Methods, apparatus, equipment, and storage media for determining fluid usage and reagent exchange.

Abstract

This application discloses a method, apparatus, device, and storage medium for determining fluid usage and reagent exchange. The method includes: sequentially introducing a first fluid without fluorescent label, a second fluid with fluorescent label, and a predetermined volume of a third fluid without fluorescent label into a fluid channel; determining the residual ratio of the second fluid in the fluid channel based on the first fluorescence parameter value, second fluorescence parameter value, and third fluorescence parameter value obtained at designated locations within the fluid channel; changing the predetermined volume of the third fluid introduced into the fluid channel and repeating the above steps until the residual ratio for all predetermined volumes is obtained; and determining a target predetermined volume based on the target residual ratio. This application, by accurately determining the usage capacity of the rinsing reagent and using a reasonable amount of reagent to complete reagent rinsing or replacement, can effectively reduce biochemical costs by ensuring that the fluid channel is thoroughly rinsed or the reagent is completely replaced without excessive reagent waste.

Description

Technical Field

[0001] This application relates to the technical field of fluid circuit systems, and more particularly to a method, apparatus, equipment, and storage medium for determining fluid usage and reagent exchange. Background Technology

[0002] Because of the flow of Poisson's leaf within the pipe, there is component diffusion when reagents of different components and concentrations flow in the pipe. Therefore, it is necessary to introduce an appropriate amount or an excessive amount of flushing reagent to ensure that the fluid channel is clean or that the reagent in the fluid channel is completely replaced.

[0003] The purpose of studying the fluid reagent replacement ratio, i.e. the reagent usage capacity, is to complete the rinsing and replacement of different reagents with the minimum reagent consumption. It is one of the important parameters of the sequencer fluid system, which is crucial to the overall consumption of fluid reagents and directly affects the sequencing cost of each run.

[0004] Current technologies rely solely on experience when determining reagent usage, adhering to the principle of "better too much than too little," leading to excessive reagent waste and increased sequencing costs. Furthermore, using too little reagent results in incomplete rinsing or replacement, severely impacting sequencing results. Therefore, rationally planning reagent usage is an urgent issue that needs to be addressed. Summary of the Invention

[0005] The main objective of this application is to provide a method, apparatus, equipment, and storage medium for determining fluid usage and reagent exchange, which can solve the problems in the prior art where the inability to accurately determine the usage volume of rinsing reagent leads to insufficient rinsing or excessive rinsing reagent causing reagent waste and increased sequencing costs.

[0006] To achieve the above objectives, the first aspect of this application provides a method for determining the fluid usage in a liquid circuit system, the method comprising:

[0007] A first fluid without fluorescent label is introduced into the fluid channel, and the first fluorescence parameter value at a specified location in the fluid channel is obtained;

[0008] A second fluid with a fluorescent label is introduced into the fluid channel to replace the first fluid, and the value of the second fluorescence parameter at a specified location in the fluid channel is obtained;

[0009] A set volume of a third fluid without fluorescent label is introduced into the fluid channel to replace the second fluid, and the value of the third fluorescence parameter at a specified location in the fluid channel is obtained.

[0010] The residual ratio of the second fluid in the fluid channel is determined based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value.

[0011] Change the set volume of the third fluid introduced into the fluid channel and repeat the above steps until the residual ratio for all set volumes is obtained;

[0012] The target set volume is determined based on the target residual ratio.

[0013] To achieve the above objectives, a second aspect of this application provides a reagent exchange method, the method comprising:

[0014] The fluid channel containing residual fifth fluid is flushed with a fourth fluid of a target set volume, so that the residual ratio of fifth fluid in the fluid channel does not exceed the allowable residual ratio threshold.

[0015] The target set volume is determined based on the method described above for determining the fluid usage in the liquid circuit system.

[0016] To achieve the above objectives, a third aspect of this application provides an apparatus for determining the fluid usage in a liquid circuit system, the apparatus comprising:

[0017] The first reagent exchange module is used to introduce a first fluid without fluorescent label into the fluid channel and to obtain the first fluorescence parameter value at a specified position in the fluid channel.

[0018] The second reagent exchange module is used to introduce a second fluid with a fluorescent label into the fluid channel to replace the first fluid, and to obtain the second fluorescence parameter value at a specified position in the fluid channel;

[0019] The third reagent exchange module is used to introduce a set volume of unlabeled third fluid into the fluid channel to replace the second fluid, and to obtain the third fluorescence parameter value at a specified position in the fluid channel.

[0020] The calculation module is used to determine the residual ratio of the second fluid in the fluid channel based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value;

[0021] The circulation module is used to change the set volume of the third fluid introduced into the fluid channel and repeat the above steps until the residual ratio under all set volumes is obtained;

[0022] The fluid usage determination module is used to determine the target set volume based on the target residual ratio.

[0023] To achieve the above objectives, a fourth aspect of this application provides a reagent exchange apparatus, the apparatus comprising:

[0024] The reagent exchange module is used to flush the fluid channel containing residual fifth fluid with a target set volume of fourth fluid, so that the residual ratio of fifth fluid in the fluid channel does not exceed the allowable residual ratio;

[0025] The target set volume is determined based on the device described above for determining the fluid usage in the liquid circuit system.

[0026] To achieve the above objectives, a fifth aspect of this application provides a computer-readable storage medium storing a computer program, which, when executed by a processor, causes the processor to perform the following steps:

[0027] A first fluid without fluorescent label is introduced into the fluid channel, and the first fluorescence parameter value at a specified location in the fluid channel is obtained;

[0028] A second fluid with a fluorescent label is introduced into the fluid channel to replace the first fluid, and the value of the second fluorescence parameter at a specified location in the fluid channel is obtained;

[0029] A set volume of a third fluid without fluorescent label is introduced into the fluid channel to replace the second fluid, and the value of the third fluorescence parameter at a specified location in the fluid channel is obtained.

[0030] The residual ratio of the second fluid in the fluid channel is determined based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value.

