CN117871847A - Colorimetric immunoassay reagent and preparation method and application thereof - Google Patents
Colorimetric immunoassay reagent and preparation method and application thereof Download PDFInfo
- Publication number
- CN117871847A CN117871847A CN202311767586.XA CN202311767586A CN117871847A CN 117871847 A CN117871847 A CN 117871847A CN 202311767586 A CN202311767586 A CN 202311767586A CN 117871847 A CN117871847 A CN 117871847A
- Authority
- CN
- China
- Prior art keywords
- mxene
- mos
- loaded
- antibody
- antigen
- 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
- 238000002360 preparation method Methods 0.000 title claims abstract description 22
- 239000003153 chemical reaction reagent Substances 0.000 title claims abstract description 9
- 238000003018 immunoassay Methods 0.000 title abstract description 4
- 108010088201 squamous cell carcinoma-related antigen Proteins 0.000 claims abstract description 37
- 239000000523 sample Substances 0.000 claims abstract description 19
- 206010041823 squamous cell carcinoma Diseases 0.000 claims abstract description 7
- 239000010931 gold Substances 0.000 claims description 103
- 239000000427 antigen Substances 0.000 claims description 30
- 102000036639 antigens Human genes 0.000 claims description 30
- 108091007433 antigens Proteins 0.000 claims description 30
- 238000002156 mixing Methods 0.000 claims description 24
- UMGDCJDMYOKAJW-UHFFFAOYSA-N thiourea Chemical compound NC(N)=S UMGDCJDMYOKAJW-UHFFFAOYSA-N 0.000 claims description 18
- 238000006243 chemical reaction Methods 0.000 claims description 14
- 239000007790 solid phase Substances 0.000 claims description 13
- 238000005406 washing Methods 0.000 claims description 13
- 239000002253 acid Substances 0.000 claims description 11
- 238000003756 stirring Methods 0.000 claims description 11
- 150000001720 carbohydrates Chemical class 0.000 claims description 10
- 238000000034 method Methods 0.000 claims description 10
- 239000011259 mixed solution Substances 0.000 claims description 10
- 206010028980 Neoplasm Diseases 0.000 claims description 9
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Natural products NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 claims description 9
- 239000011734 sodium Substances 0.000 claims description 9
- 239000011684 sodium molybdate Substances 0.000 claims description 9
- 235000015393 sodium molybdate Nutrition 0.000 claims description 9
- TVXXNOYZHKPKGW-UHFFFAOYSA-N sodium molybdate (anhydrous) Chemical compound [Na+].[Na+].[O-][Mo]([O-])(=O)=O TVXXNOYZHKPKGW-UHFFFAOYSA-N 0.000 claims description 9
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims description 8
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 claims description 8
- 102000004169 proteins and genes Human genes 0.000 claims description 8
- 108090000623 proteins and genes Proteins 0.000 claims description 8
- 229910052708 sodium Inorganic materials 0.000 claims description 8
- 238000011534 incubation Methods 0.000 claims description 7
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 6
- 239000007788 liquid Substances 0.000 claims description 6
- 238000000926 separation method Methods 0.000 claims description 6
- 241000283707 Capra Species 0.000 claims description 4
- 241000282693 Cercopithecidae Species 0.000 claims description 4
- 241000283074 Equus asinus Species 0.000 claims description 4
- 241000287828 Gallus gallus Species 0.000 claims description 4
- 241000699666 Mus <mouse, genus> Species 0.000 claims description 4
- 241000283973 Oryctolagus cuniculus Species 0.000 claims description 4
- 241001494479 Pecora Species 0.000 claims description 4
- 241000700159 Rattus Species 0.000 claims description 4
- 235000013330 chicken meat Nutrition 0.000 claims description 4
- 239000003593 chromogenic compound Substances 0.000 claims description 4
- 238000001027 hydrothermal synthesis Methods 0.000 claims description 4
- 238000011065 in-situ storage Methods 0.000 claims description 4
- 229920001184 polypeptide Polymers 0.000 claims description 4
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 4
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 4
- GEYOCULIXLDCMW-UHFFFAOYSA-N 1,2-phenylenediamine Chemical compound NC1=CC=CC=C1N GEYOCULIXLDCMW-UHFFFAOYSA-N 0.000 claims description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 2
- 108010022366 Carcinoembryonic Antigen Proteins 0.000 claims description 2
- 102000012406 Carcinoembryonic Antigen Human genes 0.000 claims description 2
- 108010037897 DC-specific ICAM-3 grabbing nonintegrin Proteins 0.000 claims description 2
- 241000283086 Equidae Species 0.000 claims description 2
- 241000283073 Equus caballus Species 0.000 claims description 2
- 102100021181 Golgi phosphoprotein 3 Human genes 0.000 claims description 2
- 101000993094 Homo sapiens Chromogranin-A Proteins 0.000 claims description 2
- 101001040734 Homo sapiens Golgi phosphoprotein 3 Proteins 0.000 claims description 2
- 101000741967 Homo sapiens Presequence protease, mitochondrial Proteins 0.000 claims description 2
- 102100033420 Keratin, type I cytoskeletal 19 Human genes 0.000 claims description 2
- 108010066302 Keratin-19 Proteins 0.000 claims description 2
- 241000699670 Mus sp. Species 0.000 claims description 2
- 102100036961 Nuclear mitotic apparatus protein 1 Human genes 0.000 claims description 2
- 102000004264 Osteopontin Human genes 0.000 claims description 2
- 108010081689 Osteopontin Proteins 0.000 claims description 2
- 108010047320 Pepsinogen A Proteins 0.000 claims description 2
- 102000012288 Phosphopyruvate Hydratase Human genes 0.000 claims description 2
- 108010022181 Phosphopyruvate Hydratase Proteins 0.000 claims description 2
- 102100038632 Presequence protease, mitochondrial Human genes 0.000 claims description 2
- 108010072866 Prostate-Specific Antigen Proteins 0.000 claims description 2
- 102100038358 Prostate-specific antigen Human genes 0.000 claims description 2
- 102100028964 Proteoglycan 3 Human genes 0.000 claims description 2
- 101710127914 Proteoglycan 3 Proteins 0.000 claims description 2
- 102100027378 Prothrombin Human genes 0.000 claims description 2
- 108010094028 Prothrombin Proteins 0.000 claims description 2
- 102000013009 Pyruvate Kinase Human genes 0.000 claims description 2
- 108020005115 Pyruvate Kinase Proteins 0.000 claims description 2
- 102000007591 Tartrate-Resistant Acid Phosphatase Human genes 0.000 claims description 2
- 108010032050 Tartrate-Resistant Acid Phosphatase Proteins 0.000 claims description 2
- 230000002159 abnormal effect Effects 0.000 claims description 2
- 102000013529 alpha-Fetoproteins Human genes 0.000 claims description 2
- 108010026331 alpha-Fetoproteins Proteins 0.000 claims description 2
- 229910052799 carbon Inorganic materials 0.000 claims description 2
