Spatial proximity chemiluminescence kit for detecting troponin I and preparation method thereof

Abstract

The invention relates to a spatial proximity chemiluminescence kit for detecting troponin I and a preparation method thereof, and relates to the technical field of biology. The kit comprises an enzyme marker, a luminous marker, a luminous auxiliary agent and a triggering agent, the enzyme marker comprises an anti-cTnI monoclonal antibody marked by horse radish peroxidase, and the luminous marker comprises an anti-cTnI monoclonal antibody marked by 9, 10-dihydroacridine. The kit adopts a space adjacent chemical double-antibody sandwich method, is simple to operate, can realize full-automatic detection, does not need magnetic beads and washing, reduces the reagent cost, shortens the detection time, adopts a homogeneous reaction, and has the advantages of high detection accuracy, high sensitivity, high sensitivity and the like by being compared with other methods such as enzyme-linked immunosorbent assay and fluorescence immunochromatography. The influence caused by the traditional chemiluminescence magnetic separation technology is avoided, so that the detection result is more accurate and stable.

Description

Technical Field

[0001] The present invention relates to the field of biotechnology, and in particular to a spatial proximity chemiluminescence kit for detecting troponin I and a preparation method thereof. Background Art

[0002] Acute myocardial infarction (AMI) is myocardial necrosis caused by acute, persistent ischemia and hypoxia in the coronary arteries. It is a sudden, high-mortality clinical disease. It typically develops on the basis of coronary atherosclerotic stenosis. When predisposing factors such as excessive emotional excitement, strenuous physical exertion, shock, and dehydration lead to a sharp decrease or even interruption in coronary blood flow, resulting in myocardial infarction. It is a leading cause of sudden death in middle-aged and elderly people. Clinical manifestations often include persistent, severe substernal pain, acute circulatory dysfunction, arrhythmias, heart failure, fever, elevated white blood cell count and serum myocardial injury marker enzymes, and progressive electrocardiographic changes indicating acute myocardial injury and necrosis. Early diagnosis and interventional treatment are crucial for saving the dying myocardium, improving prognosis, and reducing acute mortality.

[0003] Cardiac troponin (cTn) is a protein that regulates myocardial contraction. cTn is composed of three subunits from different genes: cardiac troponin T (cTnT), cardiac troponin I (cTnI), and troponin C (TnC). In cardiomyocytes, these three subunits form the cardiac troponin complex, which together carry out the essential function of calcium signaling through intracellular actin interactions. cTnI is a highly specific and sensitive marker of myocardial injury. When the integrity of the myocardial cell membrane is compromised, cTnI is first released from the cytoplasm. Because cTnI appears in the blood early, persists for a long time, and has high sensitivity and specificity, it holds great promise for clinical application. To meet the clinical and societal needs for the prevention and treatment of heart failure, the development of a test kit for cTnI protein is crucial.

[0004] Currently, common methods for detecting cTnI protein include enzyme-linked immunosorbent assay (ELISA) and chemiluminescence assay. ELISA has complex and time-consuming procedures, requires specialized personnel, and is subject to numerous factors affecting the operation, which can easily lead to false-positive and false-negative results. Chemiluminescence assays offer high sensitivity, good specificity, and excellent stability, and are non-toxic to the environment and the human body. In light of this, the present invention provides a kit for detecting cTnI using a spatial proximity chemiluminescence assay and a method for preparing the same. Summary of the Invention

[0005] The technical problem to be solved by the present invention is to provide a spatial proximity chemiluminescence assay kit for detecting troponin I and a method for preparing the same. The goal is to utilize spatial proximity chemiluminescence analysis without the need for a carrier or washing process, thereby reducing kit costs, shortening detection time, and making detection results more reliable.

[0006] The technical solution of the present invention to solve the above technical problems is as follows:

[0007] In a first aspect, a spatial proximity chemiluminescence kit for detecting cardiac troponin I is provided, the kit comprising an enzyme marker, a luminescent marker, a luminescent adjuvant, and a trigger; the enzyme marker comprises an anti-cTnI monoclonal antibody labeled with horseradish peroxidase (HRP), and the luminescent marker comprises an anti-cTnI monoclonal antibody labeled with 9,10-dihydroacridine (Acridan).

[0008] On the basis of the above technical solution, the present invention can also be improved as follows.

