CN111735961A - Acute respiratory distress syndrome detection kit - Google Patents
Acute respiratory distress syndrome detection kit Download PDFInfo
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- CN111735961A CN111735961A CN202010515456.7A CN202010515456A CN111735961A CN 111735961 A CN111735961 A CN 111735961A CN 202010515456 A CN202010515456 A CN 202010515456A CN 111735961 A CN111735961 A CN 111735961A
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- 206010001052 Acute respiratory distress syndrome Diseases 0.000 title claims abstract description 33
- 208000013616 Respiratory Distress Syndrome Diseases 0.000 title claims abstract description 32
- 201000000028 adult respiratory distress syndrome Diseases 0.000 title claims abstract description 32
- 238000001514 detection method Methods 0.000 title claims abstract description 30
- 102100040112 Tumor necrosis factor receptor superfamily member 10B Human genes 0.000 claims abstract description 42
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 35
- 210000002966 serum Anatomy 0.000 claims abstract description 34
- 238000002965 ELISA Methods 0.000 claims description 7
- 239000007788 liquid Substances 0.000 claims description 7
- 238000000034 method Methods 0.000 claims description 6
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 claims description 5
- 238000004811 liquid chromatography Methods 0.000 claims description 5
- 238000003745 diagnosis Methods 0.000 abstract description 9
- 238000005406 washing Methods 0.000 description 8
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 4
- 239000012528 membrane Substances 0.000 description 4
- 238000007789 sealing Methods 0.000 description 4
- 102100024598 Tumor necrosis factor ligand superfamily member 10 Human genes 0.000 description 3
- 239000007791 liquid phase Substances 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 2
- 230000006907 apoptotic process Effects 0.000 description 2
- 229960002685 biotin Drugs 0.000 description 2
- 235000020958 biotin Nutrition 0.000 description 2
- 239000011616 biotin Substances 0.000 description 2
- 239000003446 ligand Substances 0.000 description 2
- 239000011259 mixed solution Substances 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 238000002331 protein detection Methods 0.000 description 2
- 108020003175 receptors Proteins 0.000 description 2
- 102000005962 receptors Human genes 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 102000004091 Caspase-8 Human genes 0.000 description 1
- 108090000538 Caspase-8 Proteins 0.000 description 1
- 102000011727 Caspases Human genes 0.000 description 1
- 108010076667 Caspases Proteins 0.000 description 1
- 101000830565 Homo sapiens Tumor necrosis factor ligand superfamily member 10 Proteins 0.000 description 1
- 206010021143 Hypoxia Diseases 0.000 description 1
- 208000019693 Lung disease Diseases 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 206010037423 Pulmonary oedema Diseases 0.000 description 1
- 108010000449 TNF-Related Apoptosis-Inducing Ligand Receptors Proteins 0.000 description 1
- 102000002259 TNF-Related Apoptosis-Inducing Ligand Receptors Human genes 0.000 description 1
- 108700012411 TNFSF10 Proteins 0.000 description 1
- 101710097160 Tumor necrosis factor ligand superfamily member 10 Proteins 0.000 description 1
- 239000003150 biochemical marker Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 239000003593 chromogenic compound Substances 0.000 description 1
- 208000035850 clinical syndrome Diseases 0.000 description 1
- 239000002131 composite material Substances 0.000 description 1
- 231100000433 cytotoxic Toxicity 0.000 description 1
- 230000001472 cytotoxic effect Effects 0.000 description 1
- 230000003111 delayed effect Effects 0.000 description 1
- 238000012631 diagnostic technique Methods 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 208000018875 hypoxemia Diseases 0.000 description 1
- 230000000977 initiatory effect Effects 0.000 description 1
- 230000001404 mediated effect Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000006213 oxygenation reaction Methods 0.000 description 1
- 208000005333 pulmonary edema Diseases 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 210000004881 tumor cell Anatomy 0.000 description 1
- 238000001195 ultra high performance liquid chromatography Methods 0.000 description 1
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
- G01N2333/705—Assays involving receptors, cell surface antigens or cell surface determinants
- G01N2333/70578—NGF-receptor/TNF-receptor superfamily, e.g. CD27, CD30 CD40 or CD95
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/12—Pulmonary diseases
- G01N2800/125—Adult respiratory distress syndrome
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- General Health & Medical Sciences (AREA)
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- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention belongs to the field of acute respiratory distress syndrome detection. The invention provides application of a reagent for detecting DR5 protein in serum in preparing an acute respiratory distress syndrome detection kit. The invention also provides a corresponding acute respiratory distress syndrome detection kit. The invention can realize the rapid diagnosis of acute respiratory distress syndrome by detecting the content of DR5 protein DR5 protein in serum, and has good application prospect.
