CN104597230A - Functional polymer thin film, preparation method and application thereof - Google Patents
Functional polymer thin film, preparation method and application thereof Download PDFInfo
- Publication number
- CN104597230A CN104597230A CN201510045459.8A CN201510045459A CN104597230A CN 104597230 A CN104597230 A CN 104597230A CN 201510045459 A CN201510045459 A CN 201510045459A CN 104597230 A CN104597230 A CN 104597230A
- Authority
- CN
- China
- Prior art keywords
- biochip
- polymer
- solution
- thin film
- preparation
- 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.)
- Granted
Links
- 229920001002 functional polymer Polymers 0.000 title claims abstract description 14
- 238000002360 preparation method Methods 0.000 title claims abstract description 11
- 239000010409 thin film Substances 0.000 title abstract 6
- 239000000758 substrate Substances 0.000 claims abstract description 28
- 229920000642 polymer Polymers 0.000 claims abstract description 23
- 238000004528 spin coating Methods 0.000 claims abstract description 18
- 238000001514 detection method Methods 0.000 claims abstract description 16
- 238000003380 quartz crystal microbalance Methods 0.000 claims abstract description 5
- 238000001847 surface plasmon resonance imaging Methods 0.000 claims abstract description 4
- 238000000018 DNA microarray Methods 0.000 claims description 100
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 90
- 238000000034 method Methods 0.000 claims description 64
- 229910052757 nitrogen Inorganic materials 0.000 claims description 48
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 36
- 239000003795 chemical substances by application Substances 0.000 claims description 35
- 229920002521 macromolecule Polymers 0.000 claims description 28
- 238000004140 cleaning Methods 0.000 claims description 26
- 239000008367 deionised water Substances 0.000 claims description 25
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 claims description 19
- 239000000126 substance Substances 0.000 claims description 14
- 239000002094 self assembled monolayer Substances 0.000 claims description 13
- 239000013545 self-assembled monolayer Substances 0.000 claims description 13
- 229910021641 deionized water Inorganic materials 0.000 claims description 12
- 238000007306 functionalization reaction Methods 0.000 claims description 11
- 230000032050 esterification Effects 0.000 claims description 10
- 238000005886 esterification reaction Methods 0.000 claims description 10
- 229920001503 Glucan Polymers 0.000 claims description 9
- 229920002125 Sokalan® Polymers 0.000 claims description 9
- 239000011521 glass Substances 0.000 claims description 9
- TWNIBLMWSKIRAT-VFUOTHLCSA-N levoglucosan Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@H]2CO[C@@H]1O2 TWNIBLMWSKIRAT-VFUOTHLCSA-N 0.000 claims description 9
- 239000003814 drug Substances 0.000 claims description 8
- LSDPWZHWYPCBBB-UHFFFAOYSA-N Methanethiol Chemical compound SC LSDPWZHWYPCBBB-UHFFFAOYSA-N 0.000 claims description 7
- 239000003153 chemical reaction reagent Substances 0.000 claims description 7
- 229940079593 drug Drugs 0.000 claims description 7
- 239000012528 membrane Substances 0.000 claims description 7
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 claims description 7
- 238000005516 engineering process Methods 0.000 claims description 6
- 239000004584 polyacrylic acid Substances 0.000 claims description 6
- 125000002887 hydroxy group Chemical group [H]O* 0.000 claims description 5
- 239000011261 inert gas Substances 0.000 claims description 5
- ZHKJHQBOAJQXQR-UHFFFAOYSA-N 1H-azirine Chemical compound N1C=C1 ZHKJHQBOAJQXQR-UHFFFAOYSA-N 0.000 claims description 4
- KWOLFJPFCHCOCG-UHFFFAOYSA-N Acetophenone Chemical compound CC(=O)C1=CC=CC=C1 KWOLFJPFCHCOCG-UHFFFAOYSA-N 0.000 claims description 4
- XKRFYHLGVUSROY-UHFFFAOYSA-N Argon Chemical compound [Ar] XKRFYHLGVUSROY-UHFFFAOYSA-N 0.000 claims description 4
- 239000004593 Epoxy Substances 0.000 claims description 4
- 241001597008 Nomeidae Species 0.000 claims description 4
- 239000004793 Polystyrene Substances 0.000 claims description 4
- 125000004093 cyano group Chemical group *C#N 0.000 claims description 4
- 150000004662 dithiols Chemical class 0.000 claims description 4
- 238000002866 fluorescence resonance energy transfer Methods 0.000 claims description 4
- 230000004907 flux Effects 0.000 claims description 4
- 102000039446 nucleic acids Human genes 0.000 claims description 4
- 108020004707 nucleic acids Proteins 0.000 claims description 4
- 150000007523 nucleic acids Chemical class 0.000 claims description 4
- 229920001184 polypeptide Polymers 0.000 claims description 4
- 229920002223 polystyrene Polymers 0.000 claims description 4
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 4
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 4
- 238000004506 ultrasonic cleaning Methods 0.000 claims description 4
- 239000000020 Nitrocellulose Substances 0.000 claims description 3
- 229920001220 nitrocellulos Polymers 0.000 claims description 3
- PNEYBMLMFCGWSK-UHFFFAOYSA-N Alumina Chemical compound [O-2].[O-2].[O-2].[Al+3].[Al+3] PNEYBMLMFCGWSK-UHFFFAOYSA-N 0.000 claims description 2
- XUIMIQQOPSSXEZ-UHFFFAOYSA-N Silicon Chemical compound [Si] XUIMIQQOPSSXEZ-UHFFFAOYSA-N 0.000 claims description 2
- 125000003545 alkoxy group Chemical group 0.000 claims description 2
- 125000000304 alkynyl group Chemical group 0.000 claims description 2
- 230000009435 amidation Effects 0.000 claims description 2
- 238000007112 amidation reaction Methods 0.000 claims description 2
- 150000004056 anthraquinones Chemical class 0.000 claims description 2
- 229910052786 argon Inorganic materials 0.000 claims description 2
- 125000000852 azido group Chemical group *N=[N+]=[N-] 0.000 claims description 2
- RWCCWEUUXYIKHB-UHFFFAOYSA-N benzophenone Chemical compound C=1C=CC=CC=1C(=O)C1=CC=CC=C1 RWCCWEUUXYIKHB-UHFFFAOYSA-N 0.000 claims description 2
- 239000012965 benzophenone Substances 0.000 claims description 2
- 150000001875 compounds Chemical class 0.000 claims description 2
- 239000012954 diazonium Substances 0.000 claims description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-O diazynium Chemical compound [NH+]#N IJGRMHOSHXDMSA-UHFFFAOYSA-O 0.000 claims description 2
- 125000000118 dimethyl group Chemical group [H]C([H])([H])* 0.000 claims description 2
- 238000002875 fluorescence polarization Methods 0.000 claims description 2
- 239000007850 fluorescent dye Substances 0.000 claims description 2
- 238000001215 fluorescent labelling Methods 0.000 claims description 2
- 229920002313 fluoropolymer Polymers 0.000 claims description 2
- 239000004811 fluoropolymer Substances 0.000 claims description 2
- 239000007789 gas Substances 0.000 claims description 2
- 239000000203 mixture Substances 0.000 claims description 2
- 229930014626 natural product Natural products 0.000 claims description 2
- 238000010534 nucleophilic substitution reaction Methods 0.000 claims description 2
- 239000003960 organic solvent Substances 0.000 claims description 2
- 229920000747 poly(lactic acid) Polymers 0.000 claims description 2
- 229920003229 poly(methyl methacrylate) Polymers 0.000 claims description 2
- 239000004417 polycarbonate Substances 0.000 claims description 2
- 229920000515 polycarbonate Polymers 0.000 claims description 2
- 229920000151 polyglycol Polymers 0.000 claims description 2
- 239000010695 polyglycol Substances 0.000 claims description 2
- 239000004626 polylactic acid Substances 0.000 claims description 2
- 239000004926 polymethyl methacrylate Substances 0.000 claims description 2
- 239000010453 quartz Substances 0.000 claims description 2
- 238000010791 quenching Methods 0.000 claims description 2
- 230000000171 quenching effect Effects 0.000 claims description 2
- 150000003254 radicals Chemical class 0.000 claims description 2
- 238000000926 separation method Methods 0.000 claims description 2
- 239000010703 silicon Substances 0.000 claims description 2
- 229910052710 silicon Inorganic materials 0.000 claims description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N silicon dioxide Inorganic materials O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 2
- 229920005573 silicon-containing polymer Polymers 0.000 claims description 2
- 238000006243 chemical reaction Methods 0.000 abstract description 13
- 238000001179 sorption measurement Methods 0.000 abstract description 5
- 230000001276 controlling effect Effects 0.000 abstract description 2
- 238000004519 manufacturing process Methods 0.000 abstract description 2
- 230000001105 regulatory effect Effects 0.000 abstract description 2
- 239000003431 cross linking reagent Substances 0.000 abstract 2
- 239000012620 biological material Substances 0.000 abstract 1
- 239000011248 coating agent Substances 0.000 abstract 1
- 238000000576 coating method Methods 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 60
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 50
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical group CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 44
- 238000004132 cross linking Methods 0.000 description 21
- LMDZBCPBFSXMTL-UHFFFAOYSA-N 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide Chemical compound CCN=C=NCCCN(C)C LMDZBCPBFSXMTL-UHFFFAOYSA-N 0.000 description 16
- VHYFNPMBLIVWCW-UHFFFAOYSA-N 4-Dimethylaminopyridine Chemical compound CN(C)C1=CC=NC=C1 VHYFNPMBLIVWCW-UHFFFAOYSA-N 0.000 description 16
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 15