[0009] Furthermore, the enzyme marker also includes bovine serum albumin, sucrose and trehalose;

[0010] The concentration of the horseradish peroxidase-labeled anti-PGⅠ monoclonal antibody in the enzyme marker is 1 mg / ml;

[0011] The mass concentration of bovine serum albumin in the enzyme marker is 0.5% to 2%;

[0012] The mass concentration of sucrose in the enzyme marker is 2% to 5%;

[0013] The mass concentration of trehalose in the enzyme marker is 1% to 3%.

[0014] Furthermore, the luminescent marker also includes casein, sucrose and trehalose;

[0015] The concentration of the 9,10-dihydroacridine-labeled anti-PGⅠ monoclonal antibody in the luminescent marker is 0.5 mg / ml;

[0016] The mass concentration of casein in the luminescent marker is 0.1% to 1%;

[0017] The mass concentration of sucrose in the luminescent marker is 2% to 5%;

[0018] The mass concentration of trehalose in the luminescent marker is 1% to 3%.

[0019] Furthermore, the luminescence auxiliary agent also includes citric acid and sodium citrate;

[0020] Each 1L of the luminescence auxiliary agent includes the following components in the following amounts: 1.5g to 2g of citric acid, 5g to 11g of sodium citrate, and 0.15mL to 0.45mL of 30% hydrogen peroxide.

[0021] Furthermore, each 1L of the trigger comprises the following components: 3g to 9g of tris(hydroxymethyl)aminomethane hydrochloride, 6g to 12g of NaCl, 1mL to 4mL of concentrated HCl, and 1mL to 4mL of a surfactant;

[0022] The mass content of the concentrated HCl is 36% to 38%.

[0023] Furthermore, the surfactant includes Tween 20.

[0024] In a second aspect, a method for preparing a kit for detecting troponin I by a spatial proximity chemiluminescence method comprises the following steps:

[0025] The enzyme marker was prepared using horseradish peroxidase and anti-cTnI monoclonal antibody;

[0026] A luminescent marker was prepared using 9,10-dihydroacridine and anti-cTnI monoclonal antibody;

[0027] A luminescence auxiliary agent is prepared by using citric acid, sodium citrate and hydrogen peroxide solution;

[0028] The trigger was prepared using tris(hydroxymethylaminomethane) hydrochloride, NaCl, concentrated HCl and a surfactant.

[0029] Furthermore, the mass ratio of the horseradish peroxidase to the anti-cTnI monoclonal antibody is 1:2.

[0030] The preparation of the enzyme marker comprises the following specific steps: dissolving horseradish peroxidase in distilled water, adding sodium periodate solution, mixing, dialysis in sodium acetate buffer, adding carbonate buffer, anti-cTnI monoclonal antibody, adding sodium borohydride solution, mixing, dialysis again in PBS, taking out the dialyzate, adding 50% glycerol and storing at -20°C for future use.

[0031] Furthermore, the mass ratio of the 9,10-dihydroacridine to the anti-cTnI monoclonal antibody is 1:3.

[0032] The preparation of the luminescent marker includes the following specific steps: dissolving 9,10-dihydroacridine, adding sodium borate buffer and anti-cTnI monoclonal antibody, shaking and mixing overnight, taking out, adding 50% glycerol and storing in a -20°C refrigerator.

[0033] Furthermore, the preparation of the luminescence auxiliary agent includes the following specific steps: taking citric acid, sodium citrate and hydrogen peroxide solution, dissolving them in purified water and fixing the volume to obtain the luminescence auxiliary agent;

[0034] The preparation of the trigger comprises the following specific steps: weighing tris(hydroxymethyl)aminomethane hydrochloride), NaCl, concentrated HCl and a surfactant, dissolving them and then fixing the volume to obtain the trigger.

[0035] The present invention uses a spatial proximity chemiluminescence analysis method to detect cTnI protein. A sample, a horseradish peroxidase (HRP)-labeled anti-cTnI monoclonal antibody, and a 9,10-dihydroacridine (Acridan)-labeled anti-cTnI monoclonal antibody are reacted together to form an antigen-antibody sandwich complex, so that the horseradish peroxidase and the 9,10-dihydroacridine (Acridan) are spatially close to each other. A luminescence auxiliary agent and a triggering agent are added to generate flash chemiluminescence, while unbound free HRP-labeled antibody and Acridan-labeled antibody do not emit light. The higher the cTnI content in the sample, the greater the measured luminescence value (RLU). Therefore, within a certain concentration range, the luminescence value is positively correlated with the sample concentration. A working curve is drawn using calibrators of known concentrations and their luminescence values, and the cTnI content in the sample can be calculated based on the luminescence value of the sample.

