CN109813773A - A kind of preparation and application of the sialic acid biosensor of photoelectricity and the response of temperature dual signal based on polymerization object point - Google Patents
A kind of preparation and application of the sialic acid biosensor of photoelectricity and the response of temperature dual signal based on polymerization object point Download PDFInfo
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Abstract
The present invention discloses a kind of preparation and application of sialic acid biosensor based on the photoelectricity for polymerizeing object point and the response of temperature dual signal.The sensor utilizes fullerene (C60) and the golden substrate for boring two kinds of nano materials as sensor, mercaptophenyl boronic acid (MPBA) molecule is fixed using the bonding action of Au-S key, its is made to capture a large amount of sialic acid (SA) molecule.Utilize a titanium dioxide (R-TiO for the fan-shaped rutile-type of polymerization object point (TPP-pdot) functionalization of four phenol doping2) label antibody as probe molecule, not only with excellent photocurrent response but also having good temperature-responsive, obtain a kind of dual signal readout mode.Based on the above advantage, it is configured a kind of biosensor of dual signal response, has realized the highly sensitive detection to sialic acid, the sensor is compared to the sensor that traditional mono signal is read, accuracy is higher, and stability is more preferable, largely avoids the false positive or false negative of testing result.
Description
Technical field
The invention belongs to novel nano-material preparation and application and biological immune sensing detection field, specifically devise one kind
The biosensor of photoelectricity and the response of temperature dual signal based on polymerization object point is used for highly sensitive detection sialic acid molecule.
Background technique
Sialic acid molecule (sialic acid, SA) is that one kind is widely present in the various histiocytic acetylations of human body
Derivative, it is related to the various biological functions of organism, participate in the various physiological processes of organism.Studies have shown that human body blood
The raising of SA content and the development of tumour have close relationship in clear, therefore develop a kind of detection side that sensitive practicability is high
Method realizes that the detection to SA in serum has important meaning to the early screening of related disease and detection.Up to the present,
There is serial of methods to may be used to the detection of SA content, such as: fluorescence analysis, mass spectral analysis, high performance liquid chromatography etc..But
These detection methods require expensive detecting instrument, complicated operating process and lengthy and tedious early-stage preparations, therefore largely
On limit its application and develop.Therefore it is high to develop a kind of simple, quick, easy to operate and accuracy, detects and limits low SA's
Detection method has very major and immediate significance to clinical treatment context of detection.
Optical electro-chemistry (PEC) sensor is a kind of analysis method for newly occurring and rapidly developing in recent years, is based on photosensitive
The photoelectric conversion performance of material determines a kind of sensor of test substance content.Its basic testing principle is: in a standing wave
Under long illumination, light-sensitive material generates electric signal via electron excitation and electric charge transfer, and test substance is by physically or chemically
The change of corresponding electric signal caused by effect, to reach the measurement of target analytes.Due to detection signal-electric signal and back
Scape signal-optical signal is kept completely separate, so that the background signal value of photoelectric sensor greatly reduces, therefore compared to traditional electricity
The sensitivity of chemical sensor, photoelectric sensor is higher, and detection limit is lower.In addition to this, photoelectric sensor also has operation letter
Just, it is easy to minimize, continuously can fast and automatically changes the advantages that detection and analysis, is detected in recent years in medical treatment, environmental monitoring and food
Fields, the PEC sensors such as safety all show unique analysis advantage and wide application prospect.
Photothermal sensor is a kind of emerging analysis method that can be used for biomolecule Sensitive Detection, is based on optothermal material
The photothermal conversion characteristic of (such as Prussian blue, carbon material and gold nano-material etc.) determines a kind of sensing of testing molecule content
Device.Under the irradiation of infrared light or near infrared light, optothermal material can convert optical signals into temperature signal, and object amount to be measured changes
Become the variation for causing temperature signal, the quantitative detection to determinand may be implemented accordingly.Detect easy reading and the infrared light of signal
Energy it is small it is small to bio-molecular damage be photothermal sensor two big advantages, therefore, photothermal sensor be it is a kind of portable,
Economic, practical analysis detection tool.