[0031] Change the set volume of the third fluid introduced into the fluid channel and repeat the above steps until the residual ratio for all set volumes is obtained;

[0032] The target set volume is determined based on the target residual ratio;

[0033] Alternatively, it causes the processor to perform the following steps:

[0034] The fluid channel containing residual fifth fluid is flushed with a fourth fluid of a target set volume, so that the residual ratio of fifth fluid in the fluid channel does not exceed the allowable residual ratio threshold.

[0035] The target set volume is determined based on the method described above for determining the fluid usage in the liquid circuit system.

[0036] To achieve the above objectives, a sixth aspect of this application provides a computer device, including a memory and a processor. The memory stores a computer program, and when the computer program is executed by the processor, the processor performs the following steps:

[0037] A first fluid without fluorescent label is introduced into the fluid channel, and the first fluorescence parameter value at a specified location in the fluid channel is obtained;

[0038] A second fluid with a fluorescent label is introduced into the fluid channel to replace the first fluid, and the value of the second fluorescence parameter at a specified location in the fluid channel is obtained;

[0039] A set volume of a third fluid without fluorescent label is introduced into the fluid channel to replace the second fluid, and the value of the third fluorescence parameter at a specified location in the fluid channel is obtained.

[0040] The residual ratio of the second fluid in the fluid channel is determined based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value.

[0041] Change the set volume of the third fluid introduced into the fluid channel and repeat the above steps until the residual ratio for all set volumes is obtained;

[0042] The target set volume is determined based on the target residual ratio;

[0043] Alternatively, it causes the processor to perform the following steps:

[0044] The fluid channel containing residual fifth fluid is flushed with a fourth fluid of a target set volume, so that the residual ratio of fifth fluid in the fluid channel does not exceed the allowable residual ratio threshold.

[0045] The target set volume is determined based on the method described above for determining the fluid usage in the liquid circuit system.

[0046] The embodiments of this application have the following beneficial effects:

[0047] This application determines the optimal target usage capacity of the rinsing reagent under a set scenario based on the residual ratio and allowable residual ratio obtained from multiple rounds of testing. It accurately determines the usage capacity of the rinsing reagent and uses a reasonable amount of reagent consumption to complete the rinsing or replacement of the reagent. This ensures that the fluid channel is thoroughly rinsed or the reagent is replaced without causing excessive waste of reagents, and can effectively reduce sequencing costs. Attached Figure Description

[0048] To more clearly illustrate the technical solutions in the embodiments of this application or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0049] in:

[0050] Figure 1 This is a flowchart illustrating the method for determining the fluid usage in a liquid circuit system as described in this application.

[0051] Figure 2 This is a graph showing the relationship between the residual ratio and the set volume of fluid in a specific embodiment of this application;

[0052] Figure 3This is a structural block diagram of the device for determining the fluid usage in the liquid circuit system in the embodiments of this application;

[0053] Figure 4 This is a structural block diagram of the computer device in the embodiments of this application. Detailed Implementation

[0054] The technical solutions of 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. Based on the embodiments of this application, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of this application.

[0055] like Figure 1 As shown, in one embodiment, a method for determining the fluid usage in a fluid circuit system is provided, the method comprising:

[0056] S100: Introduce a first fluid without fluorescent label into the fluid channel and obtain the first fluorescence parameter value at a specified location within the fluid channel.

[0057] Specifically, in the defined scenario, for the same chip, it is first rinsed with a first fluid without fluorescent labeling. The defined scenario specifically refers to the chip being connected to the same fluid path system and supplied with fluid by the same system. The chip includes, but is not limited to, sequencing chips known in the field of gene sequencing.

[0058] The chip includes at least one fluid channel for fluid flow.

[0059] The first fluid without fluorescent label is introduced into at least one fluid channel of the chip through the liquid circuit system, that is, at least one fluid channel is used to conduct parallel experiments in one experiment; in the next experiment, the first fluid without fluorescent label is introduced into all fluid channels of the at least one fluid channel again.

[0060] After rinsing the chip with the first fluid, the first fluorescence parameter value at a specified location within the fluid channel is obtained.

[0061] This embodiment prioritizes rinsing the chip with the first fluid, which can eliminate the interference of the chip's own fluorescent background on the residual ratio calculation. This minimizes the interference from fluorescent contamination and inherent fluorescent properties of the chip before the experiment, making the residual ratio calculation more accurate.

[0062] In addition, the amount of the first fluid is set according to the actual situation, and this application does not impose any restrictions on it.

[0063] Preferably, the same amount of the first fluid is used in each round of experiments.

[0064] S200: Introduce a second fluid with a fluorescent label into the fluid channel to replace the first fluid, and obtain the second fluorescence parameter value at a specified location within the fluid channel.

[0065] Specifically, a second fluid with a fluorescent label is introduced into the fluid channel that has been filled with the first fluid through the liquid circuit system, so that the second fluid replaces the first fluid in the fluid channel.

[0066] The concentration of fluorescent labeling can be set according to actual conditions. Preferably, a fully fluorescently labeled rinsing solution is used as the second fluid. The chip can be rinsed multiple times with the fluorescently labeled rinsing solution to achieve a complete rinsing effect. For example, the chip can be rinsed two, three, or four times in one round of testing. In addition, using the same concentration of fluorescently labeled rinsing solution and the same number of rinsing times in each round of testing can more effectively demonstrate the rinsing effect of different volumes of unlabeled rinsing solution on the same residual amount of fluorescently labeled rinsing solution.

[0067] After rinsing the chip with the second fluid, obtain the second fluorescence parameter value at a specified location within the fluid channel.

[0068] The chip is first rinsed with a second fluid to ensure it is sufficiently contaminated with the fluorescent label. Then, a predetermined volume of a third fluid without the fluorescent label is used to rinse the contaminated chip. This tests the effectiveness or replacement degree of the third fluid in rinsing the fluorescent label. The better the third fluid's rinsing effect on the fluorescent label, the less fluorescent label remains on the chip.

[0069] S300: Introduce a set volume of a third fluid without fluorescent label into the fluid channel to replace the second fluid, and obtain the third fluorescence parameter value at a specified location within the fluid channel.