[0036] The beneficial effects of the present invention are as follows: the spatial proximity chemical double antibody sandwich method adopted in this kit is simple to operate and can be fully automatically detected compared with other methods such as enzyme-linked immunosorbent assay and fluorescent immunochromatography, without the need for magnetic beads and washing, which not only reduces the reagent cost but also shortens the detection time. In addition, it adopts a homogeneous reaction, avoiding the influence of traditional chemiluminescence magnetic separation technology, making the detection results more accurate and stable. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] Figure 1 It is the standard curve diagram of the present invention. DETAILED DESCRIPTION

[0038] The principles and features of the present invention are described below. The examples are only used to explain the present invention and are not intended to limit the scope of the present invention. Where specific techniques or conditions are not specified in the examples, the techniques or conditions described in the literature in this field or the product instructions are used. Where the manufacturers of the reagents or instruments used are not specified, they are all conventional products that can be purchased through regular channels.

[0039] Example

[0040] 1. Materials and Reagents

[0041] (1) Antigen and Antibody:

[0042] Product No.: 10066 (anti-cTnI monoclonal antibody), 10066 was purchased from Nanjing Huading Biotechnology Co., Ltd.; Product No. cTnI-Ag5, antigen was purchased from Feipeng Biotechnology Co., Ltd.

[0043] (2) Other reagents:

[0044] Horseradish peroxidase (HRP), sodium periodate, sodium borohydride, sodium acetate, sodium carbonate, sodium bicarbonate, disodium hydrogen phosphate dodecahydrate, sodium dihydrogen phosphate dihydrate, sodium chloride, tris (hydroxymethyl)aminomethane hydrochloride (Tris), hydrochloric acid, citric acid, sodium citrate, glycerol, boric acid, sodium tetraborate, sucrose, aminopyrine, Tween 20, Triton 100, potassium dihydrogen phosphate, hydrogen peroxide, and dimethylformamide were purchased from Sinopharm Chemical Reagent Co., Ltd. Bovine serum albumin, casein, Proclin 300, and polyethylene glycol 2000 were purchased from Sigma-Aldrich (Shanghai) Trading Co., Ltd. 9,10-Dihydroacridine (HRP) was purchased from Sichuan Weikeqi Biotechnology Co., Ltd.

[0045] (3) Instruments:

[0046] Guangzhou Jinde Biotechnology Co., Ltd. HomoG 100 fully automatic chemiluminescence analyzer (Guangdong Medical Device Registration No. 20212220196)

[0047] 2. Kit and Preparation Method

[0048] The spatial proximity chemiluminescence analysis method, Spatial Proximity Analyte Reagent Capture Luminescence (SPARCL), is used. The basic principle is that the analyte to be tested is simultaneously specifically bound to two antibodies labeled with horseradish peroxidase (HRP) and a chemiluminescent substrate (Acridan, 9,10-dihydroacridine), respectively, to form a complex. Horseradish peroxidase (HRP) and 9,10-dihydroacridine (Acridan) are spatially close to each other. After the addition of a trigger containing hydrogen peroxide (H2O2) and an enhancer, a flash-type chemiluminescent signal is immediately generated. This luminescent signal can be detected by a chemiluminescence instrument. The intensity of the light is proportional to the amount of the analyte to be tested in the sample.

[0049] 2.1 Spatial proximity chemiluminescence kit for detecting troponin I

[0050] A spatial proximity chemiluminescence kit for detecting cardiac troponin I comprises an enzyme marker, a luminescent marker, a luminescent auxiliary agent, and a trigger; the enzyme marker comprises an anti-cTnI monoclonal antibody labeled with horseradish peroxidase (HRP), and the luminescent marker comprises an anti-cTnI monoclonal antibody labeled with 9,10-dihydroacridine (Acridan).

[0051] Preferably, the enzyme marker further comprises bovine serum albumin, sucrose and trehalose;

[0052] The concentration of the horseradish peroxidase-labeled anti-PGⅠ monoclonal antibody in the enzyme marker is 1 mg / ml;

[0053] The mass concentration of bovine serum albumin in the enzyme marker is 0.5% to 2%;

[0054] The mass concentration of sucrose in the enzyme marker is 2% to 5%;

[0055] The mass concentration of trehalose in the enzyme marker is 1% to 3%.