- 239000007864 aqueous solution Substances 0.000 description 13
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 12
- HOGDNTQCSIKEEV-UHFFFAOYSA-N n'-hydroxybutanediamide Chemical compound NC(=O)CCC(=O)NO HOGDNTQCSIKEEV-UHFFFAOYSA-N 0.000 description 12
- RFVNOJDQRGSOEL-UHFFFAOYSA-N 2-hydroxyethyl octadecanoate Chemical compound CCCCCCCCCCCCCCCCCC(=O)OCCO RFVNOJDQRGSOEL-UHFFFAOYSA-N 0.000 description 11
- 229950007687 macrogol ester Drugs 0.000 description 11
- 101100310856 Drosophila melanogaster spri gene Proteins 0.000 description 10
- 229920002845 Poly(methacrylic acid) Polymers 0.000 description 10
- 239000010410 layer Substances 0.000 description 9
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 8
- 229960000549 4-dimethylaminophenol Drugs 0.000 description 8
- 229920002307 Dextran Polymers 0.000 description 7
- 238000010560 atom transfer radical polymerization reaction Methods 0.000 description 7
- 230000000977 initiatory effect Effects 0.000 description 7
- 239000000178 monomer Substances 0.000 description 7
- 210000002966 serum Anatomy 0.000 description 7
- 238000003756 stirring Methods 0.000 description 7
- 150000003573 thiols Chemical class 0.000 description 7
- 238000009281 ultraviolet germicidal irradiation Methods 0.000 description 7
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 6
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- 108010090804 Streptavidin Proteins 0.000 description 6
- 230000035484 reaction time Effects 0.000 description 6
- 235000020183 skimmed milk Nutrition 0.000 description 6
- 230000000694 effects Effects 0.000 description 5
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 5
- 239000010931 gold Substances 0.000 description 5
- 229910052737 gold Inorganic materials 0.000 description 5
- 230000003287 optical effect Effects 0.000 description 5
- 230000008569 process Effects 0.000 description 5
- 102000004169 proteins and genes Human genes 0.000 description 5
- 108090000623 proteins and genes Proteins 0.000 description 5
- 239000000523 sample Substances 0.000 description 5
- 239000002904 solvent Substances 0.000 description 5
- WYTZZXDRDKSJID-UHFFFAOYSA-N (3-aminopropyl)triethoxysilane Chemical compound CCO[Si](OCC)(OCC)CCCN WYTZZXDRDKSJID-UHFFFAOYSA-N 0.000 description 4
- FALRKNHUBBKYCC-UHFFFAOYSA-N 2-(chloromethyl)pyridine-3-carbonitrile Chemical compound ClCC1=NC=CC=C1C#N FALRKNHUBBKYCC-UHFFFAOYSA-N 0.000 description 4
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 4
- VYZAMTAEIAYCRO-UHFFFAOYSA-N Chromium Chemical compound [Cr] VYZAMTAEIAYCRO-UHFFFAOYSA-N 0.000 description 4
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 4
- 229960002685 biotin Drugs 0.000 description 4
- 235000020958 biotin Nutrition 0.000 description 4
- 239000011616 biotin Substances 0.000 description 4
- 210000004899 c-terminal region Anatomy 0.000 description 4
- 238000007385 chemical modification Methods 0.000 description 4
- ORTQZVOHEJQUHG-UHFFFAOYSA-L copper(II) chloride Chemical compound Cl[Cu]Cl ORTQZVOHEJQUHG-UHFFFAOYSA-L 0.000 description 4
- 238000013016 damping Methods 0.000 description 4
- UQLDLKMNUJERMK-UHFFFAOYSA-L di(octadecanoyloxy)lead Chemical compound [Pb+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O UQLDLKMNUJERMK-UHFFFAOYSA-L 0.000 description 4
- 238000001704 evaporation Methods 0.000 description 4
- 230000008020 evaporation Effects 0.000 description 4
- 239000012530 fluid Substances 0.000 description 4
- 230000003993 interaction Effects 0.000 description 4
- 239000008363 phosphate buffer Substances 0.000 description 4
- 229940014800 succinic anhydride Drugs 0.000 description 4
- 101710186708 Agglutinin Proteins 0.000 description 3
- ROFVEXUMMXZLPA-UHFFFAOYSA-N Bipyridyl Chemical compound N1=CC=CC=C1C1=CC=CC=N1 ROFVEXUMMXZLPA-UHFFFAOYSA-N 0.000 description 3
- 238000005033 Fourier transform infrared spectroscopy Methods 0.000 description 3
- 101710146024 Horcolin Proteins 0.000 description 3
- 101710189395 Lectin Proteins 0.000 description 3
- 102000004856 Lectins Human genes 0.000 description 3
- 108090001090 Lectins Proteins 0.000 description 3
- 101710179758 Mannose-specific lectin Proteins 0.000 description 3
- 101710150763 Mannose-specific lectin 1 Proteins 0.000 description 3
- 101710150745 Mannose-specific lectin 2 Proteins 0.000 description 3
- 229920001213 Polysorbate 20 Polymers 0.000 description 3
- 239000000910 agglutinin Substances 0.000 description 3
- 239000000427 antigen Substances 0.000 description 3
- 102000036639 antigens Human genes 0.000 description 3
- 108091007433 antigens Proteins 0.000 description 3
- 239000012472 biological sample Substances 0.000 description 3
- 125000000524 functional group Chemical group 0.000 description 3
- 239000002523 lectin Substances 0.000 description 3
- 238000002156 mixing Methods 0.000 description 3
- 230000004048 modification Effects 0.000 description 3
- 238000012986 modification Methods 0.000 description 3
- 238000006116 polymerization reaction Methods 0.000 description 3
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 3
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 3
- 229920000136 polysorbate Polymers 0.000 description 3
- ZAHRKKWIAAJSAO-UHFFFAOYSA-N rapamycin Natural products COCC(O)C(=C/C(C)C(=O)CC(OC(=O)C1CCCCN1C(=O)C(=O)C2(O)OC(CC(OC)C(=CC=CC=CC(C)CC(C)C(=O)C)C)CCC2C)C(C)CC3CCC(O)C(C3)OC)C ZAHRKKWIAAJSAO-UHFFFAOYSA-N 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 229960002930 sirolimus Drugs 0.000 description 3
- QFJCIRLUMZQUOT-HPLJOQBZSA-N sirolimus Chemical compound C1C[C@@H](O)[C@H](OC)C[C@@H]1C[C@@H](C)[C@H]1OC(=O)[C@@H]2CCCCN2C(=O)C(=O)[C@](O)(O2)[C@H](C)CC[C@H]2C[C@H](OC)/C(C)=C/C=C/C=C/[C@@H](C)C[C@@H](C)C(=O)[C@H](OC)[C@H](O)/C(C)=C/[C@@H](C)C(=O)C1 QFJCIRLUMZQUOT-HPLJOQBZSA-N 0.000 description 3
- 150000003384 small molecules Chemical class 0.000 description 3
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 2
- 102100026735 Coagulation factor VIII Human genes 0.000 description 2
- 229910021591 Copper(I) chloride Inorganic materials 0.000 description 2
- 239000007995 HEPES buffer Substances 0.000 description 2
- 101000911390 Homo sapiens Coagulation factor VIII Proteins 0.000 description 2
- WOBHKFSMXKNTIM-UHFFFAOYSA-N Hydroxyethyl methacrylate Chemical compound CC(=C)C(=O)OCCO WOBHKFSMXKNTIM-UHFFFAOYSA-N 0.000 description 2
- 102100027913 Peptidyl-prolyl cis-trans isomerase FKBP1A Human genes 0.000 description 2
- 108010006877 Tacrolimus Binding Protein 1A Proteins 0.000 description 2
- FJWGYAHXMCUOOM-QHOUIDNNSA-N [(2s,3r,4s,5r,6r)-2-[(2r,3r,4s,5r,6s)-4,5-dinitrooxy-2-(nitrooxymethyl)-6-[(2r,3r,4s,5r,6s)-4,5,6-trinitrooxy-2-(nitrooxymethyl)oxan-3-yl]oxyoxan-3-yl]oxy-3,5-dinitrooxy-6-(nitrooxymethyl)oxan-4-yl] nitrate Chemical compound O([C@@H]1O[C@@H]([C@H]([C@H](O[N+]([O-])=O)[C@H]1O[N+]([O-])=O)O[C@H]1[C@@H]([C@@H](O[N+]([O-])=O)[C@H](O[N+]([O-])=O)[C@@H](CO[N+]([O-])=O)O1)O[N+]([O-])=O)CO[N+](=O)[O-])[C@@H]1[C@@H](CO[N+]([O-])=O)O[C@@H](O[N+]([O-])=O)[C@H](O[N+]([O-])=O)[C@H]1O[N+]([O-])=O FJWGYAHXMCUOOM-QHOUIDNNSA-N 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 229960005070 ascorbic acid Drugs 0.000 description 2
- 235000010323 ascorbic acid Nutrition 0.000 description 2
- 239000011668 ascorbic acid Substances 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 230000002902 bimodal effect Effects 0.000 description 2
- OXBLHERUFWYNTN-UHFFFAOYSA-M copper(I) chloride Chemical compound [Cu]Cl OXBLHERUFWYNTN-UHFFFAOYSA-M 0.000 description 2
- 229960003280 cupric chloride Drugs 0.000 description 2
- 206010012601 diabetes mellitus Diseases 0.000 description 2
- 239000000284 extract Substances 0.000 description 2
- MHMNJMPURVTYEJ-UHFFFAOYSA-N fluorescein-5-isothiocyanate Chemical compound O1C(=O)C2=CC(N=C=S)=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 MHMNJMPURVTYEJ-UHFFFAOYSA-N 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 229910052751 metal Inorganic materials 0.000 description 2
- 239000002184 metal Substances 0.000 description 2
- 238000002493 microarray Methods 0.000 description 2
- 239000011259 mixed solution Substances 0.000 description 2
- 229920001223 polyethylene glycol Polymers 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 238000010025 steaming Methods 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 238000012546 transfer Methods 0.000 description 2
- 238000001291 vacuum drying Methods 0.000 description 2
- 229920000936 Agarose Polymers 0.000 description 1
- 241000283707 Capra Species 0.000 description 1
- SXRSQZLOMIGNAQ-UHFFFAOYSA-N Glutaraldehyde Chemical compound O=CCCCC=O SXRSQZLOMIGNAQ-UHFFFAOYSA-N 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 206010024769 Local reaction Diseases 0.000 description 1
- CERQOIWHTDAKMF-UHFFFAOYSA-N Methacrylic acid Chemical compound CC(=C)C(O)=O CERQOIWHTDAKMF-UHFFFAOYSA-N 0.000 description 1
- 239000004677 Nylon Substances 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 108091093037 Peptide nucleic acid Proteins 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- 108010039918 Polylysine Proteins 0.000 description 1
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 1
- 206010067584 Type 1 diabetes mellitus Diseases 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 238000011938 amidation process Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000027455 binding Effects 0.000 description 1
- 230000033228 biological regulation Effects 0.000 description 1
- 210000004027 cell Anatomy 0.000 description 1
- 230000001427 coherent effect Effects 0.000 description 1
