CN107064072B - The preparation method of surface plasma resonance instrument chip based on amphoteric ion polymethyl base cysteine modified - Google Patents
The preparation method of surface plasma resonance instrument chip based on amphoteric ion polymethyl base cysteine modified Download PDFInfo
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- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 title claims abstract description 35
- 235000018417 cysteine Nutrition 0.000 title claims abstract description 35
- 238000002360 preparation method Methods 0.000 title claims abstract description 15
- -1 mercapto alkene Chemical class 0.000 claims abstract description 34
- 239000000178 monomer Substances 0.000 claims abstract description 16
- 229920001690 polydopamine Polymers 0.000 claims abstract description 8
- 239000003999 initiator Substances 0.000 claims abstract description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 90
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 54
- 229910052757 nitrogen Inorganic materials 0.000 claims description 42
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 42
- 239000008367 deionised water Substances 0.000 claims description 34
- 229910021641 deionized water Inorganic materials 0.000 claims description 34
- 239000000243 solution Substances 0.000 claims description 31
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims description 24
- 150000002500 ions Chemical class 0.000 claims description 24
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 claims description 18
- 239000000758 substrate Substances 0.000 claims description 18
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 16
- VYFYYTLLBUKUHU-UHFFFAOYSA-N dopamine Chemical compound NCCC1=CC=C(O)C(O)=C1 VYFYYTLLBUKUHU-UHFFFAOYSA-N 0.000 claims description 16
- VYZAMTAEIAYCRO-UHFFFAOYSA-N Chromium Chemical compound [Cr] VYZAMTAEIAYCRO-UHFFFAOYSA-N 0.000 claims description 13
- QAOWNCQODCNURD-UHFFFAOYSA-N sulfuric acid Substances OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 12
- 238000007872 degassing Methods 0.000 claims description 11
- 238000004108 freeze drying Methods 0.000 claims description 10
- 239000000843 powder Substances 0.000 claims description 10
- 238000004140 cleaning Methods 0.000 claims description 8
- 229960003638 dopamine Drugs 0.000 claims description 8
- 235000019441 ethanol Nutrition 0.000 claims description 8
- 239000011259 mixed solution Substances 0.000 claims description 8
- 238000005516 engineering process Methods 0.000 claims description 7
- 239000011521 glass Substances 0.000 claims description 7
- ROFVEXUMMXZLPA-UHFFFAOYSA-N Bipyridyl Chemical group N1=CC=CC=C1C1=CC=CC=N1 ROFVEXUMMXZLPA-UHFFFAOYSA-N 0.000 claims description 6
- 229910021589 Copper(I) bromide Inorganic materials 0.000 claims description 6
- 239000007983 Tris buffer Substances 0.000 claims description 6
- ODWXUNBKCRECNW-UHFFFAOYSA-M bromocopper(1+) Chemical compound Br[Cu+] ODWXUNBKCRECNW-UHFFFAOYSA-M 0.000 claims description 6
- 238000006243 chemical reaction Methods 0.000 claims description 6
- NKNDPYCGAZPOFS-UHFFFAOYSA-M copper(i) bromide Chemical compound Br[Cu] NKNDPYCGAZPOFS-UHFFFAOYSA-M 0.000 claims description 6
- 230000008021 deposition Effects 0.000 claims description 6
- 229910001873 dinitrogen Inorganic materials 0.000 claims description 6
- 238000005566 electron beam evaporation Methods 0.000 claims description 6
- 229910052698 phosphorus Inorganic materials 0.000 claims description 6
- 239000011574 phosphorus Substances 0.000 claims description 6
- 238000003756 stirring Methods 0.000 claims description 6
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 claims description 6
- 238000000605 extraction Methods 0.000 claims description 5
- 241000252506 Characiformes Species 0.000 claims description 4
- 230000015572 biosynthetic process Effects 0.000 claims description 4
- 239000006210 lotion Substances 0.000 claims description 4
- 238000003786 synthesis reaction Methods 0.000 claims description 4
- 238000005406 washing Methods 0.000 claims description 4
- 229910052709 silver Inorganic materials 0.000 claims description 3
- 239000004332 silver Substances 0.000 claims description 3
- 239000011248 coating agent Substances 0.000 claims description 2
- 238000000576 coating method Methods 0.000 claims description 2
- 238000000034 method Methods 0.000 claims description 2
- 239000013307 optical fiber Substances 0.000 claims description 2
- 238000006116 polymerization reaction Methods 0.000 claims description 2
- DWNBOPVKNPVNQG-LURJTMIESA-N (2s)-4-hydroxy-2-(propylamino)butanoic acid Chemical compound CCCN[C@H](C(O)=O)CCO DWNBOPVKNPVNQG-LURJTMIESA-N 0.000 claims 1
- NIXOWILDQLNWCW-UHFFFAOYSA-N 2-Propenoic acid Natural products OC(=O)C=C NIXOWILDQLNWCW-UHFFFAOYSA-N 0.000 claims 1
- XYIBRDXRRQCHLP-UHFFFAOYSA-N ethyl acetoacetate Chemical compound CCOC(=O)CC(C)=O XYIBRDXRRQCHLP-UHFFFAOYSA-N 0.000 claims 1
- QQONPFPTGQHPMA-UHFFFAOYSA-N propylene Natural products CC=C QQONPFPTGQHPMA-UHFFFAOYSA-N 0.000 claims 1
- 239000012460 protein solution Substances 0.000 abstract description 8
- 230000003373 anti-fouling effect Effects 0.000 abstract description 6
- 238000001514 detection method Methods 0.000 abstract description 4
- 238000012650 click reaction Methods 0.000 abstract 1
- 238000010526 radical polymerization reaction Methods 0.000 abstract 1
- 241001494479 Pecora Species 0.000 description 18
- 108060003951 Immunoglobulin Proteins 0.000 description 16