[0056] Preferably, in this embodiment, the luminescent marker further comprises casein, sucrose and trehalose;

[0057] The concentration of the 9,10-dihydroacridine-labeled anti-PGⅠ monoclonal antibody in the luminescent marker is 0.5 mg / ml;

[0058] The mass concentration of casein in the luminescent marker is 0.1% to 1%;

[0059] The mass concentration of sucrose in the luminescent marker is 2% to 5%;

[0060] The mass concentration of trehalose in the luminescent marker is 1% to 3%.

[0061] In this embodiment, preferably, each 1 L of the luminescence auxiliary agent includes the following components: 1.5 g to 2 g of citric acid, 5 g to 11 g of sodium citrate, and 0.15 mL to 0.45 mL of 30% hydrogen peroxide.

[0062] Furthermore, each 1L of the trigger includes the following components: 3g to 9g of tris(hydroxymethyl)aminomethane hydrochloride, 6g to 12g of NaCl, 1mL to 4mL of concentrated HCl and 1mL to 4mL of a surfactant; the surfactant includes Tween 20.

[0063] The mass content of the concentrated HCl is 36% to 38%.

[0064] The following is a detailed description of the preparation process of this kit, and the prepared kit is used for experimental detection.

[0065] 2.2 Specific preparation method of the kit

[0066] (1) Preparation of enzyme markers:

[0067] Weigh 1 mg of horseradish peroxidase (HRP) and dissolve it in 200 μL of distilled water. Add 40 μL of freshly prepared 0.1 M sodium periodate and shake at room temperature for 20 min. Pour the above liquid into a dialysis bag and dialyze it against 1 mM sodium acetate buffer (pH 4.4) at 4°C overnight. Add 20 μL of 0.2 M carbonate buffer (pH 9.5). Immediately add 1 mg of anti-cTnI monoclonal antibody and stir gently at room temperature in the dark for 2 h. Add 10 μL of freshly prepared 4 mg / mL sodium borohydride solution, mix well, and let it stand at 4°C for 2 h. Pour the above liquid into a dialysis bag and dialyze it against 0.15 M PBS (pH 7.4) at 4°C overnight. Remove the dialysate, add 50% glycerol and store it at -20°C until use.

[0068] (2) Preparation of luminescent markers:

[0069] Dissolve 1 mg of the luminescent substrate 9,10-dihydroacridine (Acridan) in 500 μL of DMF, remove 41.3 μL, add 708.7 μL of 0.05 M sodium borate buffer, then add 250 μL of anti-cTnI (diluted to 1 mg / mL), invert and mix five times, let stand at room temperature for 30 minutes, place the labeled reaction tube on a shaker at 2-8°C and mix overnight, remove and add 50% glycerol and store in a refrigerator at -20°C.

[0070] (3) Luminescence auxiliary agent:

[0071] Dissolve 1.82 g of citric acid and 10.45 g of sodium citrate in purified water and dilute to 1000 mL. Add 0.3 mL of 30% hydrogen peroxide to the buffer solution, mix well, and place in a 4°C refrigerator for later use (divide into 10 mL / bottle).

[0072] (4) Trigger:

[0073] Weigh 6.06 g of tris (hydroxymethyl)aminomethane hydrochloride (Tris) and 9 g of NaCl, dissolve them in appropriate amount of purified water, add 2.1 mL of concentrated HCl and 2 mL of Tween 20, mix well, and adjust the volume to 1000 mL. Place in a 4°C refrigerator until ready for use.

[0074] (5) Calibrators:

[0075] ① Calibrator diluent: Weigh 14.1 g of potassium dihydrogen phosphate and 3.0 g of sodium dihydrogen phosphate dihydrate, dissolve them in an appropriate amount of purified water, add 0.5 mL of Proclin 300, mix well, and dilute to 1000 mL with purified water. Place in a 4°C refrigerator until ready for use.

[0076] ② Dilute recombinant cTnI to 0.02 ng / mL, 0.078 ng / mL, 0.313 ng / mL, 1.25 ng / mL, 5 ng / mL, 20 ng / mL, and 50 ng / mL with calibrator diluent and place in a 4°C refrigerator until use.

[0077] 3. Detection method:

[0078] The system requires a sample volume of 10 μL per measurement. Before applying for a test, you should prepare all the materials required for the test and carefully read the chemiluminescence immunoassay user manual to obtain information on system operating procedures, sample management, usage precautions, maintenance, and other information. The main steps for applying for a test are as follows:

[0079] (1) Calibration:

[0080] Perform a calibration test using the included kit and calibrator. Before starting the kit calibration, scan the master curve in the kit's 2D barcode and import it into the system.

