Automated sample preparation apparatus, method and biochemical analysis system
Patent Information
- Application Number
- CN202510304333.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-03-13
- Publication Date
- 2026-09-15
Smart Images
Figure CN122750451A_ABST
Abstract
Description
Technical Field
[0001] This application relates to the field of biochemical technology, and in particular to an automated sample preparation device, method and biochemical analysis system. Background Technology
[0002] In gene sequencing, sequencing samples often require nucleic acid extraction and library construction, which includes multiple steps such as lysis, nucleic acid adsorption and purification, fragmentation, adapter ligation and purification, PCR and purification. Current extraction and library construction devices are mainly based on the pipetting workstation method, which uses movable pipette modules (containing one or more pipettes) to perform liquid manipulation in different sections of the instrument for different steps, while different sections provide the reaction temperatures required for different steps.
[0003] However, because pipetting workstations are open systems, aerosols generated during different steps of the pipetting process can easily be released into the environment, potentially contaminating adjacent samples and causing false positive results. To reduce the impact of aerosol contamination at different steps, strict laboratory zoning is required. Extraction and library preparation must be performed on different instruments in different laboratories, resulting in numerous experimental devices, a large space requirement, and complex operations, making full automation difficult. Summary of the Invention
[0004] In view of this, in order to solve at least one of the above technical problems, it is necessary to propose an automated sample preparation device.
[0005] In addition, embodiments of this application also provide a sample preparation method using the aforementioned automated sample preparation device, and a biochemical analysis system using the aforementioned automated sample preparation device.
[0006] In a first aspect, embodiments of this application provide an automated sample preparation apparatus, comprising: a first storage module, a reaction module, and a drive module. The reaction module includes at least one reaction channel, and the reaction channel includes multiple interconnected reaction regions, each of which is used to perform at least one step in the sample preparation process. The inlet end of the reaction channel is connected to the first storage module, and the outlet end of the reaction channel is connected to the drive module. The drive module generates a driving force to load fluid located in the first storage module into the reaction channel and transfer it between different reaction regions.
[0007] In some possible embodiments, the plurality of reaction zones include a temperature control zone and a magnetic separation zone, and the reaction module further includes a temperature control structure disposed in the temperature control zone and a magnetic component movable in the magnetic separation zone.
[0008] In some possible embodiments, the plurality of reaction zones further include a mixing zone, wherein the mixing zone, the temperature control zone, and the magnetic separation zone are sequentially connected along the liquid inlet end to the liquid outlet end of the reaction channel.
[0009] In some possible embodiments, the automated sample preparation apparatus further includes a gas-containing module, which includes a gas-containing channel communicating with the mixing zone, the gas-containing channel being filled with a sealing liquid, and the sealing liquid not reacting with the fluid in the reaction channel.
[0010] In some possible embodiments, the mixing zone includes a main mixing channel, a first branch channel, and a second branch channel. One end of the main mixing channel is connected to the temperature control zone, and the other end is connected to the first branch channel and the second branch channel, respectively. The end of the first branch channel away from the main mixing channel is connected to the first storage module, and the end of the second branch channel away from the main mixing channel is connected to the gas flow channel.
[0011] In some possible embodiments, the main mixing channel is curved, and the first branch channel, the second branch channel and the main mixing channel are Y-shaped; or, the main mixing channel is a mixing cavity.
[0012] In some possible embodiments, the magnetic separation zone is located near the liquid inlet of the reaction channel, the temperature control zone is located near the liquid outlet of the reaction channel, and the plurality of reaction zones further include a fluid distribution module connecting the magnetic separation zone and the temperature control zone. The fluid distribution module is also connected to a second storage module and is used to select the direction of fluid transport.
[0013] In some possible embodiments, the automated sample preparation device further includes a cleaning fluid storage module and a waste fluid collection module connected in communication with the drive module.
[0014] In some possible embodiments, the automated sample preparation apparatus further includes a collection module, and the drive module is also connected to the collection module. The drive module is also used to generate a driving force to transfer reaction products located in the reaction channel to the collection module.
[0015] Secondly, embodiments of this application provide a biochemical analysis system, characterized in that it includes a detection module and an automated sample preparation device as described above, wherein the detection module is used to detect at least one reaction product formed within the reaction region.
[0016] Thirdly, embodiments of this application provide a sample preparation method using the automated sample preparation apparatus described above, comprising the following steps: providing driving force through a driving module to cause fluid located in a first storage module to enter the reaction channel and enter a specific reaction region, the reaction region being used to perform at least one step of a biochemical reaction.
[0017] In some possible embodiments, the plurality of reaction zones include a mixing zone, a temperature control zone, and a magnetic separation zone, wherein the mixing zone, the temperature control zone, and the magnetic separation zone are sequentially connected along the liquid inlet end to the liquid outlet end of the reaction channel, and the reaction module further includes a temperature control structure disposed in the temperature control zone and a magnetic component movablely disposed in the magnetic separation zone. The sample preparation method includes the following steps: biochemical reaction: connecting the first storage module to the reaction channel, and using the driving module to allow the first fluid located in the first storage module to enter the temperature control zone, the temperature control zone having a required temperature, so that the first fluid undergoes a biochemical reaction in the temperature control zone to obtain a reaction product; mixing the reaction product with a second fluid: using the driving module to allow the second fluid located in the first storage module to enter the mixing zone and mix with the reaction product to obtain a mixed fluid; magnetic separation: first, bringing the magnetic element close to or attaching it to the magnetic separation zone, and then using the driving module to transfer the mixed fluid to the magnetic separation zone, so that the magnetic beads adsorbed with biological samples are adsorbed on the inner wall of the magnetic separation zone, and discharging the fluid separated from the magnetic beads; and elution: using the driving module to inject the eluent from the first storage module into the magnetic separation zone, and removing the magnetic element, so that the magnetic beads adsorbed with biological samples are dispersed into the eluent, incubating at room temperature for a period of time, and then performing magnetic bead separation, so that the biological samples are separated from the magnetic beads and enter the eluent, and then collecting the eluent containing the biological samples.
