CN107807111A - The three-dimensional of integrated nanometer optical resonator waits the application for being used for cancer protein mark liquid biopsy from resonance sensor - Google Patents
The three-dimensional of integrated nanometer optical resonator waits the application for being used for cancer protein mark liquid biopsy from resonance sensor Download PDFInfo
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- CN107807111A CN107807111A CN201711012212.1A CN201711012212A CN107807111A CN 107807111 A CN107807111 A CN 107807111A CN 201711012212 A CN201711012212 A CN 201711012212A CN 107807111 A CN107807111 A CN 107807111A
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- 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
- G01N21/59—Transmissivity
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- 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
- G01N21/59—Transmissivity
- G01N2021/5903—Transmissivity using surface plasmon resonance [SPR], e.g. extraordinary optical transmission [EOT]
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Abstract
The invention provides one kind without frequency conversion cubical array plasma resonance sensor, including plasma resonance substrate, for the array of cavities of nanostructured, multi-layered material structure layer is covered thereon, including the first metal layer, insulating barrier and second metal layer arranged successively from bottom to up, the multi-layered material structure layer forms the optical resonator array being embedded in the plasma resonance sensor in substrate, and the optical resonator is the cavity of tapered cups.It is limited to solve prior art plasma resonance sensor detection sensitivity, and the large-scale not readily portable technical problem of device, hypersensitive, label-free biomolecule detection are realized, to realize that opening for the portable medical biology sensor of early detection disease biomarker provides basis.
Description
Technical field
The present invention relates to field of sensing technologies, especially, is related to a kind of the three-dimensional of integrated nanometer optical resonator and waits from altogether
Vibration sensor is used for the application of cancer protein mark liquid biopsy.
Background technology
Carcinomebryonic antigen (CEA) is that nineteen sixty-five Gold and Freedman have found from fetus and colon cancer tissue first.CEA
It is the polysaccharide protein complex that a kind of molecular weight is 22KD, 45% is protein.CEA encoding gene is located at No. 19 chromosomes.
Generally, CEA is as synthesized by Gastrointestinal Tract of Fetus epithelial tissue, pancreas and liver cell.The CEA in 6 months generally before gestation
Content increases, and content is very low in serum after birth.CEA concentration is less than in the non-smoking healthy adult human serums of 96%-97%
2.5 μ g/L, a large amount of smokers have 20%-40% people CEA>2.5 μ g/L, a small number of CEA>5.0μg/L.
CEA belongs to non-organ specificity tumor associated antigen, and the tumour for secreting CEA is located at hollow organ, such as stomach and intestine mostly
Road, respiratory tract, the urinary tract etc..CEA is through gastrointestinal tract under normal circumstances, and CEA during neoplastic state then enters blood and lymph
Circulation, causes change of serum C EA to increase extremely, the change of serum C EA of above-mentioned various tumor patients is had and increase.Clinically, when CEA is big
When 60 μ g/L, it is seen that in colon and rectum carcinoma, stomach cancer and lung cancer.CEA values raise, and show there is lesion remaining or progress.Such as lung
Cancer, breast cancer, carcinoma of urinary bladder and serum of ovarian cancer patients CEA amounts can be significantly raised, are shown as tumor-infiltrated mostly, wherein about 70%
For metastatic carcinoma.In general, 6 weeks after surgery excision, CEA is horizontal to recover normal, and otherwise prompting has residual tumor, if CEA is dense
The constant rise of degree, or its numerical value prompt prognosis mala more than normal 5-6 times of person.Continuous follow-up quantitatively detects change of serum C EA
Content, the tumour state of an illness is judged more significant.
Carcinomebryonic antigen is a broad spectrum activity tumor markers, and it can reflect the presence of kinds of tumors to people, to large intestine
The judgement of the effect of cancer, breast cancer and lung cancer, disease development, monitoring and prognosis estimation are a preferable tumor markerses.97%
Health adult's change of serum C EA concentration in below 2.5g/L, primary colon cancer patient CEA, which increases, accounts for 45~80%.Except primary
Beyond colon cancer, cancer of pancreas, cholangiocarcinoma, stomach cancer, the cancer of the esophagus, gland cancer, lung cancer, the tumer positive rate of breast cancer and urinary system
It is very high, typically 50~70%.Therefore the Indexs measure has greater significance for diagnosing some diseases.