[0081] The chemiluminescence immunoassay analyzer uses the results of calibrator tests to adjust the master curve and generate the calibration curve for the current system. The instrument operating software includes a calibration check function that automatically verifies the validity of the calibration curve. If calibration fails, consider the instrument settings, status, location of the calibrators (place low-value calibrators first, then high-value calibrators), and expiration dates.

[0082] A valid calibration curve is required before all tests. Recalibration is required in the following situations:

[0083] A. Use a new batch of test kit;

[0084] B. The same batch number of test kits has been used on the analyzer for more than 30 days;

[0085] C. After instrument maintenance;

[0086] D. The quality control value exceeds the specified range;

[0087] (2) Quality Control:

[0088] To ensure the reliability of test results, it is recommended that both high and low level quality control products be tested once every 24 hours. In addition, quality control testing is also recommended after each calibration test, reagent batch change, maintenance and troubleshooting.

[0089] The test values of quality control products should be within the specified range. If they are outside the specified range, the user should check the detection system, such as the location, expiration date, storage method, calibration process, instrument performance and status, and recalibrate if necessary. Sample testing can only be carried out after the quality control is qualified.

[0090] (3) Sample testing:

[0091] ① Enter the sample application interface and enter the sample number, sample rack number, location number, sample type, remarks and other information according to the test requirements;

[0092] ② Select the test item, select the sample type in the item options, enter the number of repetitions and other information;

[0093] ③ After preparing the test sample and placing it correctly, click the start button to start the test. The chemiluminescence immunoassay analyzer will perform the following operations in sequence:

[0094] A. The sample handling system transfers the sample rack to the sample aspiration position;

[0095] B. The cuvette loading and scheduling system loads the cuvette from the loading area to the sample loading position;

[0096] C. The sample injection system completes the sample injection and cleaning of the sample needle;

[0097] D. The reagent processing system provides the reagents required for the test, mixes each reagent, and sends it to the reagent aspiration position for aspiration;

[0098] E. The sample injection system completes the reagent injection and cleaning of the reagent needle;

[0099] F. Mix the reaction mixture with the counter-liquid mixing system and incubate in the reaction plate;

[0100] G. After incubation is complete, inject substrate solution into the substrate system, mix, incubate, and wait for light detection;

[0101] H. The light measurement reaction system sends each reaction cup to the signal collection position to collect signals for calculating the luminescence value;

[0102] I. Calculate the amount of analyte in the sample.

[0103] (4) Calculation

[0104] Using stored calibration data, the system software automatically determines the sample test results using a weighted four-parameter logarithmic curve (4PLC, Y-weighted) mathematical method. Results are given in pg / mL.

[0105] 4. Comparison reagents:

[0106] Beijing Regen Biotechnology Co., Ltd. High-Sensitivity Troponin I Assay Kit (Magnetic Particle Chemiluminescence Immunoassay), Beijing Medical Device Registration No. 20172400298.

[0107] 5. Results

[0108] Detection of 50 samples from patients with infectious diseases, quantitative correlation coefficient R with control reagent 2 =0.9963( Figure 1 and Table 1 ).

[0109] Table 1

[0110]

[0111]

[0112] according to Figure 1 As shown in Table 1, the two are well correlated, and the yin-yang coincidence rate is high.

[0113] (2) Blank detection limit

[0114] Using PBS buffer as a blank sample, the detection and comparison reagents prepared by the present invention were simultaneously measured 25 times. The test and comparison values from the test strips prepared by the present invention, plus twice the standard deviation, were substituted into the fitted curve for regression. The blank detection limit for the test kit prepared by the present invention was 0.015 ng / mL, and the blank detection limit for the comparison reagent was 0.016 ng / mL. The results are shown in Table 2.

[0115] Table 2

[0116]

[0117]

[0118] (3) Accuracy

[0119] A high-concentration reference substance (Solution A) was added to a low-concentration reference substance (Solution B), with the volume ratio between the high-concentration reference substance (Solution A) and the low-concentration reference substance (Solution B) being no greater than 1:9. Each sample was measured three times and the average value was obtained. Calculated according to formula (1), the recovery rate of the detection preparation method prepared by the present invention was 99.3%, indicating good accuracy. The results are shown in Table 3.

[0120]

[0121] Where: R is the recovery rate; V is the volume of solution A added; V0 is the volume of serum sample B; c is the detection concentration of serum sample after adding solution A; c0 is the detection concentration of serum sample B; c s is the concentration of solution A.