- 102000018358 immunoglobulin Human genes 0.000 description 16
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 12
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 10
- 235000013336 milk Nutrition 0.000 description 9
- 239000008267 milk Substances 0.000 description 9
- 210000004080 milk Anatomy 0.000 description 9
- 238000001179 sorption measurement Methods 0.000 description 9
- 239000008363 phosphate buffer Substances 0.000 description 7
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 6
- 229940098773 bovine serum albumin Drugs 0.000 description 6
- 235000010469 Glycine max Nutrition 0.000 description 5
- 244000068988 Glycine max Species 0.000 description 5
- 102000016943 Muramidase Human genes 0.000 description 5
- 108010014251 Muramidase Proteins 0.000 description 5
- 108010062010 N-Acetylmuramoyl-L-alanine Amidase Proteins 0.000 description 5
- 239000002253 acid Substances 0.000 description 5
- 229910021529 ammonia Inorganic materials 0.000 description 5
- 239000004325 lysozyme Substances 0.000 description 5
- 229960000274 lysozyme Drugs 0.000 description 5
- 235000010335 lysozyme Nutrition 0.000 description 5
- 230000004044 response Effects 0.000 description 5
- 210000002966 serum Anatomy 0.000 description 5
- HZAXFHJVJLSVMW-UHFFFAOYSA-N 2-Aminoethan-1-ol Chemical compound NCCO HZAXFHJVJLSVMW-UHFFFAOYSA-N 0.000 description 4
- 102000008946 Fibrinogen Human genes 0.000 description 4
- 108010049003 Fibrinogen Proteins 0.000 description 4
- 230000001143 conditioned effect Effects 0.000 description 4
- 238000001035 drying Methods 0.000 description 4
- 125000004494 ethyl ester group Chemical group 0.000 description 4
- 229940012952 fibrinogen Drugs 0.000 description 4
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 4
- 229910052737 gold Inorganic materials 0.000 description 4
- 239000010931 gold Substances 0.000 description 4
- 239000006228 supernatant Substances 0.000 description 4
- 238000009210 therapy by ultrasound Methods 0.000 description 4
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 3
- 230000008859 change Effects 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- 239000002184 metal Substances 0.000 description 3
- 229910052751 metal Inorganic materials 0.000 description 3
- CCMKPCBRNXKTKV-UHFFFAOYSA-N 1-hydroxy-5-sulfanylidenepyrrolidin-2-one Chemical compound ON1C(=O)CCC1=S CCMKPCBRNXKTKV-UHFFFAOYSA-N 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 2
- 102000006395 Globulins Human genes 0.000 description 2
- 108010044091 Globulins Proteins 0.000 description 2
- 150000001412 amines Chemical class 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 239000000427 antigen Substances 0.000 description 2
- 102000036639 antigens Human genes 0.000 description 2
- 108091007433 antigens Proteins 0.000 description 2
- 238000002242 deionisation method Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 229940088598 enzyme Drugs 0.000 description 2
- 229940031098 ethanolamine Drugs 0.000 description 2
- 238000002347 injection Methods 0.000 description 2
- 239000007924 injection Substances 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 230000010355 oscillation Effects 0.000 description 2
- 235000018102 proteins Nutrition 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 235000013322 soy milk Nutrition 0.000 description 2
- 101001065501 Escherichia phage MS2 Lysis protein Proteins 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- AMPHKYRLSOPVBX-YFKPBYRVSA-N allylcysteine Chemical compound OC(=O)[C@H](CS)NCC=C AMPHKYRLSOPVBX-YFKPBYRVSA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000007664 blowing Methods 0.000 description 1
- 238000003759 clinical diagnosis Methods 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- 235000020247 cow milk Nutrition 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 230000009878 intermolecular interaction Effects 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 239000007800 oxidant agent Substances 0.000 description 1
- 230000001590 oxidative effect Effects 0.000 description 1
- 238000007747 plating Methods 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 238000002198 surface plasmon resonance spectroscopy Methods 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/55—Specular reflectivity
- G01N21/552—Attenuated total reflection
- G01N21/553—Attenuated total reflection and using surface plasmons
Landscapes
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention discloses a kind of preparation methods of surface plasma resonance instrument chip based on amphoteric ion polymethyl base cysteine modified, methylpropenyl cysteine monomer is obtained by " mercapto alkene " click-reaction, initiator is modified by the poly-dopamine on golden film surface to surface, and then the Transfer Radical Polymerization caused by surface causes monomer in golden film surface aggregate, to obtain a kind of surface plasma resonance instrument chip based on amphoteric ion polymethyl base cysteine modified, chip is to all having very strong antifouling property in single protein solution and natural complex sample system, and there is high supported quantity and good compatibility to large biological molecule such as antibody, target molecule is highly sensitive in achievable complexity albumen system, unmarked quick detection；Have many advantages, such as very strong antifouling property, high stability and high supported quantity, there is good feasibility and practicability.
