CN106093015A - A kind of closed type bipolar electrode electrogenerated chemiluminescence cloth chip and preparation method thereof and purposes - Google Patents
A kind of closed type bipolar electrode electrogenerated chemiluminescence cloth chip and preparation method thereof and purposes Download PDFInfo
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Abstract
The invention discloses a kind of closed type bipolar electrode electrogenerated chemiluminescence cloth chip and preparation method thereof and purposes, this chip substrate is pieces of cloth, distribution electrode on pieces of cloth, and bipolar electrode is placed in the middle, a pair drive electrode distribution both sides；Positive drive electrode with bipolar electrode negative electrode by supporting that microchannel connects, feminine gender drive electrode with bipolar electrode anode by reporting that microchannel connects；The surrounding of microchannel is hydrophobic wax dam；Microchannel is hydrophilic.The chip of the present invention has fabulous flexibility, and the most potential structure surpasses flexible sensor；Additionally compared to traditional die backing material, cloth chip has the advantages such as cheap, universal, user friendly, bio-compatibility is good, biodegradable.
Description
Technical field
The invention belongs to micro-fluidic chip field, be specifically related to a kind of closed type bipolar electrode electrogenerated chemiluminescence cloth chip
And preparation method, and the purposes in detection tripropyl amine (TPA), hydrogen peroxide, glucose.
Background technology
Micro-fluidic chip, also known as micro-total analysis system, refers to utilize micro-processing technology, is formed micro-on certain backing material
The analysis system of channel design, can be used for the detection application in the fields such as biology, medical science, chemistry.
2011, cotton was incorporated into micro-fluidic chip processing neck as chip substrate material by Dendukuri group first
Territory, utilizes loom to be intertwined by line, prepares fabricbase micro-fluidic colorimetric immunoassay chip.Relative to glass, silicon chip, stone
The inorganic material such as English and PDMS (polydimethylsiloxane), PMMA (polymethyl methacrylate), PC (Merlon) are contour
The chip of molecule aggregation materials processing, fabricbase micro-fluidic chip based on organic fiber (i.e. cloth chip) has cheap easy
, it is easy to process, the advantage such as simple, convenient, be therefore developing progressively as a kind of novel analysis test platform, ground
The persons of studying carefully pay close attention to the most widely.Up to the present, cloth chip with a series of detection methods (such as colorimetric, electrochemistry, chemistry
Luminous, electrogenerated chemiluminescence (electrochemiluminescence, ECL)) couple combination to realize different testing goal.
At present, ECL can simply be divided into three electrode ECL and bipolar electrode ECL (bipolar electrode-ECL, BPE-
ECL).Being different from traditional three electrode ECL, BPE-ECL carrys out wireless type only with a pair drive electrode and excites one or several pairs
Property electrode, bipolar electrode two ends adhere to yin, yang the two poles of the earth separately, can react with direct-coupling electrochemistry and ECL.
Calendar year 2001, BPE-ECL concept is incorporated into micro-fluidic chip field by Manz et al. first.Hereafter, BPE-ECL technology
Develop very fast, define closed type BPE-ECL (closed BPE-ECL, C-BPE-ECL) and open BPE-ECL
(open BPE-ECL, O-BPE-ECL) two kinds of basic configurations.In C-BPE-ECL, position respectively, yin, yang the two poles of the earth of bipolar electrode
Current flow path between two reaction tanks, two reaction tanks only passes through bipolar electrode；In O-BPE-ECL, bipolar electrode bag
Being embedded in a fluid passage, electric current merely through bipolar electrode and does not forms path through solution.
Currently, whether C-BPE-ECL or O-BPE-ECL, that is reported is mostly to enter in micro-fluidic chip
OK.Micro-fluidic chip backing material includes glass, PDMS, or glass/PDMS heterozygote, and electrode material is typically gold, oxygen
Change indium tin thin film.Such micro-fluidic chip relates to the chip lining of complexity, tediously long, chip manufacture process not environmentally and costliness
The end and electrode material.Additionally, the detection application of these micro-fluidic chips typically requires the ancillary equipment of costliness, including commercialization ECL
Analyser is for data acquisition, mechanical pump fluid drives etc. in the chip microchannel.Therefore, novel BPE-ECL method is compeled
Being essential will be with a series of important restriction overcoming traditional BP E-ECL to be faced.Up to the present, BPE-ECL ties mutually with cloth chip
The detection method and the device that close the most are not reported.
