CN110567921A - Polymer for catalyzing p-hydroxyphenylethylamine to generate fluorescence signal by combining magnetic nanoparticles with HRP and preparation method thereof - Google Patents
Polymer for catalyzing p-hydroxyphenylethylamine to generate fluorescence signal by combining magnetic nanoparticles with HRP and preparation method thereof Download PDFInfo
- Publication number
- CN110567921A CN110567921A CN201910733328.7A CN201910733328A CN110567921A CN 110567921 A CN110567921 A CN 110567921A CN 201910733328 A CN201910733328 A CN 201910733328A CN 110567921 A CN110567921 A CN 110567921A
- Authority
- CN
- China
- Prior art keywords
- polymer
- exosome
- mixed solution
- antibody
- magnetic nanoparticles
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N2021/6417—Spectrofluorimetric devices
- G01N2021/6421—Measuring at two or more wavelengths
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
- G01N2021/6439—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" with indicators, stains, dyes, tags, labels, marks
Abstract
The invention provides a fluorescence method for generating a polymer with a fluorescence signal and detecting exosomes by using magnetic nanoparticles as substrates and using horseradish peroxidase to catalyze p-hydroxyphenylethylamine. According to the technical scheme, magnetic nanoparticles are used as a substrate, a specific antibody is modified on the magnetic nanoparticles to capture an exosome, and an antibody modified by horseradish peroxidase is introduced through a sandwich structure of 'antibody-exosome-antibody'. Under the condition that a small amount of hydrogen peroxide exists, the horseradish peroxidase can catalyze p-hydroxyphenylethylamine to generate a polymer so as to generate a fluorescence signal, the fluorescence intensity and the exosome content present a linear relation in a certain concentration range, and the exosome content can be specifically detected by detecting the fluorescence intensity.
Description
Technical Field
The invention belongs to the technical field of biosensors, and particularly relates to a polymer for detecting exosomes and capable of catalyzing p-hydroxyphenylethylamine to generate a fluorescent signal by combining magnetic nanoparticles with HRP (horse radish peroxidase) and a preparation method thereof.
background
Tumor is a serious disease which seriously affects the human life and health and has high death rate. The detection of tumor markers plays a critical role in the early diagnosis and timely treatment of tumors. The exosome is secreted by various living cells, has the diameter of about 30-100 nm, and is provided with a lipid bilayer and nano-scale cystic vesicles rich in contents. The formation of exosomes starts with endocytosis, and is widely distributed in various body fluids, such as peripheral blood, saliva, milk, urine, cerebrospinal fluid and the like, and contains abundant components such as protein, nucleic acid, lipid and the like. Since the level of exosome in the plasma of most malignant tumor patients presents an abnormal expression state, such as lung cancer, breast cancer, liver cancer and the like, the exosome has great potential to become a tumor marker. Therefore, the high-sensitivity and high-specificity detection of the exosome level in the plasma has very important significance for clinical diagnosis of tumors, particularly early diagnosis of lung cancer and breast cancer, and a new thought is provided for early diagnosis and clinical treatment of tumors.
The methods for detecting the level of exosome include protein analysis, transmission electron microscope analysis, gene level analysis and the like. However, the existing exosome level detection method still has some defects, such as poor selectivity, complex operation, high cost, low sensitivity and the like. At present, a method with good selectivity, simple operation, higher cost effectiveness and high sensitivity is urgently needed for detecting the level of exosome.
The magnetic nanoparticles have excellent physicochemical properties and biocompatibility such as high separation speed, high efficiency, simple operation, easy realization of functionalization and automation, no influence on the activity of separated substances and the like, and are widely applied to the fields of separation and sorting of cells, separation, purification and detection of biomacromolecules, targeted diagnosis and treatment and the like at present. The magnetic nano-particles are mainly used for biomacromolecule detection, and specific probes or antibodies are modified on the surfaces of the magnetic nano-particles and then can be used for specific detection of target molecules.