- 239000002299 complementary DNA Substances 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 230000008030 elimination Effects 0.000 description 1
- 238000003379 elimination reaction Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 description 1
- 238000000799 fluorescence microscopy Methods 0.000 description 1
- 150000004676 glycans Chemical class 0.000 description 1
- 229910052736 halogen Inorganic materials 0.000 description 1
- 150000002367 halogens Chemical class 0.000 description 1
- 239000000017 hydrogel Substances 0.000 description 1
- 238000005286 illumination Methods 0.000 description 1
- 238000003384 imaging method Methods 0.000 description 1
- 238000007654 immersion Methods 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 208000014674 injury Diseases 0.000 description 1
- 230000002687 intercalation Effects 0.000 description 1
- 238000009830 intercalation Methods 0.000 description 1
- 229910044991 metal oxide Inorganic materials 0.000 description 1
- 150000004706 metal oxides Chemical class 0.000 description 1
- 239000002052 molecular layer Substances 0.000 description 1
- 239000002159 nanocrystal Substances 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 229920001778 nylon Polymers 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
- 239000010452 phosphate Substances 0.000 description 1
- 229920002401 polyacrylamide Polymers 0.000 description 1
- 229920000656 polylysine Polymers 0.000 description 1
- 229920006254 polymer film Polymers 0.000 description 1
- 229920001282 polysaccharide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 125000006239 protecting group Chemical group 0.000 description 1
- 239000012460 protein solution Substances 0.000 description 1
- 238000003908 quality control method Methods 0.000 description 1
- 238000010526 radical polymerization reaction Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 230000009257 reactivity Effects 0.000 description 1
- 239000012488 sample solution Substances 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 238000001338 self-assembly Methods 0.000 description 1
- -1 shitosan Chemical compound 0.000 description 1
- 239000001632 sodium acetate Substances 0.000 description 1
- 235000017281 sodium acetate Nutrition 0.000 description 1
- 239000007974 sodium acetate buffer Substances 0.000 description 1
- 230000009870 specific binding Effects 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- RVEZZJVBDQCTEF-UHFFFAOYSA-N sulfenic acid Chemical compound SO RVEZZJVBDQCTEF-UHFFFAOYSA-N 0.000 description 1
- 238000006557 surface reaction Methods 0.000 description 1
Abstract
The invention discloses a functional polymer thin film, a preparation method of the thin film and an application of the thin film. The preparation method comprises the following steps: (1) modifying a photo-cross-linking agent on the surface of a substrate; (2) rotationally coating the surface formed in the step (1) with a polymer solution; (3) performing irradiation reaction on the polymer solution with ultraviolet lights and covalently grafting the polymer to the substrate to form the polymer thin film; and (4) further functionalizing the polymer or directly performing biological sensing application. According to the preparation method, a three-dimensional polymer surface can be simply, quickly and efficiently prepared; the density and thickness of the surface polymer can be precisely controlled by regulating and controlling the density of the photo-cross-linking agent, the molecule weight of the polymer and spin-coating parameters; moreover, the preparation method is strong in operability, excellent in repeatability and suitable for mass production. The surface of the prepared polymer thin film has the high requirement for the fixed amount of bio-molecules and the excellent anti-nonspecific adsorption ability, thus the detection performance of a biological sensor (for example, surface plasmon resonance imaging, fluorescence, quartz crystal microbalance and electrochemical transducer) can be improved.
Description
Technical field
The present invention relates to a kind of simple, quick, general macromolecule membrane, preparation method and the application in bio-sensing thereof, the application particularly in surface plasmon resonance imaging.
Background technology
Biochip is mainly through the surface chemical modification to substrate, the biomolecule such as protein, polypeptide, nucleic acid and the biological sample such as cell, tissue are fixed on chip surface, and then realize detecting accurate, quick, the large information capacity of targeted biological component, there is the features such as height collimation, diversity, microminiaturization and robotization.It is more efficient, stable that rational surface chemical modification can make biological sample fix.In order to improve the crystallized ability of biochip to biological sample, reducing non-specific adsorption, improving the signal intensity detected, and then improve instrument detection sensitivity, large quantifier elimination has been carried out in people's effects on surface chemistry aspect.
At present, self assembled monolayer is applied comparatively extensive at bio-sensing and chip technology field, its advantage is to form stable, homogeneous, fine and close molecular layer on surface, not only can in testing process at the bottom of protecting group, reduce non-specific adsorption, also can have diversified terminal functional groups, do further modification; In addition, it is very simple that self assembled monolayer modifies operation, modifies reproducible.But self assembled monolayer is as two-dimensional surface because its biomolecule fixed amount is little, and during detection bio-molecular interaction, signal is weak, makes it apply and is greatly limited.In order to strengthen the fixed amount of biochip to biomolecule further, thus improve biology sensor detection sensitivity, a lot of researchs have employed the chip surface method of modifying of three-dimensional structure.Biochip substrate builds the macromolecule membrane being rich in various reactive group that one deck has three-dimensional structure, surface prepared by this kind of modification mode is commonly referred to as three-dimensional surface, because its three dimensions surface has the binding site of large number of biological molecule, substantially increase the fixed amount of biomolecule, tens of even hundreds of times of common chip can be reached.
The existing research about three-dimensional structure finishing comprises the three-dimensional surface of various polymer film, as glucosan, polyacrylic acid, cellulose nitrate, nylon, agarose, polyacrylamide, the polyethylene glycol hydrogel of polylysine modification, micro/nano level rough surface structure and some Supramolecular self assembly three-dimensional structures etc., wherein most widely used (the Johnsson B. of glucosan three-dimensional surface, et al., Comparison of methods forimmobilization to carboxymethyl dextran sensor surfaces by analysis of the specificactivity of monoclonal antibodies.Journal of Molecular Recognition, 1995.8 (1-2): p.125-131.).In addition, the research of the polymer brush shaped polymer prepared based on atom transfer radical polymerization (ATRP) method is also because its reaction conditions is gentle, easy to operate, the advantages such as anti-non-specific adsorption ability is strong, be subject to extensive concern (Wang, J.-S.and K.Matyjaszewski, Controlled/ " living " radicalpolymerization.Atom transfer radical polymerization in the presence oftransition-metal complexes.Journal of the American Chemical Society, 1995.117 (20): p.5614-5615.).And this Biocompatibility is good, end function group is enriched, and has the good characteristics such as anti-non-specific adsorption, is widely applied in biosensor surface.
In polymer growth, strand generally has two kinds of methods with the connection at surface or interface: one is the method for " being grafted to " (graft-to), and another is the method (graft-from) of " from surface grafting "." from surface grafting " method, polymkeric substance directly grows on the surface of the substrate, usually first the initiating agent of polymkeric substance is fixed on the surface, next step polymerization is caused again by the initiating agent on surface, the method can control polymer thickness by adjustment monomer and reaction time, and controls the grafting density of polymkeric substance by the surface coverage adjusting initiating agent.But the method experimental procedure is comparatively complicated, when regulating and controlling surface aggregate thing density, repeatability is relatively poor, and production cost is higher, uses catalyzer usually to have bio-toxicity, and easily remains in surface, be unfavorable for that biomolecule activity keeps.And " being grafted to " method is simple, quick owing to preparing, use material easily to carry out antenatal Quality Control, be subject to people's favor.But polymer reaction functional group is connected by chemical bond, differential responses system to be adopted for different functional groups, relative Consideration is more, and for nano-crystal, reaction rate is usually not high, and the time is longer, macromolecule in the solution mobile compared be difficult to and self space steric hindrance large, cause local reaction limited, shortcoming is outstanding all the more.