Summary of the invention
The object of the present invention is to provide the biosensors of a kind of photoelectricity based on polymerization object point and the response of temperature dual signal
And its preparation method and application.
To realize goal of the invention, the present invention adopts the following technical scheme:
1. a kind of preparation method and application of the biosensor of photoelectricity and the response of temperature dual signal based on polymerization object point,
It is characterized in that, comprising the following steps:
(1) machinery successively is carried out with 50 nanometers and 30 nanometers of alumina powder foot couple glass-carbon electrode (GCE) on wet chamois leather
Sanding and polishing rinses the alumina powder of electrode surface remnants with deionized water, finally uses nothing until electrode surface is bright clean
Water-ethanol and secondary deionized water are thoroughly cleaned；
(2) fullerene (C that 3 μ L concentration are 3mg/mL is added dropwise60) solution to clean glassy carbon electrode surface is polished, puts to infrared
It takes out after drying in lamp box, cools down at room temperature, C is made60Modify to obtain glass-carbon electrode；
(3) gold nano (Au) solution electrode surface obtained by step (2) that 3 μ L concentration are 3mg/mL is added dropwise, puts to infrared
Drying, is cooled to room temperature in lamp box, and Au/C is made60The glass-carbon electrode of modification；
(4) electrode surface obtained by mercaptophenyl boronic acid (MPBA) solution to step (3) that 3 μ L concentration are 5.0mM, room temperature is added dropwise
It is rinsed after lower reaction 40min with secondary deionized water, removes the physical absorption of electrode surface, dry spare, obtained MBPA/
Au/C60The glass-carbon electrode of modification；
(5) sialic acid (SA) standard solution of 3 μ L various concentrations is added dropwise to step (4) electrode surface obtained, in 4 °C of ice
It is incubated for 40 min in case, is rinsed with secondary deionized water remove extra SA molecule after the reaction was completed, dried, obtain SA/MBPA/
Au/C60The glass-carbon electrode of modification, and be stored in 4 °C of refrigerators；
(6) take 3 μ L concentration be 1.0 wt.% BSA be added drop-wise to modified electrode surface obtained by step (5) and
40 min are incubated in 4 °C of refrigerators, come nonspecific activity site on enclosed-electrode surface, rinse electrode surface with deionized water
Physical absorption is washed away, and is stored in 4 °C of refrigerators；
(7) secondary antibody that the titanium dioxide label of the fan-shaped golden red type of the polymerization object point functionalization of 3 μ L same concentrations is finally added dropwise is molten
Liquid (R-TiO2@TPP-Pdot @Ab2) electrode surface obtained by step (6), after Immune discrimination 40min, gone with secondary from water
It rinses；It dries, R-TiO is made2@TPP-Pdot @Ab2/SA/MBPA/Au/C60The glass-carbon electrode of modification is electrode to be measured.
1. the titanium dioxide (R-TiO2) of above-mentioned fan-shaped rutile-type is prepared by following methods: 1g detergent alkylate
Sodium sulfonate (SDBS) is dissolved in 50 mL, 2.2 mol/L HNO3 solution, after oscillation mixes dozens of minutes on eddy mixer
1 mL isopropyl titanate (TIP) is added, then in 48 °C of condition by persistent oscillation under above-mentioned mixture for 24 hours, spend from
Sub- water and ethyl alcohol repeatedly rinse the mixture after reaction, calcine 1h under 400 °C after centrifugation, 60 °C are dried,
To remove remaining organic components in product, final product is obtained.Products therefrom is dispersed in 1 mL deionized water
(so that it is uniformly dispersed using the methods of ultrasound mixing), gained R-TiO2 solution is placed in 4 °C of refrigerators, is stored for future use.