[0070] Specifically, a set volume of a third fluid without fluorescent marking is introduced into the fluid channel that has already been filled with the second fluid through the liquid circuit system, so that the third fluid replaces the second fluid in the fluid channel.

[0071] In one specific embodiment, multiple fluid channels operate simultaneously. That is, in one experiment, the same predetermined volume of a third fluid is introduced into each of the multiple fluid channels that have already been supplied with a second fluid, and the experiments are conducted in parallel. In the next experiment, a different predetermined volume of a third fluid is introduced into each of the multiple fluid channels for parallel experiments. In the same experiment, the same predetermined volume of a third fluid is introduced into each fluid channel; however, in different rounds of experiments, the predetermined volume of the third fluid introduced is different.

[0072] Using multiple fluid channels in each experiment can make the experimental data more accurate and comprehensive.

[0073] In one specific embodiment, when only one fluid channel is used, a predetermined volume of a third fluid is introduced into the same fluid channel for each experiment. The predetermined volume of the third fluid is different for each experiment. This reduces reagent consumption during the experiment.

[0074] In one specific embodiment, multiple fluid channels operate simultaneously. However, in one experiment, different predetermined volumes of a third fluid are introduced into each of the multiple fluid channels that have already been supplied with the second fluid, allowing the fluid channels to conduct parallel experiments. In the next experiment, different predetermined volumes of the third fluid are introduced into all fluid channels in at least one fluid channel to conduct parallel experiments.

[0075] For example, a chip has 10 fluid channels. In one experiment, a third fluid with predetermined volumes of v1-v10 is introduced into each of the 10 channels. In the next experiment, a third fluid with predetermined volumes of v11-v20 is introduced into each of the 10 channels. The predetermined volumes v1-v20 are all different. In each experiment, the third fluid is introduced into each fluid channel once, and the predetermined volume of the third fluid introduced into each fluid channel is different.

[0076] In each experiment, the set volume of the third fluid introduced into each fluid channel is different, and the set volume of the third fluid introduced into the same fluid channel is also different in all rounds of experiments. In this way, experiments can be conducted simultaneously for multiple different set volumes, which can quickly complete experiments corresponding to all different set volumes, reduce the number of experiments, save reagent consumption during experiments, and speed up the determination of fluid usage in the liquid circuit system.

[0077] After rinsing the chip with the third fluid, obtain the third fluorescence parameter value at a specified location within the fluid channel.

[0078] Using different volumes of a third fluid in different tests to rinse chips that have already been rinsed with the second fluid may result in different rinsing effects or replacement levels, meaning that the amount of fluorescently labeled second fluid remaining on the chip may vary.

[0079] In this embodiment, fluorescence method is used to characterize the degree of reagent rinsing or replacement, that is, the actual amount of residual fluorescent labeling on the chip after rinsing with different volumes of rinsing solution is obtained based on the fluorescence parameter values ​​after each experiment.

[0080] Each round of experiments corresponds to a first fluorescence parameter value, a second fluorescence parameter value, and a third fluorescence parameter value.

[0081] S400: Determine the residual ratio of the second fluid in the fluid channel based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value.

[0082] Specifically, the same round of experiments includes three rounds of rinsing, obtaining the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value, respectively. Each round of experiments includes the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value corresponding to at least one fluid channel.

[0083] The first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value can all refer to the average fluorescence gray value or other image indicators that characterize the distribution of fluorescent markers.

[0084] In one specific embodiment, the residual ratio of the fluid channel in this round of experiments can be obtained based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value of the same fluid channel in the same experiment.

[0085] In another specific embodiment, the residual ratio of the chip in this round of experiments can be obtained based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value of all fluid channels in the same experiment.

[0086] A higher residue ratio indicates a higher amount of residue, resulting in a worse rinsing effect or replacement rate of the third fluid for the corresponding set volume; a lower residue ratio indicates a lower amount of residue, resulting in a better rinsing effect or replacement rate of the third fluid for the corresponding set volume.

[0087] S500: Change the set volume of the third fluid introduced into the fluid channel and repeat the above steps until the residual ratio for all set volumes is obtained.

[0088] Specifically, if the same set volume of third fluid is introduced into each fluid channel that has been introduced into the second fluid in each round, then after one round of experiment, the set volume of the third fluid is changed, and steps S100-S400 are repeated in the next round of experiment. In S300, the new set volume of third fluid is used to flush each fluid channel that has been introduced into the second fluid, and the new set volume of third fluid introduced into all fluid channels is also the same.

[0089] If each fluid channel that has been supplied with the second fluid in each round is supplied with a different set volume of the third fluid, then after one round of experiment, the set volume of the third fluid is changed, and steps S100-S400 are repeated in the next round of experiment. In S300, each fluid channel that has been supplied with the second fluid is supplied with a new set volume of the third fluid, and the new set volume of the third fluid supplied to different fluid channels is different.

[0090] By repeating this process, experiments corresponding to different set volumes can be completed.

[0091] Furthermore, in this embodiment, the same chip is rinsed in each round of experiments without replacing it. Since the chip is rinsed with a fluorescently labeled second fluid in each round of experiments, different experiments are equivalent to rinsing chips with the same level of contamination using different set volumes of third fluid, in order to observe the rinsing effect or replacement degree of various set volumes of third fluid.

[0092] S600: Determine the target set volume based on the target residual ratio.

[0093] Specifically, through multiple rounds of experiments, the flushing effect of different set volumes of the third fluid on the fluorescently labeled second fluid can be summarized by comparing the residual ratios. This allows us to determine which set volume of the third fluid, while meeting the target residual ratio (allowable residual ratio), provides both good flushing or replacement results and saves reagent usage. Thus, the optimal target usage of the fluid for reagent replacement in the specified scenario can be determined. In other words, the determined target set volume can be used as the fluid volume for reagent replacement in the liquid system.

[0094] In one embodiment, if the same chip is used in multiple rounds of experiments, step S100 is only performed in the very first round of experiments to rinse the chip with the first fluid and obtain a first fluorescence parameter value. Step S100 is not performed in subsequent experiments, thus reducing reagent waste.