Horseradish (Brassicaceae) is a cold-resistant perennial herb, mainly distributed in temperate regions of the world, and the root of the cold-resistant perennial herb has good cooking value. The root of horseradish is also a rich source of peroxidase, and contains at least 30 HRP isozyme forms, of which the content of HRP isozyme C is most abundant, and thus is collectively called horseradish peroxidase (HRP). The horseradish peroxidase is a heme enzyme which takes heme as an oxidation-reduction center and can oxidize most organic matters and inorganic matters by utilizing hydrogen peroxide. The nano-composite has the advantages of high activity, high heat resistance, good acid-base stability and the like, has high tolerance to pollutant concentration and salinity, and has little activity loss after being coupled with antigen or antibody, so the nano-composite is widely applied to the fields of sewage treatment, food industry, organic synthesis, analysis and detection and the like.
Disclosure of Invention
The invention aims to overcome the defects of an exosome detection method in the prior art, and designs a polymer for detecting exosomes and a preparation method thereof, wherein the polymer is used for catalyzing p-hydroxyphenylethylamine to generate a fluorescence signal by combining magnetic nanoparticles with HRP, so that high-selectivity and high-sensitivity detection on exosomes is realized.
In order to achieve the above object, the present invention provides a polymer for detecting exosome, which catalyzes p-hydroxyphenylethylamine to generate a fluorescent signal by combining magnetic nanoparticles with HRP, and a preparation method thereof, wherein the polymer comprises the following steps:
(1) Dissolving the carboxylated magnetic nanoparticles into a PBS buffer solution to prepare a magnetic nanoparticle mixed solution;
(2) Adding a mixed solution of EDC/NHS into the magnetic nanoparticle solution in the step (1) to activate carboxyl, then modifying with a CD63 antibody, and blocking with BAS;
(3) adding an exosome to be detected at room temperature, then adding an EpCAM antibody marked by biotin, finally adding Horse Radish Peroxidase (HRP) marked by streptavidin, and obtaining magnetic nanoparticles combined with the HRP and the exosome in an antibody-exosome-antibody form;
(4) And (3) adding the magnetic nanoparticle polymer in the step (3) into a solution containing p-hydroxyphenylethylamine and hydrogen peroxide, and performing fluorescence signal intensity measurement after reaction.
The concentration of the magnetic nanoparticle mixed solution in the step (1) is 20-200 micrograms/ml.
The preparation method of the carboxylated magnetic nanoparticles in the step (1) comprises the following steps: EDC/NHS (20 mg/5 mg) mixed solution was added to the ferroferric oxide magnetic nanoparticles, and the mixture was reacted for 30 minutes to activate carboxyl groups, followed by magnetic separation, and after the supernatant was discarded, the resultant was dissolved in 1 ml of PBS solution to prepare a carboxylated magnetic nanoparticle mixed solution (concentration: 0.1 mg/ml).
EDC in the EDC/NHS mixed solution in the step (2) is a bifunctional coupling agent, and is mixed with NHS for use, and the EDC/NHS mixed solution needs to be prepared in situ.
The time for activating the carboxyl group of the EDC/NHS mixed solution in the step (2) is 30 minutes, and the concentration of the used CD63 antibody is 20-200 micrograms/ml.
The concentration of the p-hydroxyphenylethylamine in the step (4) is 10-400 micrograms/ml, and the concentration of the hydrogen peroxide is 0.01-0.5%; when the fluorescence signal is measured, the excitation wavelength is 320nm, the emission spectrum scanning range is 370-550 nm, and the fluorescence signal intensity at 406nm (+ -2 nm) is selected as a detection value.