Summary of the invention
The primary and foremost purpose of the present invention is to provide a kind of closed type bipolar electrode electrogenerated chemiluminescence cloth chip, and this chip is tied
Close C-BPE-ECL and the advantage of two kinds of technology of cloth chip, cloth chip has been supported microchannel and report microchannel can not be same
In one plane, and its bending angle θ (after referring to that cloth chip bends along centre position, plane institute shape residing for certain microchannel and its original
The angle become) scope is 0 °-180 °；0 ° represents not bending, and i.e. two microchannels are in approximately the same plane；180 ° then represent
Two microchannels are the most overlapping.
Another object of the present invention is to provide the preparation method of above-mentioned cloth chip, required chip substrate and electrode material are honest and clean
Valency, environmental protection, the course of processing is easy, quickly.
It is still another object of the present invention to provide the application in the detection of above-mentioned cloth chip, its detection equipment easy to operate,
Low price, can be portable.
The purpose of the present invention is achieved through the following technical solutions:
A kind of cloth chip, its substrate is pieces of cloth, distribution electrode on pieces of cloth, and bipolar electrode is placed in the middle, a pair drive electrode distribution
Both sides；Positive drive electrode with bipolar electrode negative electrode by supporting that microchannel connect, feminine gender drive electrode and bipolar electrode anode by
The connection of report microchannel；The surrounding of described microchannel is hydrophobic wax dam；
Described microchannel is hydrophilic；
Two microchannels are not communicated with, and bipolar electrode is across two microchannels；
Described bipolar electrode can be divided into three parts, and its two ends are placed in supports dividing in microchannel and report microchannel
Being not negative electrode and anode, clip by wax dam in the middle of it is the 3rd part；
Preferably, described support microchannel and report microchannel, its length is equal, and width is equal；
Described electrode is made up of conductive carbon paste；
Its shape of described electrode is rectangle；Bipolar electrode is equal to the distance of drive electrode positive, negative.
The preparation method of above-mentioned cloth chip, comprises the following steps:
(1) design the pattern of microchannel and electrode, then make microchannel web plate and electrode web plate；
(2) take pieces of cloth and be close to below electrode web plate, electrode web plate is brushed conductive carbon paste, by electrode print at pieces of cloth
On；Brush complete, pieces of cloth are toasted 20-40 minute at 80-120 DEG C, makes electrode resistance tend towards stability, after baking at room temperature
Cooling；
(3) take and be printed on the pieces of cloth of electrode, microchannel web plate is close on the pieces of cloth back side (i.e. not having the one side of electrode), uses
Solid wax, smooth utensil are successively smeared on the web plate of microchannel, are milled；Then pieces of cloth are placed on heating together with the web plate of microchannel
On plate, being heated for only a few seconds at 70-130 DEG C, wax infiltration forms microchannel in pieces of cloth；
(4) pieces of cloth are cooled to room temperature, prepare closed type bipolar electrode electrogenerated chemiluminescence cloth chip；
In step (1), preferably with mapping software Adobe Illustrator CS5 design microchannel and the pattern of electrode；
In step (2), described pieces of cloth preferred white all cotton fabric.
