CN109239135A - A kind of portable biometric immunoassay method based on the building of flexible air pressure sensing - Google Patents
A kind of portable biometric immunoassay method based on the building of flexible air pressure sensing Download PDFInfo
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- CN109239135A CN109239135A CN201811241023.6A CN201811241023A CN109239135A CN 109239135 A CN109239135 A CN 109239135A CN 201811241023 A CN201811241023 A CN 201811241023A CN 109239135 A CN109239135 A CN 109239135A
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- air pressure
- building
- method based
- pressure sensor
- pressure sensing
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
- G01N27/02—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating impedance
- G01N27/04—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating impedance by investigating resistance
- G01N27/06—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating impedance by investigating resistance of a liquid
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
- G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
- G01N27/28—Electrolytic cell components
- G01N27/30—Electrodes, e.g. test electrodes; Half-cells
- G01N27/327—Biochemical electrodes, e.g. electrical or mechanical details for in vitro measurements
- G01N27/3275—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
- G01N33/57473—Immunoassay; Biospecific binding assay; Materials therefor for cancer involving carcinoembryonic antigen, i.e. CEA
Abstract
The present invention discloses a kind of portable biometric immunoassay method based on the building of flexible air pressure sensing, and the pliable pressure sensor of preparation is sensitive to air pressure change response, and is used for monitoring biological immune reaction, realizes the sensitive portable detection to Multiple Antibodies.The present invention is prepared for pliable pressure sensor as flexible substrates using carbon nano tube modified paper base electrode first, in conjunction with immune response, realizes the specific recognition to object.After injecting hydrogen peroxide, nano-probe introduce Pt nanoparticle can efficient catalytic hydrogen peroxide decompose generate oxygen, increase the air pressure in closed detection system.Raised air pressure causes pliable pressure sensor, and deformation occurs, to make the reduction of its resistance.This variation can be monitored by digital multimeter by detecting its resistance variations, to realize the quantitative detection to object.This method equipment is simple, easy to carry, easy to operate, provides a kind of simple, sensitive and stable method for portable inspectiont.
Description
Technical field
The present invention relates to a kind of portable biometric immunoassay methods based on the building of flexible air pressure sensing, belong to sensing
Instrument building and biochemical analysis technical field.
Background technique
With the development of society, environmental monitoring, food safety and medical diagnosis on disease are increasingly valued by people, build
Many detection methods based on optics, chromatography, mass spectrum and electrochemistry are found, these detection methods have accurate, sensitive, stable
The advantages that.But equipment is expensive and the reasons such as operation is multiple limit its in terms of quick detection and daily life into one
Step application, especially in remote districts.Therefore, design a kind of equipment it is simple, it is easily operated, stablize practical detection method be tool
Significant.
Air pressure has application as a kind of traditional physical phenomenon, at many aspects of life.In chemical reaction process,
There are many chemical reactions can generate bulk gas to increase the air pressure in container.And in these reactions, hydrogen peroxide point
Solution, which generates this reaction of oxygen, efficient gas production rate and the properties such as nontoxic.Meanwhile the rate energy of the reaction
It is enough greatly improved by catalyst, such as catalase, Pt nanoparticle, is that one kind is very suitable for being used as detection signal
Reaction.In many existing portable bio-analysis using air pressure as detection signal, traditional barometer and capillary column
It is usually used to as signal read-out device.In recent years, flexible device be since it can have sensitive response to external stress,
There are many researchs for electronic skin, electronic textile and stress mornitoring etc..In order to improve the sensitivity of portable sensor,
Signal the generating resource and signal supervisory instrument are most important one two o'clocks.Based on the above content, a kind of sensitive flexibility is constructed
Baroceptor and the portable immunity biosensor for being applied to vapour-pressure type must can effectively improve the sensitivity of detection
And accuracy.
Summary of the invention
The purpose of the present invention is to provide a kind of preparation methods of pliable pressure sensor, and combine antigen-antibody immune anti-
Portable pneumatic immunosensor should be constructed.Its technical principle is catalyzed by introducing Pt nanoparticle on immunological probe
The hydrogen peroxide of injection generates oxygen, and the oxygen of generation can change the air pressure in detector, and the air pressure of generation can be flexible
Pressure sensor Sensitive Detection.When not having object, air pressure increase is less, and flexible baroceptor deformation is small, resistance variations
It is smaller；In the presence of object, air pressure increase is larger, and flexible baroceptor deformation is big, and resistance variations are larger.The variation of resistance
It is positively correlated in a certain range with target concentration, and can realize portable quantitative detection by digital multimeter.
