Method, equipment and program product for detecting complement system reaction degree based on endothelial cells

Abstract

The invention relates to the field of intelligent medical treatment, in particular to a method, equipment and a program product for detecting the reaction degree of a complement system based on endothelial cells. Comprising the following steps: S1, acquiring serum of a detected object; s2, acquiring human endothelial cells, adding serum of a detected object and the human endothelial cells into the human endothelial cell culture medium, and culturing to obtain cultured endothelial cells; s3, adding a primary antibody into the cultured endothelial cells for incubation to obtain a primary antibody incubation product, wherein the primary antibody is a primary antibody of protein related to complement reaction on the endothelial cells; s4, adding a secondary antibody into the primary antibody incubation product for incubation to obtain a secondary antibody incubation product; and S5, detecting the secondary antibody incubation product through a flow cytometer or an electron microscope to obtain the fluorescence intensity, and obtaining the complement reaction level based on the fluorescence intensity. The method can reflect the complement activation degree in the tissue, is used for complement reaction detection, and has good clinical value.

Description

Technical Field

[0001] The present application relates to the field of intelligent medical treatment, and specifically to a method, device, program product and computer storage medium for detecting the degree of complement system reaction based on endothelial cells. Background Art

[0002] The complement reaction is an enzyme cascade reaction, which is a collection of blood and cell surface proteins. The complement system is composed of 30 different proteins, including serum proteins, plasma membrane proteins and cell membrane receptors, which play an important role in the innate immune system. The complement system has four major functions, including lysis of infectious organisms, activation of inflammation, conditioning and immune clearance. Complement is believed to play a major role in innate immunity (nonspecific immunity, innate immune system), that is, to produce a rapid and powerful response to invading pathogens. However, recent studies have shown that complement also plays an important role in adaptive immunity (adaptive immune systems) involving T cells and B cells. The complement system is not only involved in innate immunity and adaptive immunity, but is also related to tissue regeneration, tumor growth, etc. Among them, in the complement activation pathway: C5b can stably bind to C6 to form C5b6, which spontaneously binds to C7 to form C5b-7. C7 in the complex is initially inserted into the lipid bilayer of the target cell membrane, and then C8 binds with high affinity to C5b-7 on the inserted membrane to form a stable C5b-8 that is deeply inserted into the cell membrane. This complex can bind to C9 molecules to form C5b-9, which is the membrane attack complex (MAC). It can directly attack the cell membrane, causing cell disintegration and thus damage. At the same time, inflammatory mediators such as C3a and C5a can be released on the complement pathway to mediate inflammation. Excessive activation or imbalance of complement can lead to tissue damage and inflammation. If excessive complement activation occurs in the circulation, it can attack red blood cells, and the patient may show hypocomplementemia, manifested as paroxysmal nocturnal hemoglobinuria; if excessive complement activation occurs locally in the tissue, such as complement end products attacking endothelial cells, it will lead to corresponding organ damage, such as thrombotic microangiopathy (TMA). If the main target organ is the kidney, it is diagnosed as thrombotic microangiopathy renal injury, of which atypical hemolytic uremic syndrome is the most serious type, and also includes malignant hypertension, pregnancy-related hemolytic uremic syndrome, transplant-related TMA, lupus-mediated TMA, scleroderma-mediated TMA, antiphospholipid syndrome-mediated TMA, etc. Summary of the invention

[0003] In view of the above problems, when diagnosing complement-related diseases and looking for indications for medication, the degree of local complement activation should be targeted rather than the entire complement system. Therefore, the present invention proposes a method for detecting the degree of complement system reaction based on endothelial cells, comprising:

[0004] S1: obtaining serum from the test subject;

[0005] S2: obtaining human endothelial cells and adding the serum of the test subject and the human endothelial cells to a human endothelial cell culture medium to obtain cultured endothelial cells;

[0006] S3: adding a primary antibody to the cultured endothelial cells for incubation to obtain a primary antibody incubation product, wherein the primary antibody is a primary antibody against a protein on the endothelial cells that is associated with complement response;

[0007] S4: adding a secondary antibody to the primary antibody incubation product for incubation to obtain a secondary antibody incubation product;

[0008] S5: The secondary antibody incubation product is detected by flow cytometry or electron microscopy to obtain fluorescence intensity, and the complement reaction level is obtained based on the fluorescence intensity.