Description
Technical Field
The present invention is in the field of acute respiratory distress syndrome diagnosis.
Background
Acute Respiratory Distress Syndrome (ARDS) is a clinical syndrome characterized by refractory hypoxemia, caused by intrapulmonary and/or extrapulmonary causes, that is of great interest for high mortality.
The current standard for ARDS diagnosis is the berlin ARDS standard, which is complex and time consuming to diagnose ARDS in 4 aspects of onset time, oxygenation index, source of pulmonary edema and X-ray chest radiographs (see in particular "dongchun, tube east, criteria for acute respiratory distress syndrome diagnosis, journal of chinese medical practice, vol. 33, No. 11, 2013). Today, the condition of the patients with ARDS is delayed because the patients cannot be diagnosed in time due to the crowded medical resources. Therefore, developing an ARDS diagnostic tool that is less time consuming is of great practical significance.
Receptor 2, also known as death receptor 5(DR5), of the TNF-related apoptosis-inducing ligand, numbered O14763 in the UniProtKB database, is a receptor for the cytotoxic ligand TNFSF10/TRAIL, responsible for recruiting Caspase-8 and initiating Caspase-mediated apoptosis. Since DR5 can rapidly induce apoptosis after binding to ligand, researchers are mainly concerned about using this principle to specifically kill tumor cells.
The relationship between DR5 and lung diseases other than cancer has not been reported yet.
Disclosure of Invention
The invention aims to provide a kit for diagnosing ARDS.
The technical scheme of the invention comprises the following steps:
the application of the reagent for detecting the content of DR5 protein in serum in preparing an acute respiratory distress syndrome detection kit.
As the application, the reagent for detecting the DR5 protein content in the serum is an enzyme-linked immunosorbent assay reagent.
As mentioned above, the reagent for detecting the DR5 protein content in the serum is a reagent for liquid chromatography or a colloidal gold detection reagent.
As mentioned above, the reagent for detecting the DR5 protein content in the serum is R & D Systems Luminex liquid phase chip detection reagent.
An acute respiratory distress syndrome detection kit comprises a reagent for detecting the content of DR5 protein in serum.
As the detection kit, the reagent for detecting the DR5 protein content in the serum is an enzyme-linked immunosorbent assay reagent.
As the detection kit, the reagent for detecting the DR5 protein content in the serum is a reagent for a liquid chromatography method or a colloidal gold detection reagent.
As the detection kit, the reagent for detecting the content of the DR5 protein in the serum is an R & D systems Luminex liquid phase chip detection reagent.
The inventor finds that the content of DR5 protein in serum of patients with ARDS is significantly different from that of healthy people, and the content of DR5 protein in serum of patients with ARDS is far higher than that of healthy people. The DR5 protein can be detected by conventional protein detection methods such as liquid chromatography (such as high performance liquid chromatography, ultra-high performance liquid chromatography), enzyme-linked immunosorbent assay and colloidal gold, and the ARDS can be diagnosed by using DR5 protein level data of existing healthy people and/or ARDS patients as a reference.
