CN110824177B - Rapid detection test paper and preparation method and application thereof - Google Patents
Rapid detection test paper and preparation method and application thereof Download PDFInfo
- Publication number
- CN110824177B CN110824177B CN201910859373.7A CN201910859373A CN110824177B CN 110824177 B CN110824177 B CN 110824177B CN 201910859373 A CN201910859373 A CN 201910859373A CN 110824177 B CN110824177 B CN 110824177B
- Authority
- CN
- China
- Prior art keywords
- microspheres
- pad
- conjugate
- test strip
- membrane
- 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.)
- Active
Links
- 238000001514 detection method Methods 0.000 title claims abstract description 84
- 238000012360 testing method Methods 0.000 title claims abstract description 39
- 238000002360 preparation method Methods 0.000 title claims abstract description 30
- 239000012528 membrane Substances 0.000 claims abstract description 75
- 102000036639 antigens Human genes 0.000 claims abstract description 14
- 108091007433 antigens Proteins 0.000 claims abstract description 14
- 239000000427 antigen Substances 0.000 claims abstract description 13
- 229940079593 drug Drugs 0.000 claims abstract description 10
- 239000003814 drug Substances 0.000 claims abstract description 10
- 239000004005 microsphere Substances 0.000 claims description 188
- 238000003908 quality control method Methods 0.000 claims description 50
- 230000027455 binding Effects 0.000 claims description 24
- 229920001220 nitrocellulos Polymers 0.000 claims description 19
- 239000000020 Nitrocellulose Substances 0.000 claims description 18
- 239000011230 binding agent Substances 0.000 claims description 12
- 241000283973 Oryctolagus cuniculus Species 0.000 claims description 11
- 241000283707 Capra Species 0.000 claims description 10
- 239000000463 material Substances 0.000 claims description 10
- 239000003365 glass fiber Substances 0.000 claims description 9
- 238000010521 absorption reaction Methods 0.000 claims description 8
- 239000003086 colorant Substances 0.000 claims description 8
- 238000009597 pregnancy test Methods 0.000 claims description 7
- 239000002245 particle Substances 0.000 claims description 5
- 239000004793 Polystyrene Substances 0.000 claims description 3
- 229940127121 immunoconjugate Drugs 0.000 claims description 3
- 229920002223 polystyrene Polymers 0.000 claims description 3
- 239000002994 raw material Substances 0.000 claims description 3
- 238000004587 chromatography analysis Methods 0.000 claims description 2
- 238000004811 liquid chromatography Methods 0.000 claims description 2
- 229920006267 polyester film Polymers 0.000 claims description 2
- 230000035935 pregnancy Effects 0.000 abstract description 5
- 239000002244 precipitate Substances 0.000 description 48
- 239000000243 solution Substances 0.000 description 44
- 239000006228 supernatant Substances 0.000 description 32
- 239000011535 reaction buffer Substances 0.000 description 30
- 238000009210 therapy by ultrasound Methods 0.000 description 26
- 238000007664 blowing Methods 0.000 description 22
- 230000010355 oscillation Effects 0.000 description 22
- 238000006243 chemical reaction Methods 0.000 description 20
- 239000011248 coating agent Substances 0.000 description 20
- 238000000576 coating method Methods 0.000 description 20
- 238000003756 stirring Methods 0.000 description 20
- 239000000047 product Substances 0.000 description 17
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 description 16
- 239000003153 chemical reaction reagent Substances 0.000 description 14
- ZPUCINDJVBIVPJ-LJISPDSOSA-N ***e Chemical compound O([C@H]1C[C@@H]2CC[C@@H](N2C)[C@H]1C(=O)OC)C(=O)C1=CC=CC=C1 ZPUCINDJVBIVPJ-LJISPDSOSA-N 0.000 description 14
- BQJCRHHNABKAKU-KBQPJGBKSA-N morphine Chemical compound O([C@H]1[C@H](C=C[C@H]23)O)C4=C5[C@@]12CCN(C)[C@@H]3CC5=CC=C4O BQJCRHHNABKAKU-KBQPJGBKSA-N 0.000 description 14
- HZAXFHJVJLSVMW-UHFFFAOYSA-N 2-Aminoethan-1-ol Chemical compound NCCO HZAXFHJVJLSVMW-UHFFFAOYSA-N 0.000 description 12
- 238000004140 cleaning Methods 0.000 description 12
- 108010044467 Isoenzymes Proteins 0.000 description 11
- 239000007788 liquid Substances 0.000 description 11
- 239000000203 mixture Substances 0.000 description 11
- 102000004420 Creatine Kinase Human genes 0.000 description 10
- 108010042126 Creatine kinase Proteins 0.000 description 10
- 102000036675 Myoglobin Human genes 0.000 description 10
- 108010062374 Myoglobin Proteins 0.000 description 10
- 102100036859 Troponin I, cardiac muscle Human genes 0.000 description 10
- 101710128251 Troponin I, cardiac muscle Proteins 0.000 description 10
- 239000003085 diluting agent Substances 0.000 description 10
- 239000012491 analyte Substances 0.000 description 9
- 238000000034 method Methods 0.000 description 9
- 230000000903 blocking effect Effects 0.000 description 8
- 238000005119 centrifugation Methods 0.000 description 8
- 238000011161 development Methods 0.000 description 8
- 229960001252 methamphetamine Drugs 0.000 description 8
- MYWUZJCMWCOHBA-VIFPVBQESA-N methamphetamine Chemical compound CN[C@@H](C)CC1=CC=CC=C1 MYWUZJCMWCOHBA-VIFPVBQESA-N 0.000 description 8
- 238000005507 spraying Methods 0.000 description 8
- 239000012089 stop solution Substances 0.000 description 8
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 description 7
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 description 7
- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 description 7
- 229960003920 ***e Drugs 0.000 description 7
- 229960005181 morphine Drugs 0.000 description 7
- 239000000126 substance Substances 0.000 description 7
- 230000000007 visual effect Effects 0.000 description 7
- 102000011022 Chorionic Gonadotropin Human genes 0.000 description 6
- 108010062540 Chorionic Gonadotropin Proteins 0.000 description 6
- 230000002159 abnormal effect Effects 0.000 description 6
- 239000002390 adhesive tape Substances 0.000 description 6
- 238000010009 beating Methods 0.000 description 6
- 230000008878 coupling Effects 0.000 description 6
- 238000010168 coupling process Methods 0.000 description 6
- 238000005859 coupling reaction Methods 0.000 description 6
- 229940084986 human chorionic gonadotropin Drugs 0.000 description 6
- 238000012123 point-of-care testing Methods 0.000 description 6
- 230000001376 precipitating effect Effects 0.000 description 6
- 238000004321 preservation Methods 0.000 description 6
- KWTSXDURSIMDCE-QMMMGPOBSA-N (S)-amphetamine Chemical compound C[C@H](N)CC1=CC=CC=C1 KWTSXDURSIMDCE-QMMMGPOBSA-N 0.000 description 5
- 229940025084 amphetamine Drugs 0.000 description 5
- 238000007865 diluting Methods 0.000 description 5
- 238000002156 mixing Methods 0.000 description 5
- 239000007790 solid phase Substances 0.000 description 5
- 238000003260 vortexing Methods 0.000 description 5
- 241000218236 Cannabis Species 0.000 description 4
- 238000010586 diagram Methods 0.000 description 4
- 238000010790 dilution Methods 0.000 description 4
- 239000012895 dilution Substances 0.000 description 4
- 238000001035 drying Methods 0.000 description 4
- 238000002372 labelling Methods 0.000 description 4
- 238000004519 manufacturing process Methods 0.000 description 4
- 239000007921 spray Substances 0.000 description 4
- 230000001360 synchronised effect Effects 0.000 description 4
- 101100512901 Caenorhabditis elegans mes-4 gene Proteins 0.000 description 3
- 244000025254 Cannabis sativa Species 0.000 description 3
- 235000012766 Cannabis sativa ssp. sativa var. sativa Nutrition 0.000 description 3
- 235000012765 Cannabis sativa ssp. sativa var. spontanea Nutrition 0.000 description 3
- 101100512902 Drosophila melanogaster Mes-4 gene Proteins 0.000 description 3
- 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 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- 229920001213 Polysorbate 20 Polymers 0.000 description 3
- 239000007983 Tris buffer Substances 0.000 description 3
- 238000004220 aggregation Methods 0.000 description 3
- 230000002776 aggregation Effects 0.000 description 3
- XAGFODPZIPBFFR-UHFFFAOYSA-N aluminium Chemical compound [Al] XAGFODPZIPBFFR-UHFFFAOYSA-N 0.000 description 3
- 229910052782 aluminium Inorganic materials 0.000 description 3
- 235000009120 camo Nutrition 0.000 description 3
- 235000005607 chanvre indien Nutrition 0.000 description 3
- 238000005520 cutting process Methods 0.000 description 3
- 239000011888 foil Substances 0.000 description 3
- 239000011487 hemp Substances 0.000 description 3
- 230000005764 inhibitory process Effects 0.000 description 3
- 239000008188 pellet Substances 0.000 description 3
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 3
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 3
- 230000008569 process Effects 0.000 description 3
- 238000007789 sealing Methods 0.000 description 3
- 229910052708 sodium Inorganic materials 0.000 description 3
- 239000011734 sodium Substances 0.000 description 3
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 3
- 238000002604 ultrasonography Methods 0.000 description 3
- 238000011179 visual inspection Methods 0.000 description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 3
- 102100025036 Norrin Human genes 0.000 description 2
- 101710085992 Norrin Proteins 0.000 description 2
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 description 2
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 2
- 229930006000 Sucrose Natural products 0.000 description 2
- 239000013504 Triton X-100 Substances 0.000 description 2
- 229920004890 Triton X-100 Polymers 0.000 description 2
- 239000000969 carrier Substances 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 238000003255 drug test Methods 0.000 description 2
- 238000000338 in vitro Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 230000009871 nonspecific binding Effects 0.000 description 2
- 238000001556 precipitation Methods 0.000 description 2
- 238000003825 pressing Methods 0.000 description 2
- 238000002203 pretreatment Methods 0.000 description 2
- 230000000717 retained effect Effects 0.000 description 2
- 238000006748 scratching Methods 0.000 description 2
- 230000002393 scratching effect Effects 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 238000001179 sorption measurement Methods 0.000 description 2
- 238000003860 storage Methods 0.000 description 2
- 239000005720 sucrose Substances 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 208000035473 Communicable disease Diseases 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 241001529936 Murinae Species 0.000 description 1
- 108091000080 Phosphotransferase Proteins 0.000 description 1
- 102000013394 Troponin I Human genes 0.000 description 1
- 108010065729 Troponin I Proteins 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- -1 amphetamines Chemical compound 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000007767 bonding agent Substances 0.000 description 1
- 229910021538 borax Inorganic materials 0.000 description 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 1
- 230000000747 cardiac effect Effects 0.000 description 1