[0009] The endothelial cells include one or more of the following: human glomerular endothelial cells, human umbilical vein endothelial cells, and human dermal vascular endothelial cells.

[0010] The primary antibody against the complement reaction-related protein includes one or more of the following: C5b-9, C5b-8, C3, C4d, C5a, and the secondary antibody is selected from the secondary antibody that binds to the primary antibody and whose signal is detected by a flow cytometer, including one or more of the following: AF647-IgG, allophycocyanin-IgG, and Cy5-IgG.

[0011] The S2 further comprises culturing the endothelial cells for 4-8 generations.

[0012] The S2 further comprises adding adenosine diphosphate to the culture medium of human endothelial cells.

[0013] The object of the present invention is to provide a system for detecting the degree of complement system reaction based on endothelial cells, comprising:

[0014] Acquisition unit: acquiring serum of a test subject;

[0015] Extraction unit: obtaining human endothelial cells and adding the serum and human endothelial cells of the test subject to human endothelial cell culture medium to obtain cultured endothelial cells;

[0016] A primary incubation unit: adding a primary antibody to the cultured endothelial cells for incubation to obtain a primary antibody incubation product, wherein the primary antibody is a primary antibody against a protein on the endothelial cells that is associated with a complement reaction;

[0017] Secondary incubation unit: adding a secondary antibody to the primary antibody incubation product for incubation to obtain a secondary antibody incubation product;

[0018] Detection unit: The secondary antibody incubation product is detected by flow cytometry or electron microscopy to obtain fluorescence intensity, and the complement reaction level is obtained based on the fluorescence intensity.

[0019] The object of the present invention is to provide a computer program product, which includes a computer program or instructions, and the computer program or instructions are executed by a processor to implement the above-mentioned method for detecting the degree of complement system response based on endothelial cells.

[0020] The object of the present invention is to provide a computer device, comprising:

[0021] A memory and a processor, wherein the memory is used to store program instructions; the processor is used to call the program instructions, and when the program instructions are executed, the above-mentioned method for detecting the degree of complement system reaction based on endothelial cells is implemented.

[0022] The object of the present invention is to provide a computer-readable storage medium having a computer program thereon, comprising:

[0023] When the computer program is executed by a processor, the above-mentioned method for detecting the degree of complement system reaction based on endothelial cells is implemented.

[0024] The object of the present invention is to provide an MDT decision-making method, which implements the above-mentioned method for detecting the degree of complement system response based on endothelial cells to obtain the complement response level in endothelial cells. When it is determined that the complement response level of the patient is abnormal, a drug recommended to inhibit complement activation is added to the medication regimen for the patient.

[0025] The drug that inhibits complement activation is a drug that inhibits complement pathway activation when the activation level is greater than the normal level.

[0026] Furthermore, when the subject is determined to have an abnormal complement reaction level of C5b-9 or C5b-8, it is recommended to administer a complement inhibitor to the subject, wherein the complement inhibitor includes one or more of the following: C5 monoclonal antibody, factor B inhibitor, C3 inhibitor, factor D inhibitor.

[0027] The C5 monoclonal antibodies include eculiumab, crovalimab, PiaSky, and Ravulizumab; the factor B inhibitors include iptacopan and NTQ5082; the C3 inhibitors include Pegcetagoplan and empaveli; and the factor D inhibitors include Danicopan.

[0028] An object of the present invention is to provide a computer program product, which includes a computer program or instructions, and the computer program or instructions are executed by a processor to implement the above-mentioned MDT decision method.

[0029] The object of the present invention is to provide an MDT decision device, comprising:

[0030] A memory and a processor, wherein the memory is used to store program instructions; the processor is used to call the program instructions, and when the program instructions are executed, the above-mentioned MDT decision method is implemented.