2. between above-mentioned four phenol adulterate polymerization object point (TPP-pdot) the preparation method is as follows: by polymer
(PFBT), functional polymer is total to four phenol (TPP) between maleic anhydride (PSMA) and sensitizer and is dissolved in tetrahydrofuran (THF) respectively
In, ultrasound makes it be uniformly dispersed.Above-mentioned three kinds of solution is mixed, in mixed liquor, the concentration of PFBT is 200 μ g/mL, PSMA's
The concentration that concentration is 40 μ g/mL, TPP is 10 μ g/mL.Then by the mixed solution ultrasound 20min of acquisition, formed evenly dispersed
Solution.Under the conditions of water bath sonicator, the said mixture of 4 mL is quickly adding into the deionized water of 20 mL.Use nitrogen
THF solvent molecule in purged solution, finally by mixture under the conditions of 55 °C rotary evaporation, come sufficiently remove remnants THF
Molecule obtains the liquid of yellow transparent.
3. two corresponding anti-solution (the R-TiO of the titanium dioxide label of the fan-shaped golden red type of above-mentioned polymerization object point functionalization2@
TPP-Pdot @Ab2) the preparation method is as follows: 1) 0.5 mL concentration is the R-TiO of 5 mg/mL2Solution and 0.5 mL concentration are 5
TPP-pdot obtained by mg/mL step (2) mixes oscillation 4h at room temperature, and centrifuge washing obtains R-TiO for several times2@TPP-
It is dispersed in secondary go from water by Pdot complex solution again；2) taking volume ratio is 1- ethyl-(the 3- dimethylamino of 2:1
Base propyl) the 50 μ L of mixed solution of carbodiimide hydrochloride (EDC) and N- hydroxy thiosuccinimide sodium salt (NHS) is added
Into the above-mentioned complex solution of 200 μ L, 30min is reacted, the carboxyl polymerizeing on object point is activated；3) sialic acid of 200 μ L is anti-
Body is added in step (2) in acquired solution, using the effect of amido bond, reacts the probe point being functionalized after 40min
Son, centrifuge washing are dispersed in again in phosphate buffer solution of the pH equal to 7.4 after for several times, and R-TiO is made2@TPP-Pdot @Ab2
Probe solution, put spare to 4 °C of refrigerator cold-storage.
4. the immunosensor that the above method is prepared for a kind of photoelectricity based on polymerization object point and the response of temperature dual signal.
5. a kind of detecting step of the immunosensor of photoelectricity and the response of temperature dual signal based on polymerization object point is as follows:
(1) it is measured using electrochemical workstation using three-electrode system, a kind of above-described tool based on polymerization object point
There is photoelectricity and the modified electrode of temperature dual signal response is working electrode, Ag/AgCl is reference electrode, and platinum electrode is auxiliary electricity
Pole, the electrochemical method of selection are chronoamperometries, are tested in the PBS buffer solution of 0.1 mol/mL pH 7.4；
(2) in current time method, initial potential 0.5V, sampling interval 0.1s, runing time are 400 s, quiescent time
For 1s, current sensitivity is 1 μ A, original frequency 100000Hz, and termination frequency is 0.1 Hz, to the sialic acid of various concentration
(SA) standard solution is detected, and current information data are acquired by electrochemical workstation.Pass through photocurrent values and saliva
Relationship between sour (SA) concentration of standard solution, draws working curve 1；After Immune discrimination occurs with the laser irradiation of 808nm
Working electrode, while recording temperature change in real time with infrared radiation thermometer, marked according to the temperature of electrode surface and sialic acid (SA)
Standard working curve 2 is established between quasi- solution concentration；
(3) testing sample solution replaces sialic acid (SA) standard solution to be detected, and the result of detection can be by the standard drawn out
Working curve 1 and 2 checks in.