- QKUSRAKPUWQSJS-UHFFFAOYSA-N diazanium 3-ethyl-2H-1,3-benzothiazole-6-sulfonate Chemical compound [NH4+].[NH4+].[O-]S(=O)(=O)C1=CC=C2N(CC)CSC2=C1.[O-]S(=O)(=O)C1=CC=C2N(CC)CSC2=C1 QKUSRAKPUWQSJS-UHFFFAOYSA-N 0.000 claims description 2
- 238000001035 drying Methods 0.000 claims description 2
- 239000012634 fragment Substances 0.000 claims description 2
- 206010017758 gastric cancer Diseases 0.000 claims description 2
- 208000010749 gastric carcinoma Diseases 0.000 claims description 2
- 108010083422 gastrin-releasing peptide precursor Proteins 0.000 claims description 2
- 102000057162 human CHGA Human genes 0.000 claims description 2
- 229910052751 metal Inorganic materials 0.000 claims description 2
- 229910052757 nitrogen Inorganic materials 0.000 claims description 2
- 108010036112 nuclear matrix protein 22 Proteins 0.000 claims description 2
- 150000003905 phosphatidylinositols Chemical class 0.000 claims description 2
- 238000002331 protein detection Methods 0.000 claims description 2
- 229940039716 prothrombin Drugs 0.000 claims description 2
- 230000035484 reaction time Effects 0.000 claims description 2
- 239000007787 solid Substances 0.000 claims description 2
- 241000894007 species Species 0.000 claims description 2
- 201000000498 stomach carcinoma Diseases 0.000 claims description 2
- 229910052719 titanium Inorganic materials 0.000 claims description 2
- 150000003624 transition metals Chemical group 0.000 claims description 2
- 229910052721 tungsten Inorganic materials 0.000 claims description 2
- 229910052720 vanadium Inorganic materials 0.000 claims description 2
- 238000001514 detection method Methods 0.000 abstract description 25
- 230000004048 modification Effects 0.000 abstract description 11
- 238000012986 modification Methods 0.000 abstract description 11
- 230000000694 effects Effects 0.000 abstract description 8
- 239000000463 material Substances 0.000 abstract description 6
- 238000001179 sorption measurement Methods 0.000 abstract description 5
- 230000035945 sensitivity Effects 0.000 abstract description 3
- 239000007822 coupling agent Substances 0.000 abstract description 2
- 230000002349 favourable effect Effects 0.000 abstract description 2
- 101100043112 Homo sapiens SERPINB3 gene Proteins 0.000 abstract 1
- 102100036383 Serpin B3 Human genes 0.000 abstract 1
- 239000008055 phosphate buffer solution Substances 0.000 description 11
- 238000010521 absorption reaction Methods 0.000 description 8
- 239000000758 substrate Substances 0.000 description 8
- 238000000862 absorption spectrum Methods 0.000 description 6
- 230000003197 catalytic effect Effects 0.000 description 6
- 229910021642 ultra pure water Inorganic materials 0.000 description 6
- 239000012498 ultrapure water Substances 0.000 description 6
- 239000000243 solution Substances 0.000 description 5
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 4
- 229910052737 gold Inorganic materials 0.000 description 4
- 230000002163 immunogen Effects 0.000 description 4
- 102000006833 Multifunctional Enzymes Human genes 0.000 description 3
- 108010047290 Multifunctional Enzymes Proteins 0.000 description 3
- 238000002835 absorbance Methods 0.000 description 3
- 239000011521 glass Substances 0.000 description 3
- 239000002105 nanoparticle Substances 0.000 description 3
- 239000002245 particle Substances 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 239000010936 titanium Substances 0.000 description 3
- 239000000439 tumor marker Substances 0.000 description 3
- 238000009777 vacuum freeze-drying Methods 0.000 description 3
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 2
- -1 CA153 Proteins 0.000 description 2
- 108090000288 Glycoproteins Proteins 0.000 description 2
- 102000003886 Glycoproteins Human genes 0.000 description 2
- YBCVMFKXIKNREZ-UHFFFAOYSA-N acoh acetic acid Chemical compound CC(O)=O.CC(O)=O YBCVMFKXIKNREZ-UHFFFAOYSA-N 0.000 description 2
- 230000005540 biological transmission Effects 0.000 description 2
- 229940098773 bovine serum albumin Drugs 0.000 description 2
- 239000007853 buffer solution Substances 0.000 description 2
- 238000004364 calculation method Methods 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 230000007547 defect Effects 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 238000003745 diagnosis Methods 0.000 description 2
- 229910001873 dinitrogen Inorganic materials 0.000 description 2
- 238000002173 high-resolution transmission electron microscopy Methods 0.000 description 2
- 230000003993 interaction Effects 0.000 description 2
- 238000012417 linear regression Methods 0.000 description 2
- 238000004020 luminiscence type Methods 0.000 description 2
- 238000003760 magnetic stirring Methods 0.000 description 2
- 239000013642 negative control Substances 0.000 description 2
- 230000003647 oxidation Effects 0.000 description 2
- 238000007254 oxidation reaction Methods 0.000 description 2
- 239000008363 phosphate buffer Substances 0.000 description 2
- TYJJADVDDVDEDZ-UHFFFAOYSA-M potassium hydrogencarbonate Chemical compound [K+].OC([O-])=O TYJJADVDDVDEDZ-UHFFFAOYSA-M 0.000 description 2
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000009210 therapy by ultrasound Methods 0.000 description 2
- 125000003396 thiol group Chemical group [H]S* 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- 238000004627 transmission electron microscopy Methods 0.000 description 2
- QGZKDVFQNNGYKY-UHFFFAOYSA-O Ammonium Chemical compound [NH4+] QGZKDVFQNNGYKY-UHFFFAOYSA-O 0.000 description 1
- 206010005003 Bladder cancer Diseases 0.000 description 1
- 208000017897 Carcinoma of esophagus Diseases 0.000 description 1
- 206010008342 Cervix carcinoma Diseases 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 241000282326 Felis catus Species 0.000 description 1
- 208000009849 Female Genital Neoplasms Diseases 0.000 description 1
- 229930186217 Glycolipid Natural products 0.000 description 1
- 101000749311 Homo sapiens C-type lectin domain family 4 member M Proteins 0.000 description 1
- 206010030155 Oesophageal carcinoma Diseases 0.000 description 1
- 208000006105 Uterine Cervical Neoplasms Diseases 0.000 description 1
- 239000008351 acetate buffer Substances 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 201000001531 bladder carcinoma Diseases 0.000 description 1
- 238000004061 bleaching Methods 0.000 description 1
- 210000001124 body fluid Anatomy 0.000 description 1
- 239000010839 body fluid Substances 0.000 description 1
- 210000004027 cell Anatomy 0.000 description 1
- 201000010881 cervical cancer Diseases 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 1
- 201000005619 esophageal carcinoma Diseases 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 210000004072 lung Anatomy 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 239000002086 nanomaterial Substances 0.000 description 1