Further, the R & D Systems Luminex liquid chip used in example 1 of the present invention detected the levels of DR5 protein in the sera of ARDS patients and healthy persons, and performed ROC analysis, and the area under the curve (AUC) was 0.992, and when the detection threshold value (cut-off value) was 79.69pg/mL, the specificity was 100%, and the sensitivity was 96.2%, indicating that the possibility of misdiagnosis was 0 in theory, and the possibility of missed diagnosis was only 3.8%, and the accuracy was very high. Such high accuracy is rarely achieved in all disease diagnostic techniques that detect a single biochemical marker, indicating that the test kit of the present invention performs very well.
The kit disclosed by the invention is a kit discovered according to the above, and can realize objective, accurate and rapid diagnosis of ARDS by detecting DR5 protein in serum.
Obviously, many modifications, substitutions, and variations are possible in light of the above teachings of the invention, without departing from the basic technical spirit of the invention, as defined by the following claims.
The present invention will be described in further detail with reference to the following examples. This should not be understood as limiting the scope of the above-described subject matter of the present invention to the following examples. All the technologies realized based on the above contents of the present invention belong to the scope of the present invention.
Drawings
FIG. 1 is a scatter plot comparing the levels of serum DR5 protein in the two groups.
FIG. 2 is a ROC plot of DR5 protein concentration versus ARDS.
Detailed Description
Example 1 detection of DR5 protein in serum of patients with ARDS
1. Object to be inspected
ARDS patients (group a) 56, healthy (group B) 32. The subject population was informed and consented to prior to the experiment.
2. Method of producing a composite material
The method comprises the following steps of (1) reserving blood of a detected object, collecting serum, and carrying out R & D Systems Luminex liquid phase chip detection on DR5 protein:
(1) resuspending, uniformly mixing and diluting the detection mixed solution, and adding 50 mu L of the mixed solution into each hole of the ELISA plate;
(2) adding 50 mu L of standard substance or sample, sealing the membrane, and then incubating for 2 hours at room temperature by shaking;
(3) washing the plate for 3 times by using 100 mu L of plate washing liquid;
(4) adding 50 μ L of biotin-labeled antibody mixture into each well, sealing the membrane, and incubating at room temperature for 30 min;
(5) washing the plate for 3 times by using 100 mu L of plate washing liquid;
(6) adding 100 μ L of color developing solution into each well, shaking at room temperature, and mixing for 2 min;
(7) plates were read within 90min (Bio-Rad);
3. results
(1) The serum DR5 level of the ARDS patient is obviously higher than that of a healthy person
The detection results of the DR5 protein in A, B two groups of blood serum are shown in figure 1, the average value of DR5 in the blood serum of an ARDS patient is obviously higher than that of DR5 in the blood serum of a healthy person, and the statistical P value is less than 0.001;
(2) the serum DR5 protein has great value in the diagnosis of ARDS
The ROC analysis result of the DR5 protein content in A, B group serum is shown in FIG. 2, and the area under the curve (AUC) of ROC is as high as 0.992; when the detection threshold value (cut-off value) was 79.69pg/mL, the specificity was 100% and the sensitivity was 96.2%.
The experimental result shows that by detecting the level of DR5 protein in serum, the misdiagnosis probability is 0 theoretically, the missed diagnosis probability is only 3.8%, and the accuracy is very high; the kit of the invention has excellent performance.
Example 2 kit of the invention
1. Compositions of the kits of the invention
2. Kit using method
(1) Adding 50 mu L of standard substance or sample into each hole of a 96-hole plate, sealing the membrane, and then, shaking and incubating for 2 hours at room temperature;
(2) washing the plate for 3 times by using 100 mu L of plate washing liquid;
(3) adding 50 mu L biotin-labeled secondary antibody into each hole, sealing the membrane, and then incubating for 30min at room temperature by shaking;
(4) washing the plate for 3 times by using 100 mu L of plate washing liquid;
(5) adding 50 μ L of chromogenic substrate into each hole, shaking and mixing for 2min at room temperature;
(6) reading by an enzyme-linked immunosorbent assay;
when the DR5 protein detection value in serum was 79.69pg/mL (i.e., cut-off value) or more, it was judged as ARDS.