[0095] In another embodiment, steps S100-S400 are performed in each round of experimentation to increase experimental reliability and data accuracy.

[0096] This embodiment determines the optimal target usage of the replacement reagent under a set scenario based on the residual ratio and target residual ratio obtained from multiple rounds of testing. It accurately determines the reagent usage capacity and uses a reasonable amount of reagent consumption to complete the reagent rinsing or replacement. This ensures that the chip is thoroughly rinsed or the reagent is fully replaced without causing excessive waste of reagents, and can effectively reduce the cost of biochemical experiments.

[0097] In one embodiment, obtaining the first fluorescence parameter value at a specified location within a fluid channel includes: after the first fluid is introduced, imaging the fluorescence signal at a specified location within each target fluid channel to obtain at least one first fluorescence image corresponding to each target fluid channel, and calculating the average fluorescence grayscale value of the first fluorescence image corresponding to each target fluid channel as the first fluorescence parameter value.

[0098] And / or,

[0099] Obtaining the second fluorescence parameter value at a specified location within the fluid channel includes: after the second fluid is introduced, imaging the fluorescence signal at a specified location within each target fluid channel to obtain at least one second fluorescence image corresponding to each target fluid channel, and calculating the average fluorescence grayscale value of the second fluorescence image corresponding to each target fluid channel as the second fluorescence parameter value;

[0100] And / or,

[0101] Obtaining the third fluorescence parameter value at a specified location within the fluid channel includes: after the third fluid is introduced, imaging the fluorescence signal at a specified location within each target fluid channel to obtain at least one third fluorescence image corresponding to each target fluid channel, and calculating the average fluorescence grayscale value of the third fluorescence image corresponding to each target fluid channel as the third fluorescence parameter value.

[0102] Specifically, during the experiment, the number of fluid channels through which fluid is introduced is no less than the number of target fluid channels.

[0103] For example, a chip may have 10 fluid channels, into which a first fluid, a second fluid, and a third fluid may be sequentially introduced in 5, 8, or 10 fluid channels. This application does not impose any restrictions on this.

[0104] When fluid is introduced into the five fluid channels, the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value of each of the five fluid channels can be obtained in the same experiment. These five fluid channels are the target fluid channels.

[0105] Alternatively, the first, second, and third fluorescence parameter values ​​of two, three, or four of the five fluid channels can be obtained in the same experiment. The selected two, three, or four fluid channels are the target fluid channels.

[0106] When fluid is introduced into 10 fluid channels, the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value of each of the five fluid channels can be obtained in the same experiment. These five fluid channels are the target fluid channels.

[0107] In addition, these 5 target fluid channels can be selected from the 10 fluid channels of the chip at intervals, that is, there is a non-target fluid channel between two adjacent target fluid channels.

[0108] Of course, it is also possible to obtain the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value for each of these 10 fluid channels in the same experiment. All 10 fluid channels are target fluid channels. This application does not impose any limitations on this.

[0109] In order to more accurately represent the residual amount of fluorescent markers, this embodiment will image the fluorescence signal at a designated location in the same target fluid channel after each round of rinsing to obtain a fluorescence image. The pixel grayscale of the fluorescence image can be used to characterize the residual amount of fluorescent markers.

[0110] Imaging is achieved by taking pictures of the target's field of view using an optical imaging system. This system includes a light source, such as a laser, and a camera. The laser emits a specific wavelength of light to illuminate a designated location within the target's fluid channel, exciting a fluorescent marker at that location to fluoresce. The camera then captures an image of the fluorescent region within the designated fluid channel.

[0111] Specifically, after the first fluid flushing is completed, the fluorescence signal at a designated location within each target fluid channel is imaged to obtain at least one first fluorescence image corresponding to each target fluid channel. The average fluorescence grayscale value is calculated for all first fluorescence images corresponding to the same target fluid channel, and the obtained average fluorescence grayscale value is used as the first fluorescence parameter value of that target fluid channel.

[0112] And / or, after the second fluid flushing is completed, the fluorescence signal at a specified location within each target fluid channel will be imaged to obtain at least one second fluorescence image corresponding to each target fluid channel. The average fluorescence grayscale value of all second fluorescence images corresponding to the same target fluid channel will be calculated, and the obtained average fluorescence grayscale value will be used as the second fluorescence parameter value of that target fluid channel.

[0113] And / or, after the third fluid flushing is completed, the fluorescence signal at a specified location within each target fluid channel will be imaged to obtain at least one third fluorescence image corresponding to each target fluid channel. The average fluorescence grayscale value of all third fluorescence images corresponding to the same target fluid channel will be calculated, and the obtained average fluorescence grayscale value will be used as the third fluorescence parameter value of that target fluid channel.

[0114] This process was repeated multiple times. After each round of experiments, each target fluid channel had a corresponding first fluorescence parameter value, second fluorescence parameter value, and third fluorescence parameter value.

[0115] For example, if there are 5 target fluid channels, then after each round of experiments, each target fluid channel will have a corresponding first fluorescence parameter value, second fluorescence parameter value, and third fluorescence parameter value.

[0116] The average fluorescence grayscale value refers to the average grayscale value of all pixels in the target area of ​​the fluorescence image, or the ratio of the sum of the grayscale values ​​of all pixels in the target area to the area of ​​the target area.

[0117] In this embodiment, the average fluorescence grayscale value of the fluorescence image is used to characterize the actual residual amount of fluorescent labeling on the chip. A higher average fluorescence grayscale value indicates a higher residual amount of fluorescent labeling.

[0118] In one embodiment, calculating the average fluorescence grayscale value of the first fluorescence image corresponding to each target fluid channel includes:

[0119] The region within a preset range of each first fluorescence image is designated as the first target region;

[0120] Calculate the average fluorescence grayscale value of all pixels in the first target region corresponding to the same target fluid channel.

[0121] Specifically, to remove the influence of edge optical effects and improve the accuracy of the average grayscale value, this embodiment selects a preset range within the first fluorescence image as the first target region, removes edge pixels of the first fluorescence image, and retains pixels near the center of the first fluorescence image. For example, pixels within a 19 / 20 range in the X direction and a 9 / 10 range in the Y direction from the center are retained, while the remaining pixels far from the center are removed. The selection of the preset range can be set according to actual applications, and this application does not limit it.