Traditional measure CEA method has fluoroimmunoassay, radioimmunology, Enzyme-Linked Immunospot etc..These inspections
Effectively to technical requirements height, taken time and effort although survey method is sensitive to a certain extent, and have certain requirement to instrument,
Each there is certain limitation.
At present, low concentration biomolecule is detected, such as the label-free inductor of tumor markers, in drug development and is showed
Got most of the attention in the applications such as field care diagnostic.Optics such as photonic crystal, toroidal cavity resonator, interferometer and surface plasma
Resonate (SPR) sensor, due to its Sensitive Detection, is hopeful to replace traditional enzyme linked immunosorbent assay (ELISA) technology (ELISA)
Method.But the bulky external optical device for having precise positioning be still needs-which has limited the popularization of application.Separately
Outside, the plasma sensor based on superpower light transmission (EOT) can use incoherent light source excitating surface plasma, use
Portable spectrometer realizes detection；But the sensitivity of these devices is than traditional spr sensor order of magnitude lower.
The content of the invention
The invention provides a kind of three-dimensional wait of integrated nanometer optical resonator to be used for cancer protein mark from resonance sensor
The application of will thing liquid body biopsy, it is limited to solve prior art detection method sensitivity, operating technology and using existing on instrument
The problem of significant limitations.
To achieve the above object, the present invention proposes a kind of the three-dimensional of integrated nanometer optical resonator and waited from resonance sensor
For the application of cancer protein mark liquid biopsy, comprise the following steps：
(1) three-dimensional is waited and be immersed in from resonance sensor in 2~20mM mercaptoundecylic acid (MUA) solution, trained at room temperature
Support 12~36 hours, sensor surface forms self-assembled monolayer；
(2) the CEA antibody that concentration is 1~100 μ g/ml is passed through into carbodiimides (EDC) and n-hydroxysuccinimide
Couple (NHS), and with mercaptoundecylic acid (MUA) covalent bond；
(3) will be cultivated after step (1) processing without frequency conversion cubical array plasma resonance sensor in BAS confining liquids,
Cultivated afterwards in ethanolamine solutions；
(4) being placed in without frequency conversion cubical array plasma resonance sensor containing A samples to be tested after step (3) is cultivated
Phosphate buffer (PBS) in cultivate；
(5) spectral measurement is carried out, transmission peaks are measured in water.
Preferably, the step (2) is to carry out at ambient temperature, step (1) processing after sensor with 70% second
Alcohol cleans, in N after cleaning2Middle drying, in 400mM carbodiimides (EDC) and 100mM n-hydroxysuccinimides after drying
(NHS) 1 is pressed:Cultivate in the mixture of 1 mixing 30~120 minutes, afterwards rush sensor in phosphate buffer (PBS)
Wash, immediately with 1~100 μ g/ml monoclonal CEA antibodie culture 1~5 hour after flushing.
Preferably, the step (3) is to carry out at ambient temperature, is cultivated in 10 μ g/mL BSA confining liquids, afterwards
30~60 minutes are cultivated in 10% ethanolamine solutions to seal up unreacted n-hydroxysuccinimide coupling (NHS).
Preferably, the three-dimensional of the integrated nanometer optical resonator is waited from resonance sensor to tumor markers carcinomebryonic antigen
CEA detection is limited to 1ng/ml.
Preferably, the application also includes establishing CEA linear models, concretely comprises the following steps：
(1) three-dimensional is waited and be immersed in from resonance sensor in 10mM mercaptoundecylic acid (MUA) solution, sensor surface shape
Into self-assembled monolayer；
(2) the CEA antibody that concentration is 30 μ g/ml is coupled by carbodiimides with n-hydroxysuccinimide, and with
Mercaptoundecylic acid covalent bond；
(3) will be cultivated after step (1) processing without frequency conversion cubical array plasma resonance sensor in BAS confining liquids,
Cultivated afterwards in ethanolamine solutions；
(4) being respectively placed in without frequency conversion cubical array plasma resonance sensor containing concentration ladder after step (3) is cultivated
In the phosphate buffer (PBS) for the CEA that degree scope is 0.1ng/ml~1000ng/ml, contain mouse immuning ball protein
(Anti-MouseIgG) cultivated in phosphate buffer (PBS) and in the phosphate buffer (PBS) without CEA；
(5) spectral measurement is carried out, transmission peaks are measured in water.