[0031] The object of the present invention is to provide a computer-readable storage medium having a computer program thereon, comprising:

[0032] When the computer program is executed by a processor, the above-mentioned MDT decision method is implemented.

[0033] Advantages of the present invention:

[0034] 1. A non-invasive in vitro method for detecting tissue complement response is proposed. This method can intuitively and accurately detect whether complement activation occurs locally in the patient's tissue, and thus provide targeted treatment to the patient in a timely manner.

[0035] 2. Propose an MDT decision-making method to classify patients based on complement reactions and provide medication guidance to patients based on the classification results. During the use of drugs, the complement activation reaction is repeatedly measured and the dosage of the drugs is adjusted to avoid overdose or underdose.

[0036] 3. Complement reaction testing for patients suspected of TMA renal injury can accurately determine whether the patient has thrombotic microvascular vascular lesions, avoid renal puncture, reduce the difficulty of the examination, and reduce the patient's pain. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the drawings required for use in the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative work.

[0038] Figure 1 A schematic flow chart of a method for detecting the degree of complement system reaction based on endothelial cells provided in an embodiment of the present invention;

[0039] Figure 2 A schematic diagram of a system for detecting the degree of complement system reaction based on endothelial cells provided in an embodiment of the present invention;

[0040] Figure 3 A schematic diagram of a device for detecting the degree of complement system reaction based on endothelial cells provided in an embodiment of the present invention;

[0041] Figure 4 The complement reaction flow cytometry results of endothelial cell detection provided in the embodiment of the present invention;

[0042] Figure 5 The fluorescence results of the complement reaction detected by the electron microscope of the endothelial cells provided in the embodiment of the present invention are shown in red. The secondary antibody AF476-IgG shows the fluorescence intensity under the CY5 channel of the microscope. The higher the complement activation, the stronger the fluorescence. After using the complement inhibitor, the complement reaction stimulated by the serum in the patient's body was completely inhibited. DETAILED DESCRIPTION

[0043] In order to enable those skilled in the art to better understand the solutions of the present invention, 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.

[0044] In some of the processes described in the specification and claims of the present invention and the above-mentioned figures, multiple operations that appear in a specific order are included, but it should be clearly understood that these operations may not be executed in the order in which they appear in this article or executed in parallel. The sequence numbers of the operations, such as S101, S102, etc., are only used to distinguish between different operations, and the sequence numbers themselves do not represent any execution order. In addition, these processes may include more or fewer operations, and these operations may be executed in sequence or in parallel. It should be noted that the descriptions of "first", "second", etc. in this article are used to distinguish different messages, devices, modules, etc., do not represent the order of precedence, and do not limit the "first" and "second" to be different types.

[0045] Figure 1 A schematic diagram of a method for detecting the degree of complement system reaction based on endothelial cells provided in an embodiment of the present invention specifically includes:

[0046] S101: obtaining serum from a test subject;

[0047] In one embodiment, human endothelial cells are cultured using a cell culture medium, which mainly consists of a basic culture fluid, additives and growth factors.

[0048] The basal culture medium is the most important component of the cell culture medium. It is composed of water, amino acids, sugars, vitamins and minerals, etc. It provides cells with sufficient nutrients and growth conditions to maintain cell survival and proliferation. Common basal culture media include DMEM, MEM, RPMI-1640, etc.

[0049] Additives mainly include fetal bovine serum (FBS), human serum, sheep serum, chicken embryo serum, horse serum, soy protein hydrolyzate, etc. These additives provide many substances and growth factors required by cells to help cell growth and proliferation. FBS is one of the most commonly used additives, especially suitable for culturing tumor cells and other difficult-to-culture cells. The addition of FBS can enhance cell adhesion, proliferation rate and inhibit apoptosis.

[0050] Growth factors are important components in cell culture media. They promote cell proliferation, differentiation and function. Growth factors include EGF, FGF, NGF, PDGF, etc., which regulate cell proliferation and differentiation by activating corresponding receptors. Growth factors have different effects in different cell types, so different cell types require different growth factors.