Description
Technical field
The present invention relates to surface plasma resonance spectrometer antipollution chip preparation fields, are based on both sexes more particularly to one kind
The preparation method of the surface plasma resonance instrument chip of ion polymethyl base cysteine modified.
Background technique
Surface plasma resonance (Surface Plasmon Resonance, abbreviation SPR) instrument is that occur the 1980s
A kind of biosensor technology, when incident light is incident on low refractive index dielectric from high refractive index medium, it may occur that total reflection,
If plating one layer of metallic film (gold or silver) at dielectric interface at this time, the evanescent wave that incident light generates can cause metal surface certainly
Collective oscillation is occurred by electronics, and then forms plasma, when the frequency and wave number phase of evanescent wave and surface plasmon oscillations
Whens equal, i.e. generation covibration (SPR) causes energy to be transferred in surface plasma-wave from evanescent wave, makes the intensity of reflected light
Weaken significantly, decaying total reflection phenomenon is presented, the incidence angle when the intensity of reflected light is zero is known as resonance angle.Resonance angle and gold
It is related to belong to film interface medium refraction index, and refractive index changes with the molecular mass for being incorporated in metal film surfaces, therefore can
The information that intermolecular interaction is obtained by analysis resonance angle, since it has the characteristics that detection quickly and is not necessarily to mark,
It has been widely used in the fields such as proteomics, medicament research and development, clinical diagnosis, food safety and environmental monitoring, and has shown
Wide application prospect out.
SPR chip is the component of surface plasma resonance instrument core the most, provides the necessary physics item for generating spr signal
Part mainly includes coupled apparatus, metal film and surface matrix.However, when application surface plasma resonance instrument carries out analysis test,
The surface contamination of sensing chip is a common problem, the non-specificity of biomolecule or microorganism in sample
Absorption will reduce the accuracy of quantitative detection, generate false positive results.Therefore, exploitation is effective against the surface of non-specific adsorption
It is the matter of utmost importance for improving surface plasma resonance sensor sensitivity and accuracy.
Summary of the invention
For the deficiencies in the prior art, the present invention provides a kind of based on half Guang ammonia of amphoteric ion polymethyl base
The preparation method of the surface plasma resonance instrument chip of acid modification.
The present invention is achieved by the following technical solutions:
A kind of preparation method of the surface plasma resonance instrument chip based on amphoteric ion polymethyl base cysteine,
The following steps are included:
A) naked golden chip preparation
The layers of chrome for coating one layer of 2-3nm thickness in substrate using electron beam evaporation deposition technology, coats one in the layers of chrome
The golden film of layer 45-50nm thickness obtains naked golden chip；
B) naked golden chip pretreatment
The naked golden chip that step a) is obtained immerses in Piranha washing lotion, 1-3h is impregnated under room temperature, deionized water is used in taking-up
With being dried with nitrogen after cleaning 3 times；
C) synthesis of amphoteric ion methylpropenyl cysteine monomer
After 24.98mmol cysteine 20mL deionized water dissolving, 27.47mmol 2- acrylic acid-is added thereto
2- hydroxyl -1,3- propylene diester and 29.4 μm of ol xylyl phosphorus, are stirred to react 2-4h, the solution after reaction at 20-30 DEG C
It is extracted with ethyl acetate once, methylene chloride is extracted twice, and the amphoteric ion methylpropenyl of white powder is obtained after freeze-drying
Cysteine monomer；
D) naked golden chip surface dopamine polymerization
1-3mg/mL dopamine solution is prepared with the TRIS buffer of pH=8.5, b) step is obtained
Naked golden chip be immersed, be incubated for 1-2h at 40-60 DEG C, taking-up clean 3 times with deionized water after with being dried with nitrogen；
E) poly-dopamine chip surface bromo isobutyl acylbromide initiator is modified
The substrate of chip that d) step obtains is immersed 20ml to contain in the methylene chloride of 1.0ml triethylamine, in ice bath and
The methylene chloride that 10ml contains 0.9ml bromo isobutyl acylbromide is added dropwise under stirring condition, is stirred to react 12-24h at room temperature, takes
With being dried with nitrogen after successively cleaning 3 times with ethyl alcohol and deionized water out；
F) naked golden chip surface polymethyl base cysteine synthesis
The chip that e) step obtains is put into advance in the Shi Lunke bottle of circulating degasification three times and inflated with nitrogen；C) step is obtained
To 5mmol monomer be dissolved in 4ml deionized water, with nitrogen deaerate 10-30min, thereto be added 0.5mmol bipyridyl,
0.167mmol cuprous bromide and 0.083mmol copper bromide are ultrasonically treated 5-10min with using after nitrogen degassing 10-30min, will
The solution arrived moves into Shi Lunke bottles, so that solution submerges the chip in bottle, takes out chip after reacting 2-4h under condition of nitrogen gas,
With being dried with nitrogen after successively cleaning 3 times with ethyl alcohol and deionized water.
