Use of Angpt2 as a diagnostic marker in the preparation of a diagnostic reagent for diagnosing acute mountain sickness

Abstract

The present invention provides the use of angiopoietin-2 (Angpt2) as a diagnostic marker in the preparation of a diagnostic reagent or kit for diagnosing acute mountain sickness. The present invention discovers that plasma Angpt2 is positively correlated with the Lake Louise score of acute altitude reaction and is also positively correlated with the severity of the patient. Therefore, Angpt2 can be used as a specific diagnostic marker for acute altitude reaction. By detecting patients using the change of the protein (Angpt2) in plasma, patients with acute altitude reaction can be identified early, the success rate of diagnosis can be significantly improved, and the disease severity can be judged.

Description

Technical Field

[0001] The present invention belongs to the technical field of disease diagnosis, and particularly relates to the application of Angpt2 as a diagnostic marker in the preparation of a diagnostic reagent or kit for diagnosing acute mountain sickness. Background Art

[0002] With the development of the economy in the northwest and the construction of the "Belt and Road Initiative", more and more tourists and plateau workers enter the plateau area from the plain. Acute altitude sickness often threatens the activities of these people, and even their life and health. Acute altitude reaction is the most common acute altitude sickness, which refers to a series of non-specific clinical manifestations that occur when plain residents suddenly enter the plateau above 2500 meters from the plain, or when they suddenly enter an area with a higher altitude from the plateau. At an altitude of 2500 meters, about 10 - 25% of the people who suddenly enter will have acute altitude reaction, and at an altitude of 4500 - 5500 meters, about 50 - 85% of the people who suddenly enter will have acute altitude reaction. Above an altitude of 2500 meters, for every 1000-meter increase in altitude, the incidence of acute altitude reaction increases by 13%. The symptoms of acute altitude reaction are very atypical, mainly including fatigue, headache, dizziness, sleep disorder, loss of appetite, nausea, vomiting, etc.

[0003] Regarding the diagnosis of acute altitude reaction, the most commonly used diagnostic system internationally at present is the Lake Louise Score, and China has also applied its own military standard (GJB - 1098 - 91), but both rely on the symptoms of patients, with low sensitivity and lack of specificity. Since most patients currently have no obvious abnormalities in routine laboratory tests and examinations, in the case of no characteristic laboratory or examination indicators for specific diagnosis, missed diagnosis and misdiagnosis are very likely to occur. Moreover, 1% of patients below an altitude of 5000 meters will progress to life-threatening high altitude cerebral edema.

[0004] Therefore, exploring specific diagnostic markers for acute altitude reaction, improving the diagnostic rate, and intervening early are of great significance for alleviating altitude reaction, preventing complications, and saving the lives of patients. Summary of the Invention

[0005] Angiopoietin - 2 (Angpt2) is a secreted growth factor, which is related to inflammatory response and angiogenesis. An increase in its expression level indicates the activation of endothelial cells and the imbalance of homeostasis. Angpt2 can be used as a diagnostic marker for acute altitude reaction, assisting in scale assessment, improving the diagnosis and treatment rate of acute altitude reaction, and reducing the occurrence of complications.

[0006] The present invention provides the application of Angiopoietin - 2 (Angpt2) as a diagnostic marker in the preparation of a diagnostic reagent or kit for diagnosing acute mountain sickness.

[0007] In one embodiment according to the present invention, the acute mountain sickness is selected from any one of acute altitude reaction, high altitude pulmonary edema, and high altitude cerebral edema.

[0008] In one embodiment according to the present invention, the diagnostic reagent or kit comprises an antibody against Angpt2 protein.

[0009] In one embodiment according to the present invention, the antibody against Angpt2 protein is conjugated with a fluorescent probe or a biotin probe.

[0010] In one embodiment according to the present invention, the Angpt2 protein is the Angpt2 protein in plasma.