- 230000000536 complexating effect Effects 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 229960003624 creatine Drugs 0.000 description 1
- 239000006046 creatine Substances 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- UQGFMSUEHSUPRD-UHFFFAOYSA-N disodium;3,7-dioxido-2,4,6,8,9-pentaoxa-1,3,5,7-tetraborabicyclo[3.3.1]nonane Chemical compound [Na+].[Na+].O1B([O-])OB2OB([O-])OB1O2 UQGFMSUEHSUPRD-UHFFFAOYSA-N 0.000 description 1
- 239000003596 drug target Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 1
- 229910002804 graphite Inorganic materials 0.000 description 1
- 239000010439 graphite Substances 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 239000004816 latex Substances 0.000 description 1
- 229920000126 latex Polymers 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 238000013508 migration Methods 0.000 description 1
- 230000005012 migration Effects 0.000 description 1
- 208000010125 myocardial infarction Diseases 0.000 description 1
- 210000004165 myocardium Anatomy 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 102000020233 phosphotransferase Human genes 0.000 description 1
- 229920000728 polyester Polymers 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 235000020183 skimmed milk Nutrition 0.000 description 1
- 229910000029 sodium carbonate Inorganic materials 0.000 description 1
- 239000001509 sodium citrate Substances 0.000 description 1
- 235000010339 sodium tetraborate Nutrition 0.000 description 1
- 239000004328 sodium tetraborate Substances 0.000 description 1
- 238000010532 solid phase synthesis reaction Methods 0.000 description 1
- 238000011895 specific detection Methods 0.000 description 1
- 230000001502 supplementing effect Effects 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- HRXKRNGNAMMEHJ-UHFFFAOYSA-K trisodium citrate Chemical compound [Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O HRXKRNGNAMMEHJ-UHFFFAOYSA-K 0.000 description 1
- 229940038773 trisodium citrate Drugs 0.000 description 1
- 210000002700 urine Anatomy 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Images
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
- G01N33/94—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving narcotics or drugs or pharmaceuticals, neurotransmitters or associated receptors
- G01N33/948—Sedatives, e.g. cannabinoids, barbiturates
-
- 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
- G01N33/558—Immunoassay; Biospecific binding assay; Materials therefor using diffusion or migration of antigen or antibody
-
- 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
-
- 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
- G01N33/74—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving hormones or other non-cytokine intercellular protein regulatory factors such as growth factors, including receptors to hormones and growth factors
- G01N33/76—Human chorionic gonadotropin including luteinising hormone, follicle stimulating hormone, thyroid stimulating hormone or their receptors
-
- 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
- G01N2333/575—Hormones
- G01N2333/59—Follicle-stimulating hormone [FSH]; Chorionic gonadotropins, e.g. HCG; Luteinising hormone [LH]; Thyroid-stimulating hormone [TSH]
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Molecular Biology (AREA)
- Hematology (AREA)
- Chemical & Material Sciences (AREA)
- Urology & Nephrology (AREA)
- Biomedical Technology (AREA)
- Microbiology (AREA)
- Pathology (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- General Physics & Mathematics (AREA)
- General Health & Medical Sciences (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- Endocrinology (AREA)
- Anesthesiology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Pharmacology & Pharmacy (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Reproductive Health (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention provides a rapid detection test paper and a preparation method and application thereof, wherein the test paper comprises a chromatographic membrane and a combination pad, and one end of the chromatographic membrane is connected with the combination pad; the detection line is an antigen/antibody corresponding to different analytes fixed at certain intervals. The invention also provides application of the test paper in drug and early pregnancy detection.
Description
Technical Field
The invention belongs to the technical field of immunochemistry in-vitro detection, and particularly relates to novel rapid detection test paper as well as a preparation method and application thereof
Background
In recent years, with the development of scientific technology, the progress of medical science, and the efficient and fast-paced working mode, POCT with the miniaturization of experimental instruments, the simplification of operations, and the immediacy of report results is becoming more and more popular.
The point-of-care testing (POCT) is a testing method that is performed on a sampling site and uses a portable analyzer and a matching reagent to quickly obtain a testing result, and is all tests performed outside a traditional, core or central laboratory.
POCT is a new field for testing medical development and is one of the most potential fields in IVD industry, and represents the future development direction of IVD. As a new clinical detection means, POCT has the characteristics of rapidness, simplicity, convenience, field analysis and the like, can reduce the sample transfer process, shorten the report time, conform to the fast-paced working mode of the modern society and meet the personalized service requirement
Common POCT in-vitro diagnosis and detection reagent products mainly comprise: pregnancy series, drug series, cardiac markers and infectious disease series, etc., the most common methodologies are colloidal gold method, latex labeling method, etc. In terms of product process, four parts of materials, namely a sample adding pad, a combination pad, a chromatographic membrane and a sample absorbing pad, are generally adhered to a bottom plate in sequence, and in the production and manufacturing process, because different components are frequently connected for many times and part of the components are adhered according to directions, the directions are opposite in the operation process or the detection sample is incompletely moved; in terms of result display, a color, such as red, blue, etc., is generally displayed in the result observation region. As a rapid detection reagent, normally, the quality control line must be developed after the test is finished, but in actual use, because specific detection lines and quality control line color development local marks are not available in test strip products, the condition that the quality control line is not developed and result judgment is wrong may occur
Disclosure of Invention
The invention mainly aims to provide test paper with different colors in a result display area, and the technical problem to be solved is that the corresponding result of an analyte is correctly judged under the condition that the existing test paper strip has no corresponding detection line and quality control line position indication.
The invention also aims to provide a preparation method of the test paper, which omits a sample adding pad and an absorption pad of the prior similar products, controls the absorption capacity of a sample by selecting the length of a binding pad and components such as a surfactant in a pretreatment solution, and achieves the aim of not needing the absorption pad.
In order to achieve the above object, the present invention provides a method for labeling antigen antibodies corresponding to different analytes by using microspheres of various colors, and then mixing the labeled antigen antibodies and the labeled microspheres together and spraying the mixture onto one end of a binding pad; meanwhile, antigen and antibody corresponding to different analytes are sprayed at one end of the chromatographic membrane according to a certain interval, and a quality control line is fixed at the farthest end and used for judging whether the reaction system is normal or not.
In one aspect, the invention provides a rapid test strip, which is characterized in that: comprises a chromatographic membrane 2 and a combined pad 3, wherein one end of the chromatographic membrane 2 is connected with the combined pad 3; the combination pad 3 is pre-sprayed with antigen/antibody conjugates 4 labeled by microspheres of different colors corresponding to different analytes, the chromatographic membrane 2 is sequentially provided with a detection line 5 and a quality control line 6 along the liquid chromatography direction, and the detection line 5 is fixed at certain intervals and is used for detecting the antigens/antibodies corresponding to the different analytes. Wherein, the number of the detection lines 5 is 1-8, preferably 1-5; the number of the quality control lines is 1.
Furthermore, one end of the combination pad 3 is sprayed with the labeled antigen/antibody combination 4 marked by microspheres with different colors, and the color of the microspheres can be more than 2 of red, blue, black, orange, green, yellow and pink. Among them, the microsphere is preferably red, blue and black in color.
Further, it is characterized by not comprising a separate sample addition pad and an absorption pad.
Further, the width of the chromatographic carrier 2 and the bonding pad 3 is 3-10 mm. Wherein the width of the chromatographic carrier 2 and the bonding pad 3 is preferably 4-8 mm.
Further, the length of the combination pad 3 is 3-10 cm, and the length of the chromatographic membrane 2 is 3-8 cm. Wherein the length of the binding pad 3 is preferably 3-6 cm, and the length of the chromatographic membrane 2 is preferably 4-6 cm.
Further, the chromatographic carrier 2 and the bonding pad 3 are sequentially adhered to the base plate 1.
Further, the chromatographic carrier 2 and the bonding pad 3 adhered to the base plate 1 are fixed to the cartridge case 12-1 through the corresponding fixing holes 7 and 8, respectively.
Further, the microspheres are selected from polystyrene color microspheres, black polymer microspheres and graphite microspheres. The polystyrene color microsphere can be unmodified, carboxyl modified, amino modified and the like.
Furthermore, the particle size of the microspheres is 20nm-4 μm. Wherein the particle size of the microspheres is preferably 300-400 nm.
Further, the raw materials used in the quality control line 6 and the combination 4 are as follows: goat anti-rabbit IgG and rabbit IgG, or rabbit anti-goat IgG and goat IgG.
Further, the chromatographic membrane material is selected from nitrocellulose membrane and fluorescent nitrocellulose membrane, and the combined pad material is selected from glass fiber and polyester film. Wherein the chromatographic membrane material is preferably nitrocellulose membrane (backing), and the bonding pad material is preferably polyester membrane.
In another aspect, the invention provides the use of the rapid test strip in drug or early pregnancy detection.