- 239000002953 phosphate buffered saline Substances 0.000 description 1
- 229920001343 polytetrafluoroethylene Polymers 0.000 description 1
- 239000004810 polytetrafluoroethylene Substances 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000004393 prognosis Methods 0.000 description 1
- 229910001220 stainless steel Inorganic materials 0.000 description 1
- 239000010935 stainless steel Substances 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- MTPVUVINMAGMJL-UHFFFAOYSA-N trimethyl(1,1,2,2,2-pentafluoroethyl)silane Chemical group C[Si](C)(C)C(F)(F)C(F)(F)F MTPVUVINMAGMJL-UHFFFAOYSA-N 0.000 description 1
- 238000002604 ultrasonography Methods 0.000 description 1
- 208000010570 urinary bladder carcinoma Diseases 0.000 description 1
Classifications
-
- 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
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
-
- 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
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
-
- 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
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
- G01N33/57473—Immunoassay; Biospecific binding assay; Materials therefor for cancer involving carcinoembryonic antigen, i.e. CEA
-
- 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
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
- G01N33/57484—Immunoassay; Biospecific binding assay; Materials therefor for cancer involving compounds serving as markers for tumor, cancer, neoplasia, e.g. cellular determinants, receptors, heat shock/stress proteins, A-protein, oligosaccharides, metabolites
-
- 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
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Immunology (AREA)
- Engineering & Computer Science (AREA)
- Urology & Nephrology (AREA)
- Biomedical Technology (AREA)
- Hematology (AREA)
- Chemical & Material Sciences (AREA)
- Molecular Biology (AREA)
- Cell Biology (AREA)
- Food Science & Technology (AREA)
- General Health & Medical Sciences (AREA)
- Biotechnology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- Microbiology (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Oncology (AREA)
- Hospice & Palliative Care (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention belongs to the field of biosensing detection, and particularly relates to a colorimetric immunoassay reagent and a preparation method and application thereof. The invention prepares the Au-loaded MoS 2 The material has high specific surface area and active binding site for biological recognition molecules, can bond the antibody by utilizing the double effects of physical adsorption and Au-S bond, can improve the immobilization capacity of the antibody and the stability of antibody modification, can directly modify the antibody, does not need a coupling agent, and is favorable for maintaining the activity of the antibody, and high in sensitivity and accuracy. Provides an Au/MoS modified by an SCCA-labeled antibody for the first time 2 MXene nanobiological probes having high nanoenzyme properties and high affinity for squamous cell carcinoma antigensAnd the detection of squamous cell carcinoma antigen is realized, the detection range is 0.1ng/mL-1000ng/mL, and the detection limit is 0.042ng/mL.
Description
Technical Field
The invention belongs to the field of biosensing detection, and particularly relates to a colorimetric immunoassay reagent and a preparation method and application thereof.
Background
Tumor markers generally refer to active substances present in body fluids or tissues of tumor patients and expressed abnormally by tumor tissues and cells, and include hormones, enzymes, specific and nonspecific glycoproteins or glycolipids, and the like. Its presence or amount may suggest the nature of the tumor to aid in diagnosis, classification, prognosis, and treatment guidance of the tumor. Because the detection of the tumor markers has the characteristics of simple and convenient acquisition, small wound and the like, the tumor marker can be used as an effective way for early detection of micro-focus tumors, and can be detected by physical examination such as ultrasound, CT, MRI or PET-CT. Squamous Cell Carcinoma Antigen (SCCA), a glycoprotein tumor marker, can be used for the auxiliary diagnosis of cervical cancer, lung squamous carcinoma, esophageal carcinoma, bladder carcinoma, treatment observation and recurrence monitoring.
The existing chip for detecting squamous cell carcinoma antigen SCCA (application number: 201911416620.2, a detection chip for detecting male malignant tumor markers, a preparation method and application thereof, application number: 201911408940.3, a detection chip for detecting female malignant tumor markers, a preparation method and application thereof, application number: 201310375818.7, a gynecological tumor marker CA125, CA153, SCCA and HE4 chip detection system based on up-conversion luminescence and a reagent), and the acquired signals are fluorescent signals, so fluorescent molecules or up-conversion luminescence particles are often required to mark antibodies, and the substances are usually high in toxicity, high in price, and have the defects of photo bleaching, poor light signal stability and the like. Aiming at the defects, it is necessary to develop an instant detection method which is simple to operate, low in detection cost, stable in color development signal, capable of directly observing color development depth through naked eyes without instruments, and capable of measuring the content of an object to be detected, and is more suitable for instant detection application scenes.
Disclosure of Invention
The first aspect of the invention aims to provide an Au-loaded MoS 2 /MXene。
The second aspect of the present invention is directed to an Au-loaded MoS 2 Process for the preparation of/MXene.
The third aspect of the present invention is directed to providing an Au-loaded MoS 2 Use of/MXene.
The object of the fourth aspect of the invention is to provide a product.
The object of the fifth aspect of the present invention is to provide a probe.
The sixth aspect of the present invention is directed to a method for preparing a probe.
The seventh aspect of the present invention is directed to a kit.
In order to achieve the above purpose of the present invention, the present invention adopts the following technical scheme:
in a first aspect of the invention, there is provided an Au-loaded MoS 2 MXene, the Au is loaded on the MoS 2 Surface of/Mxene.
Preferably, the MoS 2 The MXene is in the shape of a three-dimensional flower.
Preferably, the MXene contains M and X elements, wherein M is one or more of metal elements in the transition metal group, and X is one or two of carbon and nitrogen.