[0051] S102: obtaining human endothelial cells and adding the serum of the test subject and the human endothelial cells to a human endothelial cell culture medium to culture the cultured endothelial cells;

[0052] In one embodiment, the endothelial cells include one or more of the following: human glomerular endothelial cells (HMEC-1), human umbilical vein endothelial cells (HUVEC), human dermal vascular endothelial cells (HDMEC).

[0053] In one embodiment, the S102 further comprises culturing the endothelial cells for 4-8 generations.

[0054] In one embodiment, the S102 further comprises adding adenosine diphosphate (ADP) to the culture medium of human endothelial cells.

[0055] In one embodiment, thrombotic microangiopathy (TMA) is characterized by microangiopathic hemolytic anemia, thrombocytopenia, and organ involvement due to platelet thrombi in the microcirculation.

[0056] S103: adding a primary antibody to the cultured endothelial cells for incubation to obtain a primary antibody incubation product, wherein the primary antibody is a primary antibody against a protein on the endothelial cells that is associated with a complement reaction;

[0057] In one embodiment, the primary antibody against complement response-related proteins includes one or more of the following: C5b-9 (also known as membrane attack complex, MAC), C5b-8, C3, C4d, and C5a.

[0058] S104: adding a secondary antibody to the primary antibody incubation product for incubation to obtain a secondary antibody incubation product;

[0059] In one embodiment, a flow cytometer is a device for automatically analyzing and sorting cells. It can quickly measure, store, and display a series of important biophysical and biochemical characteristic parameters of dispersed cells suspended in a liquid, and can sort out specified cell subpopulations according to a preselected parameter range. Most flow cytometers are zero-resolution instruments that can only measure indicators such as total nucleic acid content and total protein content of a cell. In actual use, the instrument must first measure the light scattering signal. When light scattering analysis is used in conjunction with fluorescent probes, stained and unstained cells in the sample can be identified. The most effective use of light scattering measurement is to identify certain subpopulations from a non-uniform population.

[0060] Electron microscope to observe fluorescence reaction:

[0061] Fluorescence signals mainly include two parts: ① spontaneous fluorescence, that is, the fluorescence emitted by the fluorescent molecules inside the cell after being irradiated with light without fluorescent staining; ② characteristic fluorescence, that is, the fluorescence emitted by the fluorescent dyes bound to the cells after being stained and exposed to light. Its fluorescence intensity is weaker and its wavelength is different from that of the irradiating laser.

[0062] In one embodiment, the maximum emission wavelength of allophycocyanin (APC) is 660 nm, and the antibody labeled with it is suitable for all flow cytometers equipped with helium-neon lasers, and the detection channel is generally the FL4 channel. FITC, PE and APC are a classic three-color combination.

[0063] In one embodiment, Cy5 is a small molecule dye with a maximum emission wavelength of 670 nm. The antibody labeled with Cy5 is suitable for all flow cytometers equipped with a 633 nm argon ion laser, and the detection channel is generally the FL4 channel.

[0064] In one embodiment, AF647, Alexa Fluor 647 is a bright red fluorescent dye with a maximum emission wavelength of 668 nm, which is generally detected in the FL4 channel of a flow cytometer. Alexa Fluor 647 has the characteristics of high light quantum yield, good photostability, wide pH range of adaptability and not easy to quench.

[0065] In one embodiment, the secondary antibody against the complement reaction-related protein includes one or more of the following: the secondary antibody is selected from the secondary antibody that binds to the primary antibody and whose signal is detected by a flow cytometer, including one or more of the following: AF647-IgG, allophycocyanin-IgG, Cy5-IgG.

[0066] Secondary antibodies that can be detected by flow cytometry can be detected by electron microscopy.

[0067] S105: The secondary antibody incubation product is detected by flow cytometry or electron microscopy to obtain fluorescence intensity, and the complement reaction level is obtained based on the fluorescence intensity.