Summary of the invention
For the problem of prior art, an object of the present invention is the preparation method providing a kind of functional polymer film, the method utilizes the mode of ultraviolet light cross-linking randomly polymkeric substance covalency to be fixed on substrate surface, obtain macromolecule membrane, then to its further functionalization or directly carry out catching and detecting of biomolecule and drug molecule etc.
To achieve these goals, present invention employs following technical scheme:
A preparation method for functional polymer film, said method comprising the steps of:
(1) self assembled monolayer of end-functionalization is formed at biochip substrate surface;
(2) photocrosslinking agent is passed through chemical bonding, be grafted to self assembled monolayer end;
(3) Polymer Solution is spun under lucifuge condition the surface that step (2) is formed, then dry;
(4) there is high molecular biochip to carry out ultraviolet lighting under inert gas shielding surperficial spin coating, under ultraviolet light conditions, carry out chemical bonding, make macromolecular grafted to surface, form macromolecule membrane;
Optionally, step (5) is carried out:
(5) macromolecule is carried out end-functionalization, form functional polymer film, it is in order to catch biomolecule and drug molecule etc., and carries out high flux detection.
The present invention, in conjunction with spin coating technique and photo-crosslinking technology, establishes a kind of simple, quick and general biochip surface chemical modification method, has prepared (function) macromolecule membrane.The present invention mainly utilizes the method for photo-crosslinking to carry out chemical modification in biochip surface, in order to form the three-dimensional surface of high density grafting, then further functionalization carried out to it and utilize the functionalization group of end on SPRi instrument, realize the micromolecular fixing and detection fixed and detect or directly carry out biomolecule or drug molecule such as biomolecule (as albumen, polypeptide, polysaccharide, nucleic acid) or medicine.
Preferably, step (1) is front carries out following pre-service to biochip substrate: biochip substrate cleaned up.
Preferably, described biochip substrate, comprises the material that all may be used for preparing biochip holder, such as glass, silicon chip and quartz; The macromolecule class films such as dimethyl silicone polymer, polystyrene, polycarbonate and polymethylmethacrylate; Metal and the metal-oxide films etc. such as gold film, silverskin and di-aluminium trioxide film.
Preferably, the cleaning way of described biochip substrate includes but not limited to organic solvent (as ethanol, methyl alcohol and N, dinethylformamide etc.) or deionized water carry out rinsing, shaking and wash or ultrasonic cleaning, also comprise and utilize plasma cleaning instrument to carry out surface clean, or both or three kinds of modes arbitrarily are wherein combinationally used.
Preferably, the reagent of self assembled monolayer includes but not limited to the mix reagent of any one or at least two kinds in single mercaptan, dithiol and silylating reagent etc. in step (1).
Preferably, in step (1), the group of end-functionalization includes but not limited to alkoxy, hydroxyl, carboxyl, amino, epoxy radicals or cyano group etc.
Preferably, described in step (2), photocrosslinking agent refers to containing photaesthesia group, the reagent of chemical reaction can be carried out, as the combination of any one in azirine, diazonium, acetophenone, benzophenone or Anthraquinones etc. or at least two kinds under specific wavelength (energy) illumination.
Preferably, the described photocrosslinking agent that makes passes through chemical bonding, the method being grafted to self assembled monolayer end adopts different bonding methods, as amidation, esterification, open loop or nucleophilic substitution etc. according to the difference of biochip substrate surface functionalization group and photocrosslinking agent functional end-group.
The adoptable photocrosslinking agent structural formula of such as the present invention as shown in Figure 2, for (a) structure, adopts esterification to be grafted to self assembled monolayer end; For (b) structure, amidation process is adopted to be grafted to self assembled monolayer end.For esterification process, carboxyl terminal photocrosslinking agent is grafted to hydroxy thiol surface, owing to containing F and N element in above-mentioned photocrosslinking agent, and the surface before graft esterification photocrosslinking agent there is no two kinds of elements and exists, so can judge photocrosslinking agent grafting is successfully according to appearing at of F and N two kinds of element peaks.As shown in Figure 3, there are F and N two obvious peaks in the surperficial XPS data display after esterification, and there is no two elements in contrast simultaneously, proves that photocrosslinking agent is successfully grafted to surface.
Preferably, described macromolecule mainly refers to have biocompatibility and the functional polymer with active end group, include but not limited to the macromolecule with good anti-non-specific binding performance, as polyglycol and derivant, fluoropolymer, polyacrylic acid, polystyrene and PLA etc.; Both sexes betaines polymkeric substance etc.; There is the macromolecule of good biocompatibility, as nitrocellulose, shitosan, glucosan and derivant thereof etc.
Preferably, step (4) described inert gas includes but not limited to nitrogen or/and inert gases such as argon gas.
Preferably, step (4) described ultraviolet wavelength is 365nm.
As shown in Figure 4, by macromolecule by after 365nm ultraviolet light cross-linking to surface, with FT-IR (plunderring angle annex), chip surface is characterized.For the surface of grafting poly-(methacrylic acid macrogol ester), 3500cm
-1there is O-H stretching vibration peak, 3000-3200cm
-1there is CH in place
2vibration bimodal, 1740cm
-1there is obvious C=O stretching vibration peak in left and right, and lower wave number region, as 1100cm
-1there is the peak of obvious C-O and C-C vibration in left and right.Similarly, for the surface of grafting glucosan, 3500cm
-1there is-OH characteristic peak, 3000-3200cm
-1there is CH in place
2vibration bimodal, 1100cm
-1there is the peak of obvious C-O-C stretching vibration in left and right.Above evidence all proves macromolecule by successful photo-crosslinking to biochip surface, also proves the feasibility of this technical scheme.
Preferably, in step (5), the group of end-functionalization includes but not limited to hydroxyl, carboxyl, aldehyde radical, amino, epoxy radicals, cyano group, alkynyl, azido or azirine etc.
Preferably, by regulation and control photocrosslinking agent density (1mM ~ 100mM), macromolecule number-average molecular weight (100,000 ~ 2,000,000), spin coating parameters (1000rpm ~ 8000rpm) etc. can the accurately density of control surface macromolecule membrane and thickness.It is simple, quick and efficient that the method prepares three-dimensional macromolecule surface.
Two of object of the present invention is to provide a kind of functional polymer film obtained by method described above.
Three of object of the present invention is the purposes providing a kind of functional polymer film as above, and its fixing and high flux for biomolecule and drug molecule detects.
Preferably, biomolecule described in the present invention includes but not limited to albumen, polypeptide, nucleic acid (DNA, RNA, cDNA and peptide nucleic acid etc.) and sugared equimolecular.Drug molecule is mainly the pharmaceutical active compounds of chemosynthesis and Separation of Natural Products and purifies the active component etc. obtained.
High-flux detection method described in the present invention, mainly comprises surface plasmon resonance imaging technique, fluorescence labeling detection technique, fluorescence intensity, fluorescence polarization, FRET (fluorescence resonance energy transfer), fluorescent quenching, QCM (Quartz Crystal Microbalance) technology and the method such as various high-throughout electrochemical measuring techniques and hot detection technique.
Compared with the prior art, the present invention has following beneficial effect:
The invention provides a kind of " being grafted to " method newly, spin coating technique and photo-crosslinking technology is utilized to obtain the fixing high molecular method of universality, can utilize ultraviolet light one step by macromolecule non-selectively covalency be fixed on surface, be directly used in preparation and there is the three-dimensional surface biochip of photo-crosslinking structure.
In addition, the method for the invention adopts photocrosslinking agent under UV-irradiation, produce the Cabbeen of high reaction activity, and its reactivity is strong, and the reaction time is short, can at random and macromolecule carry out intercalation reaction, be applicable to all macromolecules, there is universality.
In addition, the method is based on the spin coating technique of mature and reliable, and required instrument and equipment has popularization and versatility.The biochip utilizing this method to prepare all is significantly improved to the fixing of various biological detection thing and detection signal, practical, is applicable to large-scale promotion and commercial applications, has boundless application prospect.
In addition, the present invention preferably adopts relatively long ultraviolet light (365nm), and subsidiary reaction is few, and other decorating molecules of effects on surface (as mercaptan etc.) are injury not.
