Flow fluorescence-based method and sample analyzer

Abstract

The invention relates to the technical field of flow fluorescence, in particular to a method based on flow fluorescence and a sample analyzer. The method comprises the following steps: acquiring detection information of a sample, and detecting a microsphere compound prepared from the sample based on the detection information to obtain a light detection signal; identifying according to a two-dimensional signal distribution diagram formed by at least two optical signals to obtain effective item groups with the same number as the detection items; when the detection mode is a dual-fluorescence mode, processing the coordinates of the effective item group according to a preset weight; the effective item groups are sorted to obtain item numbers corresponding to the effective item groups, and the item numbers correspond to the detection items; and outputting a detection result of each detection item of the sample based on the effective item group and the corresponding item number. By determining the effective project groups, a data basis is provided for arrangement of the project groups; effective project groups and project numbers which are irregularly arranged correspond to each other through coordinate processing, so that the accuracy of a detection result is improved.

Description

Technical Field

[0001] The present invention relates to the technical field of flow fluorescence, and in particular to a method and a sample analyzer based on flow fluorescence. Background Art

[0002] Flow cytometry, also known as suspension arrays or liquid-phase chips, is a multi-indicator combined diagnostic technology that has developed over the past 20 years. Based on fluorescently encoded microspheres, this technology integrates flow cytometry principles, laser analysis, and high-speed digital signal processing, enabling parallel analysis of multiple indicators and accurate quantitative detection of up to 2-500 different biomolecules in a single tube. It boasts high throughput, high sensitivity, and parallel detection, and is applicable to a wide range of research fields, including immunoassays, nucleic acid research, enzymology, and receptor and ligand recognition analysis.

[0003] The basic idea of ​​flow cytometry is to fix specific probe molecules on each coded fluorescent microsphere to obtain capture microspheres and suspend them in a liquid phase system. The sample to be tested is labeled with a reporter fluorescence different from the coding and then placed in the above liquid phase system for a sufficient hybridization reaction. The microspheres after the hybridization reaction are detected and analyzed one by one by optical imaging or flow cytometry to determine the coded fluorescence and reporter fluorescence on each microsphere. The coded fluorescence determines the type of the molecule to be tested, and the reporter fluorescence intensity determines the content of the molecule to be tested, thereby achieving the purpose of single-sample multi-component detection.

[0004] When the microspheres are fluorescently coded, different fluorescent microspheres can be obtained by fluorescent coding of different brightness and microspheres of different particle sizes, thereby realizing simultaneous detection of different items (hereinafter referred to as the joint inspection item). For example, when performing auxiliary diagnosis of tumors, it is necessary to detect multiple tumor markers. Among them, when performing fluorescence coding of different brightness, the microspheres can be encoded by a single fluorescence of different brightness, or two different fluorescent mixed microspheres can be used for encoding. When a single fluorescence is used, the joint inspection items will be limited due to the limited distinction of a single fluorescence. When two fluorescences are used, the number of joint inspection items can be greatly increased. However, when two fluorescences are used, due to problems such as fluorescence crosstalk, the arrangement between the various project areas in the detection signal is irregular, so that the various project areas cannot correspond to the preset project numbers, and thus there is a problem of inaccurate detection results. Summary of the Invention

[0005] In view of this, the present invention provides a method and sample analyzer based on flow fluorescence to solve the technical problem that each project area cannot correspond to the preset project number, resulting in inaccurate detection results.

[0006] In a first aspect, the present invention provides a method based on flow fluorescence, the method comprising: obtaining detection information of a sample, and detecting a microsphere complex prepared from the sample based on the detection information to obtain an optical detection signal; wherein the optical detection signal comprises a plurality of optical signals, and the detection information comprises a detection mode and a detection item; identifying based on a two-dimensional signal distribution diagram composed of at least two optical signals to obtain a valid item group that is the same in number as the detection items; when the detection mode is a dual fluorescence mode, processing the coordinates of the valid item group according to a preset weight; and sorting the valid item group based on the processed coordinates to obtain an item number corresponding to the valid item group, and the item number corresponds to the detection item.

[0007] The flow fluorescence-based method provided in an embodiment of the present invention performs identification based on a two-dimensional signal distribution diagram composed of at least two optical signals, and obtains valid item clusters with the same number as the detection items, providing a data basis for the arrangement of the item clusters; after obtaining the valid item clusters, when using the dual fluorescence mode, the coordinates of the valid item clusters are processed by preset weights, thereby being able to match irregularly arranged valid item clusters with item numbers, thereby improving the accuracy of the detection results and avoiding the influence of fluorescence crosstalk.

[0008] In an optional embodiment, identification is performed based on a two-dimensional signal distribution diagram composed of at least two optical signals to obtain valid item clusters that are the same in number as the detection items, including: circle cluster identification is performed based on a two-dimensional signal distribution diagram composed of at least two optical signals to obtain multiple item clusters; and the item clusters are processed based on a comparison result of the number of item clusters and the number of detection items to obtain valid item clusters that are the same in number as the detection items.

[0009] In this embodiment, item clusters in the two-dimensional signal distribution diagram are obtained through circle cluster recognition. By comparing the item clusters with the detection items, valid item clusters with the same number of detection items are determined, thereby facilitating the determination of detection results for each detection item.