[0018] In some possible embodiments, the automated sample preparation apparatus further includes a gas-containing module, which includes a gas-containing channel communicating with the mixing zone. The gas-containing channel is filled with a sealing liquid, and the sealing liquid does not react with the fluid in the reaction channel. The step of mixing the reaction product with the second fluid includes: disconnecting the first storage module from the mixing zone and connecting the mixing zone to the gas-containing channel; temporarily storing the reaction product located in the mixing zone in the gas-containing channel by the driving module; disconnecting the gas-containing channel from the mixing zone and connecting the first storage module and the mixing zone; injecting the second fluid located in the first storage module into the mixing zone by the driving module; and connecting the gas-containing channel to the mixing zone, driving the reaction product into the mixing zone by the driving module, and mixing it with the second fluid entering the mixing zone to form a mixed fluid.
[0019] In some possible embodiments, the plurality of reaction zones include a temperature control zone and a magnetic separation zone, the magnetic separation zone being close to the liquid inlet end of the reaction channel and the temperature control zone being close to the liquid outlet end of the reaction channel. The reaction module further includes a temperature control structure disposed in the temperature control zone, a magnetic component movablely disposed in the magnetic separation zone, and a fluid distribution module connecting the magnetic separation zone and the temperature control zone. The fluid distribution module is also connected to a second storage module and is used to select the direction of fluid transport. The sample preparation method includes: a biochemical reaction: connecting the first storage module to the temperature-controlled zone via the fluid distribution module, and injecting a first fluid from the first storage module into the temperature-controlled zone via the drive module; then disconnecting the temperature-controlled zone from the fluid distribution module and the drive module to allow the first fluid to be contained within the temperature-controlled zone for a biochemical reaction, yielding reaction products; and magnetic separation: connecting the temperature-controlled zone to the second storage module via the fluid distribution module, and injecting the reaction products from the temperature-controlled zone into the second storage module via the drive module, mixing them with a magnetic bead solution in the second storage module; then connecting the temperature-controlled zone to the temperature-controlled zone via the fluid distribution module. The magnetic separation zone is connected, and the magnetic element is brought close to or attached to the magnetic separation zone. The driving module allows the magnetic bead solution containing the reaction product to enter the magnetic separation zone, so that the magnetic beads containing the reaction product are adsorbed on the inner wall of the magnetic separation zone, and the separated fluid is discharged. Elution: The first storage module is connected to the magnetic separation zone, and the magnetic element is removed. The driving module injects the eluent in the first storage module into the magnetic separation zone, and the magnetic beads containing the reaction product are dispersed in the eluent. After incubation at room temperature for a period of time, the magnetic beads are separated so that the reaction product is separated from the magnetic beads and enters the eluent. The eluent containing the reaction product is then collected.
[0020] In summary, the automated sample preparation device and method provided in this application divide the reaction channel into different reaction zones. Combined with a drive module, this allows the fluid to be processed to flow between these zones, completing various steps in sample preparation. This integrates multiple operational steps within a single reaction module, achieving a one-click, fully automated sample preparation process. This reduces human intervention, minimizes human error, saves manpower, and lowers costs. Furthermore, because the entire reaction channel is a closed system with sealed connections between different reaction zones, aerosols are not released into the environment. Therefore, the problem of false positives caused by aerosol contamination is effectively eliminated, improving the quality of the prepared samples and making the results more accurate and reliable. Attached Figure Description
[0021] Figure 1This is a module architecture diagram of an automated sample preparation apparatus provided in an embodiment of this application.
[0022] Figure 2 This is a schematic diagram of the structure of an automated sample preparation device provided in an embodiment of this application.
[0023] Figure 3 To adopt Figure 2 The flowchart shown illustrates the sample preparation process using the automated sample preparation device.
[0024] Figure 4 To adopt Figure 2 The diagram shows the specific fluid flow during sample preparation using the automated sample preparation device.
[0025] Figure 5 To adopt Figure 2 The flowchart shown illustrates the fluid mixing process in the automated sample preparation device.
[0026] Figure 6 To adopt Figure 2 The flowchart shown illustrates the mixing of reaction products and a second fluid in an automated sample preparation apparatus.
[0027] Figure 7 This is a schematic diagram of the structure of an automated sample preparation apparatus provided in another embodiment of this application.
[0028] Figure 8 To adopt Figure 7 The flowchart shown illustrates the sample preparation process using the automated sample preparation device.
[0029] Figure 9 This is a schematic diagram of the one-step library construction test process using the automated sample preparation device of this application.
[0030] Figure 10 This is a schematic diagram showing the band distribution of the library preparation products obtained by one-step library preparation and manual library preparation using the automated sample preparation device of this application.
[0031] Figure 11 This is a schematic diagram of a two-step PCR library preparation test using the automated sample preparation device described in this application.
[0032] Figure 12 This is a module architecture diagram of a biochemical analysis system provided in an embodiment of this application.
[0033] Figure 13 This is a schematic diagram of the structure of a biochemical analysis system provided in an embodiment of this application.