Preferably, M is selected from one or more of the Ti, zr, hf, V, nb, ta, cr, sc, mo, W elements.
Preferably, the MXene comprises Ti 3 C 2 。
In a second aspect of the present invention, there is provided an Au-loaded MoS in the second aspect of the present invention 2 The preparation method of the MXene is characterized in that:
(1) Preparing MoS by adopting a hydrothermal method 2 /MXene；
(2) The Au-loaded MoS is prepared by adopting an in-situ growth method 2 /MXene。
Preferably, the MoS is prepared by a hydrothermal method in the step (1) 2 the/MXene is specifically: mixing sodium molybdate, thiourea and MXene, and reacting to obtain MoS 2 /MXene。
Preferably, the reaction conditions are 180-240 ℃ for 18-30 h.
Preferably, the mass ratio of the sodium molybdate to the thiourea to the MXene is (18-22): (30-32): 1.
preferably, the mixing of sodium molybdate, thiourea and MXene is specifically as follows: firstly mixing sodium molybdate and thiourea to obtain a sodium molybdate-thiourea mixed solution; and (3) mixing the sodium molybdate-thiourea mixed solution with MXene for the second time to obtain the sodium molybdate-thiourea-MXene mixed solution.
Preferably, the time of the first mixing is 5-15 min.
Preferably, the time of the second mixing is 20-40 min.
Preferably, the first mixing and the second mixing are stirring; further, the stirring is carried out by strong magnetic force.
Preferably, the reaction further comprises the following steps: solid-liquid separation to obtain MoS 2 MXene; washing the obtained MoS 2 /Mxene。
In the step (2), an in-situ growth method is adopted to prepare the Au-loaded MoS 2 the/MXene is specifically: the MoS is subjected to 2 mixing/MXene with chloroauric acid, and reacting to obtain Au-loaded MoS 2 /MXene。
Preferably, the chloroauric acid and MoS 2 The mass ratio of the/MXene is 1: (9-45).
Preferably, the MoS 2 The mixing of/MXene and chloroauric acid is specifically as follows: adding chloroauric acid into MoS under ultrasonic condition 2 in/MXene.
Preferably, the reaction time is 12-36 hours.
Preferably, the reaction is carried out under stirring conditions; further under stirring at room temperature.
Preferably, the reaction further comprises the following steps: solid-liquid separation to obtain Au-loaded MoS 2 MXene; washing and drying.
In a third aspect of the invention, there is provided the Au-loaded MoS of the first aspect of the invention 2 Use of/MXene for the preparation of a product for detecting a protein.
Preferably, the product detects proteins by immunochemistry.
Preferably, the product comprises a biorecognition molecule and an Au-loaded MoS 2 MXene, said biological recognition molecule specifically recognizing said protein.
Preferably, the biological recognition molecule comprises at least one of an antigen, an antibody, a polypeptide; further antibodies; and further squamous cell carcinoma antibodies.
Preferably, the biological recognition molecule is supported on the Au MoS through coordination bond 2 Au binding in MXene; further through a sulfur gold bond and the Au loaded MoS 2 Au binding in MXene.
In a fourth aspect, the invention provides a protein detection product comprising a biorecognition molecule and an Au-loaded MoS according to the first aspect of the invention 2 MXene, said biological recognition molecule specifically recognizing said protein.
Preferably, the biological recognition molecule comprises at least one of an antigen, an antibody, a polypeptide; further antibodies; and further squamous cell carcinoma antibodies.
Preferably, the biological recognition molecule is supported on the Au MoS through coordination bond 2 Au binding in MXene; further through a gold sulfide bond, the Au is loaded with MoS 2 Au binding in MXene.
In a fifth aspect of the invention there is provided a probe for detecting an antigen comprising an antibody molecule Ab1 which specifically binds to said antigen and an Au-loaded MoS of the first aspect of the invention 2 /MXene。
The antibody molecule Ab1 is supported on the Au-supported MoS through a coordination bond 2 Au binding in MXene; further through a sulfur gold bond and the Au loaded MoS 2 Au binding in MXene.
Preferably, the species of antibody molecule Ab1 is selected from any one of mouse, rat, goat, rabbit, monkey, chicken, horse, donkey, and sheep.
Preferably, the antibody molecule Ab1 is capable of specifically binding to at least one of carcinoembryonic antigen, alpha fetoprotein, carbohydrate antigen 125, carbohydrate antigen 15-3, carbohydrate antigen 72-4, carbohydrate antigen 24-2, carbohydrate antigen 50, prostate specific antigen, squamous cell carcinoma antigen, cytokeratin 19 fragment, gastrin-releasing peptide precursor, neuron specific enolase, nuclear matrix protein-22, tartrate-resistant acid phosphatase 5b, tumor M2 pyruvate kinase, abnormal prothrombin, osteopontin, phosphatidylinositol proteoglycan 3, golgi protein 73, metalloprotease 1, human gastric carcinoma MG7 antigen, pepsinogen, lectin DC-SIGN, lectin DC-SIGNR, and human chromogranin A.
In a sixth aspect of the present invention, there is provided a method for preparing a probe according to the fifth aspect of the present invention, comprising the steps of: antibody molecule Ab1 was loaded with Au to MoS 2 mixing/MXene, and incubating to obtain the Au-loaded MoS modified with antibody molecule Ab1 2 /MXene。
Preferably, the incubation condition is that the incubation is performed for 8-24 hours at 4-40 ℃ under shaking.
Preferably, the antibody molecules Ab1 and Au are loaded with MoS 2 The mass ratio of the/MXene is 1: (45-55).
Preferably, the incubation further comprises the following steps: solid-liquid separation is carried out to obtain the Au-loaded MoS modified with the antibody molecule Ab1 2 MXene, washing.
In a seventh aspect of the invention there is provided a kit for detecting an antigen comprising a solid support, a chromogenic substrate reagent and a probe of the fifth aspect of the invention; the solid phase carrier loads a solid phase antibody Ab2.
Preferably, the solid phase antibody Ab2 is capable of specifically binding to the same antigen as the antibody molecule Ab 1.
Preferably, the solid phase antibody Ab2 and the antibody molecule Ab1 are directed against different epitopes of the same antigen.
Preferably, the solid phase antibody Ab2 is from any one of mice, rats, goats, rabbits, monkeys, chickens, horses, donkeys, and sheep.
Preferably, the chromogenic substrate reagent is at least one of 3,3', 5' -tetramethylbenzidine, o-phenylenediamine, 2' -diazabis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt.