Compared with the prior art, the invention has a plurality of innovation points: (1) carboxylated magnetic nanoparticles and coupling methods; (2) magnetic nanoparticles are used as a substrate, the polymer with a fluorescence signal is generated by catalyzing p-hydroxyphenylethylamine through HRP, and a fluorescence method for detecting exosomes is realized. The technical scheme has the excellent performance that the magnetic nanoparticles are used as the substrate, the exosome is captured by modifying the specific antibody on the magnetic nanoparticles, and the HRP modified antibody is introduced through a sandwich structure of 'antibody-exosome-antibody'. Under the condition that a small amount of hydrogen peroxide exists, the HRP can catalyze p-hydroxyphenylethylamine to generate a polymer so as to generate a fluorescence signal, the fluorescence intensity and the content of exosomes present a linear relation in a certain concentration range, and the content of exosomes can be specifically detected by detecting the fluorescence intensity. The polymer with the fluorescent signal is generated by catalyzing p-hydroxyphenylethylamine through HRP, and the fluorescent method for detecting exosomes is realized, so that the method has the advantages of good selectivity, simple preparation process, low preparation cost, high product sensitivity and the like, and the defects of the exosome detection method in the prior art are successfully overcome. The invention can be developed into a novel biosensor, and is suitable for high-efficiency and specific detection of exosome content in various biological samples.
Drawings
FIG. 1 is a schematic diagram of the preparation of fluorescent signals generated by magnetic nanoparticles in combination with HRP to catalyze p-hydroxyphenylethylamine.
FIG. 2 is a graph of fluorescence signals generated by magnetic nanoparticles in combination with HRP to catalyze p-hydroxyphenylethylamine.
Fig. 3 is a graph showing the comparative effect of effect example 1.
Fig. 4 is a graph showing the comparative effect of effect example 2.
Detailed Description
Effect example 1 detection of sample from liver cancer patient
Preparing an exosome: the plasma samples were removed from a refrigerator at-20 ℃ and thawed on ice, and 4. mu.l (611U/ml) of thrombin was added to the thawed plasma samples per 500. mu.l to give a final enzyme concentration of 5U/ml, and incubated at room temperature for five minutes with mixing being carried out by inversion. Centrifuging at 10000rpm for 5 min, and transferring the supernatant to a clean centrifuge tube. Adding PEG in the plasma exosome kit into the supernatant according to the ratio of 1: 4, uniformly mixing, standing at 4 ℃ for 30-60 minutes, centrifuging at the rotating speed of 1500g for 30 minutes, taking the supernatant, centrifuging at the rotating speed of 1500g for 5 minutes again, and completely sucking the supernatant. Adding 400 microliters of the heavy suspension into the precipitate, uniformly mixing, subpackaging and storing in a refrigerator at the temperature of-80 ℃.
(1) The carboxylated ferroferric oxide magnetic nanoparticles were dissolved in 1 ml of PBS buffer to prepare a magnetic nanoparticle mixed solution (concentration: 0.1 mg/ml)
(2) Adding a mixed solution of EDC/NHS (20 mg/5 mg) into the magnetic nanoparticle solution in the step (1), reacting for 30 minutes to activate carboxyl, carrying out magnetic separation, discarding the supernatant, mixing the obtained product with 1 ml of PBS solution, adding a CD63 antibody (10 micrograms/ml), reacting for 30 minutes, carrying out magnetic separation, washing with the PBS solution once, adding BAS (0.4 mg ml), reacting for 30 minutes, carrying out magnetic separation, discarding the supernatant, and mixing the obtained product with 1 ml of PBS solution.
(3) at room temperature, adding the extracted exosome to be detected, incubating for 15 minutes on a shaking table, performing magnetic separation, discarding supernatant, adding 1 ml of PBS solution, adding a biotin-labeled EpCAM antibody (10 micrograms/ml), incubating for 10 minutes, performing magnetic separation, discarding supernatant, uniformly mixing the obtained product with 1 ml of PBS solution, adding streptavidin-modified HRP (10 micrograms/ml), incubating for 10 minutes, performing magnetic separation, discarding supernatant, uniformly mixing the obtained product with 1 ml of PBS solution, adding 4-hydroxyphenylethylamine (50 micrograms/ml) and hydrogen peroxide (0.2%) to react for 15 minutes, and performing magnetic separation;
(4) And (4) measuring the intensity of the fluorescence signal of the product obtained in the step (3), wherein the excitation wavelength is 320nm, the scanning range of the emission spectrum is 370-550 nm, and the intensity of the fluorescence signal at 406nm (+/-2 nm) is selected as a detection value.