[0034] Explanation of main component symbols
[0035] Automated sample preparation device 100
[0036] First storage module 1
[0037] Reaction Module 2
[0038] First valve twenty one
[0039] Temperature control structure twenty two
[0040] Magnetic components twenty three
[0041] Second valve twenty four
[0042] Third valve 25
[0043] driver module 3
[0044] Reaction channel 4
[0045] Reaction Zone 41
[0046] Temperature-controlled area 41a
[0047] Magnetic separation region 41b
[0048] Mixing zone 41c
[0049] Main mixing channel 411
[0050] First branch flow channel 412
[0051] Second branch flow channel 413
[0052] Inlet end 42
[0053] liquid outlet end 43
[0054] Gas Capacity Module 5
[0055] airflow channel 51
[0056] Storage tank 52
[0057] Collection module 6
[0058] Cleaning fluid storage module 7
[0059] Waste liquid collection module 8
[0060] Fluid distribution module 91
[0061] Second storage module 92
[0062] Biochemical analysis system 1000
[0063] Detection module 10
[0064] The following detailed description, in conjunction with the accompanying drawings, will further illustrate this application. Detailed Implementation
[0065] The technical solutions in the embodiments of this application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of this application, and not all embodiments.
[0066] It should be noted that when a component is described as "fixed to" or "mounted to" another component, it can be directly on the other component or may be interspersed with an intermediate component. When a component is described as "set to" another component, it can be directly set on the other component or may be interspersed with an intermediate component. The term "and / or" as used herein includes all and any combination of one or more of the associated listed items.
[0067] It should be noted that although a logical order is shown in the flowchart, in some cases, the steps shown or described may be performed in a different order than that shown in the flowchart. The methods disclosed in the embodiments of this application include one or more steps or actions for implementing the method. Method steps and / or actions may be interchanged with each other without departing from the scope of the claims. Unless a specific order of steps or actions is specified, the order and / or use of specific steps and / or actions may be modified without departing from the scope of the claims.
[0068] Please see Figure 1As shown in the illustration, this application provides an automated sample preparation device 100, comprising: a first storage module 1, a reaction module 2, and a drive module 3. The reaction module 2 includes at least one reaction channel 4, which includes multiple interconnected reaction regions 41. Each reaction region 41 is used to perform at least one step in the sample preparation process. The first storage module 1 stores fluid, which may be, for example, a liquid or solution containing biological samples (such as human blood samples, tissue samples, or saliva samples) or various reagents required for biochemical analysis, or a suspension containing sample carriers (such as magnetic beads) carrying biological samples in sequencing library construction, but is not limited to these. The inlet end 42 of the reaction channel 4 is connected to the first storage module 1, and the outlet end 43 of the reaction channel 4 is connected to the drive module 3. The drive module 3 generates a driving force to load the fluid located in the first storage module 1 into the reaction channel 4 and transfer it between different reaction regions 41, thereby completing the sample preparation process. Specifically, the automated sample preparation device 100 can be used for sample preparation in gene sequencing processes, including nucleic acid extraction and sequencing library construction, and can automate the entire extraction-library construction process within a single device. It is understood that the automated sample preparation device 100 can include one reaction channel 4 as a single-channel extraction-library construction device, or it can include multiple reaction channels 4, thus expanding into a multi-channel extraction-library construction device.
[0069] The first storage module 1 may include multiple storage containers for storing various fluids, such as sample containers and reagent containers. In some embodiments, the reaction module 2 further includes a first valve 21. The first storage module 1 can be connected to the reaction channel 4 through the first valve 21 (e.g., a solenoid valve). By controlling the first valve 21, different storage containers can be connected to the reaction channel 4 to input different fluids into the reaction channel 4.
[0070] Please refer to both together. Figure 2As shown, different reaction zones 41 can be designed according to different biochemical reaction purposes, and combined with corresponding functional structures to achieve different reaction steps. Multiple reaction zones 41 may include a temperature control zone 41a and a magnetic separation zone 41b. The reaction module 2 also includes a temperature control structure 22 disposed in the temperature control zone 41a and a magnetic component 23 movable in the magnetic separation zone 41b. When the driving module 3 drives the fluid into the temperature control zone 41a, the temperature of the temperature control zone 41a can be controlled by the temperature control structure 22, for example, heating or cooling the fluid to achieve a specific biochemical reaction. When the driving module 3 drives the fluid into the magnetic separation zone 41b, the magnetic component 23 can be moved to approach or contact the flow channel of the magnetic separation zone 41b, thereby separating the magnetic carrier from the fluid. Furthermore, it is understood that there can be more than one temperature control zone 41a; by setting different temperature control zones 41a, biochemical reactions under different temperature conditions can be achieved.
[0071] The multiple reaction zones 41 may further include a mixing zone 41c, wherein the mixing zone 41c, the temperature control zone 41a, and the magnetic separation zone 41b are sequentially connected along the liquid inlet 42 to the liquid outlet 43 of the reaction channel 4. By setting the mixing zone 41c, the mixing steps of different fluids can be realized.
[0072] In some embodiments, the reaction channel 4 of the mixing zone 41c can be curved. For example, the reaction channel 4 can be wound around the same center multiple times to form a spiral shape, or the reaction channel 4 can be bent back and forth to form a wave-shaped or spiral structure, thereby extending the length of the reaction channel 4 in a limited space so that the fluid has enough path to achieve mixing in the mixing zone 41c and improve the mixing effect.
[0073] In some embodiments, the mixing zone 41c can be connected to the first storage module 1 through the first valve 21 to input different fluids into the mixing zone 41c to achieve fluid mixing.
[0074] Please refer to it again. Figure 2As shown, the automated sample preparation device 100 also includes a gas-containing module 5. The gas-containing module 5 includes a gas-containing channel 51 connected to the mixing zone 41c. The gas-containing channel 51 is filled with a sealing liquid, and the sealing liquid does not react with the fluid within the gas-containing channel 51. Specifically, the sealing liquid can be, for example, pure water. By connecting the gas-containing module 5 to the reaction channel 4, it can accommodate the air at the front end of the original fluid (e.g., solution or liquid) within the reaction channel 4. Since the gas-containing channel 51 is pre-filled with sealing liquid, and there is always sealing liquid in the gas-containing channel 51 throughout the entire workflow, the entire reaction process is enclosed within the gas-containing channel 51, preventing air from overflowing into the environment and thus avoiding the problem of aerosol spillage and pollution. The gas-containing module 5, on the one hand, can seal the entire reaction channel 4, preventing aerosol spillage; on the other hand, the fluid within the reaction channel 4 can undergo biochemical reactions or be temporarily stored within the gas-containing channel 51, which provides space for these reactions or storage.