Remarkable advantage of the invention are as follows:
(1) sensor with dual signal response modes provides a kind of reliable, convenient, quick detection method, to realize pair
The highly sensitive detection of SA content in serum.The corresponding two kinds of detection signals of the test substance of same concentration, facilitate realization to tie detection
The self calibration of fruit largely avoids the testing result of false positive or false negative, realizes the accurate analysis to determinand.
(2) the polymerization object point (TPP-pdot) of this experiment synthesis is both to the titanium dioxide (R-TiO of fan-shaped rutile-type2) light
Electroresponse has good sensitization, it is made to obtain biggish photo-signal, provides possibility to SA super sensitivity detection to realize.
In addition TPP-pdot has excellent light thermal property again, and testing result thermometer can realize quantitative analysis, to establish
A kind of portable sensor offer is possible.In addition to this, a large amount of carboxyl is contained on the polymerization object point molecule of synthesis, can be marked
A large amount of biomolecule has further widened the detection range of the sensor.
(3) titanium dioxide of the rutile-type of this experiment synthesis is a kind of good photoelectric material, in the excitation of ultraviolet light
Under, it can produce stable photo-signal, ensure that the higher stability of the biosensor and preferable reproducibility.
Detailed description of the invention
Fig. 1 is the transmission electron microscope picture (TEM) of the TPP-pdot of synthesis；
Fig. 2 is the TPP-pdot UV absorption (UV) and fluorescent emission figure (FL) of synthesis；
Fig. 3 is dual signal immunosensor to the relationship between photocurrent values and sialic acid (SA) concentration of standard solution of standard
Figure；
Fig. 4 is the standard between the temperature and sialic acid (SA) concentration of standard solution of the electrode surface of dual signal immunosensor
Working curve diagram.
Specific embodiment
The present invention is further illustrated by the following examples, but protection scope of the present invention is not limited to following reality
Apply example.
Embodiment 1
1. a kind of preparation method and application of the biosensor of photoelectricity and the response of temperature dual signal based on polymerization object point, special
Sign is, comprising the following steps:
(1) machinery successively is carried out with 50 nanometers and 30 nanometers of alumina powder foot couple glass-carbon electrode (GCE) on wet chamois leather
Sanding and polishing rinses the alumina powder of electrode surface remnants with deionized water, finally uses nothing until electrode surface is bright clean
Water-ethanol and secondary deionized water are thoroughly cleaned.
(2) fullerene (C that 3 μ L concentration are 3mg/mL is added dropwise60) solution (solvent is dimethylformamide) is clean in polishing
Glassy carbon electrode surface, put to after drying in infrared lamp box and take out, cool down at room temperature, C is made60Modify to obtain glass-carbon electrode；
(3) gold nano (Au) solution electrode surface obtained by step (2) that 3 μ L concentration are 3mg/mL is added dropwise, puts to infrared
Drying, is cooled to room temperature in lamp box, and Au/C is made60The glass-carbon electrode of modification；
(4) electrode surface obtained by mercaptophenyl boronic acid (MPBA) solution to step (3) that 3 μ L concentration are 5.0mM, room temperature is added dropwise
It is rinsed after lower reaction 40min with secondary deionized water, removes the physical absorption of electrode surface, dry spare, obtained MBPA/
Au/C60The glass-carbon electrode of modification；
(5) sialic acid (SA) standard solution of 3 μ L various concentrations is added dropwise to step (4) electrode surface obtained, in 4 °C of refrigerators
40 min of middle incubation are rinsed with secondary deionized water remove extra SA molecule after the reaction was completed, dried, obtain SA/MBPA/
Au/C60The glass-carbon electrode of modification, and be stored in 4 °C of refrigerators；
(6) take 3 μ L concentration be 1.0 wt.% BSA be added drop-wise to modified electrode surface obtained by step (5) and
40 min are incubated in 4 °C of refrigerators, come nonspecific activity site on enclosed-electrode surface, rinse electrode surface with deionized water
Physical absorption is washed away, and is stored in 4 °C of refrigerators；
(7) the titanium dioxide label of the fan-shaped golden red type of the polymerization object point functionalization of 3 μ L same concentrations (5mg/mL) is finally added dropwise
Two corresponding anti-solution (R-TiO2@TPP-Pdot @Ab2) electrode surface obtained by step (6), after Immune discrimination 40min, with two
It is secondary to go to rinse from water；It dries, R-TiO is made2@TPP-Pdot @Ab2/SA/MBPA/Au/C60The glass-carbon electrode of modification is to be measured
Electrode.