Moreover, above-mentioned substrate is glass sheet substrate or optical fiber substrate.
Moreover, above-mentioned golden film can also be silverskin or silver/Jin Fuhe film.
Moreover, above-mentioned Piranha washing lotion is the mixed solution of the 98wt.% concentrated sulfuric acid and 30wt.% hydrogen peroxide, the concentrated sulfuric acid
Volume ratio with hydrogen peroxide is 7:3.
The invention has the advantages and beneficial effects that:
1) surface plasma resonance instrument prepared by the present invention based on amphoteric ion polymethyl base cysteine modified
Chip has good antifouling property；
2) surface plasma resonance instrument prepared by the present invention based on amphoteric ion polymethyl base cysteine modified
Chip has biological functional effect, such as can detect its antigen by immobilized sheep immune globulin antibody；
3) surface plasma resonance instrument prepared by the present invention based on amphoteric ion polymethyl base cysteine modified
Chip has high stability, after storing 1 month or being stored 7 days in the phosphate buffer that pH is 7.4 in the dry state,
Antifouling property is unchanged；
4) the present invention is based on the surface plasma resonance instrument chips of amphoteric ion polymethyl base cysteine modified
Preparation method has certain universality, is not limited by base material property.
Detailed description of the invention
Fig. 1 is that the present invention is based on the surface plasma resonance instrument chips of amphoteric ion polymethyl base cysteine modified
Structural schematic diagram；
Fig. 2 a is that the present invention is based on the surface plasma resonance instrument cores of amphoteric ion polymethyl base cysteine modified
Piece is to single albumen system (lysozyme (Lysozyme), the bovine serum albumin (BSA), fibrinogen of 1mg/mL
(fibrinogen)) the non-specific adsorption amount of albumen in；
Fig. 2 b is that the present invention is based on the surface plasma resonance instrument cores of amphoteric ion polymethyl base cysteine modified
Piece is to egg in complex system (100% human serum (Blood serum), soya-bean milk (Soybean milk), milk (Cow milk))
White non-specific adsorption amount；
Wherein:
1, glass sheet substrate；2, layers of chrome；3, golden membranous layer；4, poly-dopamine coupling causes oxidant layer；5, the poly- methyl of amphoteric ion
Allyl cysteine layer.
Specific embodiment
The present invention is described in further detail by following embodiment.It should be understood that following embodiments be it is illustrative, be not
It is limited, it cannot be limited the scope of protection of the present invention with following embodiments.
Embodiment 1
The layers of chrome for being coated one layer of 2nm thickness in BK7 substrate of glass using electron beam evaporation deposition technology, is applied in the layers of chrome
The golden film for covering one layer of 48nm thickness obtains naked golden chip；Above-mentioned naked golden chip is immersed into mixing for 98% concentrated sulfuric acid and 30% hydrogen peroxide
It closes in solution, the volume ratio of 98% concentrated sulfuric acid and 30% hydrogen peroxide is 7:3, impregnates 2h under room temperature.It then takes out and uses deionization
Water is spare with being dried with nitrogen after cleaning 3 times.
After 24.98mmol cysteine 20mL deionized water dissolving, 27.47mmol 2- acrylic acid-is added thereto
2- hydroxyl -1,3- propylene diester and 29.4 μm of ol xylyl phosphorus, are stirred to react 4h, the solution acetic acid after reaction at 20 DEG C
Ethyl ester extraction is primary, and methylene chloride is extracted twice, and the half Guang ammonia of amphoteric ion methylpropenyl of white powder is obtained after freeze-drying
Acid monomers；
2mg/mL dopamine solution is prepared with the TRIS buffer of pH=8.5, above-mentioned drying is spare
Naked golden chip be immersed, be incubated for 2h at 40 DEG C, taking-up clean 3 times with deionized water after with being dried with nitrogen；It will be modified with
It includes to add dropwise under ice bath and stirring condition in the 20ml methylene chloride of 1.0ml triethylamine that the substrate of poly-dopamine, which immerses,
Enter the methylene chloride that 10ml includes 0.9ml bromo isobutyl acylbromide, conditioned response is then stirred at room temperature for 24 hours, second is used in taking-up
Pure and mild deionized water successively clean 3 times after with being dried with nitrogen, obtained chip is put into advance circulating degasification inflated with nitrogen three times
In Shi Lunke bottles, take the white powder amphoteric ion methylpropenyl cysteine monomer 5mmol obtained after above-mentioned freeze-drying molten
Solution in 4ml deionized water, with nitrogen deaerate 15min, thereto be added 0.5mmol bipyridyl, 0.167mmol cuprous bromide and
0.083mmol copper bromide, then solution is moved into Shi Lunke bottles by what is obtained with nitrogen degassing 10min, rear ultrasonic treatment 5min
In and chip submerged, take out chip after reacting 2h under condition of nitrogen gas, successively respectively cleaned 3 times with ethyl alcohol and deionized water, used
It is dried with nitrogen spare.