[0075] In some embodiments, the gas-containing module 5 further includes a liquid storage tank 52. The end of the gas-containing channel 51 away from the mixing zone 41c is connected to the liquid storage tank 52. The liquid storage tank 52 contains a sealing liquid (e.g., pure water). In this way, when air at the liquid front end in the mixing zone 41c enters the gas-containing channel 51, it will push the sealing liquid in the gas-containing channel 51 into the liquid storage tank 52, but the air will not overflow into the environment. In addition, the gas-containing channel 51 can also serve as a temporary storage space for fluids in the reaction channel 4.
[0076] In some embodiments, the airflow channel 51 is curved, for example, the airflow channel 51 can be wound around a center multiple times to form a spiral shape.
[0077] In some embodiments, the mixing zone 41c includes a main mixing channel 411, a first branch channel 412, and a second branch channel 413. One end of the main mixing channel 411 is connected to the temperature control zone 41a, and the other end is connected to the first branch channel 412 and the second branch channel 413, respectively. The end of the first branch channel 412 away from the main mixing channel 411 is connected to the first storage module 1, and the end of the second branch channel 413 away from the main mixing channel 411 is connected to the gas flow channel 51. Specifically, the first branch channel 412 is connected to the first storage module 1 through a first valve 21, and the second branch channel 413 is connected to the gas flow channel 51 through a second valve 24. The connectivity of the mixing zone 41c can be selected by controlling the opening and closing of the first valve 21 and the second valve 24. In some embodiments, the main mixing channel 411 is curved as described above, and the first branch channel 412, the second branch channel 413, and the main mixing channel 411 form a Y-shape. The mixing zone 41c adopts a Y-shaped pipeline design for mixing. The mixing principle is that the flow velocity of the two fluids differs at the bend of the flow channel, which causes the proportion of the two fluids to change when flowing through this section of the flow channel, thereby achieving more thorough mixing of the two fluids. At the same time, combined with the curved structure of the main mixing channel 411, the fluids achieve further thorough mixing of the two fluids after multiple bends.
[0078] Understandably, in other embodiments, in addition to using the aforementioned Y-shaped tubing, the mixing zone 41c of the automated sample preparation device 100 can also be replaced with a mixing chamber. After different reagents are added to the mixing chamber, the mixing is achieved by alternating the suction / discharge actions of the drive module 3.
[0079] In some embodiments, the drive module 3 may specifically be a peristaltic pump, gear pump, syringe pump, plunger pump, centrifugal pump, diaphragm pump, etc.
[0080] Please refer to it again. Figure 1 and Figure 2 As shown, the automated sample preparation device 100 further includes a collection module 6, and the drive module 3 is also connected to the collection module 6. The drive module 3 is also used to generate a driving force to transfer the reaction products located in the reaction channel 4 to the collection module 6. Specifically, the drive module 3 is connected to the reaction channel 4 and the collection module 6 through a third valve 25 (e.g., a three-way valve).
[0081] The automated sample preparation device 100 also includes a cleaning solution storage module 7 and a waste liquid collection module 8 connected to the drive module 3. The drive module 3 can inject the cleaning solution from the cleaning solution storage module 7 into the reaction channel 4 to clean the reaction channel 4, and can also drive the waste liquid in the reaction channel 4 to the waste liquid collection module 8.
[0082] Please see Figure 3 As shown, combined Figure 1 As shown in the figure, this application embodiment also provides a sample preparation method, which is performed using the aforementioned automated sample preparation device 100, and specifically includes the following steps:
[0083] Step S1: Driving force is provided by the driving module 3 to cause the fluid in the first storage module 1 to enter the reaction channel 4 and enter a specific reaction zone 41, which is used to perform at least one step of the biochemical reaction.
[0084] like Figure 2 As shown, the reaction channel 4 is divided into three zones: a temperature control zone 41a, a magnetic separation zone 41b, and a mixing zone 41c. The driving force required for liquid manipulation is provided by a drive module 3 (e.g., an injection pump). The temperature control zone 41a provides the temperature required for the biochemical reaction, the magnetic separation zone 41b separates magnetic beads during purification, and the mixing zone 41c mixes the newly added fluid with the existing fluid in the reaction channel 4. One of the pipes at the front end of the mixing zone 41c (i.e., the second branch channel 413) connects to the gas flow channel 51 to accommodate air at the front end of the existing fluid in the reaction channel 4. Because the gas flow channel 51 is pre-filled with pure water and its other end is connected to pure water in the storage tank 52, and water is always present in the gas flow channel 51 throughout the entire process, the entire reaction process is enclosed within the gas flow channel 51, preventing air from escaping into the environment and thus avoiding aerosol contamination.
[0085] Please see Figure 4 As shown, based on Figure 2 The automated sample preparation device 100 shown includes steps such as biochemical reaction, fluid mixing, magnetic separation, and elution. Each step is described in detail below.
[0086] Step S11, biochemical reaction: Connect the first storage module 1 to the reaction channel 4, and use the drive module 3 to bring the first fluid in the first storage module 1 into the temperature control zone 41a of the reaction channel 4. The temperature control zone 41a has the required temperature so that the first fluid can undergo a biochemical reaction in the temperature control zone 41a to obtain reaction products.