Accompanying drawing explanation
Fig. 1 modifies process flow diagram based on the functional polymer three-dimensional surface of photo-crosslinking method;
Fig. 2 is the photocrosslinking agent molecular structural formula in the present invention, wherein, and (a) carboxyl terminal photocrosslinking agent; (b) amino terminal photocrosslinking agent;
Fig. 3 is that x-ray photoelectron power spectrum (XPS) is to the surface-element analysis result before and after grafting photocrosslinking agent, wherein, figure (a) and (b) are respectively the XPS figure before and after the grafting of N1s element, and figure (c) and figure (d) is respectively the XPS figure before and after the grafting of F1s element;
Fig. 4 is that Fourier transform infrared spectroscopy (FT-IR) surface to photo-crosslinking grafting polymer characterizes, and (a) gathers (methacrylic acid macrogol ester); (b) glucosan);
Fig. 5 is the glucosan three-dimensional surface bio-sensing chip prepared based on photo-crosslinking method, fixes Streptavidin (SA) in the mode circulated, and detects 4nM biotin (Biotin);
Fig. 6 fixes rapamycin on the surface at poly-(methacrylic acid macrogol ester), SPRi is utilized to detect the FKBP12 (PBS with 100nM, pH7.4,0.5%T20) the interaction of albumen, wherein, rapamycin-1, rapamycin-2 and rapamycin-3 represents the sample repeating for three times to test employing respectively;
Fig. 7 is agglutinin microarray surface plasma resonance imaging figure;
Fig. 8 is that agglutinin array is to serum screening result.
Embodiment
Technical scheme of the present invention is further illustrated by embodiment below in conjunction with accompanying drawing.
Embodiment 1. dextran surface (Small molecular detects: the fixing SA of circulation, detects Biotin)
(1) layers of chrome of one deck 3nm thickness and the layer gold of one deck 47nm thickness is prepared, as the substrate of biochip by the method for hot evaporation on the glass substrate.
(2) the ethanolic solution HS-(CH of C-terminal mercaptan is prepared
2)
11-EG6-OH, concentration is 1mM.
(3) biochip substrate ethanol or washed with de-ionized water is clean, then biochip is put into plasma clean instrument cleaning 3 minutes.
(4) be immersed in by biochip in ready thiol solution, hatch 12 hours, after reaching the schedule time at 4 DEG C, taken out by biochip, the alternately cleaning of ethanol and deionized water, nitrogen dries up.
(5) prepare esterification and connect solution 20mL needed for photocrosslinking agent, carboxyl terminal photocrosslinking agent 10mM, 1-(3-dimethylamino-propyl)-3-ethyl carbodiimide (EDC) 10mM, DMAP (DMAP) 1mM, solvent is DMF (DMF).
(6) immersed by biochip in the esterification solution prepared, room temperature lucifuge reacts 4 hours, after reaching the schedule time, and cleaned with DMF, ethanol and deionized water successively by biochip, nitrogen dries up for subsequent use.
(7) be the aqueous solution that the glucosan of 2000KDa is mixed with mass concentration 40% by molecular weight, stir, removing bubble is to water white uniform solution.
(8) dextran solution prepared is paved with in biochip surface, with the rotating speed of 8000rpm, spin coating 1 minute, after spin coating, by biochip at room temperature lucifuge leave standstill within 1 hour, dry.
(9) biochip is put into ultraviolet light cross-linking instrument, under nitrogen environment, with 365nm UV-irradiation 15 minutes (2.4J/cm
2).
(10) after reaching the schedule time, repeatedly shaken with the hot water of 50 DEG C by biochip and wash 1 hour, remove the dextran molecule that the non-covalency in surface is fixing, period changes water 3-5 time, and the rear nitrogen of biochip cleaning dries up for subsequent use.
(11) biochip is immersed in the DMF solution of succinic anhydride (10mg/mL) and DMAP (15mg/mL), room temperature (25 DEG C) reaction 16 hours, after reaching predetermined reaction time, biochip is taken out, use DMF, ethanol, washed with de-ionized water successively, nitrogen dries up.
(12) sodium acetate buffer (pH=4.5) preparing 50ug/mL Streptavidin (SA) is for subsequent use.
(13) adjust SPRi optical position, SA solution is passed to biochip surface with the speed of 3uL/s, continues in conjunction with 100s.
(14) step 12 is repeated, until surperficial fixed amount reaches capacity.
(15) System Solution is changed into PBS damping fluid (containing 1%DMSO), adjustment optical position, passes into the PBS solution (1%DMSO) of the biotin containing 4nM, in conjunction with 300s, 300s, the 1:100 phosphate aqueous solution that dissociates is lived again, and 10mM NaOH aqueous solution is lived again.
The results show, has very high proteopexy amount with biochip prepared by the method, is enough to detect and protein bound Small molecular (as Fig. 5).
Embodiment 2. poly-(methacrylic acid macrogol ester) surface (array of small molecules)
(1) layers of chrome of one deck 3nm thickness and the layer gold of one deck 47nm thickness is prepared, as the substrate of biochip by the method for hot evaporation on the glass substrate.
(2) ethanolic solution (HS-(CH of the mercaptan of C-terminal and carboxyl terminal is prepared
2)
11-EG6-OH and HS-(CH
2)
11-EG6-COOH), concentration is 1mM, by above two kinds of thiol solutions according to 999:1 (v/v) mixing, for subsequent use.
(3) biochip ethanol or washed with de-ionized water is clean, then biochip is put into plasma clean instrument cleaning 3 minutes.
(4) be immersed in by biochip in the thiol solution mixed, hatch 12 hours, after reaching the schedule time at 4 DEG C, taken out by biochip, the alternately cleaning of ethanol and deionized water, nitrogen dries up.
(5) biochip surface carboxyl is activated, immerse in the aqueous solution of 1-(3-dimethylamino-propyl)-3-ethyl carbodiimide (EDC) and N-hydroxy-succinamide (NHS) (0.4M and 0.1M) again, incubated at room 30 minutes, reach the schedule time, with water cleaning, nitrogen dries up for subsequent use.
(6) biochip is immersed the N of 10mM amino terminal photocrosslinking agent, in dinethylformamide (DMF) solution, room temperature lucifuge reacts 4 hours, after reaching the schedule time, cleaned with DMF, ethanol and deionized water successively by biochip, nitrogen dries up for subsequent use.
(7) atomic radicals transfer polymerization (ATRP) method is utilized to synthesize the methanol/water solution of poly-(methacrylic acid macrogol ester) high molecular polymer.By cupric chloride (CuCl
2) solution mixes with its part second bipyridine (Bpy), stirs 15min.First monomer OEGMA and monomer HEMA is added the mixed solution of first alcohol and water (1:1, volume ratio) with 1:1 (mol ratio), ultrasonic 15min, continue to pass into nitrogen 30min.Add the aqueous ascorbic acid that 0.04M newly configures again, stir 10min and be mixed, then add the ethanolic solution of the dithiol initiating agent of 1mM, wherein the mol ratio of initiating agent and monomer is 1:25000, continues to pass into nitrogen, reacts 16 hours.
(8) ATRP macromolecule stoste step 7 obtained revolves steaming (35 DEG C), and removing methyl alcohol, remaining liq dichloromethane solvent extracts, centrifugal.Take out lower floor's dichloromethane solution part, be spin-dried for.
(9) with the macromolecule of purifying in ethanol dissolving step (8), the ethanolic solution of poly-(methacrylic acid macrogol ester) high molecular polymer of 2 μMs is obtained, for subsequent use.
(10) Polymer Solution configured is paved with in biochip surface, with the rotating speed of 8000rpm, spin coating 1 minute, after spin coating, by biochip at room temperature lucifuge leave standstill within 1 hour, dry, after reaching the schedule time, second alcohol and water is cleaning alternately, and nitrogen dries up.
(11) biochip is put into ultraviolet light cross-linking instrument, under nitrogen environment, with 365nm UV-irradiation 15 minutes (2.4J/cm
2).
(12), after reaching the schedule time, by biochip second alcohol and water ultrasonic cleaning, nitrogen dries up core for subsequent use.
(13) biochip is immersed in the DMF solution of succinic anhydride (10mg/mL) and DMAP (15mg/mL), room temperature reaction 12 hours, after reaching predetermined reaction time, biochip is taken out, use DMF, ethanol and washed with de-ionized water successively, nitrogen dries up.
(14) be layered on biochip surface with the ethanolic solution of the EG3 of 1mM, close 30min, then second alcohol and water alternately cleaning, nitrogen dries up.So far, poly-(methacrylic acid macrogol ester) surperficial chip prepared by photo-crosslinking method completes, and again utilizes photo-crosslinking method to fix Small molecular and utilizes SPRi to detect its associated protein, as follows:
(15) biochip surface is activated again, immerse in the aqueous solution of 1-(3-dimethylamino-propyl)-3-ethyl carbodiimide (EDC) and N-hydroxy-succinamide (NHS) (0.4M and 0.1M), incubated at room 30 minutes, reach the schedule time, with water cleaning, nitrogen dries up for subsequent use.
(16) biochip is immersed again the N of 10mM photocrosslinking agent, in dinethylformamide (DMF) solution, room temperature lucifuge reacts 4 hours, after reaching the schedule time, cleaned with DMF, ethanol and deionized water successively by biochip, nitrogen dries up for subsequent use.
(17) the PBST damping fluid (wherein tween 0.05%) of 100nM rapamycin (Rapamycin) small molecule solution and 100ug/mLFKBP12 albumen is prepared, for subsequent use.
(18) by the surface that small molecule solution is prepared in step (16) by point sample instrument point sample, vacuum drying.
(19) chip is put into ultraviolet light cross-linking instrument again, under nitrogen environment, with 365nm UV-irradiation 15 minutes (2.4J/cm
2), carry out photo-crosslinking.