[0068] In a specific embodiment, the reaction degree of the complement system of the subject is detected by the technical solution of the present invention, and the result obtained by flow cytometry is as follows: Figure 4 As shown in Figure a, endothelial cells stimulated with serum from complement activated patients have significant C56-9 deposition on endothelial cells. Compared with the isotype control, more endothelial cells have stronger AF-647 channel fluorescence intensity, which is the formation of C5b-9 specifically bound to the C5b-9 antibody. In Figure b, glomerular endothelial cells stimulated with different TMA patient sera can lead to different degrees of C5b-9 formation on cells.

[0069] The fluorescence results of different patients, empty bag controls, and patients after administration of complement inhibitors are shown in Figure 2. Figure 5 As shown, it can be found that patients treated with complement inhibitors show significant differences compared to untreated patients. The method of the present invention can detect the degree of complement response in the test subject based on endothelial cells, which helps to assist doctors in disease diagnosis and treatment, and further recommends treatment plans and medications to improve medical diagnosis efficiency.

[0070] Figure 2 The schematic diagram of the system for detecting the degree of complement system reaction based on endothelial cells provided in the embodiment of the present invention specifically includes:

[0071] Acquisition unit: acquiring serum of a test subject;

[0072] Extraction unit: obtaining human endothelial cells and adding the serum and human endothelial cells of the test subject to human endothelial cell culture medium to obtain cultured endothelial cells;

[0073] A primary incubation unit: adding a primary antibody to the cultured endothelial cells for incubation to obtain a primary antibody incubation product, wherein the primary antibody is a primary antibody against a protein on the endothelial cells that is associated with a complement reaction;

[0074] Secondary incubation unit: adding a secondary antibody to the primary antibody incubation product for incubation to obtain a secondary antibody incubation product;

[0075] Detection unit: The secondary antibody incubation product is detected by flow cytometry or electron microscopy to obtain fluorescence intensity, and the complement reaction level is obtained based on the fluorescence intensity.

[0076] Figure 3 A schematic diagram of a device for detecting the degree of complement system reaction based on endothelial cells provided in an embodiment of the present invention specifically includes:

[0077] A memory and a processor; the memory is used to store program instructions; the processor is used to call the program instructions, and when the program instructions are executed, any one of the above-mentioned methods for detecting the degree of complement system response based on endothelial cells is executed.

[0078] A computer-readable storage medium stores a computer program, wherein the computer program, when executed by a processor, is any one of the above-mentioned methods for detecting the degree of complement system response based on endothelial cells.

[0079] An MDT decision-making method, comprising:

[0080] The method implements the above-mentioned method for detecting the degree of complement system reaction based on endothelial cells to obtain the complement reaction level in endothelial cells. When it is determined that the complement reaction level of the subject is abnormal, the patient is given a drug recommended to be added to the medication regimen for inhibiting complement activation.

[0081] The drug that inhibits complement activation is a drug that inhibits complement pathway activation when the activation level is greater than the normal level.

[0082] The drug for inhibiting complement activation includes drugs developed in the future for inhibiting excessive activation of the complement pathway.

[0083] In one embodiment, when the subject's C5b-9 or C5b-8 complement response level is determined to be abnormal, it is recommended to administer a complement inhibitor to the subject, wherein the complement inhibitor includes one or more of the following: C5 monoclonal antibody, factor B inhibitor, C3 inhibitor, factor D inhibitor.

[0084] In one embodiment, the C5 monoclonal antibody includes but is not limited to: eculiumab, crovalimab, PiaSky, Ravulizumab; the B factor inhibitor includes but is not limited to: iptacopan, NTQ5082; the C3 inhibitor includes but is not limited to: Pegcetagoplan, empaveli; the D factor inhibitor includes but is not limited to: Danicopan.

[0085] In one embodiment, eculizumab is a monoclonal antibody IgG2 / 4K that specifically binds to complement protein C5, thereby inhibiting its cleavage into C5a (a proinflammatory anaphylatoxin) and C5b (the initiating subunit of the terminal complement complex [C5b-9]) and preventing the production of C5b-9.