In addition, the rapid test strip of the present invention comprises:
the contact part of the chromatographic carrier 2 and the binding pad 3 can be a straight edge, preferably a semicircular edge; the contact portion of the bonding pad 3 is an isosceles trapezoid or a 1/4 circle with two oblique sides, and the upper side is 2-6 mm, preferably 2-4 mm.
The corresponding shape of the fixing holes 7 and 8 on the box-shaped card shell 12-1 is preferably polygonal, the diameter is 1-3 mm, and preferably 1-2 mm; the fixing hole 7 on the conjugate pad 3 is between the sample application region and the conjugate 4, near the application region, and is 42-5 cm, preferably 2-4 cm, away from the conjugate; the fixed orifices of the chromatographic carrier 2 are not between the quality control line 6 and the binding substance 4, and are located at the most distant end of the chromatographic carrier in the chromatographic direction, and the distance is preferably 3 to 6 mm.
When the reagent is used in a sandwich product and sample detection is carried out, an analyte in a sample is firstly combined with a colored microsphere marked by an antigen antibody corresponding to the analyte to form a compound of colored microsphere-antigen-analyte, and under the action of chromatography, the compound moves forwards on a chromatographic membrane 2 and is captured by a solid phase antigen and an antibody of a detection line corresponding to the analyte. If the concentration of the analyte in the sample is not lower than the detection limit of the reagent, a set colored line appears at the position of the corresponding detection line, and a positive result is presented; if the position of the detection line does not develop color, the result is negative; the reagent is a product of an inhibition method (including a neutralization inhibition method and a competition inhibition method), and when a sample is detected, a detection line of a corresponding analyte does not develop color, namely a positive result, and when the corresponding color is displayed, the detection line of the corresponding analyte is a negative result. Whether the sample to be tested contains the corresponding analyte or not, a quality control line 6 always appears on the chromatographic membrane and is used as a standard for judging whether the sample is sufficient or not and whether the operation is correct or not.
Compared with the prior art, the invention achieves the following remarkable progress:
1. regarding the reagent assembling structure: the sample is directly added on the conjugate adsorption pad without a sample pad, so that the uneven quick release of the sample can be avoided, the product sensitivity is improved, and the product result repeatability is increased; and the water absorption paper is not arranged, so that the backflow of the sample is overcome, and the possibility of abnormal results is avoided. 2. Different multiplex analytes were labeled with different colored microspheres: the judgment of the detection result is more visual; non-specific binding between the same color label binders is avoided, and the probability of abnormal results is reduced. 1. Regarding the reagent assembling structure: the sample is directly added on the conjugate adsorption pad without a sample pad, so that the uneven quick release of the sample can be avoided, the product sensitivity is improved, and the product result repeatability is increased; and the water absorption paper is not arranged, so that the backflow of the sample is overcome, and the possibility of abnormal results is avoided. 2. Different multiplex analytes were labeled with different colored microspheres: the judgment of the detection result is more visual; non-specific binding between the same color label binders is avoided, and the probability of abnormal results is reduced.
Drawings
Fig. 1 and fig. 2 are schematic structural diagrams of an early pregnancy test strip provided in embodiment 1 of the present invention;
fig. 3 is a schematic structural diagram of a synchronous detection reagent card for cardiac troponin I/creatine kinase isoenzyme/myoglobin provided in example 2 of the present invention;
FIG. 4 is a schematic diagram showing the results of a plurality of drug test strips provided in embodiment 3 of the present invention;
FIG. 5 is a schematic structural diagram of an early pregnancy test strip provided in embodiment 4 of the present invention;
FIG. 6 is a schematic view showing the contact effect between the chromatographic carrier and the conjugate pad.
In the figure, 1 is a bottom plate, 2 is a chromatographic membrane, 3 is a binding pad, 4 is a conjugate, 5 is a detection line (specifically, 5a is a cardiac troponin I detection line, 5b is a creatine kinase isoenzyme detection line, 5c is a myoglobin detection line, 5d is a morphine detection line, 5e is a methamphetamine detection line, 5f is an amphetamine detection line, 5g is a ***e detection line, and 5h is a hemp detection line), 6 is a quality control line, 7 is a fixing pad combining hole, 8 is a chromatographic membrane fixing hole, 9 is a sample pad, 10 is a water absorption pad, 11 is a single-sided adhesive tape, and 12-1 is a box-shaped card shell.
Detailed Description
The technical solutions of the present invention will be described clearly and completely with reference to fig. 1 to 6, and it is obvious that the described embodiments are some, not all embodiments of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
In the description of the present invention, it should be noted that the terms "fixed", "connected" and "connected" are to be construed broadly and may be, for example, fixedly connected, detachably connected or contactingly connected to each other, unless expressly stated or limited otherwise. The specific meanings of the above terms in the present invention can be understood in specific cases to those skilled in the art.
In the description of the present invention, it should be noted that the terms "center", "upper", "lower", "left", "right", "vertical", "horizontal", "inner", "outer", "distal", and the like indicate orientations or positional relationships based on the orientations or positional relationships shown in the drawings, and are only for convenience of description and simplicity of description, but do not indicate or imply that the device or element being referred to must have a specific orientation, be constructed and operated in a specific orientation, and thus, should not be construed as limiting the present invention. Furthermore, the terms "first," "second," and "third" are used for descriptive purposes only and are not to be construed as indicating or implying relative importance.
The embodiments of the present invention are illustrated below by specific examples, and unless otherwise indicated, the experimental methods disclosed in the present invention are performed by using conventional techniques in the art, and reagents and raw materials used in the examples are commercially available.
Example 1
The embodiment provides an early pregnancy test strip, the structure of which is shown in fig. 1 and fig. 2, the state of the contact part between a chromatographic membrane and a binding pad is shown in fig. 6, and the test strip comprises a bottom plate 1, a chromatographic membrane 2, a binding pad 3, and a single-sided adhesive tape 11, wherein the chromatographic membrane has 2 lines: a detection line 5 and a quality control line 6. The specific preparation method of the test strip is as follows:
1. preparation of conjugate pad (3) containing conjugate (4)
1.1 preparation of Mass-control line (6) microsphere conjugate (4-2)
In this product, the control line labeled antibodies are: rabbit IgG antibody, labeled using microspheres: blue microspheres.
1.1.1 cleaning of microspheres
1mL of blue microspheres was placed in a beaker, and 0.05mpH8.0 BB16mL of reaction buffer was weighed and stirred for 2 minutes. (reaction buffer was used 16 times the amount of labeled microspheres)
Centrifuging at 12000rpm for 20 min, removing supernatant, precipitating with reaction buffer solution 0.05mPH8.0 BB16mL, repeatedly blowing and beating the pellet, vortexing, fully suspending and dissolving the microspheres, and performing ultrasonic treatment for 10 s for later use. (reaction buffer was used 16 times the amount of labeled microspheres)
1.1.2 activated microspheres
EDC and NHS were equilibrated to room temperature before use, prepared using reaction buffer 0.05mpH8.0 BB reaction buffer, EDC 2mg/mL, NHS4mg/mL (ready to use): (1.1.1 operation washing precipitation) slowly adding 2mg/mL EDC 500 microliter and then 4mg/mL NHS 500 microliter under stirring, continuing stirring for 2 minutes after the addition is finished, fully mixing the microspheres, and reacting for 30 minutes in a shaking table at 37 ℃ in a dark place.
Centrifuging at 12000rpm for 20 min, removing supernatant, precipitating, re-dissolving the precipitate with 0.05mPH8.0 BB 4 ml reaction buffer solution, repeatedly blowing and beating the microspheres, vortex oscillating to fully re-suspend and dissolve the microspheres, and performing ultrasonic treatment for 10 s (the microspheres are not aggregated by visual observation) (the reaction buffer solution is used according to 4 times of the amount of marked microspheres)
1.1.3 coupling of microspheres to antibodies
Dialyzing the coupled antibody with 0.01M PH7.4PB overnight, taking 800 microliter of the treated antibody, slowly adding the treated antibody into the redissolved microspheres in a stirring state, continuously stirring and combining for 2 minutes after the addition is finished, and carrying out shaking table photophobic reaction for 3 hours at 37 ℃. Centrifuge at 12000rpm for 20 minutes, remove supernatant, and leave a precipitate.
1.1.4 blocking of unbound surface groups on microspheres
Taking 4 ml of stop solution (2% BSA,100mM ethanolamine solution, pH 8.0), re-dissolving the precipitate, repeatedly blowing the microspheres, performing vortex oscillation to fully re-suspend and dissolve the microspheres, performing ultrasonic treatment for 10 seconds, and performing shaking table light-shielding reaction at 37 ℃ for 1 hour. (stop solution was used in an amount of 4 times the amount of labeled microspheres)
1.1.5 cleaning and preservation of the coupling-completed colloidal microspheres
Centrifuge at 12000rpm for 20 minutes, remove the supernatant, and leave the precipitate. Taking 4 ml of 0.02M PH7.4PB + 2% BSA, redissolving the precipitate, repeatedly blowing the microspheres, carrying out vortex oscillation to fully resuspend and dissolve the microspheres, carrying out ultrasonic treatment for 10 seconds, and carrying out shaking table light-shielding reaction for 10 minutes at 37 ℃. (blocking solution was used 4 times the amount of labeled microspheres)
Centrifuge at 12000rpm for 20 minutes, remove the supernatant, and leave the precipitate. Taking 4 ml of 0.05M PH8.0BB + 0.2% BSA + 0.02% NaN3 solution, dissolving the precipitate, performing vortex oscillation to fully resuspend and dissolve the microspheres, performing ultrasonic treatment for 10 seconds, and storing. (microspheres should not aggregate upon visual observation) (storage solution used 4 times the amount of labeled microspheres)
1.2 detection of line 5 microsphere conjugate 4-1 preparation
The labeled antibodies corresponding to the detection lines in the product are: the mouse anti-human alpha-HCG monoclonal antibody is marked by the following microspheres: red microspheres.