[0122] In one embodiment, calculating the average fluorescence grayscale value of the second fluorescence image corresponding to each target fluid channel includes:

[0123] The region within a preset range of each second fluorescence image is designated as the second target region;

[0124] Calculate the average fluorescence grayscale value of all pixels in the second target region corresponding to the same target fluid channel.

[0125] Specifically, to remove the influence of edge optical effects and improve the accuracy of the average grayscale value, this embodiment selects a predetermined range within the second fluorescence image as the second target region, removes edge pixels of the second fluorescence image, and retains pixels near the center of the second fluorescence image. For example, pixels within a 19 / 20 range in the X direction and a 9 / 10 range in the Y direction from the center are retained, while the remaining pixels far from the center are removed. The selection of the predetermined range can be set according to actual applications, and this application does not limit it.

[0126] In one embodiment, calculating the average fluorescence grayscale value of the third fluorescence image corresponding to each target fluid channel includes:

[0127] The region within a preset range of each third fluorescence image is designated as the third target region;

[0128] Calculate the average fluorescence grayscale value of pixels in all third target regions corresponding to the same target fluid channel.

[0129] Specifically, to remove the influence of edge optical effects and improve the accuracy of the average grayscale value, this embodiment selects a predetermined area within the third fluorescence image as the third target area, removes edge pixels of the third fluorescence image, and retains pixels near the center of the third fluorescence image. For example, pixels within a 19 / 20 range in the X direction and a 9 / 10 range in the Y direction from the center are retained, while the remaining pixels far from the center are removed. The selection of the predetermined range can be set according to actual applications, and this application does not limit it.

[0130] In one embodiment, the formula for calculating the residual ratio is shown in formula (1):

[0131] Residual ratio = (Third fluorescence parameter value - First fluorescence parameter value) / (Second fluorescence parameter value - First fluorescence parameter value) Formula (1);

[0132] In formula (1), the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value correspond to the same target fluid channel.

[0133] Specifically, the residual ratio in formula (1) is the residual ratio of a certain target fluid channel in an experiment. Each target fluid channel in each round of experiments corresponds to a residual ratio.

[0134] In one embodiment, if only steps S200-S400 are performed in each round of experiment, that is, if the chip is rinsed by the second fluid and the set volume of the third fluid in each round of experiment, then the residual ratio of the same target fluid channel in the same round of experiment is the ratio of the average fluorescence gray value of the fluorescence image obtained after rinsing with the set volume of the third fluid to the average fluorescence gray value of the fluorescence image obtained after rinsing with the second fluid.

[0135] That is, the residual ratio = the value of the third fluorescence parameter / the value of the second fluorescence parameter.

[0136] In one embodiment, the residual ratio of the chip in this round of experiments can be obtained based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value of all target fluid channels in the same experiment.

[0137] Specifically, the sum of the first fluorescence parameter values ​​of all target fluid channels in the same experiment is calculated as the first value;

[0138] The sum of the second fluorescence parameter values ​​of all target fluid channels in the same experiment is used as the second value;

[0139] Calculate the sum of the third fluorescence parameter values ​​for all target fluid channels in the same experiment, and use it as the third value;

[0140] After one round of experiments, the chip's residual ratio = (third value - first value) / (second value - first value).

[0141] or,

[0142] Calculate the average value of the first fluorescence parameter of all target fluid channels in the same experiment, and use it as the first value;

[0143] Calculate the average value of the second fluorescence parameter for all target fluid channels in the same experiment, and use it as the second value;

[0144] Calculate the average value of the third fluorescence parameter for all target fluid channels in the same experiment, and use it as the third value;

[0145] After one round of experiments, the chip's residual ratio = (third value - first value) / (second value - first value).

[0146] In one embodiment, the designated location within the fluid channel includes the target field of view area at the outlet of the target fluid channel.

[0147] Specifically, acquiring images at the outlet of the target fluid channel is not only more convenient, but the resulting images can also accurately reflect the residual level of fluorescent markers in the target fluid channel.

[0148] In one embodiment, the target field of view includes n*m fields of view, where n and m are both greater than or equal to 1.

[0149] Specifically, a chip typically includes multiple parallel fluid channels, also known as channels. These channels provide the physical space to contain liquids. Different solutions or reagents can be introduced into the fluid channels to carry out biochemical reactions, and the liquids after the reactions flow out of the fluid channels.

[0150] In this embodiment, at least one target fluid channel from multiple fluid channels of the chip can be selected for imaging of n*m fields of view. Each field of view corresponds to one image; that is, one target fluid channel corresponds to n*m first fluorescence images, n*m second fluorescence images, and n*m third fluorescence images. The number of target fluid channels and which fluid channels are selected can be set according to the actual situation. For example, one, two, or four target fluid channels can be selected, with the middle fluid channel selected as the target fluid channel, or non-adjacent fluid channels can be selected as target fluid channels, etc. This application does not limit this.

[0151] This application also provides a reagent exchange method for a chip, the chip including fluid channels, the method comprising:

[0152] The fluid channels of the chip containing residual fifth fluid are flushed with a fourth fluid of a target set volume, so that the residual ratio of fifth fluid in the fluid channels does not exceed the allowable residual ratio threshold.

[0153] The target set volume is determined according to the method for determining the fluid usage in the fluid circuit system as described above.

[0154] Specifically, preferably, the chip in this embodiment and the chip in the method for determining the amount of fluid in the liquid circuit system described above can be the same type of chip or the same batch of chips.

[0155] Preferably, the liquid circuit system connected to the chip in this embodiment is the same liquid circuit system as the liquid circuit system in the method for determining the amount of fluid in the liquid circuit system described above.