Above-mentioned cloth chip can be used for detecting TPA (tripropyl amine (TPA)), H2O2And glucose；
Detection process comprises the following steps:
(1) cloth chip is fixed on a support, below support, pads last layer pad, on the one hand be easy to fixed chip and
Keep smooth；On the other hand prevent from chip, supporting that in microchannel and report microchannel, the reaction solution of dropping connects with object stage
Touch, cause unnecessary pollution to object stage；
Described pad is emptied corresponding to the area requirement of cloth chip microchannel, makes microchannel unsettled, it is to avoid to micro-logical
Reaction in road produces impact；
(2) use conducting adhesive cloth and alligator clamp to be connected with power supply by the drive electrode of cloth chip on support, and put into together
On object stage in camera bellows；Regulation cloth chip position and CCD (Charge Coupled Device, charge-coupled image sensor) are burnt
Cloth chip image clearly is can show that away from making on computer screen；CCD relevant parameter is set (such as time of exposure, between taking pictures
Every etc.) so that the ECL image needed for Huo get ing；
(3) on chip, support microchannel adds buffer, in report microchannel, add test fluid, wait the several seconds
Make solution be full of whole microchannel, start CCD automatic camera pattern；
(4) connect DC source to drive electrode applying appropriate voltage to trigger C-BPE-ECL, use CCD realtime imaging
ECL process.For each data point, experiment is repeatedly.The image of captured in real-time by Adobe Photoshop CS4,
The softwares such as Origin are analyzed processing, thus obtain data and curves；
In above-mentioned steps, for different detection objects, buffer is all 0.1M PBS (phophate buffer
Solution, phosphate buffer) (pH value 7.4), and test fluid is different:
For detection TPA, configure certain density Ru (bpy) with 0.1M PBS (pH value 7.4)3 2+Solution and finite concentration
TPA solution, then equal-volume mixing both solution obtain detect TPA test fluid；
For detection H2O2, it is respectively configured certain density Luminol solution with certain with 0.1M NaOH and 0.1M PBS
The H of concentration2O2Solution, then equal-volume mixing both solution obtains detecting H2O2Test fluid；
For detection glucose, it is respectively configured certain density Luminol solution and one with 0.1M NaOH and 0.1M PBS
Determining the glucose solution of concentration, then equal-volume mixing both solution obtains detecting the test fluid of glucose.Test fluid adds
In report microchannel.Before testing, bipolar electrode anode has pre-fixed glucoseoxidase, and enzyme pre-fixes process and is:
Bipolar electrode anode tap drips 2 μ L glucose oxidase solution, after 2min dries, then drips 1 enzymatic solution identical for μ L, at 4 DEG C
Dry in refrigerator and save backup.
In above-mentioned steps, it is different for different detection objects, chip bending angle θ or solution dropping mode；
Being 0 ° for detection TPA, chip bending angle θ, solution dropping mode is to use liquid-transfering gun by buffer and test fluid
It is added separately in support microchannel and the report microchannel of cloth chip；
For detection H2O2And glucose, chip bending angle θ is 180 °, and solution dropping mode is: first dropped in by solution dry
Drop is formed on clean disposable PE thin film；Then hydrophilic microchannel is contacted droplet solution, will under capillary force effect
Solution is drawn to support in microchannel or report microchannel.
The present invention has such advantages as relative to prior art and effect:
1, closed type bipolar electrode electrogenerated chemiluminescence, fabricbase microfluidic chip technology are carried out organic collection by the present invention first
Become, thus build a kind of simple, cheap biochemical sensor.
2, wire mark processing method based on carbon ink and solid wax is applied on fabricbase micro-fluidic chip by the present invention, this processing
One of advantage of method is to need not costly, complicated instrument and equipment, and another important advantage is batch machining can to go out required cloth
Base micro-fluidic chip.
3, in the present invention, electrode (including drive electrode and bipolar electrode) is carbon electrode, compared to conventional electrode materials such as
Gold, indium tin oxide films, not only cost of manufacture is cheap, and can single use, it is not necessary to complicated polishing pretreatment.
4, the inventive method is compared with existing micro-fluidic chip bipolar electrode electrogenerated chemiluminescence technology, it will be apparent that advantage
One of be that chip has fabulous flexibility, the most potential structure surpasses flexible sensor；Another advantage is compared to traditional die
Backing material (such as glass, PDMS etc.), cloth chip material have cheap, universal, user friendly, bio-compatibility is good, can be biological
The advantages such as degraded.
5, the inventive method is compared with existing tradition electrogenerated chemiluminescence, fabricbase micro-fluidic electrogenerated chemiluminescence technology,
Without expensive potentiostat；It is easily achieved high throughput testing.
6, method operating process described in the invention is simple, it is not necessary to professional operates.
7, the inventive method decreases the pollution to environment, and after sample analysis completes, chip can be processed by combustion method
Fall.
8, the inventive method only needs a few minutes from being loaded onto sample analysis, can realize quick, detection by quantitative.
9, the method for the present invention is applicable to common lab, easy to spread.
10, the cloth chip detection system of the inventive method has good stability, higher detection sensitivity, wider
Detection dynamic range and the advantage such as easily controllable, can on cloth chip detection by quantitative tripropyl amine (TPA), hydrogen peroxide and glucose, this
Extremely important Research Significance is had in fields such as environmental monitoring, food safety detection, medicals diagnosis on disease.
Accompanying drawing explanation
Fig. 1 is microchannel web plate.
Fig. 2 is electrode web plate.
Fig. 3 is the pictorial diagram of closed type bipolar electrode electrogenerated chemiluminescence cloth chip of the present invention；Wherein, the 1-positive drives electricity
Pole, 2-supports microchannel, 3-bipolar electrode, 4-wax dam, and 5-reports microchannel, 6-feminine gender drive electrode.