(20) adjust SPRi optical position, FKBP12 protein solution is passed to chip surface with the speed of 3uL/s, continues in conjunction with 300s, dissociate 300s, and glycine-HCI is lived again.
The results show, has very high Small molecular fixed amount with biochip prepared by the method, is enough to the interaction (as shown in Figure 6) detecting that Small molecular is combined with its associated protein.
Embodiment 3. dextran surface (antigen-antibody fluoroscopic examination)
(1) substrate of glass ethanol or washed with de-ionized water are totally, biochip put into plasma clean instrument cleaning 3 minutes, as the substrate of biochip.
(2) prepare, in the 3-aminopropyl triethoxysilane (APES) of 1:50 acetone diluted, to take out after 20-30s, then with the APES that pure acetone solution removal is not tied, APES is combined with glass, form one deck unimolecular layer at glass surface.
(3) soak 30 minutes in the glutaraldehyde of 25%, with acetone cleaning, one end of immersion 1mM is amino one end is in the aqueous solution of the PEG of hydroxyl, incubated at room temperature 1h, after reaching the schedule time, takes out biochip, by washed with de-ionized water, nitrogen dries up.
(4) prepare esterification and connect solution 20mL needed for photocrosslinking agent, carboxyl terminal photocrosslinking agent 10mM, 1-(3-dimethylamino-propyl)-3-ethyl carbodiimide (EDC) 10mM, DMAP (DMAP) 1mM, solvent is DMF (DMF).
(5) immersed by biochip in the esterification solution prepared, room temperature lucifuge reacts 4 hours, after reaching the schedule time, and cleaned with DMF, ethanol and deionized water successively by biochip, nitrogen dries up for subsequent use.
(6) be the aqueous solution that the glucosan of 2000KDa is mixed with mass concentration 40% by molecular weight, stir, removing bubble is to water white uniform solution.
(7) dextran solution prepared is paved with in biochip surface, with the rotating speed of 8000rpm, spin coating 1 minute, after spin coating, by chip at room temperature lucifuge leave standstill within 1 hour, dry.
(8) biochip is put into ultraviolet light cross-linking instrument, under nitrogen environment, with 365nm UV-irradiation 15 minutes (2.4J/cm
2).
(9) after reaching the schedule time, repeatedly shaken with the hot water of 50 DEG C by biochip and wash 1 hour, remove the dextran molecule that the non-covalency in surface is fixing, period changes water 3-5 time, and the rear nitrogen of biochip cleaning dries up for subsequent use.
(10) biochip is immersed in the DMF solution of succinic anhydride (10mg/mL) and DMAP (15mg/mL), room temperature (25 DEG C) reaction 16 hours, after reaching predetermined reaction time, biochip is taken out, use DMF, ethanol, washed with de-ionized water successively, nitrogen dries up, for subsequent use.
(11) human IgG (solvent is the sodium acetate solution of pH value 4.5) getting the 1mg/ml of 3 μ l with liquid-transfering gun drops on the table that obtains in step (10), and the BSA being aided with 1mg/ml is negative control, incubated at room 1 hour, by the phosphate buffer clean surface twice (each 5 minutes) of the Tween-20 containing 0.0005g/ml, and with after washed with de-ionized water, nitrogen dries up.
(12) carry out after room temperature closes 1 hour with the skimmed milk solution (Skim milk) of 0.05g/ml (taking the phosphate buffer that 5g skimmed milk power is dissolved in the 0.1M of 100ml to be prepared from) effects on surface, by the phosphate buffer clean surface twice (each 5 minutes) of the Tween-20 containing 0.0005g/ml, washed with de-ionized water, nitrogen dries up.
(13) by fluorescein isothiocynate (Fluorescein isothiocyanate, FITC) the Goat anti human IgG marked is diluted in the skim milk aqueous solution of 0.005mg/ml with the extension rate of 1:50, and the chip after step 12 processes is immersed in this skim milk, room temperature places 1 hour, carry out antigen-antibody reaction, by the phosphate buffer clean surface twice (each 5 minutes) of the Tween-20 of 0.0005g/ml, and with after washed with de-ionized water, nitrogen dries up.
(14) by the chip after step 13 processes, by using Leica M-6000 fluorescent microscope with the 300ms time shutter under 480nm halogen light modulation, light intensity is carry out fluorescence imaging with 20 times of object lens under the condition of 5.
Embodiment 4. poly-(methacrylic acid macrogol ester) surface (SPRi glycoprotein array detection serum)
(1) layers of chrome of one deck 3nm thickness and the layer gold of one deck 47nm thickness is prepared, as the substrate of biochip by the method for hot evaporation on the glass substrate.
(2) ethanolic solution (HS-(CH of the mercaptan of C-terminal and carboxyl terminal is prepared
2)
11-EG6-OH and HS-(CH
2)
11-EG6-COOH), concentration is 1mM, by above two kinds of thiol solutions according to 999:1 (v/v) mixing, for subsequent use.
(3) biochip ethanol or washed with de-ionized water is clean, then biochip is put into plasma clean instrument cleaning 3 minutes.
(4) be immersed in by biochip in the thiol solution mixed, hatch 12 hours, after reaching the schedule time at 4 DEG C, taken out by biochip, the alternately cleaning of ethanol and deionized water, nitrogen dries up.
(5) biochip surface carboxyl is activated, chip is immersed in the aqueous solution of 1-(3-dimethylamino-propyl)-3-ethyl carbodiimide (EDC) and N-hydroxy-succinamide (NHS) (0.4M and 0.1M), incubated at room 30 minutes, reach the schedule time, with water cleaning, nitrogen dries up for subsequent use.
(6) biochip is immersed the N of 10mM photocrosslinking agent, in dinethylformamide (DMF) solution, room temperature lucifuge reacts 4 hours, after reaching the schedule time, cleaned with DMF, ethanol and deionized water successively by biochip, nitrogen dries up for subsequent use.
(7) atomic radicals transfer polymerization (ATRP) method is utilized to synthesize the methanol/water solution of poly-(methacrylic acid macrogol ester) high molecular polymer.By cupric chloride (CuCl
2) solution mixes with its part second bipyridine, stirs 15min.First monomer OEGMA and monomer HEMA is added the mixed solution of first alcohol and water (1:1, volume ratio) with 1:1 (mol ratio), ultrasonic 15min, continue to pass into nitrogen 30min.Add the aqueous ascorbic acid that 0.04M newly configures again, stir 10min and be mixed.Then add the ethanolic solution of the dithiol initiating agent of 1mM, wherein the mol ratio of initiating agent and monomer is 1:25000, continues to pass into nitrogen, reacts 16 hours.
(8) the ATRP macromolecule stoste that step (7) obtains is revolved steaming (35 degree), removing methyl alcohol, remaining liq dichloromethane solvent extracts, centrifugal, takes out lower floor's dichloromethane solution part, is spin-dried for.
(9) with the macromolecule of purifying in ethanol dissolving step (8), the ethanolic solution of poly-(methacrylic acid macrogol ester) high molecular polymer of purifying is obtained, for subsequent use.
(10) Polymer Solution configured is paved with in biochip surface, with the rotating speed of 8000rpm, spin coating 1 minute, after spin coating, by biochip at room temperature lucifuge leave standstill within 1 hour, dry.
(11) biochip is put into ultraviolet light cross-linking instrument, in nitrogen environment, with 365nm UV-irradiation 15 minutes (2.4J/cm
2).
(12), after reaching the schedule time, by biochip second alcohol and water ultrasonic cleaning, nitrogen dries up for subsequent use.
(13) biochip is immersed in the DMF solution of succinic anhydride (10mg/mL) and DMAP (15mg/mL), room temperature reaction 16 hours, after reaching predetermined reaction time, biochip is taken out, use DMF, ethanol, washed with de-ionized water successively, nitrogen dries up.So far, poly-(methacrylic acid macrogol ester) surperficial chip prepared by photo-crosslinking method completes, utilize point sample instrument carry out the fixing of agglutinin (lectin) and utilize SPRi to the coherent detection of type 1 diabetes (T1), diabetes B (T2) and 1.5 patients with type Ⅰ DM (LADA) human serum, and with normal human serum in contrast (C).
(14) biochip surface is activated, immerse in the aqueous solution of 1-(3-dimethylamino-propyl)-3-ethyl carbodiimide (EDC) and N-hydroxy-succinamide (NHS) (0.4M and 0.1M), incubated at room 30 minutes, reach the schedule time, with water cleaning, nitrogen dries up for subsequent use.
(15) point sample instrument is utilized by 49 kinds of different lectin point samples in the chip surface prepared, vacuum drying, stand-by.
(16) by the chip in step (15) with 2% skim milk (PBS solution) close, spend the night under 4 degree of conditions, use 10*PBST successively, 1*PBST, 0.1*PBST and water cleaning 10min, nitrogen dries up, stand-by.
(17) with HEPES damping fluid, 4 kinds of different blood serum samples are diluted with 1:4000, for subsequent use.
(18) SPRi optical position is adjusted, first pass into HEPES damping fluid 10min with the flow velocity of 4ul/s, 1:200 hydrochloric acid (adding 0.05% tween) is used to live again again surperficial 3 times, and blood serum sample solution is passed to chip surface with the speed random sequence of 3uL/s, continue in conjunction with 300s, 300s, the 1:200 hydrochloric acid (adding 0.05% tween) that dissociates is lived again.