Preferably, the kit further comprises hydrogen peroxide.
Preferably, the kit further comprises an immunogen.
Preferably, the immunogen specifically binds to the antibody molecule Ab1 and the solid phase antibody Ab2.
Preferably, the immunogen and the solid phase antibody Ab2 and the probe may form an immuno-sandwich structure.
Preferably, the immunogen comprises a standard working fluid at a concentration of 0.1-1000ng/mL.
The beneficial effects of the invention are as follows:
the invention prepares the Au-loaded MoS 2 The material has high specific surface area and active binding site for biological recognition molecules, can bond the antibody by utilizing the double effects of physical adsorption and Au-S bond, can improve the immobilization capacity of the antibody and the stability of antibody modification, can directly modify the antibody without a coupling agent, is favorable for maintaining the activity of the antibody, and improves the sensitivity and accuracy of detection. The invention provides an Au-loaded MoS based on SCCA antibody molecule modification for the first time 2 MXene nanobiological probes having high nanoenzyme properties and high affinity for Squamous Cell Carcinoma Antigen (SCCA). The invention provides a colorimetric immune detection method for squamous cell carcinoma antigen, which realizes the detection of squamous cell carcinoma antigen SCCA, the detection range is 0.1ng/mL-1000ng/mL, the detection limit calculation equation is 3σ/k, wherein σ is the standard deviation of a negative control sample, k is the slope of a linear regression equation, and DeltaOD=0.36 log [ SCCA ]]+0.528, calculated to give a detection limit of 0.042ng/mL. The method has the advantages of simple operation, strong specificity, high sensitivity and high accuracy.
Drawings
FIG. 1 is an Au-loaded MoS 2 Transmission electron microscopy and high resolution-transmission electron microscopy of/MXene: wherein a represents Au-loaded MoS 2 Transmission electron microscope results of/MXene; b represents Au-loaded MoS 2 High resolution-transmission electron microscopy results of/MXene.
FIG. 2 is an Au-loaded MoS 2 /MXene(Au/MoS 2 Comparison of catalytic Activity before and after modification of SCCA-labeled antibody (Ab 1) by MXene, au-loaded MoS 2 MXene was prepared from example 1.
FIG. 3 is an Au-loaded MoS 2 /MXene(Au/MoS 2 Comparison of catalytic Activity before and after modification of SCCA-labeled antibody (Ab 1) by MXene, au-loaded MoS 2 MXene was prepared from example 2.
Fig. 4 is a detection principle and a detection flow chart.
FIG. 5 is a graph of UV-visible absorbance spectra of a substrate system at different standard concentrations of squamous cell carcinoma antigen SCCA.
FIG. 6 is a standard working curve for different concentrations of SCCA antigen.
Detailed Description
The conception and the technical effects produced by the present invention will be clearly and completely described in conjunction with the embodiments below to fully understand the objects, features and effects of the present invention. It is apparent that the described embodiments are only some embodiments of the present invention, but not all embodiments, and that other embodiments obtained by those skilled in the art without inventive effort are within the scope of the present invention based on the embodiments of the present invention.
Wherein MXene is titanium carbide Ti 3 C 2 Purchased from Jiangsu Xianfeng nanomaterials science and technology Co., ltd; squamous cell carcinoma antigen SCCA was purchased from the division of bioengineering (Shanghai), cat: d111414-0100; SCCA antibody (tag) molecule Ab1 was purchased from bio-engineering (Shanghai) Inc., cat No.: d194022-0100, mouse origin; SCCA solid phase antibody (coated) Ab2 was purchased from bio-engineering (Shanghai) Inc., cat No.: d194021-0100, mouse origin. The rest materials can be obtained through conventional purchasing in the market.
EXAMPLE 1 Au-loaded MoS 2 Preparation of MXene Material
(1)MoS 2 Preparation of MXene
0.242g (1 mmol) of sodium molybdate (Na 2 MoO 4 ·2H 2 O) and 0.38g (5 mmol) of thiourea (CH 4 N 2 S) in 24mL of ultrapure water, carrying out strong magnetic stirring for 10min to obtain sodium molybdate-thiourea mixed solution; then adding 6mL of 2.0mg/mL MXene into the ammonium molybdate-thiourea mixed solution, and continuing strong magnetic stirring for 30min to prepare sodium molybdate-thiourea-MXene mixed solution; transferring the mixed solution into a stainless steel reaction kettle with a polytetrafluoroethylene lining, reacting for 24 hours at 210 ℃, and naturally finishing the reactionCooling to room temperature, centrifuging at 6500rpm for 10min, alternately washing with absolute ethanol and ultrapure water for three times to obtain MoS 2 the/MXene product was finally resuspended in 45mL of ultrapure water for further use.
(2) Au-loaded MoS 2 Preparation of MXene
Taking the 12mL MoS 2 Adding 300 mu L of chloroauric acid solution with the mass fraction of 1% into a glass reaction bottle by MXene (3.82 mg/mL) at room temperature under ultrasonic, continuing to carry out ultrasonic treatment for 5min, stirring for 12h at room temperature, centrifuging at 6500rpm for 10min, washing with ultrapure water for three times, and carrying out vacuum freeze drying for 48h to obtain Au-loaded MoS 2 The product of MXene was ready for use.
Au-supported MoS of this embodiment 2 As a result of transmission electron microscopy of/MXene is shown in FIG. 1a, it can be observed that Au nanoparticles are uniformly dispersed in MoS having a three-dimensional flower morphology 2 Surface of MXene, demonstrating Au-loaded MoS 2 /MXene(Au/MoS 2 (MXene). Wherein Au is loaded with MoS 2 The particle size of/MXene is about 600nm, while the size of the Au nanoparticles on the surface is about 7.5nm.
Au-supported MoS of this embodiment 2 As shown in FIG. 1 b, the high resolution transmission electron microscope results of/MXene, in which a lattice spacing d=0.25 nm was observed, consistent with the (111) interplanar spacing of gold particles, demonstrated that gold nanoparticles were present in MoS 2 Successful loading of the MXene surface.
EXAMPLE 2 Au-loaded MoS 2 Preparation of MXene Material
The same as in example 1 except (2).
(2) Au-loaded MoS 2 Preparation of MXene
Taking the 12mL MoS 2 Adding 100 mu L of chloroauric acid solution with the mass fraction of 1% into a glass reaction bottle by MXene (3.82 mg/mL) at room temperature under ultrasonic, continuing to carry out ultrasonic treatment for 5min, stirring for 12h at room temperature, centrifuging at 6500rpm for 10min, washing with ultrapure water for three times, and carrying out vacuum freeze drying for 48h to obtain Au-loaded MoS 2 The product of MXene was ready for use.