To achieve the above object, The technical solution adopted by the invention is as follows:
A kind of portable biometric immunoassay method based on the building of flexible air pressure sensing comprising the steps of:
(1) modification of paper base electrode: qualitative filter paper is cut into certain shapes, and the carbon nanotube for being immersed in 1-5wt% is molten
It in liquid (the Pluronic F-127 surfactant containing 1-10wt%), then dries, is repeated several times, until resistance reaches 20-
70 kΩ；
(2) preparation of pliable pressure sensor: paper base electrode obtained is connected with copper conductive adhesive with silver conductive adhesive, and wherein one
It is puted up with BOPP adhesive tape in face；It combines two paper base electrodes faces prepared, and in one strata dimethyl silica of outer layer
Alkane PDMS closing；
(3) pressure sensor prepared the portable immunoassay apparatus building of air-pressure type: is enclosed in a centrifugation with PDMS
Guan Zhong is connected with a conduit with detection cell at its end, in the catheter between be equipped with a filter device, prevent liquid from flowing into, institute
There is interface to be closed with PDMS.
(4) nano-probe constructs: first by 10 mL platinum acid chloride solutions (0.1-5 mM) and 1 mL ascorbic acid solution
(0.1-5 M) mixing, in 80 DEG C of 30 min of heating, be stored in 4 DEG C it is spare.Nano platinum particle solution carbon prepared by 5 mL
Sour sodium adjusts pH to 9.0, adds the object antibody (10-100 μ L) that concentration is 0.5 mg/mL, incubates at 4 DEG C overnight
Change.Finally above-mentioned solution is centrifuged 20 minutes under conditions of 4 DEG C and 14000 g centrifugal force, the precipitating that centrifugation is obtained is again
It is dispersed in the PBS (pH=7.4) that 3 mL contain 1.0 wt % BSA, 0.5 wt % Tween 20 and 0.1 wt % sodium azide
In, be stored in 4 DEG C it is spare.
(5) immune response building: being added capture antibody (10-80 μ L) in the high affine plate in 96 holes first and close overnight, then
It is immune to form antibody-antigen-antibody interlayer type to sequentially add target detection thing (10-80 μ L), nano-probe (10-80 μ L)
Compound hatches 40 min, and after being washed with phosphate buffer solution, 100 μ L hydroperoxidations are added；
(6) it exports electrical signal detection: reading resistance signal variation with digital multimeter come quantitative objective object concentration.
More specifically,
The shape specification that filter paper as described in step (1) is cut into is as shown in Figure 2.
Pliable pressure sensor specification described in step (2) is 1 × 1.5 cm2, specific preparation process such as Fig. 3 institute
Show, the pliable pressure sensor pictorial diagram of preparation is as shown in the A in Fig. 4, to air pressure response range as shown in figure 5, being 0.08-50
kPa。
The autocratic outside of detection described in step (3) is closed with PDMS increases air-tightness, and connects with digital multimeter, has
Shape is as shown in B in Fig. 4.
Concentration of hydrogen peroxide described in step (4) is 30wt%, and the reaction time is 270 s.
Resistance signal described in step (5) is that Δ R(initial resistance subtracts terminating resistor).
Advantages of the present invention is as follows:
(1) material feedstock required for the present invention is easy to get, and preparation process is simple, low in cost.
(2) the pliable pressure sensor that the present invention constructs responds air pressure sensitive, and the response time is short, and the range of linearity is wide, will
It is used to be immunoreacted detection, greatly simplifies detection device, easy to operate, responds sensitive, makes that it is suitable for portable inspectionts.
(3) present invention can be increased by a small amount of gas using air pressure as detection signal and cause huge change,
Be conducive to improve the sensitivity of detection.
(4) present invention is used as signal amplification mechanism using immune touching method and nano platinum particle catalysis, can greatly improve
Testing result accuracy and sensitivity.
Detailed description of the invention
Fig. 1 is a kind of portable biometric immunoassay method schematic diagram based on the building of flexible air pressure sensing；
Fig. 2 is the specification that filter paper is cut；
Fig. 3 is pliable pressure sensor assembling process；
Fig. 4 is pliable pressure sensor pictorial diagram and detection device pictorial diagram;A is pliable pressure sensor, and B is detection dress
It sets；
Fig. 5 is response range of the pliable pressure sensor to air pressure.
Fig. 6 is by the standard working curve that does in embodiment 1 to carcinomebryonic antigen.
Specific embodiment
Below by specific implementation example, technical scheme is described further, but this cannot be limited with this
The range of invention.
Embodiment 1
1. the modification of paper base electrode
Qualitative filter paper is cut into specification shown in Fig. 2, and is immersed in the carbon nano-tube solution of 2wt% (containing 2wt%'s
Pluronic F-127 surfactant), it then dries, is repeated several times, until resistance reaches 60 k Ω.
2. the preparation of pliable pressure sensor
Paper base electrode obtained is connected with copper conductive adhesive with silver conductive adhesive, one side is puted up with BOPP adhesive tape wherein；By two
The paper base electrodes face combination prepared, and closed in one layer of PDMS of outer layer, specific assembling process is as shown in figure 3, assembling
After the completion as shown in A in Fig. 4.
3. the portable immunoassay apparatus building of air-pressure type
The pressure sensor prepared is enclosed in a centrifuge tube with PDMS, is connected in piece conduit in its end and detection cell
Connect, in the catheter between be equipped with a filter device, prevent liquid from flowing into.It is closed at whole device structure with PDMS, guarantees air-tightness,
Pictorial diagram is as shown in B in Fig. 4.