By the surface plasma resonance chip pitch of the amphoteric ion methylpropenyl cysteine modified of above-mentioned acquisition
It fits on the prism of surface plasma resonance system, the phosphate buffer for being 7.4 using pH selects flow velocity for 10 μ as mobile phase
L/min.After baseline is steady, the protein solution of 100 μ L 1mg/mL: bovine serum albumin(BSA), bacteriolyze is injected separately into quantitative loop
Enzyme, fibrinogen, 100% serum, soymilk supernatant and milk supernatant push above-mentioned each protein solution to reach through mobile phase
Chip surface reads surface plasma resonance after 20min by surface plasma resonance system measurement resonance angle real-time change curve
Angle changing value, and calculate non-specific adsorption amount, respectively 1.40ng/cm2、0.81ng/cm2、1.89ng/cm2、12.81ng/
cm2、6.08ng/cm2、7.15ng/cm2, non-specific adsorption amount of the protein adsorption quantity far below each albumen of naked gold surface.
Embodiment 2
The layers of chrome for being coated one layer of 3nm thickness in BK7 substrate of glass using electron beam evaporation deposition technology, is applied in the layers of chrome
The golden film for covering one layer of 47nm thickness obtains naked golden chip；Above-mentioned naked golden chip is immersed into mixing for 98% concentrated sulfuric acid and 30% hydrogen peroxide
It closes in solution, the volume ratio of 98% concentrated sulfuric acid and 30% hydrogen peroxide is 7:3, impregnates 1h under room temperature.It then takes out and uses deionization
Water is spare with being dried with nitrogen after cleaning 3 times.
After 24.98mmol cysteine 20mL deionized water dissolving, 27.47mmol 2- acrylic acid-is added thereto
2- hydroxyl -1,3- propylene diester and 29.4 μm of ol xylyl phosphorus, are stirred to react 2h, the solution acetic acid after reaction at 30 DEG C
Ethyl ester extraction is primary, and methylene chloride is extracted twice, and the half Guang ammonia of amphoteric ion methylpropenyl of white powder is obtained after freeze-drying
Acid monomers；
3mg/mL dopamine solution is prepared with the TRIS buffer of pH=8.5, above-mentioned drying is spare
Naked golden chip be immersed, be incubated for 1h at 60 DEG C, taking-up clean 3 times with deionized water after with being dried with nitrogen；It will be modified with
It includes to add dropwise under ice bath and stirring condition in the 20ml methylene chloride of 1.0ml triethylamine that the substrate of poly-dopamine, which immerses,
Enter the methylene chloride that 10ml includes 0.9ml bromo isobutyl acylbromide, conditioned response 18h is then stirred at room temperature, second is used in taking-up
Pure and mild deionized water successively clean 3 times after with being dried with nitrogen, obtained chip is put into advance circulating degasification inflated with nitrogen three times
In Shi Lunke bottles, take the white powder amphoteric ion methylpropenyl cysteine monomer 5mmol obtained after above-mentioned freeze-drying molten
Solution in 4ml deionized water, with nitrogen deaerate 20min, thereto be added 0.5mmol bipyridyl, 0.167mmol cuprous bromide and
0.083mmol copper bromide, then obtained solution is moved into Shi Lunke bottles with nitrogen degassing 20min, rear ultrasonic treatment 10min
In and chip submerged, take out chip after reacting 3h under condition of nitrogen gas, successively respectively cleaned 3 times with ethyl alcohol and deionized water, used
It is dried with nitrogen spare.
By the surface plasma resonance chip pitch of the amphoteric ion methylpropenyl cysteine modified of above-mentioned acquisition
It fits on the prism of surface plasma resonance system, the phosphate buffer for being 7.4 using pH selects flow velocity for 10 μ as mobile phase
L/min.After baseline is steady, the protein solution of 100 μ L 1mg/mL: bovine serum albumin(BSA), bacteriolyze is injected separately into quantitative loop
Enzyme, fibrinogen, 100% serum, soymilk supernatant and milk supernatant push above-mentioned each protein solution to reach through mobile phase
Chip surface reads surface plasma resonance after 20min by surface plasma resonance system measurement resonance angle real-time change curve
Angle changing value, and calculate non-specific adsorption amount, respectively 1.95ng/cm2、1.02ng/cm2、2.20ng/cm2、14.25ng/
cm2、6.95ng/cm2、9.05ng/cm2, non-specific adsorption amount of the protein adsorption quantity far below each albumen of naked gold surface.