Fig. 4 is that luminous intensity is with the block diagram of liquid volume added change in cloth chip microchannel.
Fig. 5 is that luminous intensity is with driving voltage (Etot) change curve.
Fig. 6 is that luminous intensity is with bipolar electrode width (WBPE) change curve.
Fig. 7 is that luminous intensity is with bipolar electrode length (LBPE) block diagram that changes.
Fig. 8 is the block diagram that luminous intensity changes with cloth chip bending angle (θ).
Fig. 9 is that luminous intensity is with tripropyl amine (TPA) concentration curve (interior illustration is data linear matched curve figures).
Figure 10 is that luminous intensity is with concentration of hydrogen peroxide change curve (interior illustration is data linear matched curve figures).
Figure 11 is that luminous intensity is with concentration of glucose change curve (interior illustration is data linear matched curve figures).
Figure 12 is the block diagram that bipolar electrode anode does not pre-fixes that the cloth chip luminous intensity of enzyme changed with the storage time.
Figure 13 is the block diagram that bipolar electrode anode pre-fixes that the cloth chip luminous intensity of enzyme changed with the storage time.
Detailed description of the invention
Below in conjunction with embodiment and accompanying drawing, the present invention is described in further detail, but embodiments of the present invention do not limit
In this.
Embodiment 1
Closed type bipolar electrode electrogenerated chemiluminescence cloth chip preparation method, comprises the steps:
(1) use Adobe Illustrator CS5 mapping software designs support/report microchannel, male/female is driven
Moving electrode, the pattern of bipolar electrode, be then processed into 300 mesh polyester microchannel web plates (as it is shown in figure 1, white area by producer
Territory is cingens is microchannel) and electrode web plate (as in figure 2 it is shown, white is electrode)；
(2) by cotton sheet (white, cotton～10.7mg/cm2,～96 line/inches wide) be placed in below electrode web plate,
Make pieces of cloth and web plate be close to, by electrode web plate, brush conductive carbon paste (< 60 Ω/square) and by drive electrode, double property
Electrode print is on cotton sheet；
(3) brushing complete, cotton sheet separates with electrode web plate, and baking in 100 DEG C of baking ovens (model DHG-9035A)
30min, makes electrode resistance tend towards stability, at room temperature cools down after baking；
(4) take and be printed on the cotton sheet of electrode, microchannel web plate is close on the cotton sheet back side (not having the one side of electrode),
Successively smear on the web plate of microchannel with wax crayon, smooth utensil, mill；Then cotton sheet is placed on together with the web plate of microchannel and adds
On hot plate (model YH-946B), heating～3s at 85 DEG C, wax infiltration forms support microchannel based on wax dam in cotton sheet
With report microchannel；
(5) cotton sheet is separated with microchannel web plate, be cooled to room temperature, thus prepare C-BPE-ECL cloth chip.
The chip prepared is as it is shown on figure 3, its substrate is pieces of cloth, and distribution electrode on pieces of cloth, bipolar electrode 3 is placed in the middle, drives for a pair
Moving electrode distribution both sides；Positive drive electrode 1 is with bipolar electrode negative electrode by supporting that microchannel 2 connects, and negative drive electrode 6 is with double
Property electrode anode is by reporting that microchannel 5 connects；The surrounding of described microchannel is hydrophobic wax dam 4；
Two microchannels are not communicated with, and bipolar electrode 3 is across two microchannels；
Described microchannel is hydrophilic；
Described support microchannel 2 and report microchannel 5, its length is equal, and width is equal；
Described electrode is made up of conductive carbon paste；
Its shape of described electrode is rectangle；Bipolar electrode 3 is to positive drive electrode 1, the distance phase of negative drive electrode 6
Deng.