Preferably, in the step of of establishing CEA linear models (4), CEA concentration be respectively 0.1ng/ml, 1ng/ml,
10ng/ml、50ng/ml、100ng/ml、1000ng/ml。
Preferably, three-dimensional wait includes plasma resonance substrate from resonance sensor, is the array of cavities of nanostructured,
Multi-layered material structure layer, including the first metal layer, insulating barrier and the second metal layer arranged successively from bottom to up are covered thereon, should
Multi-layered material structure layer forms the optical resonator array being embedded in the plasma resonance sensor, the light in substrate
Learn the cavity that resonator is tapered cups.
Preferably, described three-dimensional etc. is 25nm from the first metal layer thickness of resonance sensor, thickness of insulating layer 80nm.
Beneficial effect：
Three-dimensional wait of integrated nanometer optical resonator provided by the invention is used for cancer protein mark from resonance sensor
The application of liquid biopsy, the detection limit to protein such as tumor markers carcinomebryonic antigen (CEA) are 1ng/mL (5pM), are realized
Overdelicate label-free biomolecule detection, this is to Current surface plasma resonance (SPR) system, and clinically
Measurement protein concentration significantly improves to the horizontal related dynamic ranges of human body diseases mark such as CEA.
The present invention detects cancer protein label, nothing using three-dimensional wait of integrated nanometer optical resonator from resonance sensor
Any fluorescence or chemiluminescent labels need to be used, does not also use secondary antibody or sandwich double antibody, with prior art phase
Than mark and secondary antibody or double antibody being eliminated, so as to accomplish the really single stage detection without additive reagent.With it is traditional
SPR inductors are different, the present invention is three-dimensional wait from resonance sensor in the case where being offset without formant by transmiting peak intensity
Change realizes RI sensings, therefore, need not have the spectroscope of high spectral resolution in detection process.For future research and development just
Formula medical bio sensor is taken to lay a good foundation.
Brief description of the drawings
Fig. 1 is that the three-dimensional of CEA detections of the present invention is waited from the surface-functionalized schematic diagram of resonance sensor；
Fig. 2 is the block diagram of each experimental group relative intensity change of CEA linear models；
Fig. 3 is the relative intensity variation diagram under the CEA concentration gradients change of the model of corresponding diagram 2；
Fig. 4 is the intensity in transmission variation diagram that 10ng/mlCEA experimental groups change over time；
Fig. 5 be in Fig. 4 the μ of part 1 shown in A g/ml CEA experimental groups and 10ng/ml CEA experimental groups with three-dimensional etc. from resonance
Intensity in transmission variation diagram during the CEA antibody bindings combined on sensor；
Fig. 6 is serum excipient control group, detects CEA in human serum sample and increases sharply (1 μ g/ml and 100ng/ml)
Block diagram；
Fig. 7 is the three-dimensional structural representation waited from resonance sensor of integrated nanometer optical resonator.
In figure：10- is three-dimensional to be waited from resonance sensor, 1- plasma resonance substrates；2- the first metal layers；3- insulating barriers；4-
Second metal layer；5- resonators；M- incident lights；N- transmitted lights.
Embodiment
In order to facilitate the understanding of the purposes, features and advantages of the present invention, it is below in conjunction with the accompanying drawings and specific real
Applying mode, the present invention is further detailed explanation.
Detection is purchased from Sigma-Aldrich companies with reagent material in the present invention.