1.2.1 cleaning of microspheres
1mL of red microspheres were placed in a beaker and the reaction buffer 0.05M pH6.0 MES16mL was weighed and stirred for 2 minutes. (reaction buffer was used 16 times the amount of labeled microspheres)
Centrifuging at 12000rpm for 20 min, removing supernatant, precipitating with reaction buffer solution 0.05M, pH6.0 MES16mL, repeatedly blowing and beating the pellet, vortexing, fully suspending and dissolving the microsphere, and performing ultrasonic treatment for 10 s for later use.
1.2.2 activated microspheres
EDC and NHS were equilibrated to room temperature before use, prepared with 0.05M PH6.0 MES reaction buffer, EDC 2mg/mL, NHS4mg/mL (ready to use) (1.2.1 operation wash precipitate) with stirring slowly add 2mg/mL EDC 500 microliter, add 4mg/mL NHS 500 microliter, add the end of the stirring for 2 minutes to make the microspheres fully mix, shake the table at 37 ℃ for 30 minutes in the dark.
Centrifuging at 12000rpm for 20 min, removing supernatant, precipitating, re-dissolving the precipitate with reaction buffer solution 0.05M, pH6.0 MES 4 ml, repeatedly blowing the microspheres, vortexing, fully re-suspending and dissolving the microspheres, and performing ultrasonic treatment for 10 s for later use. (reaction buffer was used 4 times the amount of labeled microspheres)
1.2.3 coupling of microspheres to antibodies
And taking 800 microliters of the treated antibody, slowly adding the treated antibody into the redissolved microspheres in a stirring state, continuously stirring and combining for 2 minutes after the addition is finished, and reacting for 3 hours in a shaking table at 37 ℃ in a dark place. Centrifuge at 12000rpm for 20 minutes, remove the supernatant, and leave the precipitate.
1.2.4 blocking of unbound surface groups on microspheres
Taking 4 ml of stop solution (2% BSA,100mM ethanolamine solution, pH 8.0), re-dissolving the precipitate, repeatedly blowing the microspheres, performing vortex oscillation to fully re-suspend and dissolve the microspheres, performing ultrasonic treatment for 10 seconds, and performing shaking table light-shielding reaction at 37 ℃ for 1 hour. (stop solution was used in an amount of 4 times the amount of labeled microspheres)
1.2.5 cleaning and preservation of the coupling-completed colloidal microspheres
Centrifuge at 12000rpm for 20 minutes, remove the supernatant, and leave the precipitate. Taking 4 ml of 0.02M PH7.4PB + 2% BSA, redissolving the precipitate, repeatedly blowing the microspheres, carrying out vortex oscillation to fully resuspend and dissolve the microspheres, carrying out ultrasonic treatment for 10 seconds, and carrying out shaking table light-shielding reaction for 10 minutes at 37 ℃. (blocking solution was used 4 times the amount of labeled microspheres)
Centrifuge at 12000rpm for 20 minutes, remove the supernatant, and leave the precipitate. Taking 4 ml of 0.05M PH8.0BB + 0.2% BSA + 0.02% NaN3 solution, dissolving the precipitate, performing vortex oscillation to fully resuspend and dissolve the microspheres, performing ultrasonic treatment for 10 seconds, and storing. (microspheres should not aggregate upon visual observation) (storage solution used 4 times the amount of labeled microspheres)
2. Preparation of chromatographic membrane (2) containing detection line (5) and quality control line (6)
2.1 nitrocellulose membrane: a nitrocellulose membrane (backing membrane) having a width of 5 cm was selected.
2.2 coating detection line (5), quality control line (6)
2.2.1 coating solution preparation
And (4) preparing coating liquid by corresponding dilution, and sequentially adding corresponding sample cups in sequence. Diluting goat anti-rabbit IgG polyclonal antibody to 1.5mg/mL with a first coating diluent (trehalose added to 0.04M PBS solution at pH7.4 at 3% by mass); the murine anti-human HCG monoclonal antibody A concentration was diluted to 0.4mg/mL using a second coating diluent (0.04M TB solution at pH8.0 with 0.2% BSA by mass).
2.2.2 coating
Debugging a coating machine, setting the liquid spray amount of a detection line to be 1.0 microliter/cm, setting the liquid spray amount of a quality control line to be 1.0 microliter/cm, setting the guide rail speed to be 6.0cm/sec, and setting the nozzle interval to be 5 mm; then spraying liquid or scribing a film, and simultaneously marking the C/T line direction by using the ball strokes to remove the edges at two ends and other abnormal parts: the coated nitrocellulose membrane was dried overnight at room temperature for 20 hours.
2.2.3 the air-dried nitrocellulose reaction membrane is marked as the production intermediate component HCG-S1. Sealing the aluminum foil bag, storing at room temperature, and having an expiration date of 1 year.
3. Making a conjugate pad (3) containing conjugate (4)
3.1 glass fibers were pretreated using conjugate pad treatment, wherein the sample pad composition (mass ratio) included 0.08M PH9.0TB + 0.5% PVPK30+ 1% norrin + 0.5% EDTA.Na2+ 0.5% S7+ 0.5% S16+ 0.1% sodium azide.
3.2 cutting the pretreated pad into 6 cm wide strips as carriers for diluting the conjugate
3.3 conjugates 4-1 and 4-2 were individually diluted to concentration using conjugate diluent (0.05M PH8.0BB + 0.5% BSA + 0.1% 8964+ 2% sucrose + 3% trehalose + 0.2% TW20+ 0.02% NaN3), sprayed onto pretreated glass fibers, and dried overnight at 37 ℃.
3.3 taking the quality control line binder (4-1) according to 1/4 of the total amount of the prepared solution, taking the detection line binder (4-2) according to 1/7 of the total amount of the prepared solution, supplementing the residual solution to the required total amount by using a binder diluent, uniformly mixing, spraying the mixed binder (4) on the '3.2 pretreated binding pad membrane', and drying at 37 ℃ overnight.
4. Assembly
Adhering a nitrocellulose membrane chromatographic membrane (2) with a detection line (5) (a mouse anti-HCG monoclonal antibody A) and a quality control line (6) (a goat anti-rabbit IgG polyclonal antibody) on a solid phase, and a glass fiber bonding pad (3) which is pre-adsorbed with microsphere-labeled rabbit IgG (blue) and mouse anti-HCG monoclonal antibody B (red microsphere) on a bottom plate (1), pressing the end of a bonding agent (4) on the bonding pad (3) at about 1-2 mm of the far end of the detection line (5) of the chromatographic membrane (2), and adhering the laminated parts of the bonding pad (3) and the chromatographic membrane (2) together by using a single-sided adhesive tape (11), thereby ensuring the uniform migration of a sample.
TABLE 1 test paper strip composition summary table for early pregnancy
TABLE 2 early pregnancy test paper strip test display results
| Test | Detection line | 5 | |
| Negative urine | No color development | Blue color | |
| Negative serum | No color development | Blue color | |
| Negative plasma | No color development | Blue color | |
| Negative whole blood | No color development | Blue color | |
| Human chorionic gonadotropin 10mIU/mL | Red colour | Blue color | |
| Human chorionic gonadotropin 100mIU/mL | Red colour | Blue color | |
| Human chorionic gonadotropin 1IU/mL | Red colour | Blue color | |
| Human chorionic gonadotropin 2IU/mL | Red colour | Blue color | |
| Human chorionic gonadotropin 10IU/mL | Red colour | Blue color | |
| Human chorionic gonadotropin 20IU/mL | Red colour | Blue color |
Example 2
The present embodiment provides a synchronous detection reagent card for cardiac troponin I/creatine kinase isoenzyme/myoglobin, which has a structure shown in fig. 3, and comprises a chromatographic membrane 2, a binding pad 3 adhered to a base plate (1), and a cassette shell 12-1. Wherein, 3 detection lines (5) (5 a-cardiac troponin I, 5 b-creatine kinase isozyme, 5 c-myoglobin) and a quality control line (6) are arranged on the chromatographic membrane 2; the combination pad (3) is provided with a combination substance (4), the combination substance (4) is a mixture marked by microspheres with different colors, and comprises 4-1a cardiac troponin I, 4-1b creatine kinase isoenzyme, 4-1c myoglobin and 4-2 rabbit IgG. The specific preparation method of the detection kit comprises the following steps:
1. preparation of conjugate pad (3) containing conjugate (4)
1.1 preparation of quality control line (6) microsphere conjugate 4-2
The quality control line microspheres are marked by black microspheres, and the marked antibodies of the quality control lines are rabbit IgG antibodies.
1.1.1 cleaning of microspheres
1mL of black microspheres was placed in a beaker, and 0.05M of reaction buffer, pH8.0BB 16mL, was weighed and stirred for 2 minutes.
Centrifuging at 12000rpm for 15 min, removing supernatant, retaining precipitate, precipitating with reaction buffer solution 0.05M, PH8.0BB 16mL, repeatedly blowing and beating the pellet, vortexing and shaking to fully resuspend and dissolve the microspheres, and performing ultrasonic treatment for 10 s for later use.
1.1.2 activated microspheres
EDC and NHS are balanced to room temperature before use, prepared by using reaction buffer 0.05M PH8.0BB, EDC 2mg/mL, NHS4mg/mL (ready to use) are stirred and slowly added with EDC 500 microliter of 2mg/mL and NHS 500 microliter of 4mg/mL, stirring is continued for 2 minutes after adding to fully mix the microspheres, and shaking table reaction is carried out for 30 minutes at 37 ℃ in a dark place.