In conclusion, the kit can realize rapid and accurate diagnosis of ARDS through quantitative detection of DR5 protein, and has a good application prospect.
Claims (10)
1. The application of the reagent for detecting the content of DR5 protein in serum in preparing an acute respiratory distress syndrome detection kit.
2. The use of claim 1, wherein the reagent for detecting the content of DR5 protein in serum is an enzyme-linked immunosorbent assay reagent.
3. The use according to claim 1, wherein the reagent for detecting the content of DR5 protein in serum is a reagent for a liquid chromatography method.
4. The use of claim 1, wherein the reagent for detecting the content of DR5 protein in serum is a colloidal gold detection reagent.
5. The use of claim 1, wherein the reagent for detecting the content of DR5 protein in serum is a reagent for R & DSystems Luminex liquid chip detection.
6. A kit for detecting acute respiratory distress syndrome is characterized by comprising a reagent for detecting the content of DR5 protein in serum.
7. The detection kit of claim 6, wherein the reagent for detecting the content of DR5 protein in serum is an enzyme-linked immunosorbent assay reagent.
8. The detection kit as claimed in claim 6, wherein the reagent for detecting the content of DR5 protein in serum is a reagent for liquid chromatography.
9. The detection kit as claimed in claim 6, wherein the reagent for detecting the content of DR5 protein in serum is a colloidal gold detection reagent.
10. The detection kit as claimed in claim 6, wherein the reagent for detecting the content of DR5 protein in serum is R & D Systems Luminex liquid chip detection reagent.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010515456.7A CN111735961A (en) | 2020-06-08 | 2020-06-08 | Acute respiratory distress syndrome detection kit |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010515456.7A CN111735961A (en) | 2020-06-08 | 2020-06-08 | Acute respiratory distress syndrome detection kit |
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| CN111735961A true CN111735961A (en) | 2020-10-02 |
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| CN202010515456.7A Pending CN111735961A (en) | 2020-06-08 | 2020-06-08 | Acute respiratory distress syndrome detection kit |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112611877A (en) * | 2020-12-31 | 2021-04-06 | 四川大学华西医院 | Kit for predicting acute respiratory distress syndrome illness and prognosis by using ANGPTL4 |
| CN113252896A (en) * | 2021-05-13 | 2021-08-13 | 四川大学华西医院 | Kit for predicting acute respiratory distress syndrome prognosis by using ND1 in alveolar lavage fluid |
| CN113624978A (en) * | 2021-04-06 | 2021-11-09 | 四川大学华西医院 | Kit for assessing acute respiratory distress syndrome/acute lung injury and prognosis by detecting alveolar lavage fluid and serum albumin |
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| AU2007216733A1 (en) * | 2000-04-11 | 2007-09-27 | Genentech, Inc. | Multivalent antibodies and uses therefor |
| WO2016110598A1 (en) * | 2015-01-09 | 2016-07-14 | Mabimmune Diagnostics Ag | Novel anti-fibroblast activation protein (fap) antibodies and uses derived thereof |
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2020
- 2020-06-08 CN CN202010515456.7A patent/CN111735961A/en active Pending
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112611877A (en) * | 2020-12-31 | 2021-04-06 | 四川大学华西医院 | Kit for predicting acute respiratory distress syndrome illness and prognosis by using ANGPTL4 |
| CN113624978A (en) * | 2021-04-06 | 2021-11-09 | 四川大学华西医院 | Kit for assessing acute respiratory distress syndrome/acute lung injury and prognosis by detecting alveolar lavage fluid and serum albumin |
| CN113624978B (en) * | 2021-04-06 | 2023-09-19 | 四川大学华西医院 | Kit for evaluating acute respiratory distress syndrome/acute lung injury and prognosis by detecting alveolar lavage fluid and serum albumin |
| CN113252896A (en) * | 2021-05-13 | 2021-08-13 | 四川大学华西医院 | Kit for predicting acute respiratory distress syndrome prognosis by using ND1 in alveolar lavage fluid |
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