[0087] Specifically, during the biochemical reaction, a first fluid, mixed outside the reaction channel 4, is first drawn into the temperature control zone 41a. At this time, the first valve 21 opens, the second valve 24 closes, the third valve 25 switches to position a, and the drive module 3 performs the liquid aspiration action. Once the liquid enters the designated position (i.e., temperature control zone 41a), the first valve 21 closes and the third valve 25 switches to position b. The first fluid can be a mixed solution of a biological sample (e.g., a nucleic acid sample) and corresponding reagents, and has been mixed before entering the automated sample preparation device 100.
[0088] In another embodiment, when the first fluid is a mixed solution, the different fluids can be pre-mixed in the mixing zone 41c of the reaction channel 4, and then the aforementioned biochemical reaction can be carried out. That is, before step S11, the preparation method further includes:
[0089] Step S10, fluid mixing: The driving module 3 provides driving force so that different fluids in the first storage module 1 enter the mixing zone 41c and are mixed in the mixing zone 41c to form a first fluid.
[0090] like Figure 5 As shown, the specific mixing steps are as follows:
[0091] Step D1: Connect the first storage module 1 and the mixing zone 41c, and provide driving force through the driving module 3 to allow the first fluid unit (e.g., nucleic acid sample) located in the first storage module 1 to enter the mixing zone 41c.
[0092] Step D2: Disconnect the first storage module 1 from the mixing zone 41c, and connect the mixing zone 41c with the flow channel 51. Use the drive module 3 to inject the first fluid unit located in the mixing zone 41c into the flow channel 51 for temporary storage.
[0093] Step D3: Disconnect the gas flow channel 51 from the mixing zone 41c, and connect the first storage module 1 and the mixing zone 41c. Inject the second fluid unit (e.g., reagent) located in the first storage module 1 into the mixing zone 41c through the drive module 3.
[0094] In step D4, the flow channel 51 is connected to the mixing zone 41c. Driven by the drive module 3, the first fluid unit is injected into the mixing zone 41c and mixed with the second fluid unit entering the mixing zone 41c to form the first fluid. Specifically, the uniform mixing of the mixture can be achieved by designing the pipeline of the mixing zone 41c.
[0095] Step S12, mixing of reaction products with the second fluid: the second fluid located in the first storage module 1 is brought into the reaction channel 4 by the drive module 3 and mixed with the reaction products to obtain a mixed fluid.
[0096] The fluid mixing method here is essentially the same as the fluid mixing methods described above. Please refer to [link / reference]. Figure 6 As shown, the method for mixing the reaction products with the second fluid specifically includes the following steps:
[0097] Step Q1: Disconnect the first storage module 1 from the mixing zone 41c, and connect the mixing zone 41c to the gas flow channel 51. The reaction product located in the mixing zone 41c is injected into the gas flow channel 51 for temporary storage through the driving module 3.
[0098] Step Q2: Disconnect the gas flow channel 51 from the mixing zone 41c, and connect the first storage module 1 and the mixing zone 41c. Inject the second fluid located in the first storage module 1 into the mixing zone 41c through the drive module 3.
[0099] In step Q3, the gas flow channel 51 is reconnected to the mixing zone 41c. Driven by the driving module 3, the reaction product is injected into the mixing zone 41c and mixed with the second fluid entering the mixing zone 41c to form a mixed fluid.
[0100] Specifically, when the reaction products need to be mixed with a second fluid (e.g., a new reagent or magnetic bead solution), the first fluid, after reacting in temperature control zone 41a, is first discharged into the pipeline (i.e., the second branch channel 413) connected to the second valve 24 at the front end of mixing zone 41c. At this time, the first valve 21 is closed, the second valve 24 is open, the third valve 25 is switched to position a, and the drive module 3 performs the discharge action. After the first fluid and the air at its front end in the second branch channel 413 enter the flow channel 51, the second valve 24 is closed, the first valve 21 is opened, and the drive module 3 continues to perform the suction action, drawing the second fluid from the first storage module 1 through the reagent needle into the pipeline before the first valve 21 (i.e., the first branch channel 412). Subsequently, the second valve 24 is opened, and the drive module 3 performs the suction action, and the second fluid and reaction products in the two pipelines (i.e., the first branch channel 412 and the second branch channel 413) are simultaneously drawn into the main mixing channel 411 of mixing zone 41c, achieving uniform mixing.
[0101] Understandably, when the second fluid is a newly introduced reagent, the resulting mixed fluid can be moved back to the temperature-controlled zone 41a for biochemical reactions. When the second fluid is a magnetic bead solution, the resulting mixed fluid is a mixed solution containing magnetic beads, which allows biological samples to be adsorbed onto the magnetic beads.
[0102] Step S13, magnetic separation: First, bring the magnetic component 23 close to or attach it to the magnetic separation zone 41b. Then, the mixed fluid containing magnetic beads is transferred to the magnetic separation zone 41b by the drive module 3, so that the magnetic beads with biological samples adsorbed are adsorbed on the inner wall of the magnetic separation zone 41b, and the fluid separated from the magnetic beads is discharged.
[0103] Specifically, during magnetic separation, the magnetic component 23 above the magnetic separation zone 41b is first placed tightly against the pipeline. Then, the mixed fluid containing magnetic beads is drawn into the magnetic separation zone 41b and slowly flows past the position where the magnetic component 23 is placed. To reduce the magnetic loss rate, the mixed fluid can be made to reciprocate within the pipeline. By repeatedly flowing past the position of the magnetic component 23, the magnetic beads in the mixed fluid containing magnetic beads can be fully adsorbed onto the inner wall of the magnetic separation zone 41b. During magnetic separation waste discharge, the waste liquid separated from the magnetic beads can be drawn into the pipeline between the third valve 25 and the drive module 3. At this time, the third valve 25 switches to position a, and the drive module 3 performs the liquid suction action. Then, it is discharged from the pipeline connected to position b of the third valve 25. At this time, the third valve 25 switches to position b, and the drive module 3 performs the liquid discharge action, finally discharging to the waste liquid collection module 8.
