle metal electrodes, placed in parallel and kept at a certain distance. One of the discharged metal tungsten needles is connected to the positive pole of the high voltage electricity, and one metal tungsten needle is used to connect the negative pole of the high voltage electricity. The positive pole voltage range of the high voltage electricity is in the range of +3KV-5KV. For negative high voltage, the voltage range is 0kV to -5KV. For negative high voltage, the value of positive high voltage depends on the degree of cell wall breaking desired. In order to prevent the two tungsten needles from contacting each other and short-circuiting, an insulating member is placed between the tungsten needles for support. During sampling and pre-treatment, the cells are placed in a disposable quartz lined tube for operation. For biological cells with cell walls, after high voltage electricity wall breaking, intracellular compounds will overflow, so it is necessary to sample the cell mixed solution after wall breaking. In this embodiment, the sampling process uses a three-dimensional robotic arm 7 with precise positioning. First, the robot arm can move to the designated position according to position recognition. This position can be a hole position that has been pre-treated with high-voltage electric wall breaking or a hole position that has not been pre-treated with high-voltage electric wall breaking. Finally, the robot arm moves downward along the Z axis, locates the cell page, and quantitatively extracts the cell solution according to the set value.

[0028] As a preferred embodiment, the robotic arm can realize horizontal movement of X and Y, and can be judged according to whether a quartz sleeve is placed in each hole groove. If a quartz sleeve is not placed, the system will automatically judge it as a non-sampling area. According to human needs, the sampling arm will move above the quartz sleeve where a certain solution is placed. According to the amount of solution placed on the page, the robotic arm will insert the sampling needle 8 below the page according to judgment. According to the volume to be sampled, the sampling wall is controlled to enter the extraction mode. At this time, the robotic arm enters the downward Z-axis movement. The cell solution is sampled by extracting negative pressure, wherein the sampling volume is 5-50uL and the sampling accuracy is ±2uL.

[0029] like Figure 8 As shown, the present invention also discloses an operation method of the above device, specifically, comprising continuously or independently completing one of the following steps:

[0030] Cell pretreatment steps: pre-inject the cell solution into the quartz sleeve, and place the quartz sleeve on the cell culture plate; place the cell wall breaking device in the cell solution to be broken; turn on the power switch of the cell wall breaking device for a preset time. In this embodiment, the start time is less than 1 minute. The cell structure is changed through a physical process, so that the cell wall of the cell with a cell wall is broken, and then the compound inside is released, eliminating the restraint of the cell wall, so that the compound inside the cell can be detected;

[0031] Cell sampling steps: Regardless of whether the cells have been pre-treated or not, the cells are mixed in the bottom layer of the quartz sleeve, in which the cells and the cell culture solution are uniform. At this time, the mixed solution of cells and cells is sampled through the sampling device in an automated manner. Specifically, the robotic arm is positioned with the positioning hole of the array well plate placement slot, and translates in the XOY plane to the top of the quartz sleeve; then it descends to a preset height to sample cells; specifically, the robotic arm can move to a designated position at will according to position recognition. This position can be a hole position that has been pre-treated with high-voltage electric wall breaking, or a hole position that has not been pre-treated with high-voltage electric wall breaking. Finally, the robotic arm moves downward along the Z axis, locates the cell page, and quantitatively extracts the cell solution according to the set value.

[0032] Cell injection and detection steps: The robotic arm rises to a preset height, translates in the XOY plane, and moves to the injection port of the detector 9. In this embodiment, a mass spectrometer is selected as the detector, and the difference in relative molecular mass of the compounds is used to detect different types of cells.

[0033] The above three processes are continuous and independent of each other. Any two of the above two processes can be performed without affecting other operations. 3. The use of pulsed high voltage to rupture the cell wall of the cell sample in the mixed solution can be used alone as an operation method. It can be achieved in less than 1 minute to rupture cells containing cell walls, providing a method for the subsequent detection of more compounds. By combining the processes of cell pretreatment, cell sampling, and cell detection in an orderly manner, and using a disposable quartz sleeve to hold the cell sample, there is no cross-interference between each quartz tube. For the cell pretreatment process, high voltage electricity is used to rupture the cell wall of cells containing cell walls, so that in the subsequent cell detection, the detection of compound types is limited due to the wrapping of the cell wall. Such a wall-breaking operation can provide conditions for the detection of more compounds. Unlike ultrasound, grinding and other methods that require a large amount of solution samples, it has a longer sample processing time. The main advantage is that it can process a large number of cells containing cell walls in a liquid with a volume of 5-50uL.

[0034] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A cell manipulation device based on a quartz lined tube array, characterized in that: It comprises an array well plate placement slot, a quartz sleeve, a cell wall breaking device and a cell fluid transfer mechanism. The array well plate placement slot is a cell culture plate, which is provided with uniform openings. The openings on the cell culture plate are array openings. Each opening is surrounded by positioning holes for positioning the cell fluid transfer mechanism. The quartz sleeve can be sleeved on the openings. The cell wall breaking device is used to break down the cells by high voltage electricity after approaching the cells in the solution, so that the cells can be broken, including a positive electrode discharge tungsten needle, a negative electrode discharge tungsten needle, a power supply connected to each probe and a support component for placing the needle tip, two tungsten needle metal electrodes are placed in parallel, and an insulating member is placed between the tungsten needles for support, and the voltage is set according to different types of cell walls or the degree of wall breaking; The cell solution transfer mechanism is used to extract the cell solution after cell wall breaking and transfer it. Specifically, the sampling process uses a three-dimensional mechanical arm to identify the hole position that has been treated with high-voltage electric wall breaking according to the position, and realizes the horizontal movement of X and Y. Finally, the mechanical arm moves downward in the Z axis and locates to the cell solution surface. According to the set value, the cell solution is quantitatively extracted, wherein the sampling volume is 5-50 μL, and the sampling accuracy is ±2 μL; The above-mentioned high voltage electricity has a positive voltage range of +3 KV to 5 KV, and a negative voltage range of 0 kV to -5 KV.

2. A method for operating the cell manipulation device based on the quartz lined tube array according to claim 1, characterized in that: Includes one of the following steps completed consecutively or independently: Cell pretreatment steps: pre-inject the cell solution into the quartz sleeve, and place the quartz sleeve on the cell culture plate; place the cell wall breaking device in the cell solution to be broken; turn on the power switch of the cell wall breaking device for a preset time; Cell sampling steps: the robot arm is positioned with the positioning hole of the array well plate placement slot, and moves in the XOY plane to the top of the quartz sleeve; then it descends to a preset height to perform cell sampling; Cell injection and detection steps: The robotic arm rises to a preset height, translates in the XOY plane, and moves to the injection port of the detector.

Images