Three-dimensional wait of integrated nanometer optical resonator provided by the invention is used for cancer protein mark from resonance sensor
The application of liquid biopsy, comprises the following steps：
(1) three-dimensional is waited and be immersed in from resonance sensor in 2~20mM mercaptoundecylic acid (MUA) solution, preferably 10~
In 15mM mercaptoundecylic acid (MUA) solution, at room temperature cultivate 12~36 hours, preferably 18~32 hours, sensor surface shape
Into self-assembled monolayer；
(2) the CEA antibody that concentration is 1~100 μ g/ml is passed through into carbodiimides (EDC) and n-hydroxysuccinimide
Couple (NHS), and be 20~60 μ g/ml with mercaptoundecylic acid (MUA) covalent bond, preferred concentration；
(3) will be cultivated after step (1) processing without frequency conversion cubical array plasma resonance sensor in BAS confining liquids,
Cultivated afterwards in ethanolamine solutions；
(4) being placed in without frequency conversion cubical array plasma resonance sensor containing A samples to be tested after step (3) is cultivated
Phosphate buffer (PBS) in cultivate；
(5) spectral measurement is carried out, transmission peaks are measured in water.
Wherein, step (2) is to carry out at ambient temperature, and the sensor after step (1) processing is cleaned with 70% ethanol,
In N after cleaning2Middle drying, pressed after drying in 400mM carbodiimides (EDC) and 100mM n-hydroxysuccinimides (NHS)
1:30~120 minutes, preferably 30~60 minutes are cultivated in the mixture of 1 mixing, afterwards by sensor in phosphate buffer
(PBS) rinsed in, immediately with 1~100 μ g/ml monoclonal CEA antibodie culture 1~5 hour, preferably 20~60 μ after flushing
G/ml monoclonal CEA antibodie, preferably cultivate 1~3 hour.
Wherein, the step (3) is to carry out at ambient temperature, is cultivated in 10 μ g/mL BSA confining liquids, Zhi Hou
30~60 minutes are cultivated in 10% ethanolamine solutions to seal up unreacted n-hydroxysuccinimide coupling (NHS), preferably
Culture 30~40 minutes.
Embodiment 1
Three-dimensional wait that this implementation provides a kind of integrated nanometer optical resonator is used for cancer protein mark from resonance sensor
The application of thing liquid body biopsy, for detecting tumor marker CEA concentration, as shown in figure 1, the three-dimensional is waited from resonance sensor table
Face combines MUA (11-mercaptuoudecanoic acid, mercaptoundecylic acid), by CEA antibody bindings to MUA, treats test sample
CEA antigens in this are combined with CEA, and three-dimensional grade is strong from the transmission of resonance sensor after being combined with CEA by detecting CEA antigens
Degree, measures CEA antigen concentrations.Specifically comprise the following steps：
(1) by the mercaptoundecylic acid (11-mercaptuoudecanoic that 10mM is immersed in from resonance sensor such as three-dimensional
Acid, MUA) in solution, cultivate 24 hours at room temperature, sensor surface forms self-assembled monolayer, one layer of MUA molecule in close and
Uniform fold is in sensor surface；
(2) the CEA antibody that concentration is 30 μ g/ml is coupled by carbodiimides (EDC) with n-hydroxysuccinimide
(NHS), and with mercaptoundecylic acid (MUA) covalent bond；
(3) will be cultivated after step (1) processing without frequency conversion cubical array plasma resonance sensor in BAS confining liquids,
Cultivated afterwards in ethanolamine solutions；
(4) being placed in without frequency conversion cubical array plasma resonance sensor containing sample to be tested after step (3) is cultivated
Cultivated in phosphate buffer (PBS)；
(5) spectral measurement is carried out, transmission peaks are measured in water.
Wherein, the step (2) is to carry out at ambient temperature, step (1) processing after sensor with 70% ethanol
Cleaning, in N after cleaning2Middle drying, in 400mM carbodiimides (EDC) and 100 mM n-hydroxysuccinimides after drying
(NHS) 1 is pressed:Cultivated in the mixture of 1 mixing 30 minutes, sensor is rinsed in phosphate buffer (PBS) afterwards, rinsed
Afterwards immediately with 30 μ g/ml monoclonal CEA antibodie culture 2 hours.
The step (3) is to carry out at ambient temperature, is cultivated in 10 μ g/ml BSA confining liquids, afterwards 10%
30 minutes are cultivated in ethanolamine solutions to seal up unreacted n-hydroxysuccinimide coupling (NHS).
In the present embodiment, detection sample is the human serum sample that CEA concentration is 1 μ g/ml and 100ng/ml, is assigned with serum
As a control group, testing result is as shown in fig. 6, intensity in transmission increases sharply with the growth of CEA concentration for shape agent.The present invention is fast and effective
Detect CEA change in concentration.