[0011] In one embodiment according to the present invention, the optimal cut-off value of Angpt2 in the diagnostic reagent or kit is 1.79 ng / ml.

[0012] The beneficial effects of the above technical solutions of the present invention are as follows:

[0013] Aiming at the problems of low sensitivity and specificity of existing diagnostic methods, the present invention provides a diagnostic marker that can specifically identify acute mountain sickness, especially for diagnosing patients with acute altitude reaction when there are no obvious abnormalities in routine laboratory tests and imaging examinations. Specifically, the present invention can detect the concentration of the protein angiopoietin 2 (Angpt2) in plasma, identify patients with acute altitude reaction at an early stage, significantly improve the success rate of diagnosis, and judge the severity of the disease. Brief Description of the Drawings

[0014] Figure 1 It is the change map of mouse plasma Angpt2 after acute hypobaric hypoxia in Example 2 of the present invention;

[0015] Figure 2 It is the change map of the content of Angpt2 in mouse brain tissue after acute hypobaric hypoxia;

[0016] Figure 3 It is the change map of the transcriptional level of Angpt2 in mouse brain tissue after acute hypobaric hypoxia;

[0017] Figure 4 It is the change map of Angpt2 in the endothelial cell culture medium after acute hypoxia;

[0018] Figure 5 It is the change map of the transcriptional level of Angpt2 in endothelial cells after acute hypoxia;

[0019] Figure 6 It is the change map of the content of Angpt2 in endothelial cells after acute hypoxia.

[0020] Figure 7Image of the rapid release of Angpt2 by endothelial cells through exocytosis after acute hypoxia

[0021] Figure 8 Plasma Angpt2 change atlas of patients with acute mountain sickness

[0022] Figure 9 Correlation atlas of plasma Angpt2 and Lake Louise score

[0023] Figure 10 ROC curve graph Specific implementation manners

[0024] To make the technical problems, technical solutions and advantages to be solved by the present invention clearer, the following will be described in detail with reference to the accompanying drawings and specific embodiments

[0025] Test materials and reagents

[0026]

[0027] Example 1

[0028] Detection of Angpt2 protein levels in plasma and brain tissue

[0029] 1. Use a small animal low-pressure hypoxia chamber to construct an animal model of acute low-pressure hypoxia (7000 m, for 24 hours). Control group mice are placed in an environment of normal pressure and normal oxygen, with other conditions being the same

[0030] 2. Sample collection

[0031] Anesthetize the mice with 1% sodium pentobarbital (50 mg / Kg). After anesthesia, fix the mice on the dissection board, collect blood from the inferior vena cava (anticoagulated with sodium heparin), immediately centrifuge (centrifugation conditions: 2000 g, 20 minutes, 4 °C), and retain the supernatant (plasma specimen)

[0032] 3. Expose the thoracic cavity of the mice and perfuse the left cardiac apex (40 ml of normal saline, 4 °C)

[0033] 4. Decapitate the mice, carefully isolate the brain tissue, and remove the olfactory bulb and cerebellum

[0034] 5. Weigh the brain tissue, add PBS according to the ratio of brain tissue weight (mg): PBS (phosphate buffered saline, ml) = 1:9, ultrasonically grind until there are no tissue lumps, let stand on ice for 30 minutes, and centrifuge (14000 g, 20 minutes, 4 °C)

[0035] 6. Take the supernatant and perform BCA protein quantification

[0036] 7. Perform quantitative ELISA detection of mouse Angpt2 (R&D, USA, MANG20)

[0037] 1) Reagent preparation: All reagent specimens are restored to room temperature before use.

[0038] 2) Prepare the washing solution: Add 20 ml of the washing solution to 480 ml of deionized water to prepare 500 ml of the washing solution.