Embodiment 2
The cloth chip that embodiment 1 prepares application in detection TPA, comprises the following steps:
(1) the cotton chip prepared is fixed on a PET support, pads last layer pad below support (right
The part answering microchannel is emptied, and makes the microchannel unsettled), on the one hand it is easy to fixed chip and keeps smooth；On the other hand core is prevented
Support on sheet that microchannel contacts with object stage with the reaction solution of dropping in report microchannel, cause unnecessary dirt to object stage
Dye；
(2) use the two-sided conducting adhesive cloth that cuts and alligator clamp by the positive drive electrode of cotton chip on support and the moon
Property drive electrode positive and negative electrode with DC source (model PS-305D) respectively be connected, and put into together ECL measure with point
On the object stage of analysis system；Regulation cloth chip position makes the bipolar electrode anode positive alignment in cloth chip report microchannel be included in
The macro-lens (model M1614-MP) of the CCD camera (model MC15) in camera bellows, and regulate CCD focal length and make on computer screen
Can show that cloth chip image clearly；CCD relevant parameter (such as time of exposure 200ms, take pictures interval 5s etc.) is set so that
ECL image needed for acquisition；
(3) liquid-transfering gun is used to support the 0.1M PBS (pH 7.4) of addition 5-25 μ L in microchannel on chip；Toward report
Adding 5-25 μ L test fluid in microchannel, this test fluid is by Ru (bpy)3 2+Solution (being prepared by 0.1M PBS (pH 7.4)) and TPA
Solution (being prepared by 0.1M PBS (pH 7.4)) equal-volume is obtained by mixing), wait that 10s makes solution be full of whole microchannel, start
CCD automatic camera pattern；
(4) connect DC source and drive electrode is applied appropriate voltage (i.e. driving voltage Etot) to trigger C-BPE-ECL,
Use CCD realtime imaging ECL process.For each data point, experiment is at least repeated 8 times.The image of captured in real-time is with JPEG
Form preserves, and size is 1024 pixel × 1360 pixels, and is analyzed by Adobe Photoshop CS4 batch facility, from
And obtain the picture of ECL luminous intensity maximum (representing with gray value)；The pixel that maximum gradation value is multiplied by light-emitting zone obtains
To the number of photons of light-emitting area, and import data to Origin and do data analysis, thus obtain data and curves；
Now with containing 5mM Ru (bpy)3 2+(chip is joined with the test fluid of 25mM TPA and cloth chip that bending angle θ is 0 °
Number: the length (L of microchannelwc) and width (Wwc) difference 13mm and 6mm, the length (L of drive electrodede) and width (Wde) respectively
For 8mm and 5mm, bipolar electrode length (LBPE) be 15mm, bipolar electrode anode lengths (Lae) and bipolar electrode cathode length
(Lce) it is all 5mm, bipolar electrode width (WBPE) be 2mm) as a example by, use cloth chip to test at driving voltage EtotFor under 6.5V
Relation between different liquid volume addeds and ECL luminous intensity in microchannel.
Result is as shown in Figure 4, it can be seen that liquid volume added ECL luminous intensity when 5 μ L change to 15 μ L in report microchannel
Increasing therewith, when liquid volume added continues to increase, luminous intensity is held essentially constant；Support that microchannel has similar situation.Cause
This, in order to save reagent consumption, reduce and analyze price, preferably 15 μ L are buffer and test fluid volume.Just report microchannel
For test fluid, its acceptable volume minima is at 10-15 μ L；For supporting the buffer in microchannel, micro-in report
In the case of in passage, liquor capacity is enough, it is only necessary to will support that the whole moistening in microchannel can be obtained by the most luminous existing
As, its acceptable volume minima can be 5 μ L.
Embodiment 3
On some key factors (the driving voltage E affecting cloth chip luminous intensity in embodiment 2tot, bipolar electrode width
WBPE, bipolar electrode length LBPE, bending angle θ) carry out preferably
A) preferably driving voltage Etot
1, other conditions are with embodiment 2, i.e. cloth chip θ=0 °, Lwc=13mm, Wwc=6mm, Lde=8mm, Wde=5mm,
LBPE=15mm, WBPE=2mm, Lae=Lce=5mm；In report microchannel, test fluid volume is 15 μ L, Ru (bpy)3 2+Concentration is
5mM, TPA concentration is 25mM；Support that in microchannel, PBS volume is 15 μ L, concentration is 0.1M (pH value 7.4).
2, some experimental grouies are set: driving voltage EtotBe set to several different value (3V, 3.5V, 4V, 4.5V, 5V, 5.5V,
6V、6.5V、7V)。
3, cloth chip C-BPE-ECL detection process is with embodiment 2, and test result is as shown in Figure 5.
From experimental result it can be seen that along with driving voltage EtotChange to 6.5V, ECL luminous intensity from 3.5V gradually increase
By force；As driving voltage EtotDuring less than 3.5V, it is barely perceivable ECL signal；As driving voltage EtotDuring more than 6.5V, ECL believes
Number begin to decline.It is therefore preferable that driving voltage EtotFor 6.5V, acceptable scope is 5.5-6.5V.