Embodiment 2
The present embodiment has also set up CEA linear models, concretely comprises the following steps：
(1) three-dimensional is waited and be immersed in from resonance sensor in 10mM mercaptoundecylic acid (MUA) solution, sensor surface shape
Into self-assembled monolayer；
(2) the CEA antibody that concentration is 30 μ g/ml is coupled by carbodiimides with n-hydroxysuccinimide, and with
Mercaptoundecylic acid covalent bond；
(3) will be cultivated after step (1) processing without frequency conversion cubical array plasma resonance sensor in BAS confining liquids,
Cultivated afterwards in ethanolamine solutions；
(4) being respectively placed in without frequency conversion cubical array plasma resonance sensor containing concentration ladder after step (3) is cultivated
In the phosphate buffer (PBS) for the CEA that degree scope is 0.1ng/ml~1000ng/ml, contain mouse immuning ball protein
(Anti-MouseIgG) cultivated in phosphate buffer (PBS) and in the phosphate buffer (PBS) without CEA；
(5) spectral measurement is carried out, transmission peaks are measured in water.
Wherein, the step (2) is to carry out at ambient temperature, step (1) processing after sensor with 70% ethanol
Cleaning, in N after cleaning2Middle drying, in 400mM carbodiimides (EDC) and 100 mM n-hydroxysuccinimides after drying
(NHS) 1 is pressed:Cultivated in the mixture of 1 mixing 30 minutes, sensor is rinsed in phosphate buffer (PBS) afterwards, rinsed
Afterwards immediately with 30 μ g/ml monoclonal CEA antibodie culture 2 hours.
The step (3) is to carry out at ambient temperature, is cultivated in 30 μ g/ml BSA confining liquids, afterwards 10%
30 minutes are cultivated in ethanolamine solutions to seal up unreacted n-hydroxysuccinimide coupling (NHS).
In the step (4), CEA concentration is respectively 0.1ng/ml, 1ng/ml, 10ng/ml, 50 ng/ml, 100ng/
ml、1000ng/ml.Referring to Fig. 2, it is shown that CEA concentration, antibody (IgG) control group and PBS excipient controls in detection above-mentioned 6
The intensity in transmission of group, intensity in transmission strengthen with the increase of CEA concentration, and concentration 0.1ng/ml CEA intensity in transmission is less than antibody
(IgG) intensity in transmission of control group, it is seen that CEA of the present invention detection limit is 1ng/ml (5pM).Referring to Fig. 3, it is shown that corresponding diagram
Relative intensity change under the CEA concentration gradients change of 2 models, has good identical property with linear model shown in Fig. 2.Ginseng
See Fig. 4, it is shown that the intensity in transmission variation diagram that concentration 10ng/ml CEA experimental groups change over time.Referring to Fig. 5, it is shown that dense
Degree 1000ng/ml CEA and concentration 10ng/ml CEA transmissions when fixed CEA antibody on the sensor surface is combined are strong
Degree changes with time, and with the growth of binding time, intensity in transmission strengthens.
The result shown in Fig. 2-Fig. 5, in present invention application, three-dimensional using integrated nanometer optical resonator is waited from altogether
Vibration sensor detection cancer protein mark CEA concentration, detection are limited to 1ng/ml, hence it is evident that less than the inspection of CEA in the prior art
Rising limit 3ng/ml, the three-dimensional optical sensitive characteristic waited from resonance sensor device of the present invention, makes its light transmittance for surface antigen
The combination of antibody is very sensitive, so as to reach sensitive, accurate testing result.This is to Current surface plasma resonance
(SPR) system, and clinically measure protein concentration and the horizontal related dynamic ranges of human body diseases mark such as CEA
Significantly improve.
In Example 1 and Example 2 of the present invention, three-dimensional wait of the integrated nanometer optical resonator is stood from resonance sensor
3D phasmon nanometer cup resonator structures are enough to, by being formed with golden (Au) layer, the nanostructured polymer substrate of deposition.