[0039] 3) Prepare the standard:

[0040] Add the required deionized water to the standard, gently invert it up and down to mix well, and let it stand for at least 15 minutes to make the concentration of the stock solution of the standard 25000 pg / mL. Add 50 μl of the standard stock solution to 450 μl of the detection diluent RD5-26 (RD5-26 needs to be diluted 4 times with deionized water in advance) to prepare a solution with a concentration of 2500 pg / mL, and dilute the standard in gradient.

[0041] 4) Sample dilution:

[0042] Dilute the specimens with the diluted RD5-26. Dilute the plasma 40 times and the brain tissue 10 times according to the instructions, and gently shake to mix well.

[0043] 5) Prepare the plate and add 50 μl of the diluent RD1-40 to each well.

[0044] 6) Add 50 μl of the standard, control and samples, stick on the sealing film, and incubate at room temperature on a horizontal rotary shaker for 2 hours (500 rpm ± 50 rpm).

[0045] 7) Wash the plate (plate washer), 400 μl / well, let it stand for 1 minute, for a total of 4 times, and aspirate each time.

[0046] 8) Add 100 μl of the mouse Angpt2-binding enzyme, cover with a new sealing film, and incubate on a room temperature shaker for 2 hours (500 rpm ± 50 rpm).

[0047] 9) Wash the plate, same as step 7).

[0048] 10) Add 100 μl of the freshly prepared substrate solution (50 μl of solution A and 50 μl of solution B), incubate at room temperature for 30 minutes in the dark.

[0049] 11) Add 100 μl of the termination solution, the color changes from blue to yellow, and gently tap to mix well.

[0050] 12) Detect with an enzyme-linked immunosorbent assay reader within 30 minutes, and set the dual wavelength to 450 / 540 nm.

[0051] 13) The results are as Figure 1 、 Figure 2 shown, Figure 1 which shows that the plasma Angpt2 in mice increased significantly after acute hypobaric hypoxia;Figure 2 It was shown that the content of Angpt2 in the brain tissue of mice increased significantly after acute hypobaric hypoxia. Compared with the control group, the concentration of Angpt2 in the plasma (as Figure 1 shown) and brain tissue (as Figure 2 shown) of mice increased significantly after acute hypobaric hypoxia.

[0052] Detection of the transcriptional level of Angpt2 in brain tissue

[0053] 1. Use a small animal hypobaric hypoxia chamber to construct an animal model of acute hypobaric hypoxia (7000 m, for 24 hours). The control group of mice was placed in a normal pressure and normal oxygen environment with the same other conditions.

[0054] 2. Sample collection:

[0055] Anesthetize the mice with 1% sodium pentobarbital (50 mg / Kg). After anesthesia, fix the mice on the dissection board, collect blood from the inferior vena cava (anticoagulated with sodium heparin), immediately centrifuge (centrifugation conditions: 2000 g, for 20 minutes, 4 °C), and collect the supernatant (plasma specimen).

[0056] 3. Expose the thoracic cavity of the mice and perfuse the left cardiac apex (40 ml of normal saline, 4 °C).

[0057] 4. Decapitate the mice, carefully separate the brain tissue, and remove the olfactory bulb and cerebellum.

[0058] 5. Place the brain tissues in RNase-free EP tubes (operate on ice).

[0059] 6. Add 500 μl of Buffer RL to each EP tube.

[0060] 7. Use an enzyme-free pestle soaked in diethyl pyrocarbonate (DEPC) water to crush the brain tissue until it is transparent in color, without obvious tissue clumps, and the texture is jelly-like.

[0061] 8. Transfer the lysed sample to FastPure gDNA-Filter ColumnsⅢ (FastPure gDNA-Filter ColumnsⅢ has been placed in the collection tube), and centrifuge at 12000 rpm for 30 sec. Discard FastPure gDNA-FilterColumnsⅢ and collect the filtrate.

[0062] 9. Add anhydrous ethanol with a volume 0.5 times that of the filtrate to the filtrate and mix well.