Embodiment 3
The layers of chrome for being coated one layer of 2nm thickness in BK7 substrate of glass using electron beam evaporation deposition technology, is applied in the layers of chrome
The golden film for covering one layer of 48nm thickness obtains naked golden chip；Above-mentioned naked golden chip is immersed into 98% concentrated sulfuric acid and 30% hydrogen peroxide (volume
Than 7:3) mixed solution in, the volume ratio of 98% concentrated sulfuric acid and 30% hydrogen peroxide is 7:3, impregnates 2h under room temperature.Then it takes
It is spare with being dried with nitrogen after cleaning 3 times with deionized water out.
After 24.98mmol cysteine 20mL deionized water dissolving, 27.47mmol 2- acrylic acid-is added thereto
2- hydroxyl -1,3- propylene diester and 29.4 μm of ol xylyl phosphorus, are stirred to react 4h, the solution acetic acid after reaction at 20 DEG C
Ethyl ester extraction is primary, and methylene chloride is extracted twice, and the half Guang ammonia of amphoteric ion methylpropenyl of white powder is obtained after freeze-drying
Acid monomers；
2mg/mL dopamine solution is prepared with the TRIS buffer of pH=8.5, above-mentioned drying is spare
Naked golden chip be immersed, be incubated for 2h at 40 DEG C, taking-up clean 3 times with deionized water after with being dried with nitrogen；It will be modified with
It includes to add dropwise under ice bath and stirring condition in the 20ml methylene chloride of 1.0ml triethylamine that the substrate of poly-dopamine, which immerses,
Enter the methylene chloride that 10ml includes 0.9ml bromo isobutyl acylbromide, conditioned response is then stirred at room temperature for 24 hours, second is used in taking-up
Pure and mild deionized water successively clean 3 times after with being dried with nitrogen, obtained chip is put into advance circulating degasification inflated with nitrogen three times
In Shi Lunke bottles, take the white powder amphoteric ion methylpropenyl cysteine monomer 5mmol obtained after above-mentioned freeze-drying molten
Solution in 4ml deionized water, with nitrogen deaerate 15min, thereto be added 0.5mmol bipyridyl, 0.167mmol cuprous bromide and
0.083mmol copper bromide, then solution is moved into Shi Lunke bottles by what is obtained with nitrogen degassing 10min, rear ultrasonic treatment 5min
In and chip submerged, take out chip after reacting 2h under condition of nitrogen gas, successively respectively cleaned 3 times with ethyl alcohol and deionized water, used
It is dried with nitrogen spare.
By the surface plasma resonance chip pitch of the amphoteric ion methylpropenyl cysteine modified of above-mentioned acquisition
It fits on the prism of surface plasma resonance system, the phosphate buffer for being 7.4 using pH selects flow velocity for 10 μ as mobile phase
L/min.After baseline is steady, into quantitative loop, 100 μ L 1- ethyl -3- (3- dimethylaminopropyl) phosphinylidyne of interior injection two is sub-
The mixed solution of amine (EDC, concentration 0.4M) and N- hydroxy thiosuccinimide (NHS, concentration 0.1M), pushes away through mobile phase
It moves above-mentioned mixed solution and reaches chip surface.100 μ L sheep immunoglobulin (anti-IgG, 1mg/mL) solution are injected after 15min,
It pushes sheep immunoglobulin solution to reach chip surface through mobile phase, then is closed through ethanol amine, it is immune that immobilized sheep is obtained after 30min
The surface plasma resonance sensing chip of globulin.
The surface plasma resonance sensing chip of the immobilized sheep immunoglobulin of above-mentioned acquisition is fitted into surface with pitch
On the prism of plasmon resonance system.The phosphate buffer for being 7.4 using pH selects flow velocity for 10 μ L/min as mobile phase.To base
After line is steady, 100 μ L protein solutions: lysozyme (1mg/mL), soya-bean milk (10mg/mL), milk are injected separately into quantitative loop
(30mg/mL) and 100% serum pushes above-mentioned each protein solution to reach chip surface, by surface plasma resonance through mobile phase
System measurement resonance angle real-time change curve reads surface plasma resonance angle changing value after 20min, and calculates non-specific suction
Attached amount, respectively 1.21ng/cm2、6.88ng/cm2、8.35ng/cm2、14.51ng/cm2。
Embodiment 4
The layers of chrome for being coated one layer of 2nm thickness in BK7 substrate of glass using electron beam evaporation deposition technology, is applied in the layers of chrome
The golden film for covering one layer of 48nm thickness obtains naked golden chip；Above-mentioned naked golden chip is immersed into 98% concentrated sulfuric acid and 30% hydrogen peroxide (volume
Than 7:3) mixed solution in, the volume ratio of 98% concentrated sulfuric acid and 30% hydrogen peroxide is 7:3, impregnates 2h under room temperature.Then it takes
It is spare with being dried with nitrogen after cleaning 3 times with deionized water out.