Centrifuging at 12000rpm for 15 min, removing supernatant, precipitating, re-dissolving the precipitate with 0.05M reaction buffer solution, pH8.0BB 4 ml, repeatedly blowing microspheres, vortexing, fully re-suspending and dissolving the microspheres, and performing ultrasonic treatment for 10 s (no aggregation of the microspheres observed by naked eyes)
1.1.3 coupling of microspheres to antibodies
And taking 800 microliters of the treated antibody, slowly adding the treated antibody into the redissolved microspheres in a stirring state, continuously stirring and combining for 2 minutes after the addition is finished, and reacting for 3 hours in a shaking table at 37 ℃ in a dark place. Centrifuge at 12000rpm for 15 minutes, remove supernatant, and leave a precipitate.
1.1.4 blocking of unbound surface groups on microspheres
Taking 4 ml of stop solution (2% BSA,100mM ethanolamine solution, pH 8.0), re-dissolving the precipitate, repeatedly blowing the microspheres, performing vortex oscillation to fully re-suspend and dissolve the microspheres, performing ultrasonic treatment for 10 seconds, and performing shaking table light-shielding reaction at 37 ℃ for 1 hour.
1.1.5 cleaning and preservation of the coupling-completed colloidal microspheres
Centrifuge at 12000rpm for 15 minutes, remove the supernatant, and leave the precipitate. Taking 4 ml of 0.02M PH7.4PB + 2% BSA, redissolving the precipitate, repeatedly blowing the microspheres, carrying out vortex oscillation to fully resuspend and dissolve the microspheres, carrying out ultrasonic treatment for 10 seconds, and carrying out shaking table light-shielding reaction for 10 minutes at 37 ℃.
Centrifuge at 12000rpm for 15 minutes, remove the supernatant, and leave the precipitate. Taking 4 ml of 0.05M PH8.0BB + 0.2% BSA + 0.02% NaN3 solution, dissolving the precipitate, performing vortex oscillation to fully resuspend and dissolve the microspheres, performing ultrasonic treatment for 10 seconds, and storing. (microspheres should not aggregate upon visual inspection)
1.2 detection (5) preparation of microsphere conjugate 4-1
The product needs to be marked by using microspheres with 3 different colors, which are respectively as follows: red (4-1a cardiac troponin I), blue (4-1b creatine kinase isozyme), green (4-1c myoglobin).
1.2.1 cleaning of microspheres
Microspheres were mixed well before use, 1mL of each microsphere was placed in a beaker, and 0.05M reaction buffer, pH6.0 MES16mL, was weighed and stirred for 2 minutes. (for microspheres with different particle sizes, the microspheres need to be centrifuged under the most appropriate centrifugation conditions (usually at a centrifugation speed of 10000-15000 rpm for 10-30 minutes), the supernatant is removed by centrifugation, the precipitate is retained, the reaction buffer solution PH6.0 MES16mL is used for redissolution, repeated blowing and balling are carried out, vortex oscillation is carried out, the microspheres are fully resuspended and dissolved, and ultrasound is carried out for 10 seconds for standby.
1.2.2 activated microspheres
EDC and NHS were equilibrated to room temperature before use, prepared with reaction buffer 0.05M PH6.0 MES, EDC 2mg/mL, NHS4mg/mL (ready to use) stirred slowly added 2mg/mL EDC 500 microliter then 4mg/mL NHS 500 microliter, after adding and stirring for 2 minutes to mix the microspheres thoroughly, 37 ℃ shaking table reaction for 30 minutes.
Centrifuging to remove supernatant and precipitate, removing supernatant and precipitate, re-dissolving the precipitate with 0.05M reaction buffer solution, PH6.0 MES 4 ml, repeatedly blowing microsphere, vortex oscillating to fully re-suspend and dissolve the microsphere, and subjecting to ultrasound for 10 s (no aggregation of microsphere observed by naked eye)
1.2.3 coupling of microspheres to antibodies
And taking 800 microliters of the treated antibody, slowly adding the treated antibody into the redissolved microspheres in a stirring state, continuously stirring and combining for 2 minutes after the addition is finished, and reacting for 3 hours in a shaking table at 37 ℃ in a dark place. (the red microspheres are added with the mouse anti-human cardiac troponin I monoclonal antibody B, the blue microspheres are added with the mouse anti-human creatine kinase isozyme monoclonal antibody B, the green microspheres are added with the mouse anti-human myoglobin monoclonal antibody B)
1.2.4 blocking of unbound surface groups on microspheres
After the reaction is finished, the supernatant is removed by centrifugation, 4 ml of stop solution (2% BSA,100mM ethanolamine solution, pH 8.0) is taken, the precipitate is redissolved, the microspheres are blown and repeatedly dissolved, vortex oscillation is carried out, the microspheres are fully resuspended and dissolved, the ultrasonic treatment is carried out for 10 seconds, and the shaking table is protected from light and reacts for 1 hour at 37 ℃.
1.2.5 cleaning and preservation of the coupling-completed colloidal microspheres
And centrifuging the coupled microspheres to remove supernatant and leave precipitate. Taking 4 ml of 0.02MPH7.4PB + 2% skimmed milk powder, redissolving the precipitate, repeatedly blowing the microspheres, performing vortex oscillation to fully resuspend and dissolve the microspheres, performing ultrasonic treatment for 10 seconds, and performing shaking table light-shielding reaction for 10 minutes at 37 ℃.
And centrifuging twice to remove supernatant, taking 4 ml of 0.05M of a solution of PH8.0BB + 0.2% BSA + 0.02% NaN3, dissolving the precipitate, performing vortex oscillation to fully resuspend and dissolve the microspheres, performing ultrasonic treatment for 10 seconds, and storing. (microspheres should not aggregate upon visual inspection)
2. Preparation of chromatographic membrane (2) containing detection line (5) and quality control line (6)
2.1 slitting of the nitrocellulose membrane: a backing film having a width of 5 cm was selected.
2.2 coating detection line (5), quality control line (6)
2.2.1 coating solution preparation
And (4) preparing coating liquid by corresponding dilution, and sequentially adding corresponding sample cups in sequence. Diluting goat anti-rabbit IgG polyclonal antibody to 0.5mg/mL with a first coating diluent (trehalose added to 0.04M PBS solution at pH7.4 at 3% by mass); the monoclonal antibodies of the three markers of myocardium (which are: mouse anti-human cardiac troponin I monoclonal antibody A, mouse anti-human creatine kinase isozyme monoclonal antibody A and mouse anti-human myoglobin monoclonal antibody A) were diluted to a concentration of 0.3mg/mL to 0.8mg/mL using a second coating diluent (trehalose was added to 0.05M PBS solution at pH7.4 in a mass ratio of 2%, and 3% methanol was added).
2.2.2 coating
Debugging a coating machine, setting the liquid spraying amount of a detection line to be 1.0 microliter/cm, 1.0 microliter/cm and 1.0 microliter/cm, setting the liquid spraying amount of a quality control line to be 0.9 microliter/cm, setting the guide rail speed to be 6.0cm/sec, and setting the nozzle interval to be 3.5 mm; then spraying or scratching the film, and marking the nitrocellulose membrane coated in the C/T line direction at the same time except the edges at two ends and other abnormal parts of the ball strokes for 20 hours at room temperature and airing overnight.
And (5) drying the nitrocellulose reaction membrane after drying. Sealing the aluminum foil bag, storing at room temperature, and having an expiration date of 1 year.
3. Making a conjugate pad (3) containing conjugate (4)
3.1 Pre-treatment with conjugate pad treatment solution, wherein the sample pad composition (mass ratio) comprises 4.82% Tris, 1% Triton X-100, 0.5% Tween-20, 1% sodium tetraborate, 0.5% PVP and 0.5% norproteinate sodium. 3.2 sticking the pretreated bonding pad on the bottom plate (1), cutting into 5 cm wide strips as carriers for diluting the bonding material.
3.3 the quality control line conjugate (4-1) and the 3 different substance conjugates (4-2) are respectively diluted to certain concentration by using conjugate diluent (mass ratio) (0.05M BB PH8.0+ 2% BSA + 20% trehalose), after uniform mixing, the mixed conjugate (4) is sprayed on the '3.2 pretreated conjugate pad membrane', and dried overnight at 37 ℃.
4. Assembly
A nitrocellulose membrane chromatographic membrane (2) with a solid phase detection line (5) (3 detection lines 5a/5B/ 5c) and a quality control line (6) (goat anti-rabbit IgG polyclonal antibody) is sequentially fixed on a box-shaped card shell 12-1 through a chromatographic membrane fixing hole 8 and a fixing hole 7 of a glass fiber binding pad (3) which is pre-adsorbed with microsphere labeled rabbit IgG (black microsphere) and a mouse anti-myocardial infarction marker monoclonal antibody B (red, blue and green microsphere), and one end of the binding pad (3) close to a binder (4) is pressed on the upper side of one end of the chromatographic membrane 2 close to the detection line 5.
TABLE 3 summary of the composition of synchronous detection reagent card for cardiac troponin I/creatine kinase isoenzyme/myoglobin
| Material numbering | Cardiac troponin I | Creatine kinase isoenzyme | Myoglobin | |
| Baseboard (1) | 1 | 1 | 1 | |
| Chromatographic carrier (2) | 2 | 2 | 2 | |
| Detection line (5) | | 5b | 5c | |
| Quality control line (6) | 6 | 6 | 6 | |
| Chromatographic carrier (2) fixed hole (8) | 8 | 8 | 8 | |
| Combined cushion (3) | 3 | 3 | 3 | |
| Detection line conjugate (4-1) | 4-1a | 4-1b | 4-1c | |
| Quality control line conjugate (4-2) | 4-2 | 4-2 | 4-2 | |
| Single-sided tape (11) | / | / | / | |
| Combined pad (3) fixed hole (7) | 7 | 7 | 7 | |
| Sample adding pad (9) | / | / | / | |
| Suction pad (10) | / | / | / | |
| Card case (12) | 12-1 box type card | 12-1 box type card | 12-1 box type card |
TABLE 4 detection and display results of synchronous detection reagent card for cardiac troponin I/creatine kinase isoenzyme/myoglobin
Example 3
The present embodiment provides a multi-drug test strip, which has a structure as shown in fig. 4, and includes a base plate (1), a chromatographic membrane (2), a bonding pad (3), and a single-sided tape (11). Wherein, the chromatographic membrane (2) is provided with a detection line (5) and a quality control line (6), the combination pad (3) is provided with a combination substance (4), each test strip comprises the quality control line (6), and the area of the detection line (5) is coated with different antigens at intervals of 2.5mm, and the maximum number is 5. The specific preparation method of the drug detection quilt comprises the following steps:
1. preparation of conjugate pad (3) containing conjugate (4)
1.1 preparation of Mass-control line (6) microsphere conjugate (4-2)
Wherein the microspheres used for the quality control marks are yellow microspheres.