[0063] 10. Transfer all the mixture in step 5 to FastPure RNA ColumnsⅢ (FastPure RNA ColumnsⅢ has been placed in the collection tube), centrifuge at 12,000 rpm for 30 sec, and discard the filtrate.

[0064] 11. Add 700 μl of Buffer RW1 to FastPure RNA ColumnsⅢ, centrifuge at 12,000 rpm for 30 sec, and discard the filtrate.

[0065] 12. Add 700 μl of Buffer RW2 (pre - added with absolute ethanol) to FastPure RNA ColumnsⅢ, centrifuge at 12,000 rpm for 30 sec, and discard the filtrate.

[0066] 13. Add 500 μl of Buffer RW2 (pre - added with absolute ethanol) to FastPure RNA ColumnsⅢ, centrifuge at 12,000 rpm for 2 min. Carefully remove the adsorption column from the collection tube, avoiding contact with the filtrate to prevent contamination. (If there is liquid residue on the adsorption column or it contacts the filtrate, discard the filtrate, put FastPure RNA ColumnsⅢ back into the collection tube, and centrifuge at 12,000 rpm for 1 min to prevent ethanol contamination.)

[0067] 14. Carefully transfer the adsorption column to a new 1.5 - ml RNase - free Collection Tube. Drop 50 - 200 μl of RNase - free ddH2O (pre - heated at 65℃) onto the center of the adsorption column, let it stand at room temperature for 5 min, and centrifuge at 12,000 rpm for 1 min to elute the RNA.

[0068] 15. Detection: Pipette 1 μl of the filtrate to detect the total RNA concentration.

[0069] 16. Preparation of cDNA library:

[0070] 1) Dilute the RNA to a concentration of 0.1 μg / μl.

[0071] 2) Prepare the reverse transcription system (total volume of 20 μl): Total RNA: 2 μl; SuperMix: 4 μl; gDNA Remover 1 μl; Enzyme - free water: 13 μl.

[0072] 3) Gently mix and incubate at 42℃ for 15 minutes.

[0073] 4) Inactivate gDNA Remover by heating at 85℃ for 5 seconds.

[0074] 5) Pipette 1 μl of the specimen to measure the cDNA content.

[0075] 17. q - PCR biological verification shows that the transcriptional level of Angpt2 in the brain tissue increases after acute hypobaric hypoxia exposure

[0076] 1) Adjust the cDNA to 0.1 μg / μl.

[0077] 2) Primer design: The internal reference gene is GAPDH. The primer designs for GAPDH and Angpt2 are as follows

[0078] Angpt2 forward: G G A C A G T C A T C C A A C A C C G A G

[0079] Angpt2 reverse: G A C T C T T C A C C A G C G A G G T A G

[0080] GAPDH forward: A G G T C G G T G T G A A C G G A T T T G

[0081] GAPDH reverse: G G G G T C G T T G A T G G C A A C A

[0082] 3) Centrifuge the primers at 3000 rpm for 1 minute. Add enzyme-free water according to the recommended volume, and mix well by shaking. At this time, the primer concentration is 100 μM.

[0083] 4) Mix the forward and reverse strands in advance at a 1:1 ratio.

[0084] 5) Prepare the qRT-PCR reaction system (20 μl volume, set up 3 replicates): Water: 6.8 μl; Dye: 0.4 μl; SuperMix: 10 μl; Primers: 0.8 μl; Template: 2 μl.

[0085] 6) Mix well and add to 4 wells of an 8-strip tube.

[0086] 7) Use forceps to remove the lid of the 8-strip tube, cover it with PV gloves (to avoid fluorescence contamination), tighten the lid, and centrifuge briefly to remove air bubbles.

[0087] 8) Run on the machine for detection: 50 °C: 2 minutes; 95 °C: 10 minutes; 95 °C: 15 seconds, 60 °C: 30 seconds, 40 cycles; 95 °C: 15 seconds; 60 °C: 1 minute; 95 °C: 15 seconds.