[0156] Figure 2 This is a graph showing the relationship between the residual ratio and the set fluid volume in a specific embodiment of this application. (Refer to...) Figure 2 This specific embodiment uses a 75µm long chip and a 0.5mm long common conduit as the test scenario, with an allowable residual ratio of 2% as the target. A second fluid with fluorescent labeling and a predetermined volume of a third fluid without fluorescent labeling are introduced into the common conduit and the chip in this test scenario. The residual ratio of each of the four target fluid channels is calculated under different replacement ratios. The replacement ratio is the ratio of the predetermined volume of the third fluid without fluorescent labeling to the sum of the volumes of the chip's fluid channels and the common conduit. In this embodiment, the replacement ratio is used as the equivalent predetermined volume.

[0157] from Figure 2 It can be seen that the residual ratio of target fluid channel LANE1_A is 70.73% when the replacement ratio is 1, the residual ratio of target fluid channel LANE2_A is 88.50% when the replacement ratio is 1, the residual ratio of target fluid channel LANE3_A is 89.11% when the replacement ratio is 1, and the residual ratio of target fluid channel LANE4_A is 84.47% when the replacement ratio is 1.

[0158] The residual ratio of target fluid channel LANE1_A at a replacement ratio of 2 is 9.62%, the residual ratio of target fluid channel LANE2_A at a replacement ratio of 2 is 15.34%, the residual ratio of target fluid channel LANE3_A at a replacement ratio of 2 is 17.71%, and the residual ratio of target fluid channel LANE4_A at a replacement ratio of 2 is 17.09%.

[0159] The residual ratio of target fluid channel LANE1_A at a replacement ratio of 3 is 1.98%, the residual ratio of target fluid channel LANE2_A at a replacement ratio of 3 is 3.16%, the residual ratio of target fluid channel LANE3_A at a replacement ratio of 3 is 2.78%, and the residual ratio of target fluid channel LANE4_A at a replacement ratio of 3 is 2.33%.

[0160] The residual ratio of target fluid channel LANE1_A at a replacement ratio of 3.5 is 1.13%, the residual ratio of target fluid channel LANE2_A at a replacement ratio of 3.5 is 1.19%, the residual ratio of target fluid channel LANE3_A at a replacement ratio of 3.5 is 1.90%, and the residual ratio of target fluid channel LANE4_A at a replacement ratio of 3.5 is 1.57%.

[0161] The residual ratio of target fluid channel LANE1_A at a replacement ratio of 4 is 0.44%, the residual ratio of target fluid channel LANE2_A at a replacement ratio of 4 is 0.82%, the residual ratio of target fluid channel LANE3_A at a replacement ratio of 4 is 0.45%, and the residual ratio of target fluid channel LANE4_A at a replacement ratio of 4 is 0.74%.

[0162] The residual ratio of target fluid channel LANE1_A at a replacement ratio of 5 is 0.38%, the residual ratio of target fluid channel LANE2_A at a replacement ratio of 5 is 0.43%, the residual ratio of target fluid channel LANE3_A at a replacement ratio of 5 is 0.58%, and the residual ratio of target fluid channel LANE4_A at a replacement ratio of 5 is 0.33%.

[0163] Therefore, it can be seen that the smaller the replacement ratio, i.e., the smaller the set volume of the third fluid without fluorescent labeling, the higher the residual ratio. As the replacement ratio increases, the residual ratio decreases. Moreover, the trend of residual ratio change is different for each target fluid channel. However, after the replacement ratio is greater than or equal to 3, the residual ratio of each target fluid channel decreases only slightly, indicating that simply increasing the replacement ratio cannot reduce the residual ratio indefinitely.

[0164] When the replacement ratio of the third fluid without fluorescent labeling is 1, the reagent residue is between 70% and 90%, indicating poor rinsing and ineffective reagent rinsing or replacement. When the replacement ratio is higher than 3.5, the concentration of the previous reagent remaining in the chip fluid channels is generally below the allowable residue ratio of 2%. A replacement ratio of 3.5 achieves good reagent replacement; beyond 3.5, the residue ratio shows no significant change. Based on these experimental results, the fluid replacement ratio should be at least 3.5 in the fluid design to ensure good reagent replacement while maintaining a reasonable reagent consumption level and reducing the cost of biochemical experiments.

[0165] certainly, Figure 2 This is merely a diagram showing the relationship between replacement ratio and residual ratio in a specific scenario. The relationship between replacement ratio and residual ratio is not limited to this in different application scenarios. Figure 2 As shown, this application does not impose any limitations on this. Furthermore, different residual ratios are permissible in different application scenarios, and the final selected replacement comparison, i.e., the target usage capacity of the reagent, will also differ; this application does not impose any limitations on this.

[0166] refer to Figure 3 This application also provides an apparatus for determining the fluid usage in a liquid circuit system, the apparatus comprising:

[0167] The first reagent exchange module 100 is used to introduce a first fluid without fluorescent label into the fluid channel and to obtain the first fluorescence parameter value at a specified position in the fluid channel.

[0168] The second reagent exchange module 200 is used to introduce a second fluid with a fluorescent label into the fluid channel to replace the first fluid with the second fluid, and to obtain the second fluorescence parameter value at a specified position in the fluid channel;

[0169] The third reagent exchange module 300 is used to introduce a set volume of a third fluid without fluorescent label into the fluid channel to replace the second fluid, and to obtain the third fluorescence parameter value at a specified position in the fluid channel.

[0170] The calculation module 400 is used to determine the residual ratio of the second fluid in the fluid channel based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value.

[0171] The circulation module 500 is used to change the set volume of the third fluid introduced into the fluid channel and repeat the above steps until the residual ratio under all set volumes is obtained.

[0172] The fluid usage determination module 600 is used to determine the target set volume based on the target residual ratio.

[0173] In one embodiment,

[0174] The first reagent exchange module 100 is specifically used to image the fluorescence signal at a specified position in each target fluid channel after the first fluid is introduced, to obtain at least one first fluorescence image corresponding to each target fluid channel, and to calculate the average fluorescence gray value of the first fluorescence image corresponding to each target fluid channel as the first fluorescence parameter value.