[0122] Table 3

[0123]

[0124] In summary, the spatial proximity chemical double antibody sandwich method adopted by the kit of the present invention is simple to operate and can be fully automatically detected compared to other methods such as enzyme-linked immunosorbent assay and fluorescent immunochromatography. It does not require magnetic beads and washing, which not only reduces reagent costs but also shortens detection time. In addition, it adopts a homogeneous reaction, avoiding the influence of traditional chemiluminescence magnetic separation technology, making the detection results more accurate and stable.

[0125] Although the embodiments of the present invention have been shown and described above, it will be understood that the above embodiments are illustrative and are not to be construed as limitations on the present invention. A person skilled in the art may change, modify, replace and modify the above embodiments within the scope of the present invention.

Claims

1. A spatial proximity chemiluminescence kit for detecting troponin I, characterized in that: The kit comprises an enzyme marker, a luminescent marker, a luminescent auxiliary agent and a trigger; the enzyme marker comprises an anti-cTnI monoclonal antibody marked with horseradish peroxidase, and the luminescent marker comprises an anti-cTnI monoclonal antibody marked with 9,10-dihydroacridine.

2. A spatial proximity chemiluminescence kit for detecting troponin I according to claim 1, characterized in that, The enzyme marker also includes bovine serum albumin, sucrose and trehalose; The concentration of the horseradish peroxidase-labeled anti-PGⅠ monoclonal antibody in the enzyme marker is 1 mg / ml; The mass concentration of bovine serum albumin in the enzyme marker is 0.5% to 2%; The mass concentration of sucrose in the enzyme marker is 2% to 5%; The mass concentration of trehalose in the enzyme marker is 1% to 3%.

3. A spatial proximity chemiluminescence kit for detecting troponin I according to claim 1, characterized in that: The luminescent marker also includes casein, sucrose and trehalose; The concentration of the 9,10-dihydroacridine-labeled anti-PGⅠ monoclonal antibody in the luminescent marker is 0.5 mg / ml; The mass concentration of casein in the luminescent marker is 0.1% to 1%; The mass concentration of sucrose in the luminescent marker is 2% to 5%; The mass concentration of trehalose in the luminescent marker is 1% to 3%.

4. A spatial proximity chemiluminescence kit for detecting troponin I according to claim 1, characterized in that: Each 1L of the luminescence auxiliary agent includes the following components in the following amounts: 1.5g to 2g of citric acid, 5g to 11g of sodium citrate, and 0.15mL to 0.45mL of 30% hydrogen peroxide.

5. A spatial proximity chemiluminescence kit for detecting troponin I according to claim 1, characterized in that: Each 1L of the trigger comprises the following components: 3g to 9g of tris(hydroxymethyl)aminomethane hydrochloride, 6g to 12g of NaCl, 1mL to 4mL of concentrated HCl, and 1mL to 4mL of a surfactant; The mass content of the concentrated HCl is 36% to 38%.

6. A spatial proximity chemiluminescence kit for detecting troponin I according to claim 5, characterized in that: The surfactant includes Tween.

7. A method for preparing a kit for detecting troponin I by a spatial proximity chemiluminescence method according to claims 1 to 6, characterized in that: The steps include: The enzyme marker was prepared using horseradish peroxidase and anti-cTnI monoclonal antibody; A luminescent marker was prepared using 9,10-dihydroacridine and anti-cTnI monoclonal antibody; A luminescence auxiliary agent is prepared by using citric acid, sodium citrate and hydrogen peroxide solution; The trigger was prepared using tris(hydroxymethylaminomethane) hydrochloride, NaCl, concentrated HCl and a surfactant.

8. The method for preparing a spatial proximity chemiluminescence kit for detecting troponin I according to claim 7, wherein: The mass ratio of the horseradish peroxidase to the anti-cTnI monoclonal antibody is 1:

2.

9. The method for preparing a spatial proximity chemiluminescence kit for detecting troponin I according to claim 7, wherein: The mass ratio of the 9,10-dihydroacridine to the anti-cTnI monoclonal antibody is 1:

3.

10. The method for preparing a spatial proximity chemiluminescence kit for detecting troponin I according to claim 7, wherein: The preparation of the luminescence auxiliary agent includes the following specific steps: taking citric acid, sodium citrate and hydrogen peroxide solution, dissolving them in purified water and adjusting the volume to obtain the luminescence auxiliary agent; The preparation of the trigger comprises the following specific steps: weighing tris(hydroxymethyl)aminomethane hydrochloride), NaCl, concentrated HCl and a surfactant, dissolving them and then fixing the volume to obtain the trigger.

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