[0088] 9) Calculate the ΔΔCT value.

[0089] 10) The results are as Figure 3 shown that the transcriptional level of Angpt2 in the mouse brain tissue increased significantly after acute hypobaric hypoxia.

[0090] Example 2:

[0091] 1. Use human umbilical vein endothelial cells (HUVEC) to construct an in vitro acute hypoxia model. Place the cells (seeded in 24-well plates and 6-well plates) in a triple-gas incubator (2% O2 , 5% CO 2 , 93% N 2 , (37 °C), and were subjected to hypoxic treatment for 0, 2, 4, 8, and 20 hours respectively.

[0092] 2. After the hypoxic treatment was completed, the cells were taken out in sequence, and immunofluorescence staining of cell Angpt2 and protein concentration detection of cell culture medium and lysate were performed respectively.

[0093] 3. Angpt2 immunocytofluorescence staining

[0094] 1) Discard the supernatant, add 500 μl of PBS to each well, and wash 2 times.

[0095] 2) Add 4% PFA and fix at room temperature for 20 minutes.

[0096] 3) Discard 4% PFA, add 500 μl / well of PBS, and wash 3 times.

[0097] 4) Add 400 μl / well of 1% BSA and block at room temperature for 1 hour.

[0098] 5) Add the primary antibody (mouse anti-Angpt2) 250 μl / well and incubate overnight at 4 °C.

[0099] 6) Take out the cells, recover the primary antibody, add 500 μl / well of PBS, and wash 3 times.

[0100] 7) Add the secondary antibody (1:1000, goat anti-mouse 488) 250 μl / well and incubate at room temperature for 1 hour.

[0101] 8) Discard the secondary antibody, add 500 μl / well of PBS, and wash 3 times.

[0102] 9) Add DAPI (1:10000) 250 μl / well and incubate at room temperature for 5 mins.

[0103] 10) Discard DAPI, add 500 μl / well of PBS, and wash 3 times.

[0104] 11) Add an anti-fluorescence quenching mounting medium to mount the slides, and seal the slides with nail polish.

[0105] 12) Laser confocal microscopy imaging, as Figure 6 shown.

[0106] 4. Cell protein extraction and BCA quantification

[0107] 1) Prepare 2 EP tubes for each specimen and make marks.

[0108] 2) Prepare the lysis buffer.

[0109] 3) Recover the supernatant, add 100 μl of lysis buffer, pipette to lyse the cells until the lysis buffer becomes jelly-like, transfer it into a labeled EP tube, and let it stand on ice for 30 minutes.

[0110] 4) Centrifuge at 13000 rpm for 15 minutes at 4 °C.

[0111] 5) Retain the supernatant.

[0112] 6) Perform BCA protein quantification.

[0113] 5. Detection of Angpt2 in cell culture medium and lysis buffer using ELISA kit (R&D, USA, DANG20):

[0114] 1) Reagent preparation:

[0115] Restore all reagent specimens to room temperature before use.

[0116] 2) Prepare the washing solution:

[0117] Add 20 ml of washing solution to 480 ml of deionized water to prepare 500 ml of washing solution.

[0118] 3) Prepare the standard:

[0119] Add 5 ml of deionized water to the standard, gently invert it up and down to mix well, let it stand for at least 15 mins to make the concentration of the stock solution of the standard 30000 pg / mL. Add 50 μl of the standard stock solution to 450 μl of detection diluent (RD5-5) to prepare a solution with a concentration of 3000 pg / mL, and dilute the standard in gradient.

[0120] 4) Sample dilution:

[0121] Dilute 5 times. Add 20 μl of the sample to 40 μl of detection diluent (RD5-5), and gently shake to mix well.

[0122] 5) Prepare the plate, and add 100 μl of diluent RD1-76 to each well.

[0123] 6) Add 50 μl of the standard, control, and sample, stick on the sealing film, and incubate at room temperature on a horizontal rotary shaker for 2 hours (500 rpm ± 50 rpm).