[0010] In an optional embodiment, the item clusters are processed based on a comparison result between the number of item clusters and the number of detection items to obtain valid item clusters that are the same as the number of detection items, including: determining the item area of ​​each detection item in the two-dimensional signal distribution map according to the detection items; when the number of item clusters is less than the number of detection items, filling the item clusters according to the item areas corresponding to the missing item clusters to obtain valid item clusters that are the same as the number of detection items; when the number of item clusters is greater than the number of detection items, deleting the redundant item clusters according to the item areas corresponding to each item cluster to obtain valid item clusters that are the same as the number of detection items.

[0011] In this embodiment, by completing or deleting the number of item groups, the correspondence between the number of item groups and the number of test items is achieved, thereby facilitating the determination of the test results of each test item.

[0012] In an optional embodiment, the detection information also includes the range of fluorescence signals that the microspheres can generate, and the light signal includes a first fluorescence signal and a second fluorescence signal; the first fluorescence signal and the second fluorescence signal are used to distinguish different detection items based on different signal intensities; when the detection mode is a dual fluorescence mode, the coordinates of the valid item group are processed according to the preset weights, including: when the detection mode is a dual fluorescence mode, based on the two-dimensional signal distribution map composed of the first fluorescence signal and the second fluorescence signal and the range of fluorescence signals that the microspheres can generate, the coordinates of the valid item group are processed according to the preset weights.

[0013] In an optional embodiment, the detection information also includes the range of fluorescence signals that the microspheres can generate, and the light signal includes a forward scattering signal and a fluorescence signal; the method also includes: when the detection mode is a single fluorescence mode, based on the two-dimensional signal distribution diagram composed of the forward scattering signal and the fluorescence signal and the range of fluorescence signals that the microspheres can generate, the valid project groups are sorted according to the horizontal and vertical coordinates of the valid project groups, and the sorted valid project groups correspond to the project numbers.

[0014] In this embodiment, for two different fluorescence modes, single fluorescence mode and dual fluorescence mode, valid signal regions are determined within different two-dimensional signals and processed into valid item clusters. This ensures the accuracy of valid item cluster sorting and makes the method applicable to multi-fluorescence applications. Furthermore, when using single fluorescence mode, valid item clusters are sorted according to their horizontal and vertical coordinates to ensure a correspondence between valid item clusters and item numbers.

[0015] In an optional embodiment, before performing identification based on a two-dimensional signal distribution diagram composed of at least two optical signals, the method further includes: cutting and filtering at least one optical signal among the multiple optical signals based on a signal threshold; and filtering corresponding other optical signals based on the filtered optical signal.

[0016] In this embodiment, by adopting the signal threshold to filter the single light signal and the corresponding light signal, the small bubbles are filtered out, thereby ensuring the accuracy of subsequent recognition.

[0017] In an optional embodiment, the optical signal includes a forward scattering signal and a side scattering signal. Before identification is performed based on a two-dimensional signal distribution diagram composed of at least two optical signals, the method further includes: determining the particle size of the microspheres based on the detection information; determining an effective area in the two-dimensional signal distribution diagram composed of the forward scattering signal and the side scattering signal based on the particle size, and the effective area is the range for identifying the project group.

[0018] In this embodiment, the effective area in the two-dimensional signal is delineated by the particle size of the microspheres, so that the method can be applied to application scenarios with multiple particle sizes.

[0019] In an optional embodiment, the test results of each test item of the sample are output based on the valid item group and the corresponding item number, including: obtaining the valid item group and the corresponding quantitative fluorescence value; determining the concentration according to the preset calibration curve and the quantitative fluorescence value, and outputting the concentration as the test result of the test item corresponding to each item number.

[0020] In this embodiment, by using the valid project group and project number, the concentration corresponding to the quantitative fluorescence value is determined in the preset calibration curve, thereby ensuring the accuracy of the detection result.

[0021] In a second aspect, the present invention provides a sample analyzer, which includes: a detection module for obtaining detection information of the sample, and a light detection signal obtained by detecting a microsphere complex prepared from the sample based on the detection information; wherein the light detection signal includes multiple light signals, and the detection information includes a detection mode and a detection item; a control module for identifying according to a two-dimensional signal distribution diagram composed of at least two light signals to obtain valid item clusters with the same number as the detection items; when the detection mode is a dual fluorescence mode, the coordinates of the valid item clusters are processed according to preset weights; the valid item clusters are sorted based on the processed coordinates to obtain the item numbers corresponding to the valid item clusters, and the item numbers correspond to the detection items; and the detection results of each detection item of the sample are output based on the valid item clusters and the corresponding item numbers.

[0022] In an optional embodiment, the at least two optical signals include a first fluorescent signal and a second fluorescent signal, and the first fluorescent signal and the second fluorescent signal are light signals scattered by the microsphere complex irradiated by the same laser. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the specific embodiments or the description of the prior art. Obviously, the drawings described below are some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0024] Figure 1 is a schematic flow chart of a method based on flow cytometry according to an embodiment of the present invention;

[0025] Figure 2 is a schematic diagram of an arrangement of items in a dual fluorescence mode according to an embodiment of the present invention;

[0026] Figure 3Ais a schematic diagram of a two-dimensional signal composed of a side scatter signal and a first fluorescence signal according to an embodiment of the present invention;

[0027] Figure 3B is a schematic diagram of a two-dimensional signal composed of a forward scattered signal and a side scattered signal according to an embodiment of the present invention;

[0028] Figure 3C is a schematic diagram of a two-dimensional signal composed of a first fluorescent signal and a second fluorescent signal according to an embodiment of the present invention;

[0029] Figure 4 is a schematic diagram of a process for deleting a project group according to an embodiment of the present invention;

[0030] Figure 5 is a schematic diagram of an arrangement of items in a single fluorescence mode according to an embodiment of the present invention;

[0031] Figure 6 is a structural block diagram of a sample analyzer according to an embodiment of the present invention;

[0032] Figure 7 Schematic diagram of the hardware structure of a computer device according to an embodiment of the present invention. DETAILED DESCRIPTION

[0033] To make the purpose, technical solutions, and advantages of the embodiments of the present invention more clear, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without making creative efforts shall fall within the scope of protection of the present invention.

