Urine protein marker for asthma and application thereof
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A protein, asthma technology, applied in the field of clinical medicine
Inactive Publication Date: 2021-01-05
BEIJING NORMAL UNIVERSITY
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[0003] There is currently no precise diagnostic test for asthma, usually based on the type of symptoms, response to treatment over time, and spirometry. For details, see Unknown Author, Chinese Guidel
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Embodiment 1
[0042]Example 1. Establishment of asthma model
[0043]Take 6-8 weeks old male C56BL / 6 mice, 3 mice in the experimental group and 6 mice in the control group. On day 0 and day 7, intraperitoneal injection of 200uL emulsion (containing OVA: 50ug; Alum: 2mg) was sensitized ; After sensitization, drip the nose from the 14th day to the 16th day, anesthetize the mice with ether and then drip 50ul (including OVA: 50ug). In this example, the inventors injected ovalbumin (OVA) and nasal drops into mice to create a model to simulate asthma. Urine was collected for analysis in the early, early and late stages of asthma.
[0044]1. Materials and reagents
[0045]1) Instrument:
[0046]Orbitrap Fusion Lumos Tribird mass spectrometer was purchased from Thermo Scientific; mouse metabolic cage: purchased from Beijing Jiayuan Xingye Technology Co., Ltd.
[0047]2) Main reagents:
[0048]Chromatography mass spectrometry grade water, acetonitrile, formic acid and methanol were purchased from Fisher; Iodoacetylammonium...
Embodiment 2
[0054]Example 2. Urine sample collection and protein identification
[0055]1. Urine sample collection:
[0056]After modeling, the experimental group and control group mice were placed in metabolic cages to collect urine on the 2, 7, 10, 14, and 17 days.
[0057]2. Extraction of protein in urine by ethanol precipitation
[0058]A total of 15 urine samples from 3 mice on days 2, 7, 14, 17 and 3 control mice were taken to extract protein for subsequent analysis. The method of ethanol precipitation of urine protein refers to the method of Sun W et al. (see Sun W et al. Proteomics, 2005, 5(18):4994-5001).
[0059]The specific steps are:
[0060]1) Take the mouse urine stored in the refrigerator at -80℃, thaw it at 37℃, centrifuge the urine after thawing: take 0.4mL, 12000g, 4℃, centrifuge for 10min; save the supernatant.
[0061]2) Precipitating protein: Add pre-cooled ethanol (1:4) to the retained supernatant, the total volume is about 2 mL.
[0062]3) In the refrigerator at 4°C overnight (-20°C, 2h).
[0063]4...
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Abstract
The invention relates to a urine protein marker for asthma and application thereof. The present application relates to the application of a urine protein marker selected from a group consisting of epidermal growth factor receptor substrate 15, Cathelicidin antimicrobial peptide, haptoglobin, alpha-1-acidic glycoprotein 1, prolactin-inducing protein homolog, odorant binding protein 2a, protein LEG1homolog, complement factor B, retinoid beta, beta-2 microglobulin, prothrombin, eosinophilic granulocyte cationic protein 1, protein AMBP, plasminogen, a procollagen C-endopeptidase enhancer 1, a sodium channel and gridding protein connecting agent 1 and vitamin D binding protein. According to the application, an animal model for screening the asthma-related urine protein marker is established, the asthma-related urine protein marker is obtained by using the established animal model, and an application of a detection reagent of the marker in preparation of a kit for diagnosing asthma of a subject is also established.
Description
Technical field[0001]The present invention relates to the field of clinical medicine; specifically, the present invention relates to a urine protein marker related to asthma. Specifically, the present invention relates to obtaining urine protein markers related to asthma by using a mouse model of asthma and mass spectrometry urine proteomics technology, and the use of the urine protein markers.Background technique[0002]Asthma is a chronic inflammatory airway disease caused by a combination of genetic and environmental factors. See Martinez F.D. Genes, environments, development and asthma: a reappraisal. European Respiratory Journal. 2007.29(1): 179-184. It is characterized by airway inflammation leading to restricted expiratory airflow, the intensity of which changes over time. Symptoms include wheezing, shortness of breath, chest tightness, and cough, see GINA.Global strategy for asthma management and prevention.Global Initiative for Asthma.2017.The updated asthma guidelines from G...
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