The preparation of above-mentioned gold cone nano material: before spin coating polystyrene bead, teflon film is cut into pros first
Shape (1.5x1.5cm) is simultaneously rinsed with ethyl alcohol and secondary water, is then cleaned 3 minutes with plasma water.Absorption 1mL concentration is 2.5 w/
The polystyrene bead acetone soln of v % is simultaneously centrifuged, and the volume ratio for being then transferred to ethyl alcohol and methanol is the mixed solution of 2:1
In.The surfactant (TX100) that volume fraction is 0.2 % is added in the solution, then adjusts the concentration of polystyrene bead
To about 5 w/v %.Then polystyrene bead is spin-coated on clean teflon film, and places a few minutes at room temperature, made molten
Agent is dry.Use O2Plasma etching polystyrene bead/the surface teflon certain time.Finally by thermal evaporation on the surface
The gold for coating 50nm, obtains gold nano-material.
The preparation of above-mentioned gold nano solution: taking gold nano-material dissolution made from appropriate above-mentioned experiment in deionized water,
Obtain gold nano (Au) solution of 3mg/mL.
Embodiment 2
Titanium dioxide (the R-TiO of fan-shaped rutile-type used in above-described embodiment 12) prepared by following methods: 1g dodecane
Base benzene sulfonic acid sodium salt (SDBS) is dissolved in 50 mL, 2.2 mol/L HNO3Solution, oscillation mixes tens of points on eddy mixer
1 mL isopropyl titanate (TIP) is added after clock, then in 48 °C of condition by persistent oscillation under above-mentioned mixture for 24 hours, use
Deionized water and ethyl alcohol repeatedly rinse the mixture after reaction, forge under 400 °C after centrifugation, 60 °C are dried
It burns 1h and obtains final product to remove remaining organic components in product.Products therefrom is dispersed in 1 mL deionization
(it is set to be uniformly dispersed using the methods of ultrasound mixing) in water, by gained R-TiO2Solution is placed in 4 °C of refrigerators, is stored standby
With.
Embodiment 3
Between used in above-described embodiment 1 four phenol adulterate polymerization object point (TPP-pdot) the preparation method is as follows: by polymer
(PFBT), functional polymer is total to four phenol (TPP) between maleic anhydride (PSMA) and sensitizer and is dissolved in tetrahydrofuran (THF) respectively
In, ultrasound makes it be uniformly dispersed.Above-mentioned three kinds of solution is mixed, in mixed liquor, the concentration of PFBT is 200 μ g/mL, PSMA's
The concentration that concentration is 40 μ g/mL, TPP is 10 μ g/mL.Then by the mixed solution ultrasound 20min of acquisition, formed evenly dispersed
Solution.Under the conditions of water bath sonicator, the said mixture of 4 mL is quickly adding into the deionized water of 20 mL.Use nitrogen
THF solvent molecule in purged solution, finally by mixture under the conditions of 55 °C rotary evaporation, come sufficiently remove remnants THF
Molecule obtains the liquid of yellow transparent.
[(9,9- dioctyl fluorene -2,7- diyl)-alternating-(2,1,3- benzothiazole)] poly- used in above-described embodiment 3
(PFBT), four phenol (TPP) are purchased from Xi'an Bao Laite Optoelectronics Technology between poly- (styrene-is total to maleic anhydride) (PSMA), sensitizer
Co., Ltd.