The results show, has very high lectin fixed amount with chip prepared by the method, and variety classes diabetes serum (as Fig. 7 and Fig. 8) can well be detected.
Protein microarray is prepared on embodiment 5. polyacrylic acid surface (PAA)
(1) layers of chrome of one deck 3nm thickness and the layer gold of one deck 47nm thickness is prepared, as the substrate of biochip by the method for hot evaporation on the glass substrate.
(2) ethanolic solution (HS-(CH of the mercaptan of C-terminal and carboxyl terminal is prepared
2)
11-EG6-OH and HS-(CH
2)
11-EG6-COOH), concentration is 1mM.By two kinds of thiol solutions (EG6-OH:EG6-COOH) according to 999:1 mixing, for subsequent use.
(3) biochip ethanol or washed with de-ionized water is clean, then biochip is put into plasma clean instrument cleaning 3 minutes.
(4) be immersed in by biochip in the thiol solution mixed, hatch 12 hours, after reaching the schedule time at 4 DEG C, taken out by biochip, the alternately cleaning of ethanol and deionized water, nitrogen dries up.
(5) biochip surface carboxyl is activated, biochip is immersed in the aqueous solution of 1-(3-dimethylamino-propyl)-3-ethyl carbodiimide (EDC) and N-hydroxy-succinamide (NHS) (0.4M and 0.1M), incubated at room 30 minutes, reach the schedule time, with water cleaning, nitrogen dries up for subsequent use.
(6) biochip is immersed the N of 10mM carboxyl terminal photocrosslinking agent, in dinethylformamide (DMF) solution, room temperature lucifuge reacts 4 hours, after reaching the schedule time, cleaned with DMF, ethanol and deionized water successively by biochip, nitrogen dries up for subsequent use.
(7) by molecular weight be 140000 polyacrylic acid macromolecule (PAA) be mixed with the aqueous solution of volumetric concentration 1%, stir, removing bubble is to water white uniform solution.
(8) the PAA solution configured is paved with in biochip surface, with the rotating speed of 8000rpm, spin coating 1 minute, after spin coating, by biochip at room temperature lucifuge leave standstill within 1 hour, dry.
(9) biochip is put into ultraviolet light cross-linking instrument, under nitrogen environment, with 365nm UV-irradiation 15 minutes (2.4J/cm
2).
(10), after reaching the schedule time, cleaned by biochip use water, nitrogen dries up for subsequent use.
(11) biochip surface is activated, immerse in the aqueous solution of 1-(3-dimethylamino-propyl)-3-ethyl carbodiimide (EDC) and N-hydroxy-succinamide (NHS) (0.4M and 0.1M), incubated at room 30 minutes.Reach the schedule time, with water cleaning, nitrogen dries up for subsequent use.So far, polyacrylic acid surface chip prepared by photo-crosslinking method completes, and utilizes SPRi to carry out the detection of the fixing of antigen and antibody.
(12) PBS solution of H-IgG of 1mg/ml and the PBS solution of the Goat-anti-H-IgG of 100ug/ml is configured, for subsequent use.
(13) by the chip surface that the H-IgG solution point prepared ocean prepares in step (11), dry, stand-by.
(14) close 10min by under the BSA solution room temperature of biochip 1mg/ml, PBS solution and water cleaning, nitrogen dries up, for subsequent use.
(15) adjust SPRi optical position, the PBS solution of Goat-anti-H-IgG is passed to chip surface with the speed of 3uL/s, continues in conjunction with 300s, dissociate 300s, and NaOH solution is lived again.
The results show, has very high antigen fixed amount with biochip prepared by the method, the interaction of antigen and its antibody can well be detected.
Applicant states, the present invention illustrates method detailed of the present invention by above-described embodiment, but the present invention is not limited to above-mentioned method detailed, does not namely mean that the present invention must rely on above-mentioned method detailed and could implement.Person of ordinary skill in the field should understand, any improvement in the present invention, to equivalence replacement and the interpolation of auxiliary element, the concrete way choice etc. of each raw material of product of the present invention, all drops within protection scope of the present invention and open scope.
Claims (10)
1. a preparation method for functional polymer film, is characterized in that, said method comprising the steps of:
(1) self assembled monolayer of end-functionalization is formed at biochip substrate surface;
(2) photocrosslinking agent is passed through chemical bonding, be grafted to self assembled monolayer end;
(3) Polymer Solution is spun under lucifuge condition the surface that step (2) is formed, then dry;
(4) there is high molecular biochip to carry out ultraviolet lighting under inert gas shielding surperficial spin coating, under ultraviolet light conditions, carry out chemical bonding, make macromolecular grafted to surface, form macromolecule membrane;
Optionally, step (5) is carried out:
(5) macromolecule is carried out end-functionalization, form functional polymer film.
2. the method for claim 1, is characterized in that, step (1) is front carries out following pre-service to biochip substrate: biochip substrate cleaned up.
3. method as claimed in claim 1 or 2, it is characterized in that, described biochip substrate is glass, silicon chip, quartz, dimethyl silicone polymer, polystyrene, polycarbonate, polymethylmethacrylate, golden film, silverskin or di-aluminium trioxide film;
Preferably, the cleaning way of described biochip substrate for carry out rinsing, shaking and wash or ultrasonic cleaning with organic solvent or deionized water, or utilizes plasma cleaning instrument to carry out surface clean, or its combination.
4. the method as described in one of claim 1-3, is characterized in that, the reagent of self assembled monolayer is the mix reagent of any one or at least two kinds in single mercaptan, dithiol and silylating reagent in step (1);
Preferably, in step (1), the group of end-functionalization is alkoxy, hydroxyl, carboxyl, amino, epoxy radicals or cyano group.
5. the method as described in one of claim 1-4, is characterized in that, photocrosslinking agent described in step (2) is the combination of any one or at least two kinds in azirine, diazonium, acetophenone, benzophenone or Anthraquinones;
Preferably, described in make photocrosslinking agent by chemical bonding, the method being grafted to self assembled monolayer end is amidation, esterification, open loop or nucleophilic substitution.
6. the method as described in one of claim 1-5, it is characterized in that, described macromolecule is polyglycol and derivant, fluoropolymer, polyacrylic acid, polystyrene, PLA, both sexes betaines polymkeric substance, nitrocellulose, shitosan or glucosan and derivant thereof.
7. the method as described in one of claim 1-6, is characterized in that, step (4) described inert gas is that nitrogen is or/and argon gas;
Preferably, step (4) described ultraviolet wavelength is 365nm;
Preferably, in step (5), the group of end-functionalization is hydroxyl, carboxyl, aldehyde radical, amino, epoxy radicals, cyano group, alkynyl, azido or azirine;
Preferably, photocrosslinking agent density is 1mM ~ 100mM, and macromolecule number-average molecular weight is 100,000 ~ 2,000,000, and spin coating rotating speed is 1000rpm ~ 8000rpm.
8. the functional polymer film obtained by the described method of one of claim 1-7.
9. a purposes for functional polymer film as claimed in claim 8, its fixing and high flux for biomolecule and drug molecule detects.