[0034] According to an embodiment of the present invention, a method embodiment based on flow cytometry is provided. It should be noted that the steps shown in the flowchart of the accompanying drawings can be executed in a computer system such as a set of computer-executable instructions, and although a logical order is shown in the flowchart, in some cases, the steps shown or described can be executed in an order different from that shown here.

[0035] In this embodiment, a method based on flow fluorescence is provided, which can be used in electronic devices, etc. Figure 1 is a flow chart of a method based on flow cytometry according to an embodiment of the present invention. Figure 1 As shown, the process includes the following steps:

[0036] Step S101, obtains the detection information of the sample, and the optical detection signal obtained by detecting the microsphere complex prepared from the sample based on the detection information; wherein the optical detection signal includes multiple optical signals, and the detection information includes the detection mode and the detection items. Specifically, the basic idea of ​​flow fluorescence technology is to fix specific probe molecules on each coded fluorescent microsphere to obtain capture microspheres and suspend them in a liquid phase system, label the sample to be tested with a reporter fluorescence different from the coding, and then place it in the above-mentioned liquid phase system for sufficient hybridization reaction. After the hybridization reaction, the microspheres are detected and analyzed one by one by optical imaging or flow cytometry to determine the coded fluorescence and reporter fluorescence on each microsphere. The coded fluorescence determines the type of the molecule to be tested, and the reporter fluorescence intensity determines the content of the molecule to be tested, thereby achieving the purpose of single sample multi-component detection.

[0037] The detected optical signals include one or more of forward scatter (FSC), side scatter (SSC), quantitative fluorescence (QFL), and classified fluorescence (CFL). The FSC signal characterizes the size of each microsphere in the test sample, the SSC signal characterizes the internal structure of each microsphere in the test sample, the CFL signal characterizes the type of each microsphere in the test sample, and the QFL signal characterizes the concentration of the test item on the corresponding microsphere.

[0038] The detection items in the detection information include multiple detection items when performing joint detection. For example, when performing auxiliary diagnosis of tumors, it is necessary to detect multiple tumor markers, and the detection of each tumor marker corresponds to one detection item. In this embodiment, the detection mode includes a single fluorescence mode and a dual fluorescence mode. When the fluorescence mode is a single fluorescence mode, the classified fluorescence signal includes only one fluorescence signal; when the fluorescence mode is a dual fluorescence mode, the classified fluorescence signal includes two signals, a first fluorescence signal CFL1 and a second fluorescence signal CFL2. The first fluorescence signal can provide M types of signal intensities, and the second fluorescence signal can provide N types of signal intensities. The first fluorescence signal and the second fluorescence signal can realize joint detection of M*N types of items, identify M*N types of item groups and output the detection results.

[0039] It should be noted that single fluorescence mode refers to encoding microspheres using a single color of fluorescence. For example, different brightnesses of single fluorescence are used to encode the same microsphere, or a single fluorescence of a single brightness is used to encode multiple microspheres of different sizes, or a single fluorescence of different brightness is used to encode multiple microspheres of different sizes. Dual fluorescence mode refers to encoding microspheres using a mixture of two different fluorescences. In practical applications, the microspheres can be inorganic microspheres, polymer microspheres, or magnetic microspheres. This application does not limit the type of microspheres.

[0040] Step S102, identify according to the two-dimensional signal distribution diagram composed of at least two optical signals, and obtain a valid item group that is the same as the number of detection items. Specifically, at least two optical signals are combined to form a two-dimensional signal distribution diagram. Wherein, when combining, one of the optical signals can be used as the horizontal coordinate and the other optical signal as the vertical coordinate, and then the specific numerical value corresponding to each optical signal is marked in the coordinate diagram to obtain a scatter plot containing the two optical signals, and the scatter plot is the two-dimensional signal distribution diagram corresponding to the two optical signals. By detecting and identifying the item groups in the two-dimensional signal distribution diagram, the valid item group finally obtained is the same as the number of detection items, which facilitates the determination of the detection results of each detection item. It should be noted that the item group here refers to the signal area formed by a plurality of points corresponding to each detection item in the two-dimensional signal distribution diagram gathered together, and the valid item group that is the same as the number of detection items is obtained by identifying the item group, that is, the item group obtained by identification is the valid item group.

[0041] Step S103: When the detection mode is the dual fluorescence mode, the coordinates of the valid item clusters are processed according to preset weights. Specifically, in this embodiment, the detection modes include single fluorescence mode and dual fluorescence mode. When the dual fluorescence mode is used, the arrangement of multiple valid item clusters is irregular, rather than the horizontal and vertical grid-like arrangement of the single fluorescence mode. Therefore, the valid item clusters obtained when using the dual fluorescence mode cannot be matched one-to-one with their order and item number. Therefore, the coordinates of the valid item clusters are processed based on the preset weights. The valid item clusters are reflected in the two-dimensional signal distribution map, that is, the coordinates of the valid item clusters include horizontal and vertical coordinates.