Embodiment 4
It polymerize the two corresponding anti-solution R- of the fan-shaped rutile titanium dioxide label of object point functionalization used in above-described embodiment 1
TiO2@TPP-Pdot@Ab2The preparation method is as follows: 1) 0.5 mL concentration be 5 mg/mL embodiment 2 made from R-TiO2Solution
Oscillation 4h is mixed at room temperature with TPP-pdot made from embodiment 3 of the 0.5 mL concentration for 5 mg/mL, and centrifuge washing obtains for several times
To R-TiO2It, is dispersed in secondary go from water by@TPP-Pdot complex solution again；2) taking volume ratio is the 1- second of 2:1
Base-(3- dimethylaminopropyl) carbodiimide hydrochloride (EDC) and N- hydroxy thiosuccinimide sodium salt (NHS) it is mixed
It closes 50 μ L of solution to be added in the above-mentioned complex solution of 200 μ L, reacts 30min, the carboxyl polymerizeing on object point is activated；3)
The sialic acid antibody of 200 μ L is added in step 2 in acquired solution, using the effect between amide, react obtain after 40min by
The probe molecule of functionalization, centrifuge washing are dispersed in again in phosphate buffer solution of the pH equal to 7.4 after for several times, and R- is made
TiO2@TPP-Pdot@Ab2Probe solution, put spare to 4 °C of refrigerator cold-storage.
Embodiment 5
The detecting step of sialic acid (SA):
(1) it is measured using electrochemical workstation using three-electrode system, polymerization object point is based on one kind prepared by embodiment 1
The modified electrode responded with photoelectricity and temperature dual signal be working electrode, Ag/AgCl is reference electrode, supplemented by platinum electrode
Electrode is helped, the electrochemical method of selection is chronoamperometry, is surveyed in the PBS buffer solution of 0.1 mol/mL pH 7.4
Examination；
(2) in current time method, initial potential 0.5V, sampling interval 0.1s, runing time are 400 s, quiescent time
For 1s, current sensitivity is 1 μ A, original frequency 100000Hz, and termination frequency is 0.1 Hz, to the sialic acid of various concentration
(SA) standard solution is detected, and current information data are acquired by electrochemical workstation.Pass through photocurrent values and saliva
Relationship between sour (SA) concentration of standard solution draws working curve 1(and sees Fig. 3)；Occurred with the laser irradiation of 808nm immune
Working electrode after identification, while temperature change is recorded in real time with infrared radiation thermometer, according to the temperature and sialic acid of electrode surface
(SA) standard working curve 2(is established between concentration of standard solution and see Fig. 4)；
Testing sample solution replaces sialic acid (SA) standard solution to be detected, and the result of detection can be by the standard work drawn out
Make curve 1 and 2 to check in.