When 5 exosomes of liver cancer patients and 5 exosomes of normal persons were tested according to the above method, the mean and standard deviation of the fluorescence intensities of the polymers formed by the exosomes of the 5 liver cancer patients were 605.2 ± 66.01(a.u), and the mean and standard deviation of the fluorescence intensities of the polymers formed by the exosomes of the 5 normal persons were 296.0 ± 16.31 (a.u). The result is shown in figure 3, P is less than 0.0001, which shows that the exosome of the liver cancer patient detected by the method has obvious statistical difference with the exosome of the normal person.
Effect example 2 detection of samples of patients with Lung cancer
preparing an exosome: the plasma samples were removed from a-20 ℃ freezer and thawed on ice, and the thawed samples were incubated with 4. mu.l (611U/ml) of thrombin per 500. mu.l final enzyme concentration of 5U/ml at room temperature for five minutes, with mixing being carried out by inversion. Centrifuging at 10000rpm for 5 min, and transferring the supernatant to a clean centrifuge tube. Adding PEG in the plasma exosome kit into the supernatant according to the proportion of 1: 4, uniformly mixing, standing for 30-60 minutes at 4 ℃, centrifuging for 30 minutes at the rotating speed of 1500g, taking the supernatant, centrifuging for 5 minutes again at 1500g, and completely sucking the supernatant. Adding 400 microliters of the heavy suspension into the precipitate, uniformly mixing, subpackaging and storing in a refrigerator at the temperature of-80 ℃.
(1) The carboxylated ferrimagnetic nanoparticles were dissolved in 1 ml of PBS buffer to prepare a magnetic nanoparticle mixed solution (0.1 mg/ml).
(2) Adding a mixed solution of EDC/NHS (20 mg/5 mg) into the magnetic nanoparticle solution in the step (1), reacting for 30 minutes to activate carboxyl, carrying out magnetic separation, discarding the supernatant, mixing the obtained product with 1 ml of PBS solution, adding a CD63 antibody (10 micrograms/ml), reacting for 30 minutes, carrying out magnetic separation, washing with PBS once, adding BAS (0.4 mg/ml), reacting for 30 minutes, carrying out magnetic separation, discarding the supernatant, and mixing the obtained product with 1 ml of PBS solution.
(3) at room temperature, adding the extracted exosome to be detected, incubating for 15 minutes on a shaking table, performing magnetic separation, discarding supernatant, adding 1 ml of PBS solution, adding a biotin-labeled PD-L1 antibody (10 micrograms/ml), incubating for 10 minutes, performing magnetic separation, discarding supernatant, uniformly mixing the obtained product with 1 ml of PBS solution, adding streptavidin-modified HRP (10 micrograms/ml), incubating for 10 minutes, performing magnetic separation, discarding supernatant, uniformly mixing the obtained product with 1 ml of PBS solution, adding 4-hydroxyphenylethylamine (50 micrograms/ml) and hydrogen peroxide (0.2%) to react for 15 minutes, and performing magnetic separation;
(4) And (4) measuring the intensity of the fluorescence signal of the product obtained in the step (3), wherein the excitation wavelength is 320nm, the scanning range of the emission spectrum is 370-550 nm, and the intensity of the fluorescence signal with the wavelength of 406nm (+/-2 nm) is selected as a detection value.
The 5 lung cancer patients and 5 normal human exosomes were tested according to the above method, and the mean and standard deviation of the fluorescence intensities of the polymers formed by the 5 lung cancer patients and the 5 normal human exosomes were 598.8 ± 57.57(a.u) and 244.8 ± 58.28 (a.u). The result is shown in figure 4, P is less than 0.0001, which indicates that the exosome of the lung cancer patient detected by the method has obvious statistical difference with the exosome of the normal human.