[0042] Among them, when processing the coordinates, the horizontal coordinate can be processed or the vertical coordinate can be processed. When processing the horizontal coordinate, the processed coordinate can be expressed as W×X+Y, that is, the horizontal coordinate X is multiplied by the preset weight W and then added to the vertical coordinate; when processing the vertical coordinate, the processed coordinate can be expressed as X+W×Y, that is, the vertical coordinate Y is multiplied by the preset weight W and then added to the horizontal coordinate. In actual applications, if it is necessary to prioritize the X direction, the horizontal coordinate processing method is adopted. If it is necessary to prioritize the Y direction, the vertical coordinate processing method is adopted. The priority of the X direction or the Y direction arrangement depends on the preset project number and the corresponding method of the project group. Therefore, for the preset weight, it can be set to a value greater than 1, thereby increasing the weight proportion of the corresponding direction.

[0043] Step S104: sort the valid item groups based on the processed coordinates to obtain the item numbers corresponding to the valid item groups. The item numbers correspond to the test items. Specifically, the processed coordinates can be sorted from large to small, and then the sorted valid item groups are matched with the item numbers. Figure 2 As shown, there are 33 valid project groups in total, and the sorted valid project groups correspond to numbers 101-133 respectively.

[0044] Step S105 outputs the test results for each test item in the sample based on the valid item clusters and their corresponding item numbers. Specifically, the detected light signal also includes a QFL signal representing the concentration of the test item on the corresponding microsphere. Therefore, when outputting the test results for each test item, the valid item cluster, its corresponding item number, and its concentration can be simultaneously output as the test result.

[0045] The flow fluorescence-based method provided in an embodiment of the present invention performs identification based on a two-dimensional signal distribution diagram composed of at least two optical signals, and obtains valid item clusters with the same number as the detection items, providing a data basis for the arrangement of the item clusters; after obtaining the valid item clusters, when using the dual fluorescence mode, the coordinates of the valid item clusters are processed by preset weights, thereby being able to match irregularly arranged valid item clusters with item numbers, thereby improving the accuracy of the detection results and avoiding the influence of fluorescence crosstalk.

[0046] In this embodiment, a method based on flow cytometry is provided, and the process includes the following steps:

[0047] Step S201: Obtain sample detection information and obtain a light detection signal by detecting a microsphere complex prepared from the sample based on the detection information; wherein the light detection signal includes multiple light signals, and the detection information includes a detection mode and detection items. Figure 1Step S101 of the illustrated embodiment will not be described in detail here.

[0048] Step S202, based on the signal threshold, at least one of the multiple light signals is cut and filtered, and based on the filtered light signal, the corresponding other light signals are filtered. Specifically, due to the influence of small bubbles present during detection, there may be stray light in the acquired light signal. Therefore, one of the light signals can be selected for filtering, such as the forward scattering signal, and the forward scattering signal is cut and filtered based on the signal threshold. That is, the signal threshold can be set in advance, the forward scattering signal and the signal threshold are compared, and the forward scattering signal that exceeds the signal threshold is filtered out. At the same time, when acquiring the light signal, the multiple light signals are acquired at the same time, that is, there is a one-to-one correspondence between the multiple light signals. Therefore, after filtering the forward scattering signal, the corresponding remaining light signals such as the side scattering signal, the classified fluorescence signal and the quantitative fluorescence signal are all filtered. This step can also be understood as when one signal of a microsphere is identified as a small bubble, the other signals of the microsphere are also deleted synchronously.

[0049] Step S203 : performing identification based on the two-dimensional signal distribution diagram formed by at least two optical signals to obtain valid item groups having the same number as the number of detection items.

[0050] Specifically, the above step S203 includes:

[0051] Step S2031: Cluster identification is performed based on a two-dimensional signal distribution map composed of at least two optical signals, resulting in multiple item clusters. Specifically, when using a dual-fluorescence mode, the optical signal includes a forward scatter signal, a side scatter signal, a first fluorescence signal, and a second fluorescence signal. Two of these signals can be combined to generate a corresponding two-dimensional signal distribution map. Cluster identification, i.e., item cluster identification within the two-dimensional signal distribution map, is then performed based on the resulting two-dimensional signal distribution map.

[0052] In an optional implementation, the above step S2031 includes:

[0053] Step a1, perform signal threshold filtering based on the two-dimensional signal distribution diagram composed of the side scatter signal and the first fluorescent signal to obtain the first signal area; specifically, when the two-dimensional signal distribution diagram is composed of the side scatter signal and the first fluorescent signal, the side scatter signal can be used as the horizontal coordinate and the first fluorescent signal as the vertical coordinate, and then the specific signal value is marked in the coordinate system to obtain the two-dimensional signal distribution diagram composed of the side scatter signal and the first fluorescent signal. The two-dimensional signal distribution diagram is also called the two-dimensional signal SSC-CFL1, that is, the two-dimensional signal distribution diagram of the SSC-CFL1 dimension. In this embodiment, the obtained two-dimensional signal distribution diagram is as follows Figure 3AAs shown in Figure 2 , signal threshold filtering is performed by comparing the preset side scatter threshold and the first fluorescence signal threshold with the specific signal value. This filtering process can identify the effective signal region, namely the first signal region, while also removing the background signal.