Claims (4)
1. a kind of preparation method of the sialic acid biosensor of photoelectricity and the response of temperature dual signal based on polymerization object point, special
Sign is, comprising the following steps:
(1) machinery successively is carried out with 50 nanometers and 30 nanometers of alumina powder foot couple glass-carbon electrode (GCE) on wet chamois leather
Sanding and polishing rinses the alumina powder of electrode surface remnants with deionized water, finally uses nothing until electrode surface is bright clean
Water-ethanol and secondary deionized water are thoroughly cleaned；
(2) fullerene (C that 3 μ L concentration are 3mg/mL is added dropwise60) solution to clean glassy carbon electrode surface is polished, puts to infrared
It takes out after drying in lamp box, cools down at room temperature, C is made60Modify to obtain glass-carbon electrode；
(3) gold nano (Au) solution electrode surface obtained by step (2) that 3 μ L concentration are 3mg/mL is added dropwise, puts to infrared
Drying, is cooled to room temperature in lamp box, and Au/C is made60The glass-carbon electrode of modification；
(4) electrode surface obtained by mercaptophenyl boronic acid (MPBA) solution to step (3) that 3 μ L concentration are 5.0mM, room temperature is added dropwise
It is rinsed after lower reaction 40min with secondary deionized water, removes the physical absorption of electrode surface, dry spare, obtained MBPA/
Au/C60The glass-carbon electrode of modification；
(5) sialic acid (SA) standard solution of 3 μ L various concentrations is added dropwise to step (4) electrode surface obtained, in 4 °C of ice
It is incubated for 40 min in case, is rinsed with secondary deionized water remove extra SA molecule after the reaction was completed, dried, obtain SA/MBPA/
Au/C60The glass-carbon electrode of modification, and be stored in 4 °C of refrigerators；
(6) take 3 μ L concentration be 1.0 wt.% BSA be added drop-wise to modified electrode surface obtained by step (5) and
40 min are incubated in 4 °C of refrigerators, come nonspecific activity site on enclosed-electrode surface, rinse electrode surface with deionized water
Physical absorption is washed away, and is stored in 4 °C of refrigerators；
(7) secondary antibody of the titanium dioxide label of the fan-shaped golden red type of the polymerization object point functionalization of 3 μ L concentration 5mg/mL is finally added dropwise
Solution (R-TiO2@TPP-Pdot @Ab2) electrode surface obtained by step (6), after Immune discrimination 40min, with it is secondary go from
Water rinses；It dries, R-TiO is made2@TPP-Pdot @Ab2/SA/MBPA/Au/C60The glass-carbon electrode of modification is electrode to be measured.
2. the method according to claim 1, wherein
(1) titanium dioxide (R-TiO of fan-shaped rutile-type2) prepared by following methods: 1g neopelex
(SDBS) it is dissolved in 50 mL, 2.2 mol/L HNO3Solution, 1 mL is added after mixing dozens of minutes in oscillation on eddy mixer
Isopropyl titanate (TIP), then in 48 °C of condition by persistent oscillation under above-mentioned mixture for 24 hours, with deionized water and second
Alcohol repeatedly rinses the mixture after reaction, calcines 1h under 400 °C after centrifugation, 60 °C are dried, is produced with removing
Remaining organic components, obtain final product in object；Products therefrom is dispersed in 1 mL deionized water (by gained R-
TiO2Solution is placed in 4 °C of refrigerators, is stored for future use；
(2) the polymerization object point (TPP-pdot) that four phenol adulterate between is prepared by following methods: by polymer (PFBT), function is poly-
Conjunction object is total to four phenol (TPP) between maleic anhydride (PSMA) and sensitizer and is dissolved in tetrahydrofuran (THF) respectively, and ultrasound makes its point
It dissipates uniformly, mixes above-mentioned three kinds of solution, in mixed liquor, the concentration that the concentration of PFBT is 200 μ g/mL, PSMA is 40 μ g/
The concentration of mL, TPP are 10 μ g/mL；Then by the mixed solution ultrasound 20min of acquisition, evenly dispersed solution is formed；In water
It bathes under ultrasound condition, the said mixture of 4 mL is quickly adding into the deionized water of 20 mL, in nitrogen purged solution
THF solvent molecule, finally by mixture, rotary evaporation, the next THF molecule for sufficiently removing remnants obtain yellow under the conditions of 55 °C
Transparent liquid；
(3) it polymerize two corresponding anti-solution (the R-TiO of the fan-shaped rutile titanium dioxide label of object point functionalization2@TPP-Pdot @
Ab2) it is prepared by the following method: 1) R-TiO made from the step of 0.5 mL concentration is 5 mg/mL (1)2Solution and 0.5 mL concentration
TPP-pdot obtained by the step of for 5 mg/mL (2) mixes oscillation 4h at room temperature, and centrifuge washing obtains R-TiO for several times2@
It is dispersed in secondary go from water by TPP-Pdot complex solution again；2) taking volume ratio is 1- ethyl-(the 3- diformazan of 2:1
Base aminopropyl) carbodiimide hydrochloride (EDC) and N- hydroxy thiosuccinimide sodium salt (NHS) 50 μ L of mixed solution
It is added in the above-mentioned complex solution of 200 μ L, reacts 30min, the carboxyl polymerizeing on object point is activated；3) saliva of 200 μ L
Sour antibody is added in step 2 in acquired solution, using the effect of amido bond, reacts the probe being functionalized after 40min
Molecule, centrifuge washing are dispersed in again in phosphate buffer solution of the pH equal to 7.4 after for several times, and R-TiO is made2@TPP-Pdot @
Ab2Probe solution, put spare to 4 °C of refrigerator cold-storage.