[0124] 7) Wash the plate (using a plate washer), 400 μl / well, let it stand for 1 min, for a total of 4 times, and aspirate each time.

[0125] 8) Add 200 μl of human Angpt2 conjugate enzyme, cover with a new sealing film, and incubate on a room temperature shaker for 2 hours (500 rpm ± 50 rpm).

[0126] 9) Wash the plate, same as step 7).

[0127] 10) Add 200 μl of freshly prepared bottom solution (100 μl of Solution A and 100 μl of Solution B), incubate at room temperature for 30 minutes in the dark.

[0128] 11) Add 50 μl of stop solution, the color changes from blue to yellow, gently tap to mix evenly.

[0129] 12) Detect with an enzyme-linked immunosorbent assay (ELISA) reader within 30 minutes, set the dual wavelength to 450 / 540 nm.

[0130] 6. Detection of the q-PCR transcriptional level of cell lysate Angpt2

[0131] 1) Add 500 μl of lysis solution to each well, let it stand for 3 minutes, transfer the sample to FastPure gDNA-FilterColumnsⅢ (FastPure gDNA-Filter ColumnsⅢ has been placed in the collection tube), centrifuge at 12,000 rpm for 30 seconds. Discard FastPure gDNA-Filter ColumnsⅢ and collect the filtrate.

[0132] 2) Add anhydrous ethanol with a volume 0.5 times that of the filtrate to the filtrate and mix well.

[0133] 3) Transfer all of the mixture from step 5 to FastPure RNA ColumnsⅢ (FastPure RNA ColumnsⅢ has been placed in the collection tube), centrifuge at 12,000 rpm for 30 seconds, and discard the filtrate.

[0134] 4) Add 700 μl of Buffer RW1 to FastPure RNA ColumnsⅢ, centrifuge at 12,000 rpm for 30 seconds, and discard the filtrate.

[0135] 5) Add 700 μl of Buffer RW2 (anhydrous ethanol has been added) to FastPure RNA ColumnsⅢ, centrifuge at 12,000 rpm for 30 seconds, and discard the filtrate.

[0136] 6) Add 500 μl of Buffer RW2 (anhydrous ethanol has been added) to FastPure RNA ColumnsⅢ, centrifuge at 12,000 rpm for 2 minutes, carefully remove the adsorption column from the collection tube, avoiding contact with the filtrate to cause contamination. (If there is liquid residue in the adsorption column or it comes into contact with the filtrate, discard the filtrate, put FastPure RNA ColumnsⅢ back into the collection tube, and centrifuge at 12,000 rpm for 1 minute to prevent ethanol contamination.)

[0137] 7) Carefully transfer the adsorption column to a new 1.5 ml centrifuge tube of RNase-free Collection Tubes. Drop 50 - 200 μl of RNase-free ddH2O (preheated at 65 °C) onto the center of the adsorption column in a suspended manner, let it stand at room temperature for 5 min, and centrifuge at 12,000 rpm for 1 min to elute the RNA.

[0138] 8) Detection: Pipette 1 μl of the filtrate to detect the total RNA concentration.

[0139] 9) Preparation of the cDNA library (the same method as above)

[0140] 10) q-PCR biological verification shows that the transcriptional level of Angpt2 in the brain tissue increases after acute hypobaric hypoxia exposure (the same method as above)

[0141] 11) The results part is as Figures 4 - 7 shown:

[0142] Figure 4 It shows that Angpt2 in the endothelial cell culture medium increases significantly after acute hypoxia.

[0143] Figure 5 It shows that the transcriptional level of Angpt2 in endothelial cells increases significantly after acute hypoxia.

[0144] Figure 6 It shows that the content of Angpt2 in endothelial cells increases significantly after acute hypoxia.