B) bipolar electrode width WBPE
1, other conditional likelihoods are in a), i.e. cloth chip θ=0 °, Lwc=13mm, Wwc=6mm, Lde=8mm, Wde=5mm,
LBPE=15mm, Lae=Lce=5mm；Driving voltage Etot=6.5V；In report microchannel, test fluid volume is 15 μ L, Ru
(bpy)3 2+Concentration be 5mM, TPA concentration be 25mM；Support that in microchannel, PBS volume is 15 μ L, concentration is 0.1M (pH value 7.4).
2, some experimental grouies are set: bipolar electrode width WBPEBe set to several different value (0.5mm, 1mm, 2mm, 3mm,
4mm)。
3, cloth chip C-BPE-ECL detection process is with embodiment 2, and test result is as shown in Figure 6.
From experimental result it can be seen that along with bipolar electrode width WBPE3mm, ECL luminous intensity is changed to bright from 0.5mm
Aobvious enhancing.But, bipolar electrode width WBPEWhen 3mm increases to 4mm further, ECL luminous intensity slightly increases and (is less than
6%), meanwhile the carbon electrode materials consumed are more than 30% in theory.Additionally, it is possible to owing to the excess of electroactive material disappears
Consumption, the bad stability of C-BPE-ECL signal.True based on these, preferably bipolar electrode width WBPEFor 3mm, acceptable model
Enclosing is 2.5-4mm.
C) bipolar electrode length LBPE
1, other conditional likelihoods are in b), i.e. cloth chip θ=0 °, Lwc=13mm, Wwc=6mm, Lde=8mm, Wde=5mm,
WBPE=3mm, Lae=Lce=5mm；Driving voltage Etot=6.5V；In report microchannel, test fluid volume is 15 μ L, Ru
(bpy)3 2+Concentration be 5mM, TPA concentration be 25mM；Support that in microchannel, PBS volume is 15 μ L, concentration is 0.1M (pH value 7.4).
2, some experimental grouies are set: bipolar electrode length LBPEBe set to several different value (11mm, 13mm, 15mm, 17mm,
19mm)。
3, cloth chip C-BPE-ECL detection process is with embodiment 2, and test result is as shown in Figure 7.
From experimental result it can be seen that bipolar electrode length LBPEChange (i.e. report microchannel and support between microchannel double
Property electrode part length change) to ECL luminous intensity produce negligible effect: in every case, averagely send out
The relative deviation of light intensity is less than 7.5%；Relative standard deviation is less than 5.3%.In order to make cloth chip compact small and exquisite, consume few with
And make two microchannels of cloth chip be prone to not affect detection performance, based on different along the bending of bipolar electrode middle position
Cloth chip detection application purpose, bipolar electrode length LBPECan be preferably 13mm or 15mm, acceptable scope is 11-19mm.
D) bending angle θ
1, other conditional likelihoods are in c), i.e. cloth chip Lwc=13mm, Wwc=6mm, Lde=8mm, Wde=5mm, LBPE=
15mm、WBPE=3mm, Lae=Lce=5mm；Driving voltage Etot=6.5V；Support that in microchannel, PBS volume is 15 μ L, concentration is
0.1M (pH value 7.4).
2, in report microchannel, test fluid volume is 15 μ L, Ru (bpy)3 2+Concentration be 0.5mM, TPA concentration be 2.5mM；
3, some experimental grouies are set: bending angle θ is set to several different value (0 °, 45 °, 90 °, 135 °, 180 °).
4, cloth chip C-BPE-ECL detection process is with embodiment 2, and test result is as shown in Figure 8.
From experimental result it can be seen that (cloth chip does not make any angle folding when being 0 ° relative to cloth chip bending angle θ
Time), the cloth chip making to bend demonstrates an analogous C-BPE-ECL luminous intensity.Therefore, C-BPE-ECL cloth chip is not
Only there is fabulous flexibility, and show stronger luminescence.Perhaps, this superperformance can widen the application that cloth chip is potential
Field, improve the potential integrated level of cloth chip.