1.1.1 cleaning of microspheres
1mL of yellow microspheres was placed in a beaker, and 0.05M reaction buffer, pH8.0BB 16mL, was weighed and stirred for 2 minutes. Centrifuging to remove supernatant and leave precipitate, using reaction buffer solution 0.05M, PH8.0BB 16mL for precipitation, repeatedly blowing and beating the microspheres in a redissolution manner, carrying out vortex oscillation to fully resuspend and dissolve the microspheres, and carrying out ultrasonic treatment for 10 seconds for later use.
1.1.2 activated microspheres
EDC and NHS were equilibrated to room temperature before use, prepared with reaction buffer 0.05M PH8.0BB, 2mg/mL EDC and NHS4mg/mL (ready to use) were stirred slowly and 2mg/mL EDC 500. mu.L and then 4mg/mL NHS 500. mu.L were added, stirring was continued for 2 min to mix the microspheres thoroughly, and shaking table was protected from light at 37 ℃ for 30 min.
Centrifuging to remove supernatant and leave precipitate, re-dissolving the precipitate with 0.05M reaction buffer solution, PH8.0BB 4 ml, repeatedly blowing and beating the microspheres, performing vortex oscillation to fully re-suspend and dissolve the microspheres, and performing ultrasonic treatment for 10 seconds for later use (the microspheres are not aggregated by visual observation).
1.1.3 coupling of microspheres to antibodies
And taking 800 microliters of the treated antibody, slowly adding the treated antibody into the redissolved microspheres in a stirring state, continuously stirring and combining for 2 minutes after the addition is finished, and reacting for 3 hours in a shaking table at 37 ℃ in a dark place. The supernatant was centrifuged off, leaving a precipitate.
1.1.4 blocking of unbound surface groups on microspheres
Taking 4 ml of stop solution (2% BSA,100mM ethanolamine solution, pH 8.0), re-dissolving the precipitate, repeatedly blowing the microspheres, performing vortex oscillation to fully re-suspend and dissolve the microspheres, performing ultrasonic treatment for 10 seconds, and performing shaking table light-shielding reaction at 37 ℃ for 1 hour.
1.1.5 cleaning and preservation of the coupling-completed colloidal microspheres
Centrifuge at 12000rpm for 15 minutes, remove the supernatant, and leave the precipitate. Taking 4 ml of 0.02M PH7.4PB + 2% BSA, redissolving the precipitate, repeatedly blowing the microspheres, carrying out vortex oscillation to fully resuspend and dissolve the microspheres, carrying out ultrasonic treatment for 10 seconds, and carrying out shaking table light-shielding reaction for 10 minutes at 37 ℃.
Centrifuge at 12000rpm for 15 minutes, remove the supernatant, and leave the precipitate. Taking 4 ml of 0.05M PH8.0BB + 0.2% BSA + 0.02% NaN3 solution, dissolving the precipitate, performing vortex oscillation to fully resuspend and dissolve the microspheres, performing ultrasonic treatment for 10 seconds, and storing. (visual observation that the microspheres did not aggregate)
The marked quality control line binders are numbered as follows: 4-2.
1.2 detection of line (5) preparation of microsphere conjugate (4-1)
In the product, 5 drug targets to be detected are as follows: morphine, methamphetamine, amphetamines, ***e, and cannabis. The corresponding monoclonal antibodies were labeled with red, blue, black, green, pink microspheres, respectively.
1.2.1 cleaning of microspheres
Microspheres were mixed well before use, 1mL of each microsphere was placed in a beaker, and 0.05M reaction buffer, pH6.0 MES16mL, was weighed and stirred for 2 minutes. (for microspheres with different particle sizes, the microspheres need to be centrifuged under the most appropriate centrifugation conditions (usually at a centrifugation speed of 10000-15000 rpm for 10-30 minutes), the supernatant is removed by centrifugation, the precipitate is retained, the reaction buffer solution PH6.0 MES16mL is used for redissolution, repeated blowing and balling are carried out, vortex oscillation is carried out, the microspheres are fully resuspended and dissolved, and ultrasound is carried out for 10 seconds for standby.
1.2.2 activated microspheres
EDC and NHS are balanced to room temperature before use, prepared with reaction buffer PH6.0 MES, EDC 2mg/mL NHS4mg/mL (ready to use) and stirred slowly added with EDC 500 microliter 2mg/mL and NHS 500 microliter 4mg/mL, after the completion of the stirring, the microspheres are fully mixed for 2 minutes, and the mixture is subjected to shaking table light-shielding reaction at 37 ℃ for 30 minutes.
Centrifuging to remove supernatant and precipitate, removing supernatant and precipitate, re-dissolving the precipitate with reaction buffer solution pH6.0 MES 4 ml, repeatedly blowing microsphere, vortex oscillating to fully re-suspend and dissolve the microsphere, and ultrasonic treating for 10 s (no aggregation of microsphere observed by naked eye)
1.2.3 coupling of microspheres to antibodies
And taking 800 microliters of the treated antibody, slowly adding the treated antibody into the redissolved microspheres in a stirring state, continuously stirring and combining for 2 minutes after the addition is finished, and reacting for 3 hours in a shaking table at 37 ℃ in a dark place.
Remarking: adding red microspheres: adding the morphine monoclonal antibody and the blue microsphere: adding the methamphetamine monoclonal antibody and the black microspheres, wherein the methamphetamine monoclonal antibody and the green microspheres are added: adding a ***e monoclonal antibody and pink microspheres: cannabis monoclonal antibodies.
1.2.4 blocking of unbound surface groups on microspheres
After the reaction is finished, the supernatant is removed by centrifugation, 4 ml of stop solution (2% BSA,100mM ethanolamine solution, pH 8.0) is taken, the precipitate is redissolved, the microspheres are blown and repeatedly dissolved, vortex oscillation is carried out, the microspheres are fully resuspended and dissolved, the ultrasonic treatment is carried out for 10 seconds, and the shaking table is protected from light and reacts for 1 hour at 37 ℃.
1.2.5 cleaning and preservation of the coupling-completed colloidal microspheres
And centrifuging the coupled microspheres to remove supernatant and leave precipitate. Taking 4 ml of 0.02MPH7.4PB + 2% BSA, redissolving the precipitate, repeatedly blowing the microspheres, carrying out vortex oscillation to fully resuspend and dissolve the microspheres, carrying out ultrasonic treatment for 10 seconds, and carrying out shaking table light-shielding reaction for 10 minutes at 37 ℃.
And centrifuging twice to remove supernatant, taking 4 ml of 0.05M of a solution of PH8.0BB + 0.2% BSA + 0.02% NaN3, dissolving the precipitate, performing vortex oscillation to fully resuspend and dissolve the microspheres, performing ultrasonic treatment for 10 seconds, and storing. (microspheres should not aggregate upon visual inspection)
The binders corresponding to the different analytes are numbered respectively: 4-1d morphine, 4-1e methamphetamine, 4-1f amphetamine, 4-1g ***e and 4-1h hemp.
2. Preparation of chromatographic membrane (2) containing detection line (5) and quality control line (6)
2.1 slitting of the nitrocellulose membrane: a backing film having a width of 5-8 cm was selected.
The product is expected to detect 5 drug analytes, so the conjugate protein antigens (morphine, methamphetamine, amphetamine, ***e and cannabis) of the 5 analytes need to be sequentially sprayed at the position of the detection line (5) at intervals of 2.5 millimeters.
2.2 coating detection line (5), quality control line (6)
2.2.1 coating liquid preparation: and (4) preparing coating liquid by corresponding dilution, and sequentially adding corresponding sample cups in sequence. Diluting goat anti-rabbit IgG polyclonal antibody to 0.5mg/mL with a first coating diluent (trehalose added to 0.04M PBS solution at pH7.4 at 3% by mass); the antigen concentrations of 5 drug analytes were diluted with the second coated diluent to 0.3-1.0mg/mL morphine 5d, methamphetamine 5e, amphetamine 5f, ***e 5g, cannabis 5 h).
2.2.2 coating: debugging a coating machine, setting the liquid spray amount of a detection line to be 1.0 microliter/cm, setting the liquid spray amount of a quality control line to be 0.9 microliter/cm, setting the guide rail speed to be 6.0cm/sec, and setting the nozzle interval to be 2.5 mm; then spraying or scratching the film, and airing the coated nitrocellulose film at room temperature overnight for 20 hours.
2.2.3 the air-dried nitrocellulose reaction membrane is marked as the production intermediate component DOA-S1. Sealing the aluminum foil bag, storing at room temperature, and having an expiration date of 1 year.
3. Making a conjugate pad (3) containing conjugate (4)
3.1 pretreating the pad membrane with pad treatment solution, wherein the sample pad composition (mass ratio) comprises 2.42% Tris, 1% S17, 0.5% S6, 1% trisodium citrate, 1% PVP and 0.5% complexing protein sodium.
3.2, adhering the pretreated bonding pad film on the bottom plate (1), and cutting into 5 cm laths.