[0054] Step a2, performing signal threshold filtering on the first signal area according to the two-dimensional signal distribution diagram composed of the forward scattering signal and the side scattering signal to obtain the second signal area. Specifically, similar to step S2031, when forming a two-dimensional signal distribution diagram based on the forward scattering signal and the side scattering signal, the forward scattering signal can be used as the horizontal coordinate and the side scattering signal as the vertical coordinate, and then the specific signal value is marked in the coordinate system to obtain a two-dimensional signal distribution diagram composed of the forward scattering signal and the side scattering signal. The two-dimensional signal distribution diagram is also called a two-dimensional signal FSC-SSC, that is, a two-dimensional signal distribution diagram of the FSC-SSC dimension. In this embodiment, the obtained two-dimensional signal distribution diagram is as follows: Figure 3B As shown in the figure, during filtering, a first signal region is selected. Then, signals exceeding the forward scatter signal threshold and the side scatter signal threshold in the first signal region are deleted, resulting in a second signal region. This second signal region is the optimized region of the first signal region. This filtering process can remove discrete signals.

[0055] Step a3, threshold filtering is performed on the project group in the second signal area according to the two-dimensional signal distribution diagram composed of the first fluorescent signal and the second fluorescent signal. Specifically, when the two-dimensional signal distribution diagram is composed of the first fluorescent signal and the second fluorescent signal, the first fluorescent signal can be used as the horizontal coordinate and the second fluorescent signal as the vertical coordinate, and then the specific signal value is marked in the coordinate system to obtain the two-dimensional signal distribution diagram composed of the first fluorescent signal and the second fluorescent signal. The two-dimensional signal distribution diagram is also called the two-dimensional signal CFL1-CFL2, that is, the two-dimensional signal distribution diagram of the CFL1-CFL2 dimension. In this embodiment, the obtained two-dimensional signal distribution diagram is as follows Figure 3C As shown in the figure, during filtering, the second signal region is first selected, and then signals within the second signal region that exceed the first and second fluorescence signal thresholds are removed. After this filtering process, the signal concentration areas in the figure are more obvious, indicating that multiple project clusters can be distinguished. This step allows the identification of project clusters while removing background signals and discrete signals.

[0056] Step S2032: Process the item clusters based on the comparison result between the number of item clusters and the number of test items to obtain valid item clusters equal to the number of test items. Specifically, this step includes determining the item area of ​​each test item in the two-dimensional signal distribution map based on the test items; when the number of item clusters is less than the number of test items, completing the item clusters based on the item areas corresponding to the missing item clusters to obtain valid item clusters equal to the number of test items; when the number of item clusters is greater than the number of test items, deleting the redundant item clusters based on the item areas corresponding to each item cluster to obtain valid item clusters equal to the number of test items.

[0057] like Figure 4 As shown, after differentiating and obtaining multiple project groups, it is necessary to determine whether the number of multiple project groups is the same as the number of test items. When the number of project groups is less than the number of test items, the area where each project group is located is matched with the project area in turn, and then the project area that is not matched is screened out. This area is the project area corresponding to the missing project group, and this area is used as the missing project group to fill in the missing project group, and the valid project group with the same number as the test items can be obtained; when the number of project groups is greater than the number of test items, the area where each project group is located is also matched with the project area in turn. If there are at least two project groups in a project area, the project group with the smaller area in the area is deleted, and only the project group with the largest area is retained, thereby ensuring that there is only one project group in each preset area, and finally obtaining the valid project group with the same number as the test items.

[0058] It should be noted that the project area refers to a closed area that is pre-set according to the detection project and corresponds to each detection project. The area is specifically composed of a square structure determined by two first fluorescent signal horizontal coordinates and two second fluorescent signal vertical coordinates. Thus, after determining the detection project, the project area corresponding to each project can be defined by pre-setting the values ​​of two horizontal coordinates and two vertical coordinates according to the detection data corresponding to each detection project and the retrieval project, such as historical detection data. When the area where each project group is located is matched with the project area, it can be determined whether the area where each project group is located is within the project area. When the area where the project group is located falls within the project area, the project group corresponds to the corresponding project area.

[0059] Furthermore, the steps for filtering project groups provided in this embodiment are primarily for the dual-fluorescence mode. That is, when using the dual-fluorescence mode, the identification method described in the steps above can be used, or other identification methods can be used. When using the single-fluorescence mode, the identification method can be determined based on the actual situation, such as identification based on a two-dimensional signal composed of forward scatter and side scatter signals, or based on a two-dimensional signal composed of forward scatter and fluorescence signals, etc.

[0060] Step S204: When the detection mode is the dual fluorescence mode, the coordinates of the valid item group are processed according to the preset weights; see Figure 1 Step S103 of the illustrated embodiment will not be described in detail here.

[0061] Step S205: sort the valid item groups based on the processed coordinates to obtain the item numbers corresponding to the valid item groups. The item numbers correspond to the test items. Figure 1 Step S104 of the illustrated embodiment will not be described in detail here.

[0062] Step S206: Output the test results of each test item of the sample based on the valid project group and the corresponding project number. Figure 1 Step S105 of the illustrated embodiment will not be described in detail here.

[0063] In this embodiment, a method based on flow cytometry is provided, which includes the following steps:

[0064] Step S301, obtaining the detection information of the sample and the light detection signal obtained by detecting the microsphere complex prepared from the sample based on the detection information; wherein the light detection signal includes multiple light signals, and the detection information includes the detection mode and the detection items; for details, please refer to Figure 1 Step S101 of the illustrated embodiment will not be described in detail here.