[0175] And / or,

[0176] The second reagent exchange module 200 is specifically used to image the fluorescence signal at a specified position in each target fluid channel after the second fluid is introduced, to obtain at least one second fluorescence image corresponding to each target fluid channel, and to calculate the average fluorescence gray value of the second fluorescence image corresponding to each target fluid channel as the second fluorescence parameter value.

[0177] And / or,

[0178] The third reagent exchange module 300 is specifically used to image the fluorescence signal at a specified location in each target fluid channel after the third fluid is introduced, to obtain at least one third fluorescence image corresponding to each target fluid channel, and to calculate the average fluorescence gray value of the third fluorescence image corresponding to each target fluid channel as the third fluorescence parameter value.

[0179] In one embodiment,

[0180] Calculate the average fluorescence grayscale value of the first fluorescence image corresponding to each target fluid channel, including:

[0181] The target region localization module locates a region within a preset range in each first fluorescence image as the first target region;

[0182] The average value calculation module calculates the average fluorescence grayscale value of all pixels in the first target region corresponding to the same target fluid channel.

[0183] In one embodiment, the formula for calculating the residual ratio is shown in formula (1):

[0184] Residual ratio = (Third fluorescence parameter value - First fluorescence parameter value) / (Second fluorescence parameter value - First fluorescence parameter value) Formula (1);

[0185] In formula (1), the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value correspond to the same target fluid channel.

[0186] In one embodiment, the designated location within the fluid channel includes the target field of view area at the outlet of the target fluid channel.

[0187] In one embodiment, the target field of view includes n*m fields of view, where n and m are both greater than or equal to 1.

[0188] This application also provides a reagent exchange device, which includes:

[0189] The reagent exchange module is used to flush the fluid channel containing residual fifth fluid with a target set volume of fourth fluid, so that the residual ratio of fifth fluid in the fluid channel does not exceed the allowable residual ratio;

[0190] The target set volume is determined by the device for determining the fluid usage in the liquid circuit system according to claim 8.

[0191] Figure 4 An internal structural diagram of a computer device in one embodiment is shown. This computer device can specifically be a terminal or a server. Figure 4As shown, the computer device includes a processor, memory, and network interface connected via a system bus. The memory includes a non-volatile storage medium and internal memory. The non-volatile storage medium stores an operating system and may also store a computer program. When executed by the processor, this computer program causes the processor to perform the steps in the above-described method embodiments. The internal memory may also store a computer program, which, when executed by the processor, causes the processor to perform the steps in the above-described method embodiments. Those skilled in the art will understand that... Figure 4 The structure shown is merely a block diagram of a portion of the structure related to the present application and does not constitute a limitation on the computer device to which the present application is applied. Specific computer devices may include more or fewer components than those shown in the figure, or combine certain components, or have different component arrangements.

[0192] In one embodiment, a computer device is provided, including a memory and a processor, the memory storing a computer program that, when executed by the processor, causes the processor to perform the following steps:

[0193] A first fluid without fluorescent label is introduced into the fluid channel, and the first fluorescence parameter value at a specified location in the fluid channel is obtained;

[0194] A second fluid with a fluorescent label is introduced into the fluid channel to replace the first fluid, and the value of the second fluorescence parameter at a specified location in the fluid channel is obtained;

[0195] A set volume of a third fluid without fluorescent label is introduced into the fluid channel to replace the second fluid, and the value of the third fluorescence parameter at a specified location in the fluid channel is obtained.

[0196] The residual ratio of the second fluid in the fluid channel is determined based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value.

[0197] Change the set volume of the third fluid introduced into the fluid channel and repeat the above steps until the residual ratio for all set volumes is obtained;

[0198] The target set volume is determined based on the target residual ratio;

[0199] Alternatively, it causes the processor to perform the following steps:

[0200] The fluid channel containing residual fifth fluid is flushed with a fourth fluid of a target set volume, so that the residual ratio of fifth fluid in the fluid channel does not exceed the allowable residual ratio threshold.

[0201] The target set volume is determined based on the method described above for determining the fluid usage in the liquid circuit system.

[0202] In one embodiment, a computer-readable storage medium is provided storing a computer program that, when executed by a processor, causes the processor to perform the following steps:

[0203] A first fluid without fluorescent label is introduced into the fluid channel, and the first fluorescence parameter value at a specified location in the fluid channel is obtained;

[0204] A second fluid with a fluorescent label is introduced into the fluid channel to replace the first fluid, and the value of the second fluorescence parameter at a specified location in the fluid channel is obtained;

[0205] A set volume of a third fluid without fluorescent label is introduced into the fluid channel to replace the second fluid, and the value of the third fluorescence parameter at a specified location in the fluid channel is obtained.

[0206] The residual ratio of the second fluid in the fluid channel is determined based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value.

[0207] Change the set volume of the third fluid introduced into the fluid channel and repeat the above steps until the residual ratio for all set volumes is obtained;

[0208] The target set volume is determined based on the target residual ratio;

[0209] Alternatively, it causes the processor to perform the following steps:

[0210] The fluid channel containing residual fifth fluid is flushed with a fourth fluid of a target set volume, so that the residual ratio of fifth fluid in the fluid channel does not exceed the allowable residual ratio threshold.

[0211] The target set volume is determined based on the method described above for determining the fluid usage in the liquid circuit system.

[0212] Those skilled in the art will understand that all or part of the processes in the methods of the above embodiments can be implemented by a computer program instructing related hardware. The program can be stored in a non-volatile computer-readable storage medium, and when executed, it can include the processes of the embodiments of the above methods. Any references to memory, storage, databases, or other media used in the embodiments provided in this application can include non-volatile and / or volatile memory. Non-volatile memory may include read-only memory (ROM), programmable ROM (PROM), electrically programmable ROM (EPROM), electrically erasable programmable ROM (EEPROM), or flash memory. Volatile memory may include random access memory (RAM) or external cache memory. By way of illustration and not limitation, RAM is available in various forms, such as static RAM (SRAM), dynamic RAM (DRAM), synchronous DRAM (SDRAM), dual data rate SDRAM (DDRSDRAM), enhanced SDRAM (ESDRAM), synchronous link DRAM (SLDRAM), RAMbus direct RAM (RDRAM), direct memory bus dynamic RAM (DRDRAM), and RAMbus dynamic RAM (RDRAM), etc.