3. a kind of photoelectricity and the response of temperature dual signal based on polymerization object point of any method preparation of claim 1-2
Biosensor.
4. the immunosensor of the photoelectricity and the response of temperature dual signal as claimed in claim 3 based on polymerization object point, feature exist
In for sialic acid (SA) detection, detecting step is as follows:
(1) it is measured using electrochemical workstation using three-electrode system, polymer is based on one kind described in claim 5
The modified electrode of point responded with photoelectricity and temperature dual signal is working electrode, and Ag/AgCl is reference electrode, and platinum electrode is
Auxiliary electrode, the electrochemical method of selection are chronoamperometries, are carried out in the PBS buffer solution of 0.1 mol/mL pH 7.4
Test；
(2) in current time method, initial potential 0.5V, sampling interval 0.1s, runing time are 400 s, quiescent time
For 1s, current sensitivity is 1 μ A, original frequency 100000Hz, and termination frequency is 0.1 Hz, to the sialic acid of various concentration
(SA) standard solution is detected, and current information data are acquired by electrochemical workstation；Pass through photocurrent values and saliva
Relationship between sour (SA) concentration of standard solution, draws working curve 1；After Immune discrimination occurs with the laser irradiation of 808nm
Working electrode, while recording temperature change in real time with infrared radiation thermometer, marked according to the temperature of electrode surface and sialic acid (SA)
Standard working curve 2 is established between quasi- solution concentration；
(3) testing sample solution replaces sialic acid (SA) standard solution to be detected, and the result of detection can be by the mark drawn out
Quasi- working curve 1 is obtained with looking into 2.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110133075A (en) * | 2019-05-29 | 2019-08-16 | 福建师范大学 | A kind of sialic acid immunosensor based on the induction signals in situ amplification of silver iodide nanoparticle photo-thermal effect |
| CN110220957A (en) * | 2019-07-06 | 2019-09-10 | 福建师范大学 | Based on NiFe2O4Double mode electrogenerated chemiluminescence-temperature immunosensor of the different luminol global function probe of nanotube |
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2019
- 2019-03-04 CN CN201910161493.XA patent/CN109813773A/en active Pending
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110133075A (en) * | 2019-05-29 | 2019-08-16 | 福建师范大学 | A kind of sialic acid immunosensor based on the induction signals in situ amplification of silver iodide nanoparticle photo-thermal effect |
| CN110133075B (en) * | 2019-05-29 | 2021-07-16 | 福建师范大学 | Sialic acid immunosensor based on silver iodide nanoparticle photothermal effect induction in-situ signal amplification |
| CN110220957A (en) * | 2019-07-06 | 2019-09-10 | 福建师范大学 | Based on NiFe2O4Double mode electrogenerated chemiluminescence-temperature immunosensor of the different luminol global function probe of nanotube |
| CN110220957B (en) * | 2019-07-06 | 2021-11-09 | 福建师范大学 | Based on NiFe2O4Dual-mode electrochemiluminescence-temperature immunosensor of nanotube isoluminol full-function probe |
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