3.3 according to certain concentration, using conjugate diluent ((0.1M BB + 20% sucrose + 10% trehalose + 0.2% Norrin sodium + 0.3% BSA + 0.1% Tween20)) to dilute the labeled antibody conjugates (morphine 4-1d, methamphetamine 4-1e, amphetamine 4-1f, ***e 4-1g, hemp 4-1g) and quality control line conjugate (4-2)) corresponding to 5 different analytes, after mixing uniformly, spraying on pretreated glass fiber, drying overnight at 37 ℃.
4. Assembly
A nitrocellulose membrane chromatographic membrane (2) with a detection line (5) (5 lines) and a quality control line (6) (1 line) on a solid phase, and a glass fiber binding pad (3) which adsorbs microsphere-labeled rabbit IgG antibodies with different colors and monoclonal antibodies of mouse anti-drug analytes in advance are stuck on a bottom plate (1), the end of a binding substance 4 is pressed at the far end of the detection line (5) of the chromatographic membrane (2) by 1-2 mm, and meanwhile, a single-sided adhesive tape (11) is used for sticking the binding pad (3) and the chromatographic membrane (2) together, wherein the binding substance (4) is arranged below the single-sided adhesive tape (11).
TABLE 5A List of test strips for detecting various drugs
TABLE 6 test paper strip for detecting and displaying multiple drugs
Example 4
The present embodiment provides a test strip for detecting early pregnancy, the structure of which is shown in fig. 5, and the test strip comprises a bottom plate (1), a chromatographic membrane (2), a binding pad (3), a sample addition pad (9) and a sample suction pad (10), wherein there are 2 lines on the chromatographic membrane: a detection line (5) and a quality control line (6). The specific preparation method of the test strip is as follows:
1. preparation of conjugate pad (3) containing conjugate (4)
1.1 preparation of Mass-control line (6) microsphere conjugate (4-2)
In this product, the control line labeled antibodies are: rabbit IgG antibody, labeled using microspheres: blue microspheres.
The labeling method of the quality control line (6) corresponding to the blue microspheres is the same as 'example 1'.
1.2 detection of line (5) preparation of microsphere conjugate (4-1)
The labeled antibodies corresponding to the detection lines in the product are: the mouse anti-human alpha-HCG monoclonal antibody is marked by the following microspheres: red microspheres.
The labeling method of the detection line (5) corresponding to the red microspheres is the same as that of 'example 1'.
2. Preparation of chromatographic membrane (2) containing detection line (5) and quality control line (6)
A nitrocellulose membrane (backing) of 2 cm width was selected.
The solid phase method comprising detection line (5) and quality control line (6) was the same as "example 1".
3. Preparation of conjugate 4
3.1 Pre-treatment of the conjugate pad membrane with conjugate pad treatment solution, wherein the sample pad composition (mass ratio) comprises 2.42% Tris, 1% Triton X-100, 0.5% Tween-20, 1% sodium carbonate, 1% PVP and 0.1% PVA.
3.2 conjugates (4-1) and (4-2) were diluted to a certain concentration using conjugate dilutions, sprayed on pretreated glass fibers, and dried overnight at 37 ℃.
4. Assembly
The sample adding pad (10), the combining pad (3), the chromatographic membrane (2) and the sample absorbing pad (9) are sequentially stuck on the bottom plate (1). Firstly, sticking a nitrocellulose membrane chromatographic membrane (2) with a solid phase detection line (5) (a mouse anti-HCG monoclonal antibody A) and a quality control line (6) (a goat anti-rabbit IgG polyclonal antibody) on a bottom plate (1); secondly, sticking a sample absorbing pad (10) at one end close to the quality control line (6), and pressing the sample absorbing pad (10) on the chromatographic membrane (2) by 1-3 mm; thirdly, the binding pad containing the binder (4) is pressed against the end of the chromatographic membrane near the detection line (5) for 1-3 mm, and fourthly, the sample addition pad (9) is pressed against the binding pad for 1-2 mm. Sticking the laminated part of the bonding pad (3) and the chromatographic carrier (2) together; finally, the conjugate pad and the chromatographic carrier are adhered by using a single-sided adhesive tape (11).
TABLE 7 reagent set List of early pregnancy test paper
| Material number | Early pregnancy test paper |
| Baseboard (1) | 1 |
| Chromatographic carrier (2) | 2 |
| Detection line (5) | 5 |
| Quality control line (6) | 6 |
| Chromatographic carrier (2) fixed hole (8) | / |
| Combined cushion (3) | 3 |
| Detection line conjugate (4-1) | 4-1 |
| Quality control line conjugate (4-2) | 4-2 |
| Single-sided tape (11) | 11 |
| Combined cushion fixed hole (7) | / |
| Sample adding pad (9) | 9 |
| Suction pad (10) | 10 |
| Card case (12) | / |
Table 8 shows the results of examination
It should be understood that the above examples are only for clarity of illustration and are not intended to limit the embodiments. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. And are neither required nor exhaustive of all embodiments. And obvious variations or modifications therefrom are within the scope of the invention.
Claims (9)
1. A rapid detection test paper is characterized in that: the kit comprises a chromatographic membrane (2) and a binding pad (3) which are sequentially stuck on a bottom plate (1), wherein one end of the chromatographic membrane (2) is connected with the binding pad (3); antigen/antibody conjugates (4) corresponding to different analytes marked by microspheres with different colors are pre-mixed and sprayed on one end of the binding pad (3), a detection line (5) and a quality control line (6) are sequentially arranged on the chromatographic membrane (2) along the liquid chromatography direction, and the detection line (5) is fixed at certain intervals and is used for detecting antigens/antibodies corresponding to different analytes;
wherein the rapid test strip does not comprise a sample addition pad and an absorption pad structure;
wherein, the number of the detection lines (5) is 1-5, and the number of the quality control lines (6) is 1;
wherein the particle size of the microsphere is 300-400nm, the microsphere is unmodified, carboxyl-modified or amino-modified polystyrene microsphere, and the color of the microsphere is more than 2 of red, blue, black, orange, green, yellow and pink;
wherein the width of the chromatographic membrane (2) and the width of the combined pad (3) are 4-8 mm; the length of the combination pad (3) is 3-6 cm, and the length of the chromatographic membrane (2) is 4-6 cm;
wherein, the contact part of the chromatographic membrane (2) is a semicircular edge at the connecting part of the chromatographic membrane (2) and the combining pad (3); the contact part of the combination pad (3) is isosceles trapezoid, and the upper side is 2-6 mm.
2. The rapid test strip of claim 1, wherein the microspheres are red, blue and black in color.
3. The rapid test strip according to claim 1, further comprising a cartridge-shaped case (12-1), wherein the chromatographic carrier (2) and the binding pad (3) adhered to the base plate (1) are fixed to the cartridge-shaped case (12-1) through the corresponding fixing holes, respectively.
4. The rapid test strip according to claim 1, wherein the contact portion of the conjugate pad (3) has an upper side of 2 to 4 mm.
5. The rapid test strip according to claim 3, wherein the fixing hole of the cartridge-type case (12-1) is polygonal and has a diameter of 1-3 mm, and the fixing hole of the conjugate pad (3) is located between the sample application region and the conjugate (4) and is 2-5 cm from the conjugate (4) near the sample application region; the fixed hole on the chromatographic membrane (2) is not positioned between the quality control line (6) and the binder (4), is positioned at the farthest end of the chromatographic direction of the chromatographic membrane, and is 3-6 mm away from the farthest end.
6. The rapid test strip according to claim 5, wherein the fixing hole of the cartridge-type case (12-1) is formed in a polygonal shape with a diameter of 1-2 mm, and the fixing hole of the conjugate pad (3) is formed between the sample application region and the conjugate (4) near the sample application region and is 2-4 cm from the conjugate (4).
7. The rapid test strip according to claim 1, wherein the quality control line (6) and the binder (4) are prepared from the following raw materials: goat anti-rabbit IgG and rabbit IgG, or rabbit anti-goat IgG and goat IgG.
8. The rapid test strip of claim 1, wherein the chromatography membrane material is nitrocellulose membrane, and the binding pad material is selected from glass fiber or polyester film.