[0104] Step S14, elution: The eluent in the first storage module 1 is injected into the magnetic separation zone 41b by the drive module 3, and the magnetic component 23 is removed, so that the magnetic beads adsorbed with biological samples are dispersed into the eluent. After incubation at room temperature for a period of time, the magnetic beads are separated so that the biological samples are separated from the magnetic beads and enter the eluent. Then the eluent containing the biological samples is collected.
[0105] Specifically, when eluting biological samples (e.g., nucleic acid samples) adsorbed by magnetic beads, the first storage module 1 is connected to the reaction channel 4, and the eluent located in the first storage module 1 is drawn into the magnetic separation zone 41b. At this time, the magnetic component 23 is removed, the first valve 21 is opened, the second valve 24 is closed, the third valve 25 is switched to position a, and the drive module 3 performs the liquid aspiration action. When the eluent reaches the magnetic bead adsorption position, it can be made to reciprocate within the reaction channel 4, washing the magnetic beads off the inner wall of the magnetic separation zone 41b and dispersing them into the eluent. After incubation at room temperature for a period of time, the magnetic bead separation action described above is performed, and the eluent containing the biological sample is discharged from the pipeline connected to position b of the third valve 25 to the collection module 6 for collection, thus obtaining the final purified product.
[0106] It is understandable that one or more of the above four steps can be selected, but the selected steps must be consecutive. It is also understandable that one or more of the above steps can be performed once or multiple times, and are not limited to the above order.
[0107] For the gene sequencing process, the aforementioned automated sample preparation device 100 can realize all operations of sample extraction and library construction before sequencing within the reaction channel 4, achieving a one-click fully automated sample preparation process, reducing human intervention, reducing human operation errors, saving manpower, and reducing costs.
[0108] Please refer to the following: Figure 1 and Figure 7 As shown, another embodiment of this application provides an automated sample preparation device 200. This automated sample preparation device 200 has a structure basically the same as the aforementioned automated sample preparation device 100, with the main difference being that the reaction channel 4 only includes a temperature control zone 41a and a magnetic separation zone 41b. The magnetic separation zone 41b is located near the liquid inlet end 42 of the reaction channel 4, and the temperature control zone 41a is located near the liquid outlet end 43 of the reaction channel 4. The automated sample preparation device 200 also includes a fluid distribution module 91 connecting the magnetic separation zone 41b and the temperature control zone 41a. The fluid distribution module 91 is also connected to a second storage module 92, and the fluid distribution module 91 is used to select the direction of fluid transport. The second storage module 92 may include a container storing reagents required for the biochemical reaction or an empty container.
[0109] In some embodiments, the fluid distribution module 91 may be a rotary valve.
[0110] like Figure 7 The automated sample preparation device 200 shown is a test fixture for verifying the feasibility of the entire process of nucleic acid extraction and library construction for gene sequencing. This automated sample preparation device 200 only includes a temperature-controlled zone 41a and a magnetic separation zone 41b. The drive module 3 provides the power for liquid manipulation, and the fluid distribution module 91 and the drive module 3 are used to seal the temperature-controlled zone 41a. This automated sample preparation device 200 can test operations such as lysis, enzyme digestion, adapter ligation, PCR, and purification. Because it only tests a single sample, there is no need to consider aerosol contamination between samples.
[0111] Please see Figure 8 As shown, based on Figure 7 The automated sample preparation device 200 shown in this application embodiment also provides a sample preparation method, which may include steps such as biochemical reaction, magnetic bead separation, and elution. The specific steps are described below.
[0112] Step S21, biochemical reaction: The first storage module 1 is connected to the temperature control zone 41a through the fluid distribution module 91, and the first fluid in the first storage module 1 is injected into the temperature control zone 41a through the drive module 3. Then the temperature control zone 41a is disconnected from the fluid distribution module 91 and the drive module 3 so that the first fluid is sealed in the temperature control zone 41a to carry out the biochemical reaction.
[0113] Specifically, during heating, the fluid distribution module 91 is switched to the orifice communicating with the temperature control zone 41a (e.g., orifice 13 in the figure), thus connecting the magnetic separation zone 41b, the temperature control zone 41a, and the first storage module 1. The drive module 3 is then switched to the orifice communicating with the temperature control zone 41a (e.g., orifice O in the figure), drawing liquid from the first storage module 1 into the temperature control zone 41a. Afterward, the fluid distribution module 91 is switched to the adjacent orifice (e.g., orifice 11) of the orifice (orifice 13) connected to the temperature control zone 41a. The drive module 3 pushes in a section of air or pure water in the reverse direction to create pressure. Then, the drive module 3 is switched to the adjacent orifice (e.g., orifice B or I) to complete the sealing of the temperature control zone 41a. The purpose of this pressure-reducing operation is to increase the gas pressure within the reaction channel 4, thereby reducing evaporation during the liquid heating process.
[0114] Step S22, magnetic separation: The temperature control zone 41a is connected to the second storage module 92 through the fluid distribution module 91, and the reaction product of the temperature control zone 41a is injected into the second storage module 92 through the drive module 3 and mixed with the magnetic bead solution in the second storage module 92; then, the temperature control zone 41a is connected to the magnetic separation zone 41b through the fluid distribution module 91, and the magnetic component 23 is brought close to or attached to the magnetic separation zone 41b. The magnetic bead solution with the adsorbed reaction product is brought into the magnetic separation zone 41b through the drive module 3, so that the magnetic beads with the adsorbed reaction product are adsorbed on the inner wall of the magnetic separation zone 41b, and the separated fluid is discharged.