Embodiment 4
The optimum condition groped with embodiment 3 utilizes cloth chip detection TPA
A () uses embodiment 3 more preferred cloth chip parameter i.e. θ=0 °, Lwc=13mm, Wwc=6mm, Lde=8mm,
Wde=5mm, LBPE=13mm, WBPE=3mm, Lae=Lce=5mm.It addition, driving voltage Etot=6.5V；Report is surveyed in microchannel
Test solution volume is 15 μ L, Ru (bpy)3 2+Concentration is 0.5mM；Support that in microchannel, PBS volume is 15 μ L, concentration is 0.1M (pH value
7.4)。
B () arranges some experimental grouies: TPA concentration be set to several different value (0mM, 0.1mM, 0.25mM, 0.5mM,
1.25mM、2.5mM、5mM、10mM)。
C () cloth chip C-BPE-ECL detection process is with embodiment 2, test result is as shown in Figure 9.
From experimental result it can be seen that C-BPE-ECL luminous intensity raises along with TPA concentration and increases.TPA concentration from
When 0mM changes to 2.5mM, C-BPE-ECL luminous intensity (representing with Y) and TPA concentration (representing with X) are in certain linear pass
System, linear equation can be expressed as Y=0.500X-0.014 (R2=0.9910).The computational methods that detection limit uses are: YL=Yb+
3Sb(YbFor average canbdle power value, S during blankbFor standard deviation during blank) (repeating experiment for eight times), by institute
The Y obtainedLThe concentration of value correspondence obtains TPA detectable limit.In this embodiment, TPA detectable limit is 0.085mM.
Embodiment 5
Some optimum conditions groped with embodiment 3 carry out super soft cloth chip detection H2O2
A () uses more preferred cloth chip parameter i.e. θ=180 ° of embodiment 3 (maximum bending angle), Lwc=13mm, Wwc
=6mm, Lde=8mm, Wde=5mm, LBPE=15mm, WBPE=3mm, Lae=Lce=5mm.
(b) driving voltage Etot=5V；Report microchannel in test fluid (the Luminol solution configured by 0.1M NaOH and
The H of 0.1M PBS configuration2O2Solution equal-volume is obtained by mixing) volume be 15 μ L, Luminol concentration be 5mM；Support in microchannel
PBS volume is 15 μ L, concentration is 0.1M (pH value 7.4).
C () arranges some experimental grouies: H2O2Concentration be set to several different value (0mM, 0.025mM, 0.125mM, 0.25mM,
0.5mM、2.5mM)。
D () cloth chip C-BPE-ECL detection similar process is in embodiment 2, slightly different: (1) for bending chip,
It is difficult to liquid feeding in camera bellows, thus after being pre-adjusted CCD focal length, in cloth chip microchannel, drips corresponding solution, connect crocodile
Fish is pressed from both sides, then places cloth chip in camera bellows；(2) solution dropping mode is: first solution drop in clean disposable PE thin film
Upper formation drop；Then hydrophilic fabricbase microchannel is contacted droplet solution, be drawn to solution prop up under capillary force effect
Hold in microchannel or report microchannel；
Start CCD automatic camera pattern, and CCD parameter (time of exposure is 200ms, takes pictures and is spaced apart 0.5s etc.) be set,
Wait that 60s makes solution be full of whole microchannel.Test result is as shown in Figure 10.
From experimental result it can be seen that C-BPE-ECL luminous intensity is along with H2O2Concentration raises and increases.At H2O2Concentration
When 0.025mM changes to 2.5mM, C-BPE-ECL luminous intensity (representing with Y) and H2O2The logarithm (representing with X) of concentration is in one
Constant linear relation, linear equation can be expressed as Y=1.145X+1.943 (R2=0.9930).The computational methods that detection limit uses
It is: YL=Yb+3Sb(YbFor average canbdle power value, S during blankbFor standard deviation during blank) (repeat for eight times real
Test), by the Y of gainedLThe concentration of value correspondence obtains H2O2Detectable limit.In this embodiment, H2O2Detectable limit is
0.024mM。
Embodiment 6
When super soft cloth chip C-BPE-ECL detects glucose, except in cloth chip report microchannel, bipolar electrode anode is pre-solid
Determine in glucoseoxidase and test fluid containing glucose different outside, other all with embodiment 5 in detection H2O2Condition
Identical, these conditions include test process, chip parameter, driving voltage etc..
A in () cloth chip report microchannel, bipolar electrode anode pre-fixes glucose oxidase procedure and is: at double property electricity
Pole anode tap drips 2 μ L glucose oxidase solution (5unit/ μ L), after 2min dries, then drips 1 enzymatic solution identical for μ L,
Preservation is dried in 4 DEG C of refrigerators.
The Luminol solution that configured by 0.1M NaOH of test fluid of (b) detection glucose and the Fructus Vitis viniferae of 0.1M PBS configuration
Sugar juice equal-volume is obtained by mixing.