EXAMPLE 3 Au-loaded MoS 2 Preparation of MXene Material
The same as in example 1 except (2).
(2) Au-loaded MoS 2 Preparation of MXene
Taking the 12mL MoS 2 Adding 500 μL chloroauric acid solution with mass fraction of 1% into glass reaction bottle at room temperature under ultrasonic, continuing ultrasonic for 5min, stirring at room temperature for 36h, centrifuging at 6500rpm for 10min, washing with ultrapure water for three times, and vacuum freeze drying for 48h to obtain Au-loaded MoS 2 The product of MXene was ready for use.
EXAMPLE 4 preparation of SCCA-labeled antibody (Ab 1) labeled modified nanobiological Probe
Taking 700 mu L of Au-loaded MoS with 1mg/mL 2 In EP tube, 200. Mu.L of 100mM potassium carbonate solution was used to adjust the pH to 8.5, 70. Mu.L of 200. Mu.g/mL SCCA antibody molecule (Ab 1) was added, incubated overnight in a 37℃shaker, centrifuged at 5000rpm for 10min, washed three times with PBS, and finally resuspended in Phosphate Buffer (PBS) at pH 7.4, 10mM to obtain a modified Au-loaded MoS of SCCA antibody molecule (Ab 1) 2 The MXene nano biological probe is stored in a refrigerator at 4 ℃ for standby.
The embodiment loads MoS on Au 2 After detecting the catalytic activity of the MXene modified SCCA antibody molecule (Ab 1), 194 mu L of acetic acid-acetate (pH=4) buffer solution with the concentration of 1M is taken and added into a micro-porous plate, 2 mu L of hydrogen peroxide solution with the concentration of 1M and 2 mu L of 3,3', 5' -tetramethyl benzidine solution with the concentration of 50mM are sequentially added, and 2 mu L of Au loaded MoS without Ab1 modification with the concentration of 1.0mg/mL are respectively added 2 Au-loaded MoS modified by MXene and Ab1 (20. Mu.g/mL) 2 mixing/MXene nano biological probe at room temperature for 10min; and then placing the microplate in a multifunctional enzyme-labeled instrument for testing, recording absorption spectrums under different wavelengths, and setting parameters: the wavelength is 350nm-850nm, and the step length is 2nm.
As shown in FIG. 2, au without Ab1 modification was used to support MoS 2 Au-loaded MoS modified by MXene and Ab1 (20. Mu.g/mL) 2 Absorption peaks at 375nm and 652nm were observed in the UV-visible absorption spectrum of the/MXene substrate system, which are the blue oxidation products (3, 3',5,UV-visible absorption characteristic peaks of charge complexes of 5' -tetramethylbenzene dinitrogen (TMBDI), illustrating Au-loaded MoS 2 MXene has peroxidase-like activity. Whereas Ab1 (20. Mu.g/mL) modified Au-loaded MoS 2 The absorption intensity of the MXene substrate system at 375nm and 652nm is lower than that of Au-loaded MoS without Ab1 modification 2 The absorption intensity of MXene shows that the interaction between amino and sulfhydryl groups of Ab1 antibody and nano gold reduces Au-loaded MoS 2 The catalytic site on the surface of/MXene also demonstrates that Ab1 antibody has been successfully immobilized on Au-loaded MoS by Au-N/Au-S bond and electrostatic adsorption 2 MXene surface.
EXAMPLE 5 preparation of SCCA-labeled antibody (Ab 1) labeled modified nanobiological probes
Taking 700 mu L of Au-loaded MoS with 1mg/mL 2 In EP tube, 200. Mu.L of 100mM potassium carbonate solution was used to adjust the pH to 8.5, 70. Mu.L of 200. Mu.g/mL SCCA antibody molecule (Ab 1) was added, incubated overnight in a 37℃shaker, centrifuged at 5000rpm for 10min, washed three times with PBS, and finally resuspended in Phosphate Buffer (PBS) at pH 7.4, 10mM to obtain a modified Au-loaded MoS of SCCA antibody molecule (Ab 1) 2 The MXene nano biological probe is stored in a refrigerator at 4 ℃ for standby.
The embodiment loads MoS on Au 2 After detecting the catalytic activity of the MXene modified SCCA antibody molecule (Ab 1), 194 mu L of acetic acid-acetate (pH=4) buffer solution with the concentration of 1M is taken and added into a micro-porous plate, 2 mu L of hydrogen peroxide solution with the concentration of 1M and 2 mu L of 3,3', 5' -tetramethyl benzidine solution with the concentration of 50mM are sequentially added, and 2 mu L of Au loaded MoS without Ab1 modification with the concentration of 1.0mg/mL are respectively added 2 Au-loaded MoS modified by MXene and Ab1 (20. Mu.g/mL) 2 mixing/MXene nano biological probe at room temperature for 10min; and then placing the microplate in a multifunctional enzyme-labeled instrument for testing, recording absorption spectrums under different wavelengths, and setting parameters: the wavelength is 350nm-850nm, and the step length is 2nm.
As shown in FIG. 3, au without Ab1 modification was used to support MoS 2 Au-loaded MoS modified by MXene and Ab1 (20. Mu.g/mL) 2 Observation in the UV-visible absorption Spectrum of the MXene substrate SystemBy the time of the occurrence of absorption peaks at 375nm and 652nm, which are characteristic peaks of the UV-visible absorption of the blue oxidation product (charge complex of 3,3', 5' -tetramethylbenzene dinitrogen (TMBDI)) of the substrate 3,3', 5' -Tetramethylbenzidine (TMB), illustrating Au-loaded MoS 2 MXene has peroxidase-like activity. Whereas Ab1 (20. Mu.g/mL) modified Au-loaded MoS 2 The absorption intensity of the MXene substrate system at 375nm and 652nm is lower than that of Au-loaded MoS without Ab1 modification 2 The absorption intensity of MXene shows that the interaction between amino and sulfhydryl groups of Ab1 antibody and nano gold reduces Au-loaded MoS 2 The catalytic site on the surface of/MXene also demonstrates that Ab1 antibody has been successfully immobilized on Au-loaded MoS by Au-N/Au-S bond and electrostatic adsorption 2 MXene surface.