[0065] Step S302, determine the particle size of the microspheres according to the detection information; determine the effective area in the two-dimensional signal distribution diagram composed of the forward scattering signal and the side scattering signal according to the particle size, and the effective area is the range for identifying the project group. Specifically, the particle size of the microspheres in the detection information can be the diameter of the microspheres. After determining the particle size of the microspheres, the forward scattering signal is used as the horizontal coordinate and the side scattering signal is used as the vertical coordinate, and then the specific signal value is marked in the coordinate system to obtain a two-dimensional signal distribution diagram composed of the forward scattering signal and the side scattering signal. Then, circle the effective area corresponding to the particle size of the microspheres in the two-dimensional signal distribution diagram. It should be noted that since the forward scattering signal is used to characterize the size of each microsphere in the test sample, marking the effective area in the two-dimensional signal distribution diagram composed of the forward scattering signal and the side scattering signal can obtain a more accurate and stable effective area. Subsequent identification of the project group can be performed in this effective area.

[0066] Step S303: Identify the two-dimensional signal distribution diagram composed of at least two optical signals to obtain a valid item group with the same number as the detection items; see Figure 1 Step S102 of the illustrated embodiment will not be described in detail here.

[0067] Step S304: When the detection mode is dual fluorescence mode, the coordinates of the valid item cluster are processed according to preset weights. Specifically, when the detection mode is dual fluorescence mode, the coordinates of the valid item cluster are processed according to preset weights based on the two-dimensional signal distribution diagram composed of the first and second fluorescence signals and the range of the fluorescent signal that can be generated by the microspheres. The range of the fluorescent signal that can be generated by the microspheres can be extracted from the detection information.

[0068] In this embodiment, when using dual-fluorescence mode, valid signal regions are identified from a two-dimensional signal distribution diagram (CFL1-CFL2) constructed based on the first and second fluorescent signals. After the two-dimensional signal distribution diagram is constructed, regions outside the range of fluorescent signals generated by the microspheres are deleted, and the remaining signal-concentrated regions are treated as valid signal regions for effective project group processing. Alternatively, a two-dimensional signal distribution diagram can be directly constructed based on the first and second fluorescent signals within the range of fluorescent signals generated by the microspheres, with the signal-concentrated regions within this two-dimensional signal distribution diagram being the valid signal regions.

[0069] Step S305: sort the valid item groups based on the processed coordinates to obtain the item numbers corresponding to the valid item groups. The item numbers correspond to the test items. Figure 1 Step S104 of the illustrated embodiment will not be described in detail here.

[0070] Step S306, when the detection mode is single fluorescence mode, based on the two-dimensional signal distribution diagram composed of the forward scattering signal and the fluorescence signal and the range of the fluorescence signal generated by the microspheres, the valid project groups are sorted according to the horizontal and vertical coordinates of the valid project groups, and the sorted valid project groups correspond to the project numbers.

[0071] In this embodiment, when using a single fluorescence mode, valid signal regions are identified from a two-dimensional signal distribution diagram (CFL1-CFL2) constructed based on the first and second fluorescence signals. After the two-dimensional signal distribution diagram is constructed, regions outside the range of fluorescence signals generated by the microspheres are deleted, and the remaining signal-concentrated regions are treated as valid signal regions for effective project group processing. Alternatively, a two-dimensional signal distribution diagram can be directly constructed based on the first and second fluorescence signals within the range of fluorescence signals generated by the microspheres, with the signal-concentrated regions within this two-dimensional signal distribution diagram being the valid signal regions.

[0072] Specifically, if Figure 5As shown, when the detection mode is a single fluorescence mode, the arrangement of the effective project groups in the effective signal area presents a regular arrangement, or in other words, it presents a grid-like horizontal and vertical arrangement. At this time, the effective project groups can be directly sorted according to the horizontal and vertical coordinates of the effective project groups. For example, if it is necessary to prioritize arrangement in the Y direction, the sizes of the horizontal coordinates of all the effective project groups can be compared first, and the effective project groups can be arranged. Then, for the effective project groups with the same horizontal coordinates, the sizes of their vertical coordinates can be compared, and the effective project groups with the same horizontal coordinates can be arranged based on the vertical coordinates, thereby achieving the sorting of the effective project groups. If it is necessary to prioritize arrangement in the X direction, the sizes of the vertical coordinates of all the effective project groups can be compared first, and the effective project groups can be arranged. Then, for the effective project groups with the same vertical coordinates, the sizes of their horizontal coordinates can be compared, and the effective project groups with the same vertical coordinates can be arranged based on the horizontal coordinates, thereby achieving the sorting of the effective project groups. Whether to prioritize arrangement in the X direction or arrangement in the Y direction depends on the pre-set item number and item group correspondence method. Finally, the sorted effective project groups and item numbers are matched one by one.

[0073] Step S307: output the test result of each test item of the sample based on the valid item group and the corresponding item number.

[0074] Specifically, the above step S307 includes:

[0075] Step S3071: Obtain valid project groups and corresponding quantitative fluorescence values.

[0076] Step S3072: Determine the concentration based on the preset calibration curve and the quantitative fluorescence value, and output the concentration as the test result of the test item corresponding to each item number.

[0077] Specifically, after determining the valid project group and the corresponding project number, the corresponding quantitative fluorescence signal, i.e., the quantitative fluorescence value, can be obtained based on the microspheres corresponding to each valid project group. Then, based on a calibration curve mapping the quantitative fluorescence value and microsphere concentration, the microsphere concentration corresponding to the quantitative fluorescence value of each valid project group can be determined. Then, based on the project number corresponding to the valid project, the microsphere concentration is output as the test result for the test item corresponding to the project number. The calibration curve is a pre-set mapping curve between existing quantitative fluorescence values ​​and corresponding concentrations.

[0078] This embodiment also provides a sample analyzer for implementing the aforementioned embodiments and preferred implementations. Details already described will not be repeated. As used below, the term "module" may refer to a combination of software and / or hardware that implements a predetermined function. While the sample analyzer described in the following embodiments is preferably implemented using software, implementation using hardware, or a combination of software and hardware, is also possible and contemplated.

[0079] This embodiment provides a sample analyzer, such as Figure 6 Shown, including:

[0080] Detection module 61, for obtaining detection information of the sample and detecting a microsphere complex prepared from the sample based on the detection information to obtain an optical detection signal; wherein the optical detection signal includes multiple optical signals, and the detection information includes a detection mode and a detection item;

[0081] The control module 62 is used to identify based on a two-dimensional signal distribution diagram composed of at least two optical signals to obtain valid item groups with the same number as the detection items; when the detection mode is a dual fluorescence mode, the coordinates of the valid item groups are processed according to preset weights; the valid item groups are sorted based on the processed coordinates to obtain the item numbers corresponding to the valid item groups, and the item numbers correspond to the detection items; and the detection results of each detection item of the sample are output based on the valid item groups and the corresponding item numbers.

[0082] In an optional embodiment, the at least two optical signals include a first fluorescent signal and a second fluorescent signal, each of which is generated by irradiating the microsphere complex with the same laser and resulting in scattered light. Specifically, when the same laser is used to excite different fluorescent dyes, such as two fluorescent dyes, the optical signals (the first fluorescent signal and the second fluorescent signal) generate more severe crosstalk. Therefore, the aforementioned coordinate processing scheme for valid item groups can reduce costs (i.e., using the same laser to excite two classified fluorescent dyes) while improving accuracy.

[0083] The further functional description of each of the above modules is the same as that of the above corresponding embodiments and will not be repeated here.

[0084] The embodiment of the present invention also provides a computer device having the above Figure 6 Sample analyzer shown.

[0085] See also Figure 7 , Figure 7 is a structural diagram of a computer device provided by an optional embodiment of the present invention, such as Figure 7As shown, the computer device includes: one or more processors 10, memory 20, and interfaces for connecting various components, including high-speed interfaces and low-speed interfaces. Various components utilize different buses to communicate with each other and can be installed on a common mainboard or installed in other ways as needed. The processor can process the instructions executed in the computer device, including instructions stored in the memory or on the memory to display the graphical information of the GUI on an external input / output device (such as, a display device coupled to the interface). In some optional embodiments, if necessary, multiple processors and / or multiple buses can be used together with multiple memories and multiple memories. Equally, multiple computer devices can be connected, and each device provides part of the necessary operations (for example, as a server array, a group of blade servers, or a multi-processor system). Figure 7 A processor 10 is taken as an example.

[0086] The processor 10 may be a central processing unit, a network processor, or a combination thereof. The processor 10 may further include a hardware chip. The hardware chip may be an application-specific integrated circuit, a programmable logic device, or a combination thereof. The programmable logic device may be a complex programmable logic device, a field programmable gate array, a general purpose array logic, or any combination thereof.

[0087] The memory 20 stores instructions that can be executed by at least one processor 10, so as to enable at least one processor 10 to execute the method shown in the above embodiment.

[0088] The memory 20 may include a program storage area and a data storage area, wherein the program storage area may store an operating system, an application required for at least one function; the data storage area may store data created based on the use of a computer device for displaying a small program landing page, etc. In addition, the memory 20 may include a high-speed random access memory, and may also include a non-transient memory, such as at least one disk storage device, a flash memory device, or other non-transient solid-state storage device. In some optional embodiments, the memory 20 may optionally include a memory remotely located relative to the processor 10, and these remote memories may be connected to the computer device via a network. Examples of the above-mentioned network include, but are not limited to, the Internet, an intranet, a local area network, a mobile communication network, and a combination thereof.

[0089] The memory 20 may include a volatile memory, such as a random access memory; the memory may also include a non-volatile memory, such as a flash memory, a hard disk or a solid-state drive; the memory 20 may also include a combination of the above types of memory.

[0090] The computer device further includes a communication interface 30 for the computer device to communicate with other devices or a communication network.

[0091] The embodiment of the present invention also provides a computer-readable storage medium. The above-mentioned method according to the embodiment of the present invention can be implemented in hardware, firmware, or implemented as a computer code that can be recorded in a storage medium, or implemented as a computer code that is originally stored in a remote storage medium or a non-temporary machine-readable storage medium and downloaded through a network and will be stored in a local storage medium, so that the method described herein can be stored in such software processing on a storage medium using a general-purpose computer, a dedicated processor, or programmable or dedicated hardware. Among them, the storage medium can be a magnetic disk, an optical disk, a read-only storage memory, a random access memory, a flash memory, a hard disk or a solid-state drive, etc.; further, the storage medium can also include a combination of the above-mentioned types of memory. It can be understood that a computer, a processor, a microprocessor controller or programmable hardware includes a storage component that can store or receive software or computer code. When the software or computer code is accessed and executed by a computer, a processor or hardware, the method shown in the above embodiment is implemented.