After 24.98mmol cysteine 20mL deionized water dissolving, 27.47mmol 2- acrylic acid-is added thereto
2- hydroxyl -1,3- propylene diester and 29.4 μm of ol xylyl phosphorus, are stirred to react 4h, the solution acetic acid after reaction at 20 DEG C
Ethyl ester extraction is primary, and methylene chloride is extracted twice, and the half Guang ammonia of amphoteric ion methylpropenyl of white powder is obtained after freeze-drying
Acid monomers；
2mg/mL dopamine solution is prepared with the TRIS buffer of pH=8.5, above-mentioned drying is spare
Naked golden chip be immersed, be incubated for 2h at 40 DEG C, taking-up clean 3 times with deionized water after with being dried with nitrogen；It will be modified with
It includes to add dropwise under ice bath and stirring condition in the 20ml methylene chloride of 1.0ml triethylamine that the substrate of poly-dopamine, which immerses,
Enter the methylene chloride that 10ml includes 0.9ml bromo isobutyl acylbromide, conditioned response is then stirred at room temperature for 24 hours, second is used in taking-up
Pure and mild deionized water successively clean 3 times after with being dried with nitrogen, obtained chip is put into advance circulating degasification inflated with nitrogen three times
In Shi Lunke bottles, take the white powder amphoteric ion methylpropenyl cysteine monomer 5mmol obtained after above-mentioned freeze-drying molten
Solution in 4ml deionized water, with nitrogen deaerate 15min, thereto be added 0.5mmol bipyridyl, 0.167mmol cuprous bromide and
0.083mmol copper bromide, then deaerating 10min with nitrogen will be in Shi Lunke bottles of obtained solution immigration afterwards with ultrasonic treatment 5min
And submerge chip, chip is taken out after reacting 2h under condition of nitrogen gas, is successively respectively cleaned 3 times with ethyl alcohol and deionized water, uses nitrogen
Air-blowing is done spare.
By the surface plasma resonance chip pitch of the amphoteric ion methylpropenyl cysteine modified of above-mentioned acquisition
It fits on the prism of surface plasma resonance system.The phosphate buffer for being 7.4 using pH selects flow velocity for 10 μ as mobile phase
L/min.After baseline is steady, into quantitative loop, 100 μ L 1- ethyl -3- (3- dimethylaminopropyl) phosphinylidyne of interior injection two is sub-
The mixed solution of amine (EDC, concentration 0.4M) and N- hydroxy thiosuccinimide (NHS, concentration 0.1M), pushes away through mobile phase
It moves above-mentioned mixed solution and reaches chip surface.100 μ L sheep immunoglobulin (anti-IgG, 1mg/mL) solution are injected after 15min,
It pushes sheep immunoglobulin solution to reach chip surface through mobile phase, then is closed through ethanol amine, obtain immobilized sheep after 30min and exempt from
The surface plasma resonance sensing chip of epidemic disease globulin is injected separately into the protein solution of 100 μ L various concentrations: lysozyme (1mg/
ML), soya-bean milk (10mg/mL), sheep immunoglobulin (15nM), sheep immunoglobulin (75nM), sheep immunoglobulin (150nM),
Sheep immunoglobulin (300nM), sheep immunoglobulin (450nM), sheep immunoglobulin (700nM), sheep immunoglobulin
The mixed liquor of (15nM) and lysozyme (1mg/mL) and the mixed liquor of sheep immunoglobulin (15nM) and soya-bean milk (10mg/mL),
The standard curve of sheep immunoglobulin (IgG) detection is obtained by SPR response signal.
Surface plasma resonance instrument core prepared by the present invention based on amphoteric ion polymethyl base cysteine modified
Piece has good antifouling property；With biological functional effect, can be detected by immobilized sheep immune globulin antibody
Its antigen；With high stability, stores 1 month or stored 7 days in the phosphate buffer that pH is 7.4 in the dry state
Afterwards, antifouling property is unchanged；And preparation method has certain universality, is not limited by base material property.
Illustrative description has been done to the present invention above, it should explanation, the case where not departing from core of the invention
Under, any simple deformation, modification or other skilled in the art can not spend the equivalent replacement of creative work equal
Fall into protection scope of the present invention.