Example 6A method of colorimetric immunodetection of squamous cell carcinoma antigen
The detection principle and detection flow of the embodiment are shown in fig. 4, and the specific steps are as follows:
(1) 100. Mu.L of squamous cell carcinoma SCCA coated antibody (Ab 2) at 10. Mu.g/mL was added to the microwell plate and incubated overnight in a 4 degree refrigerator;
(2) After washing three times by adding 200. Mu.L of Phosphate Buffer Solution (PBS) with pH of 7.4 to the coated microwells, 100. Mu.L of Bovine Serum Albumin (BSA) with mass fraction of 1% was added, and incubated for 2.52h with a 37 degree shaker to block non-specific adsorption sites;
(3) After further three washes with 200. Mu.L of Phosphate Buffered Saline (PBS) at pH 7.4, 100. Mu.L of squamous cell carcinoma antigen (0.1 ng/mL-1000 ng/mL) at different concentration gradients was added and incubated for 1h at 37℃on a shaker;
(4) After the washing three times with 200. Mu.L of Phosphate Buffer Solution (PBS) having a pH of 7.4, 100. Mu.L of Au-loaded MoS modified with 0.1mg/mL Ab 1-labeled antibody was added 2 MXene nanoprobe (prepared in example 4), 37℃shaking table incubation for 1h;
(5) After further washing three times with 200. Mu.L of Phosphate Buffer Solution (PBS) having a pH of 7.4, an immunocompetent structure was prepared and stored in a 4℃refrigerator for further use.
(6) 100 μl of 4 is added to microwells containing an immunocompetent structure0mM hydrogen peroxide (H) 2 O 2 ) And 3.0mm of 3,3', 5' -Tetramethylbenzidine (TMB) in acetate buffer (ph=4.0) for 10min at room temperature;
(7) Detecting the micropores by using a multifunctional enzyme-labeled instrument, recording the absorbance at 652nm, and setting test parameters: the wavelength range is 350-850nm, and the step length is 2nm.
The UV-visible absorbance spectra of the substrate system at different concentrations of SCCA antigen are shown in FIG. 4, from which it can be seen that the absorbance intensity of the substrate system increases with increasing concentration of squamous cell carcinoma antigen SCCA (0.1, 1, 10, 100, 250, 500, 1000 ng/mL), indicating a positive correlation between the two.
A series of SCCA (0.1, 1, 10, 100, 250, 500, 1000 ng/mL) antigens with different concentrations are used as standard substances, and the absorbance change delta OD and log [ SCCA ] at the wavelength of 652nm is used]Establishing a standard curve, as shown in FIG. 5, wherein DeltaOD has a good linear relationship with the logarithm of squamous cell carcinoma antigen SCCA concentration, the linear range is 0.1-1000ng/mL, the detection limit calculation equation is 3σ/k, wherein σ is the standard deviation of a negative control sample, k is the slope of a linear regression equation, deltaOD=0.36 log [ SCCA ]]+0.528，R 2 =0.988, and the detection limit was calculated to be 0.042ng/mL.
Claims (10)
1. Au-loaded MoS 2 MXene, the Au is loaded on the MoS 2 Surface of/Mxene;
preferably, the MoS 2 MXene is in three-dimensional flower shape;
preferably, the MXene contains M and X elements, wherein M is one or more of metal elements in a transition metal group, and X is one or two of carbon and nitrogen;
preferably, the M is selected from one or more of Ti, zr, hf, V, nb, ta, cr, sc, mo, W elements;
preferably, the MXene comprises Ti 3 C 2 。
2. The Au-supported MoS of claim 1 2 The preparation method of the MXene is characterized in that:
(1) Preparing MoS by adopting a hydrothermal method 2 /MXene；
(2) The Au-loaded MoS is prepared by adopting an in-situ growth method 2 /MXene。
3. The preparation method according to claim 2, characterized in that:
preparing MoS by adopting a hydrothermal method in the step (1) 2 the/MXene is specifically: mixing sodium molybdate, thiourea and MXene, and reacting to obtain MoS 2 /MXene；
Preferably, the reaction condition is 180-240 ℃ for 18-30 h;
preferably, the mass ratio of the sodium molybdate to the thiourea to the MXene is (18-22): (30-32): 1, a step of;
preferably, the mixing of sodium molybdate, thiourea and MXene is specifically as follows: firstly mixing sodium molybdate and thiourea to obtain a sodium molybdate-thiourea mixed solution; mixing the sodium molybdate-thiourea mixed solution with MXene for the second time to obtain a sodium molybdate-thiourea-MXene mixed solution;
preferably, the time of the first mixing is 5-15 min;
preferably, the time of the second mixing is 20-40 min;
preferably, the first mixing and the second mixing are stirring; further stirring by strong magnetic force;
preferably, the reaction further comprises the following steps: solid-liquid separation to obtain MoS 2 MXene; washing the obtained MoS 2 /Mxene。
4. The preparation method according to claim 2, characterized in that:
in the step (2), an in-situ growth method is adopted to prepare the Au-loaded MoS 2 the/MXene is specifically: the MoS is subjected to 2 mixing/MXene with chloroauric acid, and reacting to obtain Au-loaded MoS 2 /MXene；
Preferably, the chloroauric acid and MoS 2 The mass ratio of the/MXene is 1: (9-45);
preferably, the MoS 2 The mixing of/MXene and chloroauric acid is specifically as follows: adding chloroauric acid into MoS under ultrasonic condition 2 in/MXene;
preferably, the reaction time is 12-36 hours;
preferably, the reaction is carried out under stirring conditions; further under stirring at room temperature;
preferably, the reaction further comprises the following steps: solid-liquid separation to obtain Au-loaded MoS 2 MXene; washing and drying.
5. The Au-supported MoS of claim 1 2 Use of/MXene for the preparation of a product for detecting a protein;
preferably, the product detects proteins by immunochemistry;
preferably, the product comprises a biorecognition molecule and an Au-loaded MoS 2 MXene, said biological recognition molecule specifically recognizing said protein;
preferably, the biological recognition molecule comprises at least one of an antigen, an antibody, a polypeptide; further antibodies; still further squamous cell carcinoma antibodies;
preferably, the biological recognition molecule is supported on the Au MoS through coordination bond 2 Au binding in MXene; further through a sulfur gold bond and the Au loaded MoS 2 Au binding in MXene.
6. A protein detection product comprising a biological recognition molecule and the Au-loaded MoS of claim 1 2 /MXene，
The biological recognition molecule specifically recognizes the protein;
preferably, the biological recognition molecule comprises at least one of an antigen, an antibody, a polypeptide; further antibodies; still further squamous cell carcinoma antibodies;
preferably, the biological recognition molecule is supported on the Au MoS through coordination bond 2 Au binding in MXene; further through a gold sulfide bond, the Au is loaded with MoS 2 Au binding in MXene.