Specifically, three-dimensional wait includes plasma resonance substrate from resonance sensor, it is the array of cavities of nanostructured, covers thereon
Multi-layered material structure layer, including the first metal layer, insulating barrier and the second metal layer arranged successively from bottom to up, the multilayer material
Structure sheaf forms the optical resonator array being embedded in the plasma resonance sensor, the optical resonator in substrate
For the cavity of tapered cups.The first metal layer is Au layers, and thickness 25nm, the insulating barrier is CdS or SiO2 or TiO2
Layer, thickness 80nm.As shown in Figure 7, it is shown that the sandwich construction and illumination direction of the plasma resonance sensor, shown in M
For incident light, N show transmitted light.
For the embedded optical resonator in the array of cavities of nanostructured, CdS or SiO2 or TiO2 layers are placed on first
On layer Au, then along with second layer Au.The reason for selecting CdS layer is that its RI is high in visible region, and extinction coefficient (k) is low, and
Au layers provide the electromagnetic field with strong restraint force on nanoscale.The present invention is by this unique integrated nanometer optical resonator
Three-dimensional wait be referred to as multi-layer nano Lycurgus cup array (ML-namoLCA) from resonance sensor because lower floor's nanometer cup structure
The Lycurgus cup made to roman has similar optical property.It is a discovery of the invention that by high-performance Conventional nano
The array of cavities of nanostructured is added in plasma sensor, it can be seen that brand-new optics and spectrum property.The present invention introduces
A kind of RI changes will not cause peak wavelength shift, but RI changes can by high sensitivity using peak strength change come
Novel three-dimensional of detection etc. is from resonance sensor.
The embodiment of the present invention is described in detail above, specific case used herein to the principle of the present invention and
Embodiment is set forth, and the explanation of above example is only intended to help the method and its core concept for understanding the present invention；
Meanwhile for those of ordinary skill in the art, according to the thought of the present invention, can in specific embodiments and applications
There is change part, in summary, this specification content should not be construed as limiting the invention.
Claims (8)
1. a kind of three-dimensional wait of integrated nanometer optical resonator is used for cancer protein mark liquid biopsy from resonance sensor
Using, it is characterised in that comprise the following steps：
(1) three-dimensional is waited and be immersed in from resonance sensor in 2~20mM mercaptoundecylic acid (MUA) solution, cultivate 12 at room temperature
~36 hours, sensor surface formed self-assembled monolayer；
(2) the CEA antibody that concentration is 1~100 μ g/ml is coupled by carbodiimides (EDC) with n-hydroxysuccinimide
(NHS), and with mercaptoundecylic acid (MUA) covalent bond；
(3) will be cultivated after step (1) processing without frequency conversion cubical array plasma resonance sensor in BAS confining liquids, afterwards
Cultivated in ethanolamine solutions；
(4) phosphoric acid containing A samples to be tested is placed in after step (3) is cultivated without frequency conversion cubical array plasma resonance sensor
Cultivated in salt buffer (PBS)；
(5) spectral measurement is carried out, transmission peaks are measured in water.
2. three-dimensional wait of integrated nanometer optical resonator according to claim 1 is used for cancer protein mark from resonance sensor
The application of will thing liquid body biopsy, it is characterised in that the step (2) is to carry out at ambient temperature, the biography after step (1) processing
Sensor is cleaned with 70% ethanol, in N after cleaning2Middle drying, in 400mM carbodiimides (EDC) and 100mM N- after drying
HOSu NHS (NHS) presses 1:Cultivate 30~120 minutes in the mixture of 1 mixing, afterwards delay sensor in phosphate
Rinse in fliud flushing (PBS), immediately with 1~100 μ g/ml monoclonal CEA antibodie culture 1~5 hour after flushing.
3. three-dimensional wait of integrated nanometer optical resonator according to claim 1 is used for cancer protein mark from resonance sensor
The application of will thing liquid body biopsy, it is characterised in that the step (3) is to carry out at ambient temperature, is sealed in 10 μ g/mL BSA
Close in liquid and cultivate, cultivate 30~60 minutes in 10% ethanolamine solutions afterwards to seal up unreacted N- hydroxysuccinimidyls acyl Asia
Amine couples (NHS).
4. three-dimensional wait of integrated nanometer optical resonator according to claim 1 is used for cancer protein mark from resonance sensor
The application of will thing liquid body biopsy, it is characterised in that the three-dimensional of the integrated nanometer optical resonator is waited from resonance sensor to swollen
The detection of tumor markers CEA is limited to 1ng/ml.