[0115] Specifically, during magnetic separation, the reacted product solution is first discharged from the temperature control zone 41a into a container (e.g., a reagent tube) in the second storage module 92. This container is pre-filled with a magnetic bead solution. The suction / discharge action of the drive module 3 is used to agitate the solution, mixing the reaction product with the magnetic bead solution to ensure the magnetic beads fully adsorb the reaction product. After mixing, the mixed liquid containing the magnetic beads is driven to the magnetic separation zone 41b. Before the mixed liquid reaches the magnetic separation zone 41b, the magnetic component 23 (e.g., an N52 magnetic rod) is attached to the pipe of the magnetic separation zone 41b. When the mixed liquid flows past the magnetic component 23, some of the magnetic beads are adsorbed. By repeatedly passing the mixed liquid past the magnetic component 23, it can be ensured that the vast majority (>99%) of the magnetic beads are adsorbed.
[0116] Step S23, elution: Connect the first storage module 1 to the magnetic separation zone 41b, remove the magnetic component 23, and inject the eluent in the first storage module 1 into the magnetic separation zone 41b through the drive module 3, and disperse the magnetic beads adsorbed with biological samples into the eluent. After incubating at room temperature for a period of time, perform magnetic bead separation so that the biological samples are separated from the magnetic beads and enter the eluent. Then collect the eluent containing the biological samples.
[0117] Specifically, during the elution step of purification, the magnetic component 23 is removed, and the eluent is drawn into the magnetic separation zone 41b and reciprocated to evenly disperse the magnetic beads adsorbed on the inner wall of the tube in the liquid. After incubation at room temperature is complete, the magnetic component 23 is placed tightly against the tube of the magnetic separation zone 41b, and the liquid is flowed through the magnetic component 23 multiple times to complete the adsorption of magnetic beads. Finally, the liquid is discharged and collected.
[0118] It is understandable that one or more of the above three steps can be selected, but the selected steps must be consecutive. It is also understandable that each of the above steps can be performed once or multiple times.
[0119] Based on existing library construction reagent kits, PCR and purification tests were performed using this automated sample preparation device 200. The test procedure is as follows: Figure 9 As shown, the procedure includes PCR, purification, and quantification steps, yielding a purified product concentration of 6.3 ng / µL. The library preparation product was analyzed by agarose gel electrophoresis and compared with that prepared manually (using the same reagent kit). The band distribution is shown below. Figure 10 As shown. The results indicate that the band distribution of the library constructed using the automated sample preparation device 200 of this application is consistent with that of the manual library construction, and is consistent with the average length of the library.
[0120] like Figure 11 As shown, the test procedure for two-step PCR library construction using the automated sample preparation device 200 of this application includes lysis, two PCRs, and two purifications, with the purified product concentration being 3 ng / uL.
[0121] Understandably, the automated sample preparation device 100 (200) of this application can be used as a standalone device or as a separate module integrated into a sequencer to realize an integrated extraction-library construction-sequencing machine.
[0122] Please see Figure 12 As shown, this application embodiment also provides a biochemical analysis system 1000, which includes the aforementioned automated sample preparation device 100 (200) and a detection module 10. The detection module 10 is used to detect the products after the biochemical reaction. Specifically, the aforementioned automated sample preparation device 100 (200) can be applied to IVD molecular diagnostics, for example, performing real-time PCR after nucleic acid extraction from blood samples or pharyngeal swabs. In some embodiments, a detection module 10 (e.g., an optical module) can be added above the temperature control zone 41a on the basis of the automated sample preparation device 200, so that the fluorescence intensity emitted in the reaction channel 4 can be detected in real time using a photomultiplier tube PMT, thereby realizing the function of qPCR.
[0123] The automated sample preparation device 100 (200) provided in this application divides the reaction channel 4 into different reaction zones 41. Simultaneously, the drive module 3 enables the fluid to be processed to flow between these zones, completing different steps in sample preparation. This integrates multiple operational steps within a single reaction module 2, achieving a one-click, fully automated sample preparation process. This reduces human intervention, minimizes human error, saves manpower, and lowers costs. Furthermore, since the entire reaction channel 4 is a closed channel with sealed connections between different reaction zones, aerosols are not released into the environment. Therefore, it effectively eliminates the problem of false positives caused by aerosol contamination, improving the quality of prepared samples and making the results more accurate and reliable.
[0124] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of this application and are not intended to limit it. Although this application has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of this application without departing from the spirit and scope of the technical solutions of this application.
Claims
1. An automated sample preparation device, characterized by, include: The first storage module, the reaction module, and the drive module, wherein the reaction module includes at least one reaction channel, the reaction channel includes multiple interconnected reaction regions, and each reaction region is used to perform at least one step in the sample preparation process; The inlet end of the reaction channel is connected to the first storage module, and the outlet end of the reaction channel is connected to the drive module. The drive module is used to generate driving force to load the fluid located in the first storage module into the reaction channel and transfer it between different reaction regions.
2. The automated sample preparation device of claim 1, wherein, The reaction zones include a temperature control zone and a magnetic separation zone. The reaction module also includes a temperature control structure disposed in the temperature control zone and a magnetic component that is movable in the magnetic separation zone.
3. The automated sample preparation apparatus as described in claim 2, characterized in that, The multiple reaction zones also include a mixing zone, and the mixing zone, the temperature control zone, and the magnetic separation zone are connected sequentially from the liquid inlet end to the liquid outlet end of the reaction channel.
4. The automated sample preparation apparatus as described in claim 3, characterized in that, It also includes a gas-containing module, which includes a gas-containing channel communicating with the mixing zone. The gas-containing channel is filled with a sealing liquid, and the sealing liquid does not react with the fluid in the reaction channel.
5. The automated sample preparation apparatus as described in claim 4, characterized in that, The mixing zone includes a main mixing channel, a first branch channel, and a second branch channel. One end of the main mixing channel is connected to the temperature control zone, and the other end is connected to the first branch channel and the second branch channel, respectively. The end of the first branch channel away from the main mixing channel is connected to the first storage module, and the end of the second branch channel away from the main mixing channel is connected to the gas flow channel.