7. A probe for detecting an antigen comprising an antibody molecule Ab1 that specifically binds to the antigen and the Au-loaded MoS of claim 1 2 /MXene；
The antibody molecule Ab1 is supported on the Au-supported MoS through a coordination bond 2 Au binding in MXene; further through a sulfur gold bond and the Au loaded MoS 2 Au binding in MXene;
preferably, the species of the antibody molecule Ab1 is selected from any one of mouse, rat, goat, rabbit, monkey, chicken, horse, donkey, and sheep;
preferably, the antigen comprises at least one of carcinoembryonic antigen, alpha fetoprotein, carbohydrate antigen 125, carbohydrate antigen 15-3, carbohydrate antigen 72-4, carbohydrate antigen 24-2, carbohydrate antigen 50, prostate specific antigen, squamous cell carcinoma antigen, cytokeratin 19 fragment, gastrin-releasing peptide precursor, neuron specific enolase, nuclear matrix protein-22, tartrate-resistant acid phosphatase 5b, tumor M2-type pyruvate kinase, abnormal prothrombin, osteopontin, phosphatidylinositol proteoglycan 3, golgi protein 73, metalloprotease 1, human gastric carcinoma MG7 antigen, pepsinogen, lectin DC-SIGN, and human chromogranin a.
8. The method for preparing the probe according to claim 7, comprising the steps of:
antibody molecule Ab1 was loaded with Au to MoS 2 mixing/MXene, and incubating to obtain the Au-loaded MoS modified with antibody molecule Ab1 2 /MXene；
Preferably, the incubation condition is that the shaking incubation is carried out for 8-24 hours at the temperature of 4-40 ℃;
preferably, the antibody molecules Ab1 and Au are loaded with MoS 2 The mass ratio of the/MXene is 1: (45-55);
preferably, the incubation further comprises the following steps: solid-liquid separation is carried out to obtain the Au-loaded MoS modified with the antibody molecule Ab1 2 MXene, washing.
9. A kit for detecting an antigen comprising a solid support, a chromogenic substrate reagent, and the probe of claim 7;
the solid phase carrier loads a solid phase antibody Ab2, the solid phase antibody Ab2 specifically binds to the antigen, and the solid phase antibody Ab2 and the antibody molecule Ab1 aim at different epitopes of the same antigen;
preferably, the solid phase antibody Ab2 is from any one of mice, rats, goats, rabbits, monkeys, chickens, horses, donkeys, and sheep.
10. The kit of claim 9, wherein:
the chromogenic substrate reagent comprises at least one of 3,3', 5' -tetramethyl benzidine, o-phenylenediamine and 2,2' -diazabis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt;
preferably, the kit further comprises hydrogen peroxide.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311767586.XA CN117871847A (en) | 2023-12-20 | 2023-12-20 | Colorimetric immunoassay reagent and preparation method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311767586.XA CN117871847A (en) | 2023-12-20 | 2023-12-20 | Colorimetric immunoassay reagent and preparation method and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN117871847A true CN117871847A (en) | 2024-04-12 |
Family
ID=90580310
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202311767586.XA Pending CN117871847A (en) | 2023-12-20 | 2023-12-20 | Colorimetric immunoassay reagent and preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117871847A (en) |
-
2023
- 2023-12-20 CN CN202311767586.XA patent/CN117871847A/en active Pending
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN110763834B (en) | Method, reagent and kit for detecting content of immune marker | |
| US20190112356A1 (en) | High-affinity immunopolymers | |
| CN105651990A (en) | Chemiluminiscence detection kit for 17alpha-hydroxyprogesterone and preparation method and application thereof | |
| KR101486149B1 (en) | Method of detecting a marker for diagnosing a desease using a competetive immunoassay based on a surface-enhanced raman scattering | |
| KR101597413B1 (en) | High-sensitive lateral flow immunoassay chip using enzyme-mimetic nanoparticles and Methods sensing using thereof | |
| Chi et al. | In situ amplified QCM immunoassay for carcinoembryonic antigen with colorectal cancer using horseradish peroxidase nanospheres and enzymatic biocatalytic precipitation | |
| JPS61217763A (en) | Immunological testing method | |
| CN112698041A (en) | Compound, growth differentiation factor 15 detection kit thereof and application | |
| CN107356743B (en) | Assay kit for detecting myoglobin | |
| TW201802472A (en) | Anti-human hemoglobin monoclonal antibody or antibody kit, insoluble carrier particle to which anti-human hemoglobin monoclonal antibody is immobilized, and measurement reagent and measurement method using same | |
| Liang et al. | Ratiometric fluorescence enzyme-linked immunosorbent assay based on carbon dots@ SiO 2@ CdTe quantum dots with dual functionalities for alpha-fetoprotein | |
| US20150185213A1 (en) | Method of internal correction in one chip assay and method for measuring test substance using said method | |
| Liu et al. | Fluorescent microsphere immunochromatographic assays for detecting bone alkaline phosphatase based on biolayer interferometry-selected antibody | |
| CN106645756A (en) | Kit for detecting NMP22 (Nuclear Matrix Protein 22) and preparation method thereof | |
| Fan et al. | Precisely designed growth of dual-color quantum dots bilayer nanobeads for ratiometric fluorescent immunoassay | |
| CN112986586A (en) | Amino-terminal brain natriuretic peptide detection method and kit | |
| CN117871847A (en) | Colorimetric immunoassay reagent and preparation method and application thereof | |
| Wu et al. | Preparation of bimetal–polydopamine organic frameworks with core–shell structure and their application in HER2 detection | |
| CN111781357A (en) | Kit for simultaneously detecting four tumor markers of lung cancer and application thereof | |
| CN111693689A (en) | Nano enzyme for enzymatic chemiluminescence detection and application thereof | |
| CN116183910A (en) | Immunofluorescence chromatography test paper for rapidly detecting human whole blood P-tau-181 protein and preparation method thereof | |
| EP2707720A2 (en) | Diagnosis and prognosis of triple negative breast and ovarian cancer | |
| EP4119945A1 (en) | Highly sensitive immunoconjugate, preparing method thereof, in vitro diagnostic reagent and in vitro diagnostic kit including the same | |
| EP2660603B1 (en) | Immunological assay method | |
| CN115290883A (en) | Synchronous detection method for multiple exosome proteins |
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 |