10. purposes as claimed in claim 9, it is characterized in that, described biomolecule is albumen, polypeptide, nucleic acid and sugar; Described drug molecule is that the pharmaceutical active compounds of chemosynthesis and Separation of Natural Products are purified the active component obtained;
Preferably, described high flux detects and comprises surface plasmon resonance imaging technique, fluorescence labeling detection technique, fluorescence intensity, fluorescence polarization, FRET (fluorescence resonance energy transfer), fluorescent quenching, QCM (Quartz Crystal Microbalance) technology, high-throughout electrochemical measuring technique and high-throughout hot detection technique.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510045459.8A CN104597230B (en) | 2015-01-29 | 2015-01-29 | A kind of functional polymer film, preparation method and applications |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510045459.8A CN104597230B (en) | 2015-01-29 | 2015-01-29 | A kind of functional polymer film, preparation method and applications |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104597230A true CN104597230A (en) | 2015-05-06 |
| CN104597230B CN104597230B (en) | 2017-04-05 |
Family
ID=53123149
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510045459.8A Expired - Fee Related CN104597230B (en) | 2015-01-29 | 2015-01-29 | A kind of functional polymer film, preparation method and applications |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104597230B (en) |
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104897617A (en) * | 2015-05-19 | 2015-09-09 | 国家纳米科学中心 | Micro-array biochip, and preparation method and application of micro-array biochip |
| CN106383224A (en) * | 2016-08-31 | 2017-02-08 | 中国科学院长春应用化学研究所 | Biological detection element and preparation method thereof |
| CN106896087A (en) * | 2017-03-31 | 2017-06-27 | 丁利 | The preparation method of the surface plasma resonance instrument chip based on hyperbranched amphion polymethyl base cysteine modified |
| CN106990079A (en) * | 2017-03-20 | 2017-07-28 | 中国科学院化学研究所 | A kind of surface multifunctional coating and preparation method and application |
| WO2017144359A1 (en) | 2016-02-22 | 2017-08-31 | Boehringer Ingelheim Vetmedica Gmbh | Method for the immobilization of biomolecules |
| CN109900814A (en) * | 2017-12-08 | 2019-06-18 | 中国科学院大连化学物理研究所 | It is a kind of based on glycosidic bond mass spectrum can fragmentation type chemical cross-linking agent analysis method and application |
| CN110282997A (en) * | 2019-07-10 | 2019-09-27 | 浙江理工大学 | A kind of polystyrene laminated film and its preparation method and application |
| CN111497366A (en) * | 2020-04-07 | 2020-08-07 | 上海交通大学 | Interface-controllable non-layered multi-level graphene conformal folds and preparation method thereof |
| CN112982012A (en) * | 2021-02-09 | 2021-06-18 | 清华大学 | Polyacrylamide derivative modified filter paper and application thereof in nucleic acid separation and enrichment |
| CN115677922A (en) * | 2022-09-28 | 2023-02-03 | 深圳市曙芯生物科技有限公司 | Difunctional polymer and modification method |
Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0590661A1 (en) * | 1992-09-30 | 1994-04-06 | Matsushita Electric Industrial Co., Ltd. | A method for measuring concentrations of substrates in a sample liquid by using a biosensor |
| CN101078730A (en) * | 2006-05-22 | 2007-11-28 | 杜宏武 | Enzyme labeled chip method capable of in situ detecting multiple ordinary small molecule drugs |
| CN102242053A (en) * | 2011-04-01 | 2011-11-16 | 沈越 | Biochip with polymer three-dimensional nanostructure |
| CN102439076A (en) * | 2009-05-26 | 2012-05-02 | 国立大学法人东京工业大学 | Self-supporting thin polymer film |
| WO2012173071A1 (en) * | 2011-06-13 | 2012-12-20 | 国立大学法人 新潟大学 | Transmitted light control device |
| CN102901809A (en) * | 2012-09-27 | 2013-01-30 | 中国科学院半导体研究所 | Production method for microporous polymer film biochip |
| CN103409809A (en) * | 2013-07-17 | 2013-11-27 | 国家纳米科学中心 | Small molecule drug screening chip, construction method and application thereof |
| CN103792345A (en) * | 2014-02-18 | 2014-05-14 | 国家纳米科学中心 | Small-molecule microarray and preparation method thereof |
-
2015
- 2015-01-29 CN CN201510045459.8A patent/CN104597230B/en not_active Expired - Fee Related
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0590661A1 (en) * | 1992-09-30 | 1994-04-06 | Matsushita Electric Industrial Co., Ltd. | A method for measuring concentrations of substrates in a sample liquid by using a biosensor |
| CN101078730A (en) * | 2006-05-22 | 2007-11-28 | 杜宏武 | Enzyme labeled chip method capable of in situ detecting multiple ordinary small molecule drugs |
| CN102439076A (en) * | 2009-05-26 | 2012-05-02 | 国立大学法人东京工业大学 | Self-supporting thin polymer film |
| CN102242053A (en) * | 2011-04-01 | 2011-11-16 | 沈越 | Biochip with polymer three-dimensional nanostructure |
| WO2012173071A1 (en) * | 2011-06-13 | 2012-12-20 | 国立大学法人 新潟大学 | Transmitted light control device |
| CN102901809A (en) * | 2012-09-27 | 2013-01-30 | 中国科学院半导体研究所 | Production method for microporous polymer film biochip |
| CN103409809A (en) * | 2013-07-17 | 2013-11-27 | 国家纳米科学中心 | Small molecule drug screening chip, construction method and application thereof |
| CN103792345A (en) * | 2014-02-18 | 2014-05-14 | 国家纳米科学中心 | Small-molecule microarray and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| 曾戎等: "《生物医用仿生高分子材料》", 31 October 2010, 华南理工大学出版社 * |
| 顾长志等: "《微纳加工及在纳米材料与器件研究中的应用》", 30 June 2013, 科学出版社 * |
Cited By (16)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104897617A (en) * | 2015-05-19 | 2015-09-09 | 国家纳米科学中心 | Micro-array biochip, and preparation method and application of micro-array biochip |
| WO2017144359A1 (en) | 2016-02-22 | 2017-08-31 | Boehringer Ingelheim Vetmedica Gmbh | Method for the immobilization of biomolecules |
| CN106383224A (en) * | 2016-08-31 | 2017-02-08 | 中国科学院长春应用化学研究所 | Biological detection element and preparation method thereof |
| CN106990079A (en) * | 2017-03-20 | 2017-07-28 | 中国科学院化学研究所 | A kind of surface multifunctional coating and preparation method and application |
| CN106990079B (en) * | 2017-03-20 | 2019-06-25 | 中国科学院化学研究所 | A kind of surface multifunctional coating and the preparation method and application thereof |
| CN106896087A (en) * | 2017-03-31 | 2017-06-27 | 丁利 | The preparation method of the surface plasma resonance instrument chip based on hyperbranched amphion polymethyl base cysteine modified |
| CN106896087B (en) * | 2017-03-31 | 2019-04-23 | 丁利 | The preparation method of surface plasma resonance instrument chip based on hyperbranched amphoteric ion polymethyl base cysteine modified |
| CN109900814B (en) * | 2017-12-08 | 2021-06-08 | 中国科学院大连化学物理研究所 | Analysis method and application of fragmentable chemical cross-linking agent based on glycosidic bond mass spectrum |
| CN109900814A (en) * | 2017-12-08 | 2019-06-18 | 中国科学院大连化学物理研究所 | It is a kind of based on glycosidic bond mass spectrum can fragmentation type chemical cross-linking agent analysis method and application |
| CN110282997A (en) * | 2019-07-10 | 2019-09-27 | 浙江理工大学 | A kind of polystyrene laminated film and its preparation method and application |
| CN111497366A (en) * | 2020-04-07 | 2020-08-07 | 上海交通大学 | Interface-controllable non-layered multi-level graphene conformal folds and preparation method thereof |
| CN111497366B (en) * | 2020-04-07 | 2021-06-15 | 上海交通大学 | Interface-controllable non-layered multi-level graphene conformal folds and preparation method thereof |
| CN112982012A (en) * | 2021-02-09 | 2021-06-18 | 清华大学 | Polyacrylamide derivative modified filter paper and application thereof in nucleic acid separation and enrichment |
| CN112982012B (en) * | 2021-02-09 | 2022-05-10 | 杭州梓晶生物有限公司 | Polyacrylamide derivative modified filter paper and application thereof in nucleic acid separation and enrichment |
| CN115677922A (en) * | 2022-09-28 | 2023-02-03 | 深圳市曙芯生物科技有限公司 | Difunctional polymer and modification method |
| CN115677922B (en) * | 2022-09-28 | 2024-01-26 | 深圳市曙芯生物科技有限公司 | Dual-functional polymer and modification method |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104597230B (en) | 2017-04-05 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104597230B (en) | A kind of functional polymer film, preparation method and applications | |
| KR101140029B1 (en) | Preparation method of antigen-immobilized immuno- fluorescence slide and the immuno-fluoroscence slide made by the method | |
| US7728094B2 (en) | Selfassembled grafted polymeric layer for use in biosensor technology | |
| Wang et al. | Fabrication and anti-fouling properties of photochemically and thermally immobilized poly (ethylene oxide) and low molecular weight poly (ethylene glycol) thin films | |
| JP2010530955A5 (en) | ||
| JP2010530955A (en) | Dual functional non-polluting surfaces and materials | |
| JP4818056B2 (en) | Structure having a binding functional group on a substrate for binding a capture molecule for capturing a target substance | |
| WO2005111618A1 (en) | Substance-fixing agents, method of fixing substance with the same, and substrate having substance fixed with the same | |
| Ivanov et al. | Interferometric detection of chloramphenicol via its immunochemical recognition at polymer-coated nano-corrugated surfaces | |
| Xu et al. | Spatially well-defined binary brushes of poly (ethylene glycol) s for micropatterning of active proteins on anti-fouling surfaces | |
| Sut et al. | Supported lipid bilayer coatings: Fabrication, bioconjugation, and diagnostic applications | |
| Lin et al. | An extremely simple method for fabricating 3D protein microarrays with an anti-fouling background and high protein capacity | |
| Gosecka et al. | Hydrophilic polymers grafted surfaces: preparation, characterization, and biomedical applications. Achievements and challenges | |
| JP5361221B2 (en) | Method for producing mixed SAM | |
| CN107003306A (en) | Porous membrane, its method and the purposes being grafted with polymer | |
| JP2007057458A (en) | Biosensor | |
| Shrestha et al. | Application of printable antibody ink for solid-phase immobilization of ABO antibody using photoactive hydrogel for surface plasmon resonance imaging | |
| Bernand‐Mantel et al. | Protein‐functionalized ultrathin glycidyl methacrylate polymer grafts on gold for the development of optical biosensors: an SPR investigation | |
| JP2000146976A (en) | Metal thin film for spr sensor, its manufacture and measuring method usint it | |
| JP2006266741A (en) | Biosensor | |
| US20110171070A1 (en) | Surface-modified sensor device and method for surface-modifying the same | |
| JP2006266742A (en) | Biosensor | |
| JP5344438B2 (en) | Substrate for fixing substance, substrate for fixing substance, and analysis method | |
| KR101195254B1 (en) | Preparation method of antigen-immobilized immuno- fluorescence slide and the immuno-fluoroscence slide made by the method | |
| KR101195253B1 (en) | Preparation method of antigen-immobilized immuno- fluorescence slide and the immuno-fluoroscence slide made by the method |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20170405 |