4. the preparation of nano-probe
10 mL platinum acid chloride solutions (1 mM) and 1 mL ascorbic acid solution (0.4 M) are mixed first, heat 30 at 80 DEG C
Min, be stored in 4 DEG C it is spare.The nano platinum particle solution sodium carbonate of 5 mL preparation is adjusted into pH to 9.0, adds 50 μ L
Concentration is the carcinomebryonic antigen antibody of 0.5 mg/mL, is being incubated overnight at 4 DEG C.Finally by above-mentioned solution in 4 DEG C and 14000 g
It is centrifuged 20 minutes under conditions of centrifugal force, the precipitating that centrifugation obtains is dispersed in 3 mL again and contains 1.0 wt % BSA, 0.5
In the PBS (pH=7.4) of wt % Tween 20 and 0.1 wt % sodium azide, be stored in 4 DEG C it is spare.
5. the detection of pair object Carcinoembryonic Antigen CEA (the present embodiment carcinomebryonic antigen is as model target analyte)
Fig. 1 is analysis method process schematic in the present invention.Firstly, it is 10 μ that 50 μ L concentration, which are added, in the high affine plate in 96 holes
The CEA antibody of g/mL, hatching overnight at 4 DEG C add the BSA that 300 μ L concentration are 1.0 wt % and close 1 hour.It is molten with PBS
After liquid washing, the CEA standard sample of 50 μ L and the nano-probe of 50 μ L are sequentially added, reacts 40 min respectively.Remove supernatant
The H that 100 μ L concentration are 30wt% is added after liquid2O2, 270 s are reacted, read digital multimeter resistance variations to quantify.Its result
To the standard working curve of CEA as shown in fig. 6, the range of linearity is 0.5-60 ng/mL, detection is limited to resulting resistance variations
0.167 ng/mL。
The foregoing is merely presently preferred embodiments of the present invention, all equivalent changes done according to scope of the present invention patent with
Modification, is all covered by the present invention.
Claims (7)
1. a kind of portable biometric immunoassay method based on the building of flexible air pressure sensing, it is characterised in that: the method packet
Include following steps:
(1) carbon nano tube modified paper base electrode preparation；
(2) preparation of pliable pressure sensor and the building of immunoassay apparatus；
(3) preparation of the nano-probe of the synthesis and its label of Pt nanoparticle；
(4) antibody-antigene-nano-probe immuno-sandwich structure is formed in high affine 96 hole elisa Plates；
(5) hydrogen peroxide of addition is catalyzed by the Pt nanoparticle of label to generate oxygen, made in closed detection system
Pressure rise so that the electric conductivity of pliable pressure sensor changes, and uses digital multimeter as read-out device
To realize the purpose of portable immunoassay.
2. a kind of portable biometric immunoassay method based on the building of flexible air pressure sensing according to claim 1: step
Suddenly carbon nanotube diameter described in (1) is 10-20 nm, and length is 1-2 μm；Paper base electrode uses the side of direct immersion-drying
Method is modified, and the carbon nano-tube solution of the Pluronic F-127 surfactant containing 1-10wt% of 1-5wt% is immersed in
In, it then dries, until resistance reaches 20-70 k Ω.
3. a kind of portable biometric immunoassay method based on the building of flexible air pressure sensing according to claim 1,
Be characterized in that: pliable pressure sensor described in step (2) is by the paper base electrode, copper conductive adhesive and poly dimethyl silicon that prepare
Oxygen alkane PDMS assembles；Immunoassay apparatus detection cell, gas-guide tube, pliable pressure sensor and digital multimeter assembling and
At.
4. a kind of portable biometric immunoassay method based on the building of flexible air pressure sensing according to claim 1,
Be characterized in that: for Pt nanoparticle from ascorbic acid reduction, partial size is 35 nm in step (3)；Nano-probe is relied on by antibody
Electrostatic Absorption is made on Pt nanoparticle.
5. a kind of portable biometric immunoassay method based on the building of flexible air pressure sensing according to claim 1,
Be characterized in that: ELISA Plate described in step (4) is the detection cell in detection device；Detection identification process is anti-by antibody-is formed
Original-nano-probe interlayer type immune complex is realized.
6. a kind of portable biometric immunoassay method based on the building of flexible air pressure sensing according to claim 1,
It is characterized in that: step (4) process specifically: capture antibody 10-80 μ L is added in the high affine plate in 96 holes and closes overnight, then according to
Secondary addition target detection thing 10-80 μ L, nano-probe 10-80 μ L form antibody-antigen-antibody interlayer type immune complex
Hatch 40 min.
7. a kind of portable biometric immunoassay method based on the building of flexible air pressure sensing according to claim 1,
Be characterized in that: concentration of hydrogen peroxide described in step (5) is 30wt%, and the catalysis time reaction time is 270 s；Measured is soft
Property pressure sensor resistance variations numerical value with target concentration increase and reduce.
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Cited By (1)
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| CN109975365A (en) * | 2019-03-12 | 2019-07-05 | 复旦大学 | A kind of wearable liquid sensor and preparation method thereof |
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