Claims (6)
1. A polymer for detecting exosomes and catalyzing p-hydroxyphenylethylamine to generate a fluorescent signal by combining magnetic nanoparticles with HRP, comprising the following preparation steps:
(1) Dissolving the carboxylated magnetic nanoparticles into a PBS buffer solution to prepare a magnetic nanoparticle mixed solution;
(2) Adding a mixed solution of EDC/NHS into the magnetic nanoparticle solution in the step (1) to activate carboxyl, then modifying with a CD63 antibody, and blocking with BAS;
(3) adding an exosome to be detected at room temperature, then adding an EpCAM antibody marked by biotin, finally adding Horse Radish Peroxidase (HRP) marked by streptavidin, and obtaining magnetic nanoparticles combined with the HRP and the exosome in an antibody-exosome-antibody form;
(4) And (3) adding the magnetic nanoparticle compound in the step (3) into a solution containing p-hydroxyphenylethylamine and hydrogen peroxide, and measuring the intensity of a fluorescence signal after reaction.
2. the polymer according to claim 1, wherein the concentration of the magnetic nanoparticle mixed solution in the step (1) is 20-200 μ g/ml.
3. The polymer according to claim 1, wherein the carboxylated magnetic nanoparticles of step (1) are prepared by a process comprising: EDC/NHS (20 mg/5 mg) mixed solution was added to the ferroferric oxide magnetic nanoparticles, and the mixture was reacted for 30 minutes to activate carboxyl groups, followed by magnetic separation, and after the supernatant was discarded, the resultant was dissolved in 1 ml of PBS solution to prepare a carboxylated magnetic nanoparticle mixed solution (concentration: 0.1 mg/ml).
4. the polymer of claim 1, wherein the EDC/NHS mixed solution in step (2) is a bifunctional coupling agent, and is used after mixing with NHS, and the EDC/NHS mixed solution is ready for use.
5. the polymer of claim 1, wherein the EDC/NHS mixed solution in step (2) has a carboxyl group activating time of 30 minutes and a CD63 antibody concentration of 20-200. mu.g/ml.
6. The polymer as claimed in claim 1, wherein the concentration of p-hydroxyphenylethylamine in the step (4) is 10-400 micrograms/ml, and the concentration of hydrogen peroxide is 0.01-0.5%; when the fluorescence signal is measured, the excitation wavelength is 320nm, the emission spectrum scanning range is 370-550 nm, and the fluorescence signal intensity at 406nm (+ -2 nm) is selected as a detection value.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910733328.7A CN110567921A (en) | 2019-08-09 | 2019-08-09 | Polymer for catalyzing p-hydroxyphenylethylamine to generate fluorescence signal by combining magnetic nanoparticles with HRP and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910733328.7A CN110567921A (en) | 2019-08-09 | 2019-08-09 | Polymer for catalyzing p-hydroxyphenylethylamine to generate fluorescence signal by combining magnetic nanoparticles with HRP and preparation method thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN110567921A true CN110567921A (en) | 2019-12-13 |
Family
ID=68775099
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201910733328.7A Pending CN110567921A (en) | 2019-08-09 | 2019-08-09 | Polymer for catalyzing p-hydroxyphenylethylamine to generate fluorescence signal by combining magnetic nanoparticles with HRP and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN110567921A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114280016A (en) * | 2021-12-07 | 2022-04-05 | 广州兆瑞医学生物科技有限公司 | Exosome detection method |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106226513A (en) * | 2016-08-02 | 2016-12-14 | 北京乐普医疗科技有限责任公司 | A kind of method of immunomagnetic beads detection by quantitative related antigen and application thereof |
| CN108387746A (en) * | 2018-03-02 | 2018-08-10 | 浙江大学 | A kind of superparamagnetic nanoparticle and preparation method thereof of capture excretion body and specific excretion body electrochemiluminescent immunoassay immue quantitative detection reagent box |
-
2019
- 2019-08-09 CN CN201910733328.7A patent/CN110567921A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106226513A (en) * | 2016-08-02 | 2016-12-14 | 北京乐普医疗科技有限责任公司 | A kind of method of immunomagnetic beads detection by quantitative related antigen and application thereof |