[0145] Figure 7 It shows that after acute hypoxia, endothelial cells rapidly release Angpt2 by exocytosis and rapidly produce it. It is found that endothelial cells rapidly release and produce Angpt2 after acute hypoxia.

[0146] Example 3:

[0147] 1. Collect plasma specimens from patients who have rapidly ascended to high altitude and seek medical treatment due to acute mountain sickness.

[0148] 2. Detect the plasma Angpt2 content using a quantitative ELISA kit for human Angpt2 (R&D, USA, DANG20):

[0149] 1) Reagent preparation:

[0150] All reagent specimens are restored to room temperature before use.

[0151] 2) Prepare the washing solution:

[0152] Add 20 ml of the washing solution to 480 ml of deionized water to prepare 500 ml of the washing solution.

[0153] 3) Prepare the standard product:

[0154] Add 5 ml of deionized water to the standard product, gently invert it up and down to mix well, and let it stand for at least 15 mins to make the concentration of the stock solution of the standard product 30000 pg / mL. Add 50 μl of the standard product stock solution to 450 μl of the detection diluent (RD5-5) to prepare a solution with a concentration of 3000 pg / mL, and dilute the standard product in gradients.

[0155] 4) Sample dilution:

[0156] Dilute 5 times. Add 20 μl of the sample to 40 μl of the detection diluent (RD5-5), and gently shake to mix well.

[0157] 5) Prepare the plate and add 100 μl of the diluent RD1-76 to each well.

[0158] 6) Add 50 μl of the standard product, control, and sample, stick on the sealing film, and incubate at room temperature on a horizontal rotary shaker for 2 hours (500 rpm ± 50 rpm).

[0159] 7) Wash the plate, 400 μl / well, let it stand for 1 min, for a total of 4 times, and aspirate each time.

[0160] 8) Add 200 μl of human Angpt2 binding enzyme, cover with a new sealing film, and incubate on a shaker at room temperature for 2 hours (500 rpm ± 50 rpm).

[0161] 9) Wash the plate, same as step 7).

[0162] 10) Add 200 μl of the freshly prepared bottom solution (100 μl of solution A and 100 μl of solution B), and incubate at room temperature for 30 minutes in the dark.

[0163] 11) Add 50 μl of the termination solution, the color changes from blue to yellow, and gently tap to mix well.

[0164] 12) Detect with an enzyme-linked immunosorbent assay reader within 30 minutes, and set the dual wavelength to 450 / 540 nm.

[0165] 13) The results are as Figures 8 - 9 shown:

[0166] Figure 8 It shows that the plasma Angpt2 in patients with acute mountain sickness increases significantly.

[0167] Figure 9 It shows that there is a positive correlation between plasma Angpt2 and the Lake Louise score. Thus, it is concluded that Angpt2 can be used for the clinical diagnosis and prediction of acute mountain sickness, that is: Angpt2 can be used as a diagnostic marker for patients with acute mountain sickness.

[0168] 3. According to the plasma Angpt2 levels of healthy people and patients with acute altitude reaction, draw an ROC curve (asFigure 10 As shown, it can be seen that AUC = 0.793, the sensitivity is 70%, the specificity is 86.7%, and the optimal cut-off value of Angpt2 is 1.79 ng / ml.

[0169] The above is the preferred embodiment of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle described in the present invention, several improvements and refinements can be made, and these improvements and refinements should also be regarded as the protection scope of the present invention.

Claims

1. Use of a reagent for detecting angiopoietin 2 (Angpt2) protein in plasma in the preparation of a kit for diagnosing acute mountain sickness; the acute mountain sickness is acute altitude reaction.

2. The use according to claim 1, wherein, the kit comprises an antibody against Angpt2 protein.

3. The use according to claim 2, wherein, the antibody against Angpt2 protein is conjugated with a fluorescent probe or a biotin probe.

4. The use according to claim 1, wherein, the optimal cut-off value of Angpt2 in the kit is 1.79 ng / ml.

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