5. three-dimensional wait of integrated nanometer optical resonator according to any one of claims 1 to 4 is used for cancer from resonance sensor
The application of disease protein marker liquid biopsy, it is characterised in that also include establishing CEA linear models, concretely comprise the following steps：
(1) three-dimensional is waited and be immersed in from resonance sensor in 10mM mercaptoundecylic acid (MUA) solution, sensor surface is formed certainly
Assemble monofilm；
(2) the CEA antibody that concentration is 30 μ g/ml is coupled by carbodiimides with n-hydroxysuccinimide, and and sulfydryl
Undecanoic acid covalent bond；
(3) will be cultivated after step (1) processing without frequency conversion cubical array plasma resonance sensor in BAS confining liquids, afterwards
Cultivated in ethanolamine solutions；
(4) being respectively placed in without frequency conversion cubical array plasma resonance sensor containing concentration gradient model after step (3) is cultivated
Enclose in the phosphate buffer (PBS) of the CEA for 0.1ng/ml~1000ng/ml, containing mouse immuning ball protein (Anti-
MouseIgG cultivated in phosphate buffer (PBS)) and in the phosphate buffer (PBS) without CEA；
(5) spectral measurement is carried out, transmission peaks are measured in water.
6. three-dimensional wait of integrated nanometer optical resonator according to claim 5 is used for cancer protein mark from resonance sensor
The application of will thing liquid body biopsy, it is characterised in that in the step (4), CEA concentration be respectively 0.1ng/ml, 1ng/ml,
10ng/ml、50ng/ml、100ng/ml、1000ng/ml。
7. it is used for according to Claims 1-4,6 any described three-dimensionals of integrated nanometer optical resonator etc. from resonance sensor
The application of cancer protein mark liquid biopsy, it is characterised in that three-dimensional wait includes plasma resonance from resonance sensor
Substrate, it is the array of cavities of nanostructured, covers multi-layered material structure layer, including the first gold medal arranged successively from bottom to up thereon
Belong to layer, insulating barrier and second metal layer, the multi-layered material structure layer is formed in substrate is embedded in the plasma resonance sensing
Optical resonator array in device, the optical resonator are the cavity of tapered cups.
8. three-dimensional wait of integrated nanometer optical resonator according to claim 7 is used for cancer protein mark from resonance sensor
The application of will thing liquid body biopsy, it is characterised in that the first metal layer thickness is 25nm, thickness of insulating layer 80nm.
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| CN110286104A (en) * | 2019-06-28 | 2019-09-27 | 量准（上海）医疗器械有限公司 | Local surface plasma resonance nano-sensor and its protein quantitation methods |
| WO2020018060A1 (en) * | 2018-07-18 | 2020-01-23 | Институт Физики Полупроводников Им. В.Е.Лашкарёва Национальной Академи Наук Украины | Method of producing a sensor element of an immunosensor based on the phenomenon of surface plasmon resonance for diagnosing leukemia in cattle |
| CN111781370A (en) * | 2020-05-20 | 2020-10-16 | 量准(武汉)生命科技有限公司 | Method for measuring molecular binding capacity by utilizing enzyme-labeling instrument and nano-gold particle-enhanced SPR sensor |
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| CN103792211A (en) * | 2014-01-27 | 2014-05-14 | 暨南大学 | Surface plasma resonance biochip as well as preparation method and application thereof |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2020018060A1 (en) * | 2018-07-18 | 2020-01-23 | Институт Физики Полупроводников Им. В.Е.Лашкарёва Национальной Академи Наук Украины | Method of producing a sensor element of an immunosensor based on the phenomenon of surface plasmon resonance for diagnosing leukemia in cattle |
| CN110286104A (en) * | 2019-06-28 | 2019-09-27 | 量准（上海）医疗器械有限公司 | Local surface plasma resonance nano-sensor and its protein quantitation methods |
| CN111781370A (en) * | 2020-05-20 | 2020-10-16 | 量准(武汉)生命科技有限公司 | Method for measuring molecular binding capacity by utilizing enzyme-labeling instrument and nano-gold particle-enhanced SPR sensor |
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Application publication date: 20180316 |