| CN108387746A (en) * | 2018-03-02 | 2018-08-10 | 浙江大学 | A kind of superparamagnetic nanoparticle and preparation method thereof of capture excretion body and specific excretion body electrochemiluminescent immunoassay immue quantitative detection reagent box |
Non-Patent Citations (3)
| Title |
|---|
| KAORU MATSUOKA ET AL: "Fluorscence Enzyme Immunoassay for Insulin using Peroxidase-Tyramine-Hydrogen Peroxide", 《CHEM.PHARM.BULL》 * |
| SANGMOO JEONG ET AL: "Integrated Magneto-Electrochemical Sensor for Exosome Analysis", 《ACS NANO》 * |
| 荆浩: "羧基磁珠捕获血清外泌体及双抗体夹心法在检测外泌体蛋白中的应用", 《中国优秀博硕士学位论文全文数据库（硕士） 医药卫生科技辑》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114280016A (en) * | 2021-12-07 | 2022-04-05 | 广州兆瑞医学生物科技有限公司 | Exosome detection method |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| JPS63500264A (en) | Methods for determining the presence and monitoring of pathological conditions in mammals and alternative test kits | |
| CN102352311A (en) | Electrochemical sensor detection method of thrombin based on aptamer-gold nanoparticle-enzyme composite | |
| CN107841527A (en) | It is a kind of to utilize aptamer and the fluorescence detection method of magnetic material detection fibrin ferment | |
| Ji et al. | Enhanced luminol chemiluminescence with oxidase-like properties of FeOOH nanorods for the sensitive detection of uric acid | |
| CN111411104A (en) | Magnetic bead for detecting neocoronal pneumonia and preparation method thereof | |
| CN106932371B (en) | Fluorescence imaging method of intracellular glutathione | |
| CN106198962B (en) | Method for closing biomagnetic beads | |
| CN112979530B (en) | Nucleic acid detection probe and preparation method thereof | |
| CN110567921A (en) | Polymer for catalyzing p-hydroxyphenylethylamine to generate fluorescence signal by combining magnetic nanoparticles with HRP and preparation method thereof | |
| CN108445213B (en) | Nano composite probe, composition and fluorescent quantitative kit for high-sensitivity fluorescent quantitative detection of serum tumor marker | |
| Yan et al. | An iron oxide nanoworm hybrid on an interdigitated microelectrode silica surface to detect abdominal aortic aneurysms | |
| CN108918873B (en) | Preparation method and application of photoelectrochemical thrombin aptamer sensor based on dual inhibition of PS @ Au on ZnCdS | |
| CN111024945B (en) | Method for synchronously detecting VEGF and CEA by using chemiluminescence enzyme-linked aptamer | |
| Lai et al. | Based devices for capturing exosomes and exosomal nucleic acids from biological samples | |
| Ronzhin et al. | Modified nanodiamonds as a new carrier for developing reusable enzymatic test-systems for determination of physiologically important substances | |
| CN115711877A (en) | Sensor for triggering spherical nucleic acid self-assembly to rapidly activate CRISPR-Cas12a signal switch by target | |
| CN104880556A (en) | An aptamer-antibody method for detecting a trace of prostate-specific antigen (PSA) in blood and a kit | |
| CN112179875B (en) | Preparation and application of type I hyaluronidase fluorescent nano sensor | |
| Kong et al. | CRISPR/Cas detection with nanodevices: moving deeper into liquid biopsy | |
| JP2019205353A (en) | Method for analyzing nucleic acid | |
| CN112250719B (en) | Tyrosine rare earth luminescent probe and preparation method and application thereof | |
| CN113403300B (en) | Method for enriching circulating free nucleic acid in body fluid by using MOF material and application thereof | |
| CN114199844B (en) | Gold nanocluster and application thereof in preparation of alkaline phosphatase fluorescent probe | |
| Xiluan et al. | Chemiluminescent detection of platelet derived growth factor-BB based on sandwich label-free aptasensor and biotin–streptavidin strategy | |
| RU2229717C1 (en) | Method for evaluating proinflammatory activity of synovia at diseases of large articulations |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20191213 |
|
| RJ01 | Rejection of invention patent application after publication |