[0086] In one embodiment, Crovalimab is a humanized complement inhibitor C5 monoclonal antibody. It is a new generation of C5 inhibitor independently developed by Roche through continuous monoclonal antibody recovery technology, engineering transformation, and independent research and development. It can block the cleavage of complement C5 into C5a and C5b and inhibit complement activation. The drug combines isoelectric point, neonatal Fc receptor (FcRn) and pH-dependent affinity technology. These characteristics not only allow it to efficiently bind to C5, enhance the absorption of C5-crovalimab by endothelial cells, and process C5 in vivo, but also enable the efficient recycling of crovalimab mediated by neonatal Fc receptors.

[0087] In one embodiment, ABP 959 is a biosimilar candidate to the eculizumab reference product (RP), which has received orphan drug designation for the treatment of a group of rare diseases, including paroxysmal nocturnal hemoglobinuria (PNH), atypical hemolytic uremic syndrome (aHUS), generalized myasthenia gravis (gMG), and neuromyelitis optica spectrum disorder (NMOSD). The development of biosimilars approved for rare disease indications must provide a scientific rationale based on the totality of evidence (TOE). ABP 959 and eculizumab RP have highly similar inhibitory effects on C5 function, regardless of the serum type used. Confirmation of functional similarity between ABP 959 and eculizumab RP contributes to the TOE of biosimilarity and provides support for extrapolation based on inhibition of C5 function to additional rare disease indications approved for eculizumab RP.

[0088] In one embodiment, Tesidolumab (LFG316) is a fully human IgG1 / Δ anti-c5 monoclonal antibody with a molecular weight of 143 kDa (without glycosylation). Tesidolumab (LFG316) blocks the cleavage of C5, preventing the subsequent formation of the membrane attack complex.

[0089] In one embodiment, Pozelimab (REGN3918) is a human IgG4 anti-complement C5 monoclonal antibody. Pozelimab binds C5 and C5 variants with high affinity, blocking complement-mediated hemolysis. Pozelimab can be used for the study of complement-mediated diseases.

[0090] A computer program product comprising a computer program or instructions comprising:

[0091] The computer program or instruction is executed by the processor to implement the above-mentioned MDT decision method.

[0092] An MDT decision-making device, comprising:

[0093] A memory and a processor; the memory is used to store program instructions; the processor is used to call the program instructions, and when the program instructions are executed, the above-mentioned MDT decision method is implemented.

[0094] A computer-readable storage medium stores a computer program or instruction, wherein the computer program or instruction is executed by a processor to implement the above-mentioned MDT decision method.

[0095] The verification results of this verification embodiment show that assigning inherent weights to indications can improve the performance of the method relative to the default setting. Those skilled in the art can clearly understand that for the convenience and simplicity of description, the specific working processes of the systems, devices and units described above can refer to the corresponding processes in the aforementioned method embodiments, and will not be repeated here. In the several embodiments provided in this application, it should be understood that the disclosed systems, devices and methods can be implemented in other ways. For example, the device embodiments described above are only schematic, for example, the division of the units is only a logical function division, and there may be other division methods in actual implementation, such as multiple units or components can be combined or integrated into another system, or some features can be ignored or not executed. Another point is that the coupling or direct coupling or communication connection between each other shown or discussed can be through some interfaces, indirect coupling or communication connection of devices or units, which can be electrical, mechanical or other forms. The unit described as a separate component may or may not be physically separated, and the component displayed as a unit may or may not be a physical unit, that is, it may be located in one place, or it may be distributed on multiple network units. Some or all of the units can be selected according to actual needs to achieve the purpose of the scheme of this embodiment. In addition, each functional unit in each embodiment of the present invention may be integrated into one processing unit, or each unit may exist physically separately, or two or more units may be integrated into one unit. The above-mentioned integrated unit may be implemented in the form of hardware or in the form of software functional units. A person of ordinary skill in the art may understand that all or part of the steps in the various methods of the above-mentioned embodiments may be completed by instructing the relevant hardware through a program, and the program may be stored in a computer-readable storage medium, and the storage medium may include: a read-only memory (ROM), a random access memory (RAM), a disk or an optical disk, etc.

[0096] A person skilled in the art can understand that all or part of the steps in the above-mentioned embodiment method can be implemented by instructing related hardware through a program, and the program can be stored in a computer-readable storage medium. The above-mentioned medium storage can be a read-only memory, a disk or an optical disk, etc.