Some experimental grouies are set and investigate the sensitivity of the method detection by quantitative glucose, wherein Fructus Vitis viniferae in test fluid further
Sugar concentration is respectively 0mM, 0.25mM, 0.5mM, 1mM, 1.25mM, 2.5mM, 5mM, and Figure 11 shows C-BPE-ECL luminous intensity
Relation with concentration of glucose.
As seen from Figure 11 the logarithm (representing with X) of C-BPE-ECL luminous intensity (representing with Y) and concentration of glucose in
Some linear, linear equation can be expressed as Y=0.927X+0.754 (R2=0.9956).The computational methods that detection limit uses
It is: YL=Yb+3Sb(YbFor average canbdle power value, S during blankbFor standard deviation during blank) (repeat for eight times real
Test), by the Y of gainedLThe concentration of value correspondence obtains the glucose detection limit.In this embodiment, the glucose detection limit is
0.195mM。
Embodiment 7
The storage stability evaluation of cloth chip
A, bipolar electrode anode do not pre-fix the cloth chip of enzyme
A () uses the cloth chip parameter of embodiment 4, i.e. θ=0 °, Lwc=13mm, Wwc=6mm, Lde=8mm, Wde=5mm,
LBPE=13mm, WBPE=3mm, Lae=Lce=5mm.It addition, driving voltage Etot=6.5V；Test fluid volume in report microchannel
It is 15 μ L, Ru (bpy)3 2+Concentration be 5mM, TPA concentration be 25mM；Support that in microchannel, PBS volume is 15 μ L, concentration is 0.1M
(pH value 7.4)；Cloth chip is stored in room temperature, air.
B () arranges some experimental grouies: cloth chip memory time interval be set to several different value (0 day, 2 days, 4 days, 6 days,
8 days, 10 days, 12 days, 14 days, 16 days, 18 days, 20 days).
C () cloth chip C-BPE-ECL detection process is with embodiment 2, test result is as shown in figure 12.
From experimental result it can be seen that C-BPE-ECL luminous intensity only occurs slight change within the storage time.Cloth chip
After storing 10 days and 20 days, its luminous intensity drops to the 95.5% and 90.2% of initial luminescence values (0 day) respectively.
B, bipolar electrode anode pre-fix the cloth chip of enzyme
A () uses the cloth chip parameter of embodiment 6 (namely embodiment 5), i.e. θ=180 °, Lwc=13mm, Wwc=6mm,
Lde=8mm, Wde=5mm, LBPE=15mm, WBPE=3mm, Lae=Lce=5mm.It addition, driving voltage Etot=5V；Report micro-logical
In road test fluid volume be 15 μ L, Luminol concentration be 5mM, concentration of glucose be 2mM；Support that in microchannel, PBS volume is 15
μ L, concentration are 0.1M (pH value 7.4).
B in () cloth chip report microchannel, bipolar electrode anode pre-fixes glucose oxidase procedure and containing glucose
Test fluid configuration process be all same as embodiment 6；The sealing of cloth chip, stored dry are in 4 DEG C of temperature.
C () arranges some experimental grouies: cloth chip memory time interval be set to several different value (0 day, 2 days, 4 days, 6 days,
8 days, 10 days, 12 days, 14 days, 16 days, 18 days, 20 days).
C () cloth chip C-BPE-ECL detection process is with embodiment 6, test result is as shown in figure 13.
From experimental result it can be seen that relative to C-BPE-ECL initial luminescence values (0 day), cloth in storing first 4 days of time
There is any change hardly in chip light emitting intensity.After cloth chip memory 10 days and 20 days, at the beginning of its luminous intensity drops to respectively
Originate the 94.5% and 87.7% of light value.
In sum, cloth chip whether bipolar electrode anode does not pre-fixes enzyme and still pre-fixes enzyme, and they all have pole
Good storage stability, it is therefore possible to be adapted to backwoodsman detection application.
Above-described embodiment is the present invention preferably embodiment, but embodiments of the present invention are not by above-described embodiment
Limit, the change made under other any spirit without departing from the present invention and principle, modify, substitute, combine, simplify,
All should be the substitute mode of equivalence, within being included in protection scope of the present invention.
Claims (10)
1. a cloth chip, it is characterised in that: substrate is pieces of cloth, distribution electrode on pieces of cloth, and bipolar electrode is placed in the middle, a pair driