Applications, reagents, kits and systems for multi-cytokine detection reagents

Abstract

The present invention provides an application of a multi-cytokine detection reagent, a multi-cytokine detection reagent, a kit, and a cytokine detection system. The multi-cytokine detection reagent provided by the present invention is used to prepare a trauma degree detection reagent, wherein the multi-cytokine detection reagent is used to detect the concentration of multiple cytokines in a solution to be detected, and the corresponding trauma degree is determined based on the concentration of the multiple cytokines. The multiple cytokines include at least three of: IL-6, IL0, APRIL, Angiopoietin-2, FGF-basic, MPO, and Adiponectin. The present invention utilizes a method based on magnetic beads and antibodies combined with multi-channel detection of small molecules to simultaneously detect multiple cytokines, and combines the concentrations of multiple cytokines to detect the patient's trauma degree, thereby significantly improving the accuracy, sensitivity, and specificity of trauma degree detection.

Description

Technical Field

[0001] The present invention relates to the fields of basic biomedical research and biological detection, and in particular to the application of a multi-cytokine detection reagent, a multi-cytokine detection reagent, a kit, and a cytokine detection system. Background Art

[0002] Accurate diagnosis of trauma is of great value in improving the success rate of treatment and alleviating post-traumatic sequelae. Currently, clinical diagnosis of trauma mainly relies on the following methods:

[0003] 1. Inquiry about the patient's condition: mainly includes inquiring about the cause and mechanism of injury, the time of injury, the location and degree of pain, and through verbal communication, understanding whether the patient is conscious and willful after the injury.

[0004] 2. Basic examination: This mainly involves basic observation and examination of the patient, such as understanding the patient's abrasions and burns, and whether there are obvious fractures in the limbs. These two examinations are less dependent on instruments and equipment and can help doctors gain a preliminary understanding of the patient's trauma.

[0005] 3. Imaging examination: mainly includes ultrasound examination, CT examination, etc., which use professional imaging equipment to further assess the patient's injuries.

[0006] 4. Blood test: Understand the trauma situation by quantitatively testing the components in the peripheral blood.

[0007] Diagnoses determined through medical inquiries and basic examinations are subjective to both the doctor and the patient, potentially inaccurate, and difficult to detect internal organ damage. While imaging tests are more accurate and can examine internal organs, they can easily cause secondary trauma to the patient and are expensive, making them difficult for patients to accept. Furthermore, CT scans can cause cell mutations and are not suitable for pregnant women or adults trying to conceive. Blood tests, which can measure lymphocyte ratios and key blood components, are ideal, low-cost, and minimally impactful.

[0008] In the related art, there are three main categories of blood test components. The first category is the patient's blood coagulation-related factors, which are mainly used to evaluate the patient's blood coagulation status. The second category is the components related to the specific detection of the patient's liver function and kidney function. The detection of these two categories of components is targeted at specific functions (blood coagulation status, liver function, and kidney function), and it is difficult to use them for the overall assessment of trauma conditions represented by multiple injuries. The third category is the detection of acute proteins (such as C-reactive protein) and some interleukins. However, these two components are related to a variety of diseases, including infection, tumors, and some cardiovascular diseases. Infection will significantly increase these two components. In the clinical diagnosis of trauma, when the content of these two components in the blood is significantly increased, it is generally believed that the patient has an infection and is usually not used to diagnose the degree of trauma. Moreover, mild trauma can also cause a significant increase in acute proteins, so acute proteins often cannot reflect the degree of trauma of the patient.

[0009] Therefore, there is a need to develop new cytokine detection reagents for the preparation of trauma degree detection reagents to improve the accuracy, sensitivity and specificity of trauma degree detection. Summary of the Invention

[0010] To solve the problems existing in the above-mentioned background technology, the present invention provides an application of a multi-cytokine detection reagent, a multi-cytokine detection reagent, a kit, and a cytokine detection system cell to improve the accuracy, sensitivity and specificity of trauma degree detection.

[0011] In the first aspect, the present invention provides an application of a multi-cytokine detection reagent, which is used to prepare a trauma degree detection reagent. The multi-cytokine detection reagent is used to detect the concentration of multiple cytokines in a solution to be detected, and the corresponding trauma degree is determined based on the concentration of the multiple cytokines. The multiple cytokines include: at least three of IL-6, IL10, APRIL, Angiopoietin-2, FGF-basic, MPO, and Adiponectin.

[0012] In a second aspect, the present invention provides a multi-cytokine detection reagent for preparing a trauma degree detection reagent, the multi-cytokine detection reagent comprising: cytokine binding-specific magnetic beads, a cytokine high-affinity monoclonal antibody, and a fluorescent dye PE, wherein the antibody is linked to biotin, and the fluorescent dye PE is linked to streptavidin. The cytokine binding-specific magnetic beads are used to adsorb cytokines contained in a solution to be detected, the cytokine high-affinity monoclonal antibody is used to bind to the cytokines, the biotin linked to the antibody is used to bind to streptavidin, and the fluorescence intensity of the fluorescent dye PE is used to determine the concentration of the cytokines in the solution to be detected;

[0013] The multi-cytokine detection reagent is the multi-cytokine detection reagent described in any one of the first aspects above, and the multi-cytokine detection reagent is used to detect the concentration of multiple cytokines in the solution to be detected, and determine the corresponding degree of trauma based on the concentration of the multiple cytokines.

[0014] In a third aspect, the present invention provides a kit comprising a multi-cytokine detection reagent for preparing a wound degree detection reagent, the kit further comprising: purified cytokine dry powder and a buffer solution, wherein the purified cytokine dry powder is used to determine a fluorescence intensity-concentration standard curve;

[0015] The multi-cytokine detection reagent is the multi-cytokine detection reagent described in any one of the first aspects above, and the multi-cytokine detection reagent is used to detect the concentration of multiple cytokines in the solution to be detected, and determine the corresponding degree of trauma based on the concentration of the multiple cytokines.

[0016] In a fourth aspect, the present invention provides a cytokine detection system, comprising: the kit described in the third aspect above, and a flow analyzer, wherein the kit is used to determine a fluorescence intensity-concentration standard curve, and the kit is further used to display the concentration of the cytokine in the form of fluorescence intensity; the flow analyzer is used to determine the value of the fluorescence intensity, and the concentration of the cytokine is determined based on the fluorescence intensity-concentration standard curve and the fluorescence intensity value;

[0017] The kit includes a multi-cytokine detection reagent used to prepare a trauma degree detection reagent. The multi-cytokine detection reagent is the multi-cytokine detection reagent described in any one of the first aspects above. The multi-cytokine detection reagent is used to detect the concentration of multiple cytokines in the solution to be detected, and the corresponding trauma degree is determined based on the concentration of the multiple cytokines.

[0018] Compared with the prior art, the present invention has the following advantages:

[0019] In an embodiment of the present invention, a multi-cytokine detection reagent is used to detect multiple cytokines in a solution to be tested, and the degree of trauma of the patient is detected in combination with the concentration of multiple cytokines, thereby accurately determining the degree of trauma of the patient. In an embodiment of the present invention, the AUC of the cytokine combination for monitoring the degree of trauma reaches 0.948 or above, which can improve the accuracy, sensitivity and specificity of trauma detection. In addition, the detection of multiple cytokines can be incorporated into the routine blood test process without the need for additional testing work, thereby conveniently determining the degree of trauma of the patient without causing harm to the patient. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1This is a graph showing the differential expression of cytokines IL-6, IL10, APRIL, Angiopoietin-2, FGF-basic, MPO, and Adiponectin in the peripheral blood of patients with severe trauma and those with mild trauma;

[0021] Figure 2 is the ROC curve of cytokines IL-10, Angiopoietin-2, and FGF-basic in determining the degree of trauma;

[0022] Figure 3 is the ROC curve of cytokines APRIL, Angiopoietin-2, and FGF-basic in determining the degree of trauma;

[0023] Figure 4 is the ROC curve of cytokines IL-10, Angiopoietin-2, and MPO in determining the degree of trauma;

[0024] Figure 5 is the ROC curve of cytokines APRIL, Angiopoietin-2, and MPO in determining the degree of trauma;

[0025] Figure 6 is the ROC curve for determining the degree of trauma using cytokines IL-10, Angiopoietin-2, FGF-basic, and MPO;

[0026] Figure 7 is the ROC curve of cytokines APRIL, Angiopoietin-2, FGF-basic, and Adiponectin in determining the degree of trauma;

[0027] Figure 8 is the ROC curve of cytokines APRIL, Angiopoietin-2, FGF-basic, and MPO in determining the degree of trauma;

[0028] Figure 9 is the ROC curve for determining the degree of trauma using cytokines IL-10, Angiopoietin-2, FGF-basic, and Adiponectin;

[0029] Figure 10 is the ROC curve for determining the degree of trauma using cytokines IL-6, APRIL, Angiopoietin-2, FGF-basic, and MPO;

[0030] Figure 11is the ROC curve for determining the degree of trauma using cytokines IL-6, IL-10, Angiopoietin-2, FGF-basic, and MPO;

[0031] Figure 12 is the ROC curve diagram for determining the degree of trauma using cytokines IL-10, Angiopoietin-2, FGF-basic, MPO, and Adiponectin;

[0032] Figure 13 is the ROC curve for determining the degree of trauma using cytokines APRIL, Angiopoietin-2, FGF-basic, MPO, and Adiponectin;

[0033] Figure 14 ROC curves of cytokines IL-6, IL-10, Angiopoietin-2, FGF-basic, MPO, and Adiponectin for determining the degree of trauma;

[0034] Figure 15 This is the ROC curve diagram of cytokines IL-6, IL-10, APRIL, Angiopoietin-2, FGF-basic, MPO, and Adiponectin in determining the degree of trauma. DETAILED DESCRIPTION

[0035] The following examples are provided for a better understanding of the present invention and are not intended to limit the best mode of implementation. They do not limit the content and scope of protection of the present invention. Any product identical or similar to the present invention obtained by anyone under the guidance of the present invention or by combining the features of the present invention with other prior arts shall fall within the scope of protection of the present invention.

[0036] If no specific experimental steps or conditions are specified in the examples, the conventional experimental steps or conditions described in the literature in this field can be used. The reagents and other instruments used, if the manufacturers are not specified, are all conventional reagents that can be purchased on the market.

[0037] Cytokines are a class of small proteins with a wide range of biological activities that are synthesized and secreted by immune cells (such as monocytes, macrophages, T cells, B cells, and NK cells) and certain non-immune cells (endothelial cells, epidermal cells, fibroblasts, and glial cells) upon stimulation. Cytokines are numerous and play a vital role in inflammatory responses, hematopoiesis, immune responses, and wound healing. Cytokines vary with the course of disease. Testing for multiple cytokines in trauma patients holds the potential to identify cytokines that can be used to assess the severity of trauma.

[0038] Based on this, in order to solve the problems existing in the related art, the inventors detected more than 100 cytokines in the peripheral blood of 20 patients with severe trauma (ISS score ≥ 15 points) and 35 patients with mild trauma. 18 cytokines were found, and the significance analysis (unpaired t-test) showed that there were significant differences in the concentrations in the peripheral blood of patients with mild and severe diseases (P < 0.05). For several other factors, P was slightly greater than 0.05. The embodiment of the present invention randomly arranged and combined a total of 27 factors, and the receiver operating characteristic curve (ROC curve) analysis results showed that 7 of the factors could obtain 4 three-factor combinations, 4 four-factor combinations, 4 five-factor combinations, 1 six-factor combination, and 1 seven-factor combination, with an AUC greater than 0.948 (as shown in Table 1 below). The use of these multi-factor combinations for trauma degree detection can improve the accuracy, sensitivity and specificity of trauma degree detection.

[0039] Table 1 AUC values ​​of multiple cytokine combinations

[0040]

[0041]

[0042] Based on this, the first aspect of the present invention proposes an application of a multi-cytokine detection reagent, which is used to prepare a trauma degree detection reagent. The multi-cytokine detection reagent is used to detect the concentration of multiple cytokines in the solution to be detected, and the corresponding trauma degree is determined based on the concentration of the multiple cytokines. The multiple cytokines include: at least three of IL-6, IL10, APRIL, Angiopoietin-2, FGF-basic, MPO, and Adiponectin.

[0043] In an optional embodiment of the present invention, the multi-cytokine detection reagent is a three-factor detection reagent, and the multiple cytokines include: IL-10, Angiopoietin-2, FGF-basic, or APRIL, Angiopoietin-2, FGF-basic, or IL-10, Angiopoietin-2, MPO, or APRIL, Angiopoietin-2, MPO.

[0044] Specifically, the method for determining the degree of trauma using the three-factor detection reagent is shown in Table 2 below.

[0045] Table 2 Severe trauma assessment criteria for the three-factor test reagent (unit: pg / ml peripheral blood serum test)

[0046]

[0047]

[0048] In an optional embodiment of the present invention, the multi-cytokine detection reagent is a four-factor detection reagent, and the multiple cytokines include: IL-10, Angiopoietin-2, FGF-basic, MPO, or APRIL, Angiopoietin-2, FGF-basic, MPO, or IL-10, Angiopoietin-2, FGF-basic, Adiponectin, or APRIL, Angiopoietin-2, FGF-basic, Adiponectin.

[0049] Specifically, the method for determining the degree of trauma using the four-factor detection reagent is shown in Table 3 below.

[0050] Table 3 Criteria for severe trauma assessment using the four-factor test reagent (unit: pg / ml peripheral blood serum test)

[0051]

[0052]

[0053] In an optional embodiment of the present invention, the multi-cytokine detection reagent is a five-factor detection reagent, and the multiple cytokines include: IL-6, IL-10, Angiopoietin-2, FGF-basic, MPO, or IL-6, APRIL, Angiopoietin-2, FGF-basic, MPO, or IL-10, Angiopoietin-2, FGF-basic, MPO, Adiponectin, or APRIL, Angiopoietin-2, FGF-basic, MPO, Adiponectin.

[0054] Specifically, the method for determining the degree of trauma using the five-factor detection reagent is shown in Table 4 below.

[0055] Table 4 Criteria for severe trauma assessment using the five-factor assay (pg / ml peripheral blood serum test)

[0056]

[0057]

[0058] In an optional embodiment of the present invention, the multi-cytokine detection reagent is a six-factor detection reagent, and the multiple cytokines include: IL-6, IL-10, Angiopoietin-2, FGF-basic, MPO, and Adiponectin.

[0059] Specifically, the method for determining the degree of trauma using the six-factor detection reagent is shown in Table 5 below.

[0060] Table 5 Six-factor test reagent severe trauma judgment standard (unit pg / ml peripheral blood serum test)

[0061]

[0062] In an optional embodiment of the present invention, the multi-cytokine detection reagent is a seven-factor detection reagent, and the cytokines include: IL-6, IL-10, APRIL, Angiopoietin-2, FGF-basic, MPO, and Adiponectin.

[0063] Specifically, the method for determining the degree of trauma using the seven-factor detection reagent is shown in Table 6 below.

[0064] Table 6 Severe trauma assessment criteria for the seven-factor test reagent (unit: pg / ml peripheral blood serum test)

[0065]

[0066] In order to enable those skilled in the art to better understand the present invention, the application of the multi-cytokine detection reagent provided by the present invention is described below through multiple specific examples.

[0067] Example 1: Experimental study on the difference in peripheral blood concentrations of seven factors between patients with severe trauma and patients with mild trauma

[0068] In an embodiment of the present invention, the cytokine concentration is detected based on a method of combining magnetic beads with antibodies and multi-channel detection of small molecules. This method can use magnetic beads of various sizes and multiple fluorescence intensities at the same time, and each magnetic bead detects a specific molecule. The specific size of the magnetic beads and the intensity of a certain wavelength of fluorescence (fluorescence A) are the labels of the magnetic beads. This parameter can be directly read by flow cytometry analysis technology to directly know which molecule a certain magnetic bead detects. The detected molecule will then be further bound by an antibody with another fluorescence (fluorescence B). The concentration of the target molecule can be known by the intensity of fluorescence B, and then the fluorescence intensity-concentration standard curve can be used for absolute quantification. For specific detection methods, please refer to the mature and feasible detection methods in the relevant technology.

[0069] Based on this detection method, the concentrations of seven cytokines (IL-6, IL10, APRIL, Angiopoietin-2, FGF-basic, MPO, and Adiponectin) in the peripheral blood of 20 patients with severe trauma (ISS score ≥ 15 points) and 35 patients with mild trauma were detected in the present embodiment. The test results were analyzed to obtain Figure 1 ,Depend on Figure 1 It can be seen that the concentrations of six of these seven factors in the peripheral blood of patients with mild or severe disease were significantly different (P < 0.05). In order to further improve the accuracy, sensitivity and specificity of trauma degree detection, in the embodiment of the present invention, any three of these seven cytokines can be selected and combined for trauma degree detection.

[0070] Example 2: Experiment on the difference in concentration of multi-cytokine detection reagents in peripheral blood of patients with severe trauma and patients with mild trauma

[0071] In the present embodiment, a method based on magnetic beads and antibodies combined with multi-channel detection of small molecules was used to detect the concentration of each cytokine combination in the peripheral blood of 20 patients with severe trauma (ISS score ≥ 15 points) and 35 patients with mild trauma. The test results were analyzed to obtain the receiver operating characteristic curve (ROC curve) of each cytokine combination. The analysis results showed that ( Figure 2-15 ), these cytokine combinations have high accuracy, sensitivity and specificity for detecting the degree of trauma.

[0072] In an embodiment of the present invention, 14 cytokine combinations with an AUC greater than 0.948 are provided. Based on these 14 cytokine combinations, 14 multi-cytokine detection reagents can be obtained respectively. These 14 multi-cytokine detection reagents are used to prepare trauma degree detection reagents, which can improve the accuracy, sensitivity and specificity of trauma degree detection.

[0073] Based on the same inventive concept, in a second aspect, the present invention provides a multi-cytokine detection reagent for preparing a trauma degree detection reagent, the multi-cytokine detection reagent comprising: cytokine binding-specific magnetic beads, a cytokine high-affinity monoclonal antibody, and a fluorescent dye PE, wherein the antibody is linked to biotin, and the fluorescent dye PE is linked to streptavidin. The cytokine binding-specific magnetic beads are used to adsorb the cytokines contained in the solution to be detected, the cytokine high-affinity monoclonal antibody is used to bind to the cytokines, the biotin linked to the antibody is used to bind to streptavidin, and the fluorescence intensity of the fluorescent dye PE is used to determine the concentration of the cytokines in the solution to be detected;

[0074] The multi-cytokine detection reagent is the multi-cytokine detection reagent described in any one of the first aspects above, and the multi-cytokine detection reagent is used to detect the concentration of multiple cytokines in the solution to be detected, and determine the corresponding degree of trauma based on the concentration of the multiple cytokines.

[0075] In a third aspect, the present invention provides a kit comprising a multi-cytokine detection reagent for preparing a wound degree detection reagent, the kit further comprising: purified cytokine dry powder and a buffer solution, wherein the purified cytokine dry powder is used to determine a fluorescence intensity-concentration standard curve;

[0076] The multi-cytokine detection reagent is the multi-cytokine detection reagent described in any one of the first aspects above, and the multi-cytokine detection reagent is used to detect the concentration of multiple cytokines in the solution to be detected, and determine the corresponding degree of trauma based on the concentration of the multiple cytokines.

[0077] In a fourth aspect, the present invention provides a cytokine detection system, comprising: the kit described in the third aspect above, and a flow analyzer, wherein the kit is used to determine a fluorescence intensity-concentration standard curve, and the kit is further used to display the concentration of the cytokine in the form of fluorescence intensity; the flow analyzer is used to determine the value of the fluorescence intensity, and the concentration of the cytokine is determined based on the fluorescence intensity-concentration standard curve and the fluorescence intensity value;

[0078] The kit includes a multi-cytokine detection reagent used to prepare a trauma degree detection reagent. The multi-cytokine detection reagent is the multi-cytokine detection reagent described in any one of the first aspects above. The multi-cytokine detection reagent is used to detect the concentration of multiple cytokines in the solution to be detected, and the corresponding trauma degree is determined based on the concentration of the multiple cytokines.

[0079] The above describes in detail the application of a multi-cytokine detection reagent, a multi-cytokine detection reagent, a kit, and a cytokine detection system provided by the present invention. Specific examples are used herein to illustrate the principles and implementation methods of the present invention. The description of the above embodiments is only used to help understand the method of the present invention and its core concept. At the same time, for those skilled in the art, according to the concept of the present invention, there may be changes in the specific implementation methods and application scopes. In summary, the contents of this specification should not be understood as limiting the present invention.

Claims

1. Use of a multi-cytokine detection reagent in the preparation of a wound degree detection reagent, characterized in that: The multi-cytokine detection reagent is used to detect the concentration of multiple cytokines in a solution to be detected, and the corresponding degree of trauma is determined based on the concentration of the multiple cytokines, wherein the multiple cytokines include at least three of IL-6, IL-10, APRIL, Angiopoietin-2, FGF-basic, MPO, and Adiponectin; In the case where the multi-cytokine detection reagent is a three-factor detection reagent, the multiple cytokines include: IL-10, Angiopoietin-2, FGF-basic, or APRIL, Angiopoietin-2, FGF-basic, or IL-10, Angiopoietin-2, MPO, or APRIL, Angiopoietin-2, MPO; or When the multi-cytokine detection reagent is a four-factor detection reagent, the multiple cytokines include: IL-10, Angiopoietin-2, FGF-basic, MPO, or APRIL, Angiopoietin-2, FGF-basic, MPO, or IL-10, Angiopoietin-2, FGF-basic, Adiponectin, or APRIL, Angiopoietin-2, FGF-basic, Adiponectin.

2. Use of the multi-cytokine detection reagent according to claim 1 in preparing a wound degree detection reagent, characterized in that: The multi-cytokine detection reagent is a five-factor detection reagent, and the multiple cytokines include: IL-6, IL-10, Angiopoietin-2, FGF-basic, MPO, or IL-6, APRIL, Angiopoietin-2, FGF-basic, MPO, or IL-10, Angiopoietin-2, FGF-basic, MPO, Adiponectin, or APRIL, Angiopoietin-2, FGF-basic, MPO, Adiponectin.

3. Use of the multi-cytokine detection reagent according to claim 1 in preparing a wound degree detection reagent, characterized in that: The multi-cytokine detection reagent is a six-factor detection reagent, and the multiple cytokines include: IL-6, IL-10, Angiopoietin-2, FGF-basic, MPO, and Adiponectin.

4. Use of the multi-cytokine detection reagent according to claim 1 in preparing a wound degree detection reagent, characterized in that: The multi-cytokine detection reagent is a seven-factor detection reagent, and the multiple cytokines include: IL-6, IL-10, APRIL, Angiopoietin-2, FGF-basic, MPO, and Adiponectin.

5. Use of the multi-cytokine detection reagent according to any one of claims 1 to 4 in the preparation of a wound degree detection reagent, characterized in that: The multi-cytokine detection reagent is a blood sample detection reagent, and the solution to be detected is a blood sample.

6. A multi-cytokine detection reagent for preparing a wound degree detection reagent, characterized in that: The multi-cytokine detection reagent includes: cytokine binding-specific magnetic beads, cytokine high-affinity monoclonal antibodies, and fluorescent dye PE, wherein the antibodies are connected to biotin, and the fluorescent dye PE is connected to streptavidin. The cytokine binding-specific magnetic beads are used to adsorb cytokines contained in the solution to be detected, the cytokine high-affinity monoclonal antibodies are used to bind to the cytokines, and the biotin connected to the antibodies is used to bind to streptavidin. The fluorescence intensity of the fluorescent dye PE is used to determine the concentration of the cytokines in the solution to be detected; The multi-cytokine detection reagent is the multi-cytokine detection reagent according to any one of claims 1 to 4, and is used to detect the concentrations of multiple cytokines in a solution to be detected, and determine the corresponding degree of trauma based on the concentrations of the multiple cytokines.

7. A kit, characterized in that The kit includes a multi-cytokine detection reagent for preparing a wound degree detection reagent, and the kit also includes: purified cytokine dry powder and a buffer solution, wherein the purified cytokine dry powder is used to determine a fluorescence intensity-concentration standard curve; The multi-cytokine detection reagent is the multi-cytokine detection reagent according to any one of claims 1 to 4, and is used to detect the concentrations of multiple cytokines in a solution to be detected, and determine the corresponding degree of trauma based on the concentrations of the multiple cytokines.

8. A cytokine detection system, characterized in that: The cytokine detection system comprises: a flow analyzer and a kit according to claim 7, wherein the kit is used to determine a fluorescence intensity-concentration standard curve, and the kit is further used to display the concentration of the cytokine in the form of fluorescence intensity, and the flow analyzer is used to determine the value of the fluorescence intensity, and the concentration of the cytokine is determined based on the fluorescence intensity-concentration standard curve and the value of the fluorescence intensity; The kit includes a multi-cytokine detection reagent used to prepare a trauma degree detection reagent, wherein the multi-cytokine detection reagent is the multi-cytokine detection reagent according to any one of claims 1 to 4, and the multi-cytokine detection reagent is used to detect the concentration of multiple cytokines in the solution to be detected, and the corresponding trauma degree is determined based on the concentration of the multiple cytokines.

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