[0097] The above is a detailed introduction to a computer device provided by the present invention. For a person skilled in the art, according to the concept of the embodiments of the present invention, there may be changes in the specific implementation method and application scope. In summary, the content of this specification should not be understood as limiting the present invention.

Claims

1. A method for detecting the degree of complement system response based on endothelial cells, It is characterized in that The method comprises: S1: obtaining serum from the test subject; S2: obtaining human endothelial cells and adding the serum of the test subject and the human endothelial cells to a human endothelial cell culture medium to obtain cultured endothelial cells; S3: adding a primary antibody to the cultured endothelial cells for incubation to obtain a primary antibody incubation product, wherein the primary antibody is a primary antibody against a protein on the endothelial cells that is associated with complement response; S4: adding a secondary antibody to the primary antibody incubation product for incubation to obtain a secondary antibody incubation product; S5: The secondary antibody incubation product is detected by flow cytometry or electron microscopy to obtain fluorescence intensity, and the complement reaction level is obtained based on the fluorescence intensity.

2. The method for detecting the degree of complement system reaction based on endothelial cells according to claim 1, It is characterized in that The endothelial cells include one or more of the following: human glomerular endothelial cells, human umbilical vein endothelial cells, and human dermal vascular endothelial cells.

3. The method for detecting the degree of complement system reaction based on endothelial cells according to claim 1, It is characterized in that The primary antibody against the complement reaction-related protein includes one or more of the following: C5b-9, C5b-8, C3, C4d, C5a, and the secondary antibody is selected from the secondary antibody that binds to the primary antibody and whose signal is detected by a flow cytometer, including one or more of the following: AF647-IgG, allophycocyanin-IgG, and Cy5-IgG.

4. The method for detecting the degree of complement system reaction based on endothelial cells according to claim 1, It is characterized in that The S2 further comprises adding adenosine diphosphate to the culture medium of human endothelial cells.

5. A MDT decision-making method, It is characterized in that include: The method implements the method for detecting the degree of complement system reaction based on endothelial cells as described in any one of claims 1 to 4 to obtain the complement reaction level in endothelial cells. When it is determined that the complement reaction level of the subject is abnormal, the subject is given a drug recommended in the medication regimen to inhibit complement activation.

6. The MDT decision method according to claim 5, It is characterized in that The drug that inhibits complement activation is a drug that inhibits complement pathway activation when the activation level is greater than the normal level; Optionally, when it is determined that the C5b-9 or C5b-8 complement reaction level of the subject is abnormal, it is recommended to administer a complement inhibitor to the subject; Optionally, the complement inhibitor includes one or more of the following: C5 monoclonal antibody, factor B inhibitor, C3 inhibitor, factor D inhibitor.

7. The MDT decision method according to claim 6, It is characterized in that The C5 monoclonal antibodies include eculizumab, crovalimab, PiaSky, and Ravulizumab; Optionally, the factor B inhibitor includes iptacopan, NTQ5082; Optionally, the C3 inhibitors include Pegcetagoplan and empaveli; Optionally, the Factor D inhibitor comprises Danicopan.

8. A computer program product comprising a computer program or instructions, It is characterized in that The computer program or instructions are executed by a processor to implement the method for detecting the degree of complement system response based on endothelial cells as described in any one of claims 1 to 4, or to implement the MDT decision-making method as described in claim 5.

9. A computer device comprising a memory, a processor and a computer program or instructions stored in the memory, It is characterized in that The processor is used to call a computer program or instruction, and when the program instruction is executed, the method for detecting the degree of complement system reaction based on endothelial cells described in any one of claims 1-4 is implemented, or the MDT decision-making method described in claim 5 is implemented.

10. A computer-readable storage medium having a computer program or instructions stored thereon, It is characterized in that When the computer program is executed by a processor, it implements the method for detecting the degree of complement system reaction based on endothelial cells as described in any one of claims 1 to 4, or implements the MDT decision-making method as described in claim 5.

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