Claims (4)
1. a kind of preparation side of the surface plasma resonance instrument chip based on amphoteric ion polymethyl base cysteine modified
Method, which comprises the following steps:
A) naked golden chip preparation
The layers of chrome for coating one layer of 2-3nm thickness in substrate using electron beam evaporation deposition technology coats one layer of 45- in the layers of chrome
The golden film of 50nm thickness obtains naked golden chip；
B) naked golden chip pretreatment
The naked golden chip that step a) is obtained immerses in Piranha washing lotion, 1-3h is impregnated under room temperature, taking-up cleans 3 with deionized water
With being dried with nitrogen after secondary；
C) synthesis of amphoteric ion methylpropenyl cysteine monomer
After 24.98mmol cysteine 20mL deionized water dissolving, 27.47mmol 2- acrylic acid -2- hydroxyl is added thereto
Base -1,3- propylene diester and 29.4 μm of ol xylyl phosphorus, are stirred to react 2-4h, the solution second after reaction at 20-30 DEG C
Acetoacetic ester extraction is primary, and methylene chloride is extracted twice, and half Guang of amphoteric ion methylpropenyl of white powder is obtained after freeze-drying
Propylhomoserin monomer；
D) naked golden chip surface dopamine polymerization
1-3mg/mL dopamine solution is prepared with the TRIS buffer of pH=8.5, b) step is obtained naked
Golden chip is immersed, and is incubated for 1-2h at 40-60 DEG C, taking-up clean 3 times with deionized water after with being dried with nitrogen；
E) poly-dopamine chip surface bromo isobutyl acylbromide initiator is modified
The substrate for the chip that d) step obtains is immersed 20ml to contain in the methylene chloride of 1.0ml triethylamine, in ice bath and stirring
Under the conditions of the methylene chloride that 10ml contains 0.9ml bromo isobutyl acylbromide is added dropwise, be stirred to react 12-24h at room temperature, take out and use
Ethyl alcohol and deionized water successively clean 3 times after with being dried with nitrogen；
F) naked golden chip surface polymethyl base cysteine synthesis
The chip that e) step obtains is put into advance in the Shi Lunke bottle of circulating degasification three times and inflated with nitrogen；C) step is obtained
5mmol monomer is dissolved in 4ml deionized water, with nitrogen deaerate 10-30min, thereto be added 0.5mmol bipyridyl,
0.167mmol cuprous bromide and 0.083mmol copper bromide are ultrasonically treated 5-10min with using after nitrogen degassing 10-30min, will
The solution arrived moves into Shi Lunke bottles, so that solution submerges the chip in bottle, takes out chip after reacting 2-4h under condition of nitrogen gas,
With being dried with nitrogen after successively cleaning 3 times with ethyl alcohol and deionized water.
2. preparation method according to claim 1, which is characterized in that the substrate is glass sheet substrate or optical fiber base
Bottom.
3. preparation method according to claim 1, which is characterized in that the golden film can also be silverskin or silver/Jin Fuhe
Film.
4. preparation method according to claim 1, which is characterized in that the Piranha washing lotion be the 98wt.% concentrated sulfuric acid with
The volume ratio of the mixed solution of 30wt.% hydrogen peroxide, the concentrated sulfuric acid and hydrogen peroxide is 7:3.
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101065665A (en) * | 2004-11-24 | 2007-10-31 | 康宁股份有限公司 | Polymer-coated substrates for binding biomolecules and methods of making and using thereof |
| CN103728278A (en) * | 2013-12-11 | 2014-04-16 | 王丽红 | Function of surfactant, and signal amplification method for SPR (Surface Plasma Resonance) detection on low molecular weight substances |
| CN104155453A (en) * | 2014-07-11 | 2014-11-19 | 天津大学 | Hyaluronic acid modified surface plasma resonance spectrometer chip and preparation method thereof |
| CN105044359A (en) * | 2015-07-14 | 2015-11-11 | 天津大学 | Surface plasma resonator chip based on modification of hyaluronic acid assisted by dopamine and preparation method of chip |
| CN105403540A (en) * | 2015-12-04 | 2016-03-16 | 天津大学 | Surface plasmon resonance instrument chip based on zwitterion heptapeptide modification and preparation method thereof |
-
2017
- 2017-03-31 CN CN201710209598.9A patent/CN107064072B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101065665A (en) * | 2004-11-24 | 2007-10-31 | 康宁股份有限公司 | Polymer-coated substrates for binding biomolecules and methods of making and using thereof |
| CN103728278A (en) * | 2013-12-11 | 2014-04-16 | 王丽红 | Function of surfactant, and signal amplification method for SPR (Surface Plasma Resonance) detection on low molecular weight substances |
| CN104155453A (en) * | 2014-07-11 | 2014-11-19 | 天津大学 | Hyaluronic acid modified surface plasma resonance spectrometer chip and preparation method thereof |
| CN105044359A (en) * | 2015-07-14 | 2015-11-11 | 天津大学 | Surface plasma resonator chip based on modification of hyaluronic acid assisted by dopamine and preparation method of chip |
| CN105403540A (en) * | 2015-12-04 | 2016-03-16 | 天津大学 | Surface plasmon resonance instrument chip based on zwitterion heptapeptide modification and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| Design and mechanisms of antifouling materials for surface plasmon resonance sensors;Boshi Liu 等;《Acta Biomaterialia》;20161231;100-118 |
| 牛血清白蛋白-三聚氰胺偶联物修饰的表面等离子体激元共振芯片检测三聚氰胺含量;刘珊珊 等;《分析化学》;20140531;第42卷(第5期);695-700 |
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