6. The automated sample preparation apparatus as described in claim 5, characterized in that, The main mixing channel is curved, and the first branch channel, the second branch channel and the main mixing channel are Y-shaped. Alternatively, the main mixing channel may be a mixing cavity.
7. The automated sample preparation apparatus as described in claim 2, characterized in that, The magnetic separation zone is located near the liquid inlet of the reaction channel, and the temperature control zone is located near the liquid outlet of the reaction channel. The reaction module also includes a fluid distribution module that connects the magnetic separation zone and the temperature control zone. The fluid distribution module is also connected to the second storage module. The fluid distribution module is used to select the direction of fluid transport.
8. The automated sample preparation apparatus as described in claim 1, characterized in that, It also includes a cleaning fluid storage module and a waste fluid collection module that are connected to the drive module.
9. The automated sample preparation apparatus as described in claim 1, characterized in that, It also includes a collection module, and the drive module is also connected to the collection module. The drive module is also used to generate a driving force to transfer the reaction products located in the reaction channel to the collection module.
10. A biochemical analysis system, characterized in that, The device includes a detection module and an automated sample preparation apparatus as described in any one of claims 1 to 9, wherein the detection module is used to detect at least one reaction product formed within the reaction region.
11. A sample preparation method, characterized in that, The sample preparation method, applied to the automated sample preparation apparatus as described in claim 1, comprises the following steps: A driving force is provided by a driving module to cause fluid located in a first storage module to enter the reaction channel and then into a specific reaction region for performing at least one step of a biochemical reaction.
12. The sample preparation method according to claim 11, characterized in that, The reaction modules include a mixing zone, a temperature control zone, and a magnetic separation zone, which are sequentially connected along the liquid inlet to the liquid outlet of the reaction channel. The reaction module also includes a temperature control structure disposed in the temperature control zone and a magnetic component movable in the magnetic separation zone. The sample preparation method includes the following steps: Biochemical reaction: The first storage module is connected to the reaction channel, and the first fluid located in the first storage module is brought into the temperature control zone through the driving module. The temperature control zone has the required temperature so that the first fluid can carry out a biochemical reaction in the temperature control zone to obtain reaction products. The reaction product is mixed with the second fluid: the second fluid located in the first storage module is brought into the mixing zone by the driving module and mixed with the reaction product to obtain a mixed fluid; Magnetic separation: First, the magnetic component is brought close to or attached to the magnetic separation zone. Then, the mixed fluid is transferred to the magnetic separation zone via the driving module, so that the magnetic beads adsorbed with biological samples are adsorbed onto the inner wall of the magnetic separation zone, and the fluid separated from the magnetic beads is discharged; and Elution: The driving module injects the eluent from the first storage module into the magnetic separation zone and removes the magnetic components, dispersing the magnetic beads adsorbed with biological samples into the eluent. After incubation at room temperature for a period of time, the magnetic beads are separated to separate the biological samples from the magnetic beads and allow them to enter the eluent. The eluent containing the biological samples is then collected.
13. The sample preparation method according to claim 12, characterized in that, The automated sample preparation device further includes a gas-containing module, which includes a gas-containing channel communicating with the mixing zone. The gas-containing channel is filled with a sealing liquid, and the sealing liquid does not react with the fluid in the reaction channel. The step of mixing the reaction product with the second fluid includes: Disconnect the first storage module from the mixing zone and connect the mixing zone to the gas flow channel. Inject the reaction product located in the mixing zone into the gas flow channel for temporary storage via the driving module. Disconnect the gas flow channel from the mixing zone, and connect the first storage module and the mixing zone; inject the second fluid located in the first storage module into the mixing zone via the drive module; and The gas flow channel is connected to the mixing zone. Driven by the driving module, the reaction product is injected into the mixing zone and mixed with the second fluid entering the mixing zone to form a mixed fluid.
14. The sample preparation method according to claim 11, characterized in that, The multiple reaction zones include a temperature control zone and a magnetic separation zone. The magnetic separation zone is located near the liquid inlet of the reaction channel, and the temperature control zone is located near the liquid outlet of the reaction channel. The reaction module also includes a temperature control structure disposed in the temperature control zone, a magnetic component movable in the magnetic separation zone, and a fluid distribution module connecting the magnetic separation zone and the temperature control zone. The fluid distribution module is also connected to a second storage module and is used to select the direction of fluid transport. The sample preparation method includes: Biochemical reaction: The first storage module is connected to the temperature control zone through the fluid distribution module, and the first fluid in the first storage module is injected into the temperature control zone through the drive module. Then the temperature control zone is disconnected from the fluid distribution module and the drive module, so that the first fluid is sealed in the temperature control zone to carry out the biochemical reaction and obtain the reaction product. Magnetic separation: The temperature control zone is connected to the second storage module via the fluid distribution module, and the reaction product in the temperature control zone is injected into the second storage module via the drive module, where it mixes with the magnetic bead solution in the second storage module; then, the temperature control zone is connected to the magnetic separation zone via the fluid distribution module, and the magnetic component is brought close to or attached to the magnetic separation zone; the magnetic bead solution containing the reaction product is then introduced into the magnetic separation zone via the drive module, causing the magnetic beads containing the reaction product to adhere to the inner wall of the magnetic separation zone, and the separated fluid is discharged; and Elution: Connect the first storage module to the magnetic separation zone and remove the magnetic component. Use the driving module to inject the eluent from the first storage module into the magnetic separation zone and disperse the magnetic beads adsorbed with the reaction products into the eluent. After incubating at room temperature for a period of time, perform magnetic bead separation to separate the reaction products from the magnetic beads and allow them to enter the eluent. Then collect the eluent containing the reaction products.