[0213] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0214] The embodiments described above are merely illustrative of several implementation methods of this application, and while the descriptions are specific and detailed, they should not be construed as limiting the scope of this patent application. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of this application, and these all fall within the protection scope of this application. Therefore, the protection scope of this patent application should be determined by the appended claims.

Claims

1. A method for determining the fluid usage in a liquid circuit system, characterized in that, The method includes: A first fluid without fluorescent label is introduced into a fluid channel, and the first fluorescence parameter value at a specified location within the fluid channel is obtained; A second fluid with a fluorescent label is introduced into the fluid channel to replace the first fluid, and the second fluorescence parameter value at the designated location within the fluid channel is obtained; A set volume of a third fluid without fluorescent label is introduced into the fluid channel to replace the second fluid, and the value of the third fluorescence parameter at the specified position in the fluid channel is obtained; The residual ratio of the second fluid in the fluid channel is determined based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value. Change the set volume of the third fluid introduced into the fluid channel and repeat the above steps until the residual ratio for all set volumes is obtained; The target set volume is determined based on the target residual ratio.

2. The method according to claim 1, characterized in that, The step of obtaining the first fluorescence parameter value at a specified location within the fluid channel includes: after the first fluid is introduced, imaging the fluorescence signal at a specified location within each target fluid channel to obtain at least one first fluorescence image corresponding to each target fluid channel, and calculating the average fluorescence grayscale value of the first fluorescence image corresponding to each target fluid channel as the first fluorescence parameter value; And / or, The step of obtaining the second fluorescence parameter value at the specified location within the fluid channel includes: after the second fluid is introduced, imaging the fluorescence signal at the specified location within each target fluid channel to obtain at least one second fluorescence image corresponding to each target fluid channel, and calculating the average fluorescence grayscale value of the second fluorescence image corresponding to each target fluid channel as the second fluorescence parameter value; And / or, The step of obtaining the third fluorescence parameter value at the specified location within the fluid channel includes: after the third fluid is introduced, imaging the fluorescence signal at the specified location within each target fluid channel to obtain at least one third fluorescence image corresponding to each target fluid channel, and calculating the average fluorescence grayscale value of the third fluorescence image corresponding to each target fluid channel as the third fluorescence parameter value.

3. The method according to claim 2, characterized in that, The calculation of the average fluorescence grayscale value of the first fluorescence image corresponding to each of the target fluid channels includes: The region within a preset range of each of the first fluorescence images is designated as the first target region; Calculate the average fluorescence grayscale value of all pixels in the first target region corresponding to the same target fluid channel.

4. The method according to claim 2, characterized in that, The calculation of the average fluorescence grayscale value of the second fluorescence image corresponding to each of the target fluid channels includes: The region within a preset range of each second fluorescence image is designated as the second target region; Calculate the average fluorescence grayscale value of all pixels in the second target region corresponding to the same target fluid channel.

5. The method according to claim 2, characterized in that, The calculation of the average fluorescence grayscale value of the third fluorescence image corresponding to each of the target fluid channels includes: The region within a preset range of each third fluorescence image is designated as the third target region; Calculate the average fluorescence grayscale value of pixels in all third target regions corresponding to the same target fluid channel.

6. The method according to claim 2 or 3, characterized in that, The formula for calculating the residual ratio is shown in formula (1): Residual ratio = (Third fluorescence parameter value - First fluorescence parameter value) / (Second fluorescence parameter value - First fluorescence parameter value) Formula (1); In formula (1), the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value correspond to the same target fluid channel.

7. The method according to claim 1, characterized in that, The designated location within the fluid channel includes the target field of view area at the outlet of the target fluid channel.

8. The method according to claim 7, characterized in that, The target field of view region includes n*m fields of view, where n and m are both greater than or equal to 1.

9. A reagent exchange method, characterized in that, The method includes: The fluid channel containing residual fifth fluid is flushed with a fourth fluid of a target set volume, such that the residual ratio of the fifth fluid in the fluid channel does not exceed the allowable residual ratio threshold. The target set volume is determined by the method for determining the fluid usage in a fluid circuit system according to any one of claims 1-6.

10. A device for determining the fluid usage in a liquid circuit system, characterized in that, The device includes: The first reagent exchange module is used to introduce a first fluid without fluorescent label into the fluid channel and to obtain the first fluorescence parameter value at a specified position in the fluid channel; The second reagent exchange module is used to introduce a second fluid with a fluorescent label into the fluid channel to replace the first fluid with the second fluid, and to obtain the second fluorescence parameter value at the specified position in the fluid channel; The third reagent exchange module is used to introduce a set volume of a third fluid without fluorescent label into the fluid channel to replace the second fluid, and to obtain the third fluorescence parameter value at the specified position in the fluid channel; The calculation module is used to determine the residual ratio of the second fluid in the fluid channel based on the first fluorescence parameter value, the second fluorescence parameter value, and the third fluorescence parameter value; The circulation module is used to change the set volume of the third fluid introduced into the fluid channel and repeat the above steps until the residual ratio under all set volumes is obtained. The fluid usage determination module is used to determine the target set volume based on the target residual ratio.

11. A reagent exchange device, characterized in that, The device includes: The reagent exchange module is used to flush the fluid channel containing residual fifth fluid with a target set volume of fourth fluid, so that the residual ratio of the fifth fluid in the fluid channel does not exceed the allowable residual ratio; The target set volume is determined by the device for determining the fluid usage in the liquid circuit system according to claim 10.

12. A computer-readable storage medium storing a computer program, characterized in that, When the computer program is executed by a processor, it causes the processor to perform the steps of the method as described in any one of claims 1 to 8, or causes the processor to perform the steps of the method as described in claim 9.

13. A computer device, comprising a memory and a processor, characterized in that, The memory stores a computer program that, when executed by the processor, causes the processor to perform the steps of the method as described in any one of claims 1 to 8, or causes the processor to perform the steps of the method as described in claim 9.

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