[0092] A portion of the present invention may be applied as a computer program product, such as a computer program instruction, which, when executed by a computer, can call or provide the method and / or technical solution according to the present invention through the operation of the computer. Those skilled in the art should understand that the form in which the computer program instruction exists in a computer-readable medium includes, but is not limited to, a source file, an executable file, an installation package file, etc. Accordingly, the way in which the computer program instruction is executed by the computer includes, but is not limited to: the computer directly executes the instruction, or the computer compiles the instruction and then executes the corresponding compiled program, or the computer reads and executes the instruction, or the computer reads and installs the instruction and then executes the corresponding installed program. Here, the computer-readable medium may be any available computer-readable storage medium or communication medium that can be accessed by the computer.

[0093] Although the embodiments of the present invention have been described with reference to the accompanying drawings, those skilled in the art may make various modifications and variations without departing from the spirit and scope of the present invention. Such modifications and variations are all within the scope defined by the appended claims.

Claims

1. A method based on flow fluorescence, characterized in that, The method comprises: Acquiring detection information of the sample and an optical detection signal obtained by detecting a microsphere complex prepared from the sample based on the detection information; wherein the optical detection signal includes multiple optical signals, and the detection information includes a detection mode and a detection item; Performing identification based on a two-dimensional signal distribution diagram formed by at least two of the optical signals to obtain a valid item group having the same number as the detection items; When the detection mode is a dual fluorescence mode, the coordinates of the valid item group are processed according to a preset weight; sorting the valid item groups based on the processed coordinates to obtain item numbers corresponding to the valid item groups, where the item numbers correspond to the detection items; Output the test result of each test item of the sample based on the valid item group and the corresponding item number.

2. The method according to claim 1, characterized in that Identification is performed based on a two-dimensional signal distribution diagram formed by at least two of the optical signals to obtain a valid item group having the same number as the number of detection items, including: performing circle identification based on a two-dimensional signal distribution diagram formed by at least two of the optical signals to obtain a plurality of item groups; The item group is processed based on a comparison result between the number of the item group and the number of the detection items to obtain a valid item group having the same number as the detection items.

3. The method according to claim 2, characterized in that The item group is processed based on a comparison result between the number of the item group and the number of the test items to obtain a valid item group having the same number as the test items, including: Determine, according to the detection items, a project area of ​​each detection item in a two-dimensional signal distribution map; When the number of the project groups is less than the number of the test items, the project groups are supplemented according to the project areas corresponding to the missing project groups to obtain valid project groups with the same number as the test items; When the number of project groups is greater than the number of the detection items, the redundant project groups are deleted according to the project areas corresponding to each project group to obtain valid project groups with the same number as the detection items.

4. The method according to claim 1, wherein The detection information also includes the range of fluorescent signals that the microspheres can generate, and the optical signals include a first fluorescent signal and a second fluorescent signal; the first fluorescent signal and the second fluorescent signal are used to distinguish different detection items based on different signal intensities; When the detection mode is the dual fluorescence mode, processing the coordinates of the valid item group according to a preset weight includes: When the detection mode is a dual fluorescence mode, based on the two-dimensional signal distribution diagram composed of the first fluorescence signal and the second fluorescence signal and the fluorescence signal range generated by the microspheres, the coordinates of the valid item group are processed according to preset weights.

5. The method according to claim 1, wherein The detection information also includes a range of fluorescent signals that can be generated by the microspheres, and the optical signal includes a forward scattering signal and a fluorescent signal; the method further includes: When the detection mode is a single fluorescence mode, a two-dimensional signal distribution diagram and microspheres composed of the forward scattering signal and the fluorescence signal can generate a fluorescence signal range, and the valid item groups are sorted according to the horizontal and vertical coordinates of the valid item groups, and the sorted valid item groups correspond to the item numbers.

6. The method according to claim 1, characterized in that Before performing identification based on the two-dimensional signal distribution diagram formed by at least two optical signals, the method further includes: Cutting and filtering at least one optical signal among the plurality of optical signals based on a signal threshold; Based on the filtered optical signal, corresponding other optical signals are filtered.

7. The method according to claim 1, characterized in that The optical signal includes a forward scattered signal and a side scattered signal. Before performing identification based on a two-dimensional signal distribution diagram formed by at least two of the optical signals, the method further includes: determining the particle size of the microspheres according to the detection information; An effective area is determined in a two-dimensional signal distribution diagram composed of the forward scattering signal and the side scattering signal according to the particle size, and the effective area is a range for identifying the project group.

8. The method according to claim 1, characterized in that Outputting the test result of each test item of the sample based on the valid item group and the corresponding item number includes: Obtain valid project groups and corresponding quantitative fluorescence values; The concentration is determined according to a preset calibration curve and a quantitative fluorescence value, and the concentration is output as a detection result of the detection item corresponding to each item number.

9. A sample analyzer, characterized in that: The sample analyzer comprises: A detection module, configured to obtain detection information of a sample and an optical detection signal obtained by detecting a microsphere complex prepared from the sample based on the detection information; wherein the optical detection signal includes multiple optical signals, and the detection information includes a detection mode and a detection item; A control module is configured to identify, based on a two-dimensional signal distribution diagram composed of at least two optical signals, valid item clusters equal in number to the number of detection items; when the detection mode is a dual fluorescence mode, process the coordinates of the valid item clusters according to preset weights; sort the valid item clusters based on the processed coordinates to obtain item numbers corresponding to the valid item clusters, wherein the item numbers correspond to the detection items; and output the detection results of each detection item of the sample based on the valid item clusters and the corresponding item numbers.

10. The sample analyzer according to claim 9, characterized in that: The at least two optical signals include a first fluorescent signal and a second fluorescent signal, wherein the first fluorescent signal and the second fluorescent signal are optical signals scattered by the microsphere complex when irradiated by the same laser.

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