9. Use of the rapid test strip according to any one of claims 1 to 8 in the preparation of a drug or an early pregnancy test kit.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910859373.7A CN110824177B (en) | 2019-09-11 | 2019-09-11 | Rapid detection test paper and preparation method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910859373.7A CN110824177B (en) | 2019-09-11 | 2019-09-11 | Rapid detection test paper and preparation method and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN110824177A CN110824177A (en) | 2020-02-21 |
| CN110824177B true CN110824177B (en) | 2021-06-11 |
Family
ID=69547980
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201910859373.7A Active CN110824177B (en) | 2019-09-11 | 2019-09-11 | Rapid detection test paper and preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN110824177B (en) |
Families Citing this family (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111157749B (en) * | 2020-01-13 | 2021-06-22 | 润和生物医药科技(汕头)有限公司 | Rapid detection test paper and preparation method and application thereof |
| CN111579782B (en) * | 2020-05-25 | 2023-10-10 | 山西瑞豪生物科技有限公司 | Biomedical detection method for intelligent fluorescent multi-marker |
| CN111638327B (en) * | 2020-07-01 | 2023-10-13 | 成都赛普克生物科技股份有限公司 | Treatment fluid and treatment method for colloidal gold immunochromatographic test paper gold-labeled pad and sample pad |
| CN112698042A (en) * | 2020-12-17 | 2021-04-23 | 北京赛诺浦生物技术有限公司 | Fluorescent immunochromatography test strip for detecting human growth differentiation factor-15 and preparation method and application thereof |
| CN112986567A (en) * | 2021-01-27 | 2021-06-18 | 佛山微侦警用科技有限公司 | Reagent component ratio and preparation method of multi-project rapid quantitative drug detection box |
| CN114236119A (en) * | 2021-11-08 | 2022-03-25 | 润和生物医药科技(汕头)有限公司 | Colloidal gold rapid detection test paper for new crown total antibody and neutralizing antibody and preparation method thereof |
Citations (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU614294B2 (en) * | 1987-07-13 | 1991-08-29 | Abbott Laboratories | Process for immunochromatography with colloidal particles |
| JP2003513244A (en) * | 1999-10-27 | 2003-04-08 | ジェノシス リミテッド | Lateral flow devices utilizing particulate carriers |
| CN1580778A (en) * | 2004-05-21 | 2005-02-16 | 王继华 | Colour latex chromato graphic analysis diagnositc test paper tape and its preparing method |
| EP1826566A1 (en) * | 2005-01-12 | 2007-08-29 | Sysmex Corporation | Immunochromatographic test device |
| CN102565388A (en) * | 2012-01-16 | 2012-07-11 | 宁波天润生物药业有限公司 | Widal test card |
| CN102928600A (en) * | 2012-11-07 | 2013-02-13 | 深圳市宝凯仑科技有限公司 | Method for preparing fast detecting test paper of illegal cooking oil |
| CN104076142A (en) * | 2014-03-05 | 2014-10-01 | 广东医学院附属医院 | Fluorescent microsphere lateral chromatographic detection strip for multiple joint inspection of trace target substances as well as preparation method and application thereof |
| CN104991067A (en) * | 2014-08-20 | 2015-10-21 | 苏州和锐医药科技有限公司 | Preparation method and applications of anti-ASCA antibody test paper |
| CN105137066A (en) * | 2015-07-31 | 2015-12-09 | 江苏巨珩新材料科技有限公司 | Polystyrene latex based cancer marker immuno-chromatography kit and application method thereof |
| CN205333649U (en) * | 2016-01-21 | 2016-06-22 | 成都微瑞生物科技有限公司 | Pig circovirus antibody test card |
| CN106290838A (en) * | 2015-05-13 | 2017-01-04 | 上海凯创生物技术有限公司 | A kind of dengue virus IgG/IgM antibody emulsion technique detection kit |
| CN107328926A (en) * | 2017-06-29 | 2017-11-07 | 海格德生物科技（深圳）有限公司 | Reagent card, preparation method and the detection method of the multinomial heart failure mark of quick detection |
| CN108267592A (en) * | 2017-01-03 | 2018-07-10 | 普迈德（北京）科技有限公司 | A kind of immune microsphere chromatography detects heparin-binding protein Test paper |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US7390674B2 (en) * | 2005-03-14 | 2008-06-24 | Kimberly-Clark Worldwide, Inc. | Lateral flow devices using reactive chemistry |
-
2019
- 2019-09-11 CN CN201910859373.7A patent/CN110824177B/en active Active
Patent Citations (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU614294B2 (en) * | 1987-07-13 | 1991-08-29 | Abbott Laboratories | Process for immunochromatography with colloidal particles |
| JP2003513244A (en) * | 1999-10-27 | 2003-04-08 | ジェノシス リミテッド | Lateral flow devices utilizing particulate carriers |
| CN1580778A (en) * | 2004-05-21 | 2005-02-16 | 王继华 | Colour latex chromato graphic analysis diagnositc test paper tape and its preparing method |
| EP1826566A1 (en) * | 2005-01-12 | 2007-08-29 | Sysmex Corporation | Immunochromatographic test device |
| CN102565388A (en) * | 2012-01-16 | 2012-07-11 | 宁波天润生物药业有限公司 | Widal test card |
| CN102928600A (en) * | 2012-11-07 | 2013-02-13 | 深圳市宝凯仑科技有限公司 | Method for preparing fast detecting test paper of illegal cooking oil |
| CN104076142A (en) * | 2014-03-05 | 2014-10-01 | 广东医学院附属医院 | Fluorescent microsphere lateral chromatographic detection strip for multiple joint inspection of trace target substances as well as preparation method and application thereof |
| CN104991067A (en) * | 2014-08-20 | 2015-10-21 | 苏州和锐医药科技有限公司 | Preparation method and applications of anti-ASCA antibody test paper |
| CN106290838A (en) * | 2015-05-13 | 2017-01-04 | 上海凯创生物技术有限公司 | A kind of dengue virus IgG/IgM antibody emulsion technique detection kit |
| CN105137066A (en) * | 2015-07-31 | 2015-12-09 | 江苏巨珩新材料科技有限公司 | Polystyrene latex based cancer marker immuno-chromatography kit and application method thereof |
| CN205333649U (en) * | 2016-01-21 | 2016-06-22 | 成都微瑞生物科技有限公司 | Pig circovirus antibody test card |
| CN108267592A (en) * | 2017-01-03 | 2018-07-10 | 普迈德（北京）科技有限公司 | A kind of immune microsphere chromatography detects heparin-binding protein Test paper |
| CN107328926A (en) * | 2017-06-29 | 2017-11-07 | 海格德生物科技（深圳）有限公司 | Reagent card, preparation method and the detection method of the multinomial heart failure mark of quick detection |
Also Published As
| Publication number | Publication date |
|---|---|
| CN110824177A (en) | 2020-02-21 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN110824177B (en) | Rapid detection test paper and preparation method and application thereof | |
| KR101540608B1 (en) | Assay strip having variable control line, and diagnosis kit using the same | |
| JP2638600B2 (en) | Immunodiagnosis device | |
| KR101947884B1 (en) | Immunochromatographic assay method | |
| JPH01503174A (en) | Test method | |
| JP2006215017A (en) | Device for discontinuous immunoassay and method of immunoassay utilizing it | |
| JPH04506117A (en) | Biological assay device and assay method using it | |
| JP4800739B2 (en) | Assay medium and assay method | |
| US11619630B2 (en) | Immunochromatography device | |
| WO2002037108A1 (en) | Reagent and method for detecting substance | |
| JP2008268043A (en) | Method for forming detection part of examination device, and examination device for measuring lateral flow immunity | |
| EP1197754A1 (en) | Methods and device for detecting prostate specific antigen (PSA) | |
| CN106526166A (en) | Rapid detection of lean meat powder in pork | |
| JP2002148266A (en) | Detector and its manufacturing method | |
| CN111157749B (en) | Rapid detection test paper and preparation method and application thereof | |
| JP4365329B2 (en) | Simple assay device and method for detecting a plurality of objects to be detected | |
| CN102388312B (en) | Analysis method, sample analyzing tool, method of preventing backflow of sample solution and method of preventing background rise | |
| JP3841559B2 (en) | Immunological examination method and immunological examination kit | |
| WO2008086845A1 (en) | Lateral-flow test device providing improved test quality | |
| EP3885767B1 (en) | Test piece for immunochromatography and immunochromatographic detection kit | |
| JP2001305139A (en) | Specific bond body | |
| JP2000028612A (en) | Immunological inspection method and immunological inspection kit thereof | |
| CN215415460U (en) | Rapid detection test strip and rapid detection card | |
| CN114113615B (en) | Immunochromatography detection test strip for screening universal monoclonal antibodies and detection method | |
| JP2002328130A (en) | Testing piece for immunological measurement method |
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 | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CP01 | Change in the name or title of a patent holder |
Address after: Room 116, Floor 1, Transportation Bureau Building, No. 95, Yingbin Avenue, Huacheng Street, Huadu District, Guangzhou, Guangdong Province, 510801 Patentee after: Guangzhou Iconas Biomedical Technology Co.,Ltd. Address before: Room 116, Floor 1, Transportation Bureau Building, No. 95, Yingbin Avenue, Huacheng Street, Huadu District, Guangzhou, Guangdong Province, 510801 Patentee before: Guangzhou Iconas Biomedical Technology Co.,Ltd. |
|
| CP01 | Change in the name or title of a patent holder | ||
| CP03 | Change of name, title or address |
Address after: Room 116, Floor 1, Transportation Bureau Building, No. 95, Yingbin Avenue, Huacheng Street, Huadu District, Guangzhou, Guangdong Province, 510801 Patentee after: Guangzhou Iconas Biomedical Technology Co.,Ltd. Address before: 515063 in Rongsheng science and Technology Park, University Road, Shantou City, Guangdong Province Patentee before: RUNBIO BIOTECH Co.,Ltd. |
|
| CP03 | Change of name, title or address | ||
| CI03 | Correction of invention patent |
Correction item: Patentee|Address Correct: RUNBIO BIOTECH Co.,Ltd.|515063 in Rongsheng science and Technology Park, University Road, Shantou City, Guangdong Province False: Guangzhou Iconas Biomedical Technology Co.,Ltd.|Room 116, Floor 1, Transportation Bureau Building, No. 95, Yingbin Avenue, Huacheng Street, Huadu District, Guangzhou, Guangdong Province, 510801 Number: 10-02 Volume: 39 |
|
| CI03 | Correction of invention patent | ||
| CP01 | Change in the name or title of a patent holder |
Address after: Room 116, Floor 1, Transportation Bureau Building, No. 95, Yingbin Avenue, Huacheng Street, Huadu District, Guangzhou, Guangdong Province, 510801 Patentee after: Guangdong Iconas Biomedical Technology Co.,Ltd. Address before: Room 116, Floor 1, Transportation Bureau Building, No. 95, Yingbin Avenue, Huacheng Street, Huadu District, Guangzhou, Guangdong Province, 510801 Patentee before: Guangdong Iconas Biomedical Technology Co.,Ltd. |
|
| CP01 | Change in the name or title of a patent holder | ||
| CP03 | Change of name, title or address |
Address after: Room 116, Floor 1, Transportation Bureau Building, No. 95, Yingbin Avenue, Huacheng Street, Huadu District, Guangzhou, Guangdong Province, 510801 Patentee after: Guangdong Iconas Biomedical Technology Co.,Ltd. Address before: 515063 in Rongsheng science and Technology Park, University Road, Shantou City, Guangdong Province Patentee before: RUNBIO BIOTECH Co.,Ltd. |
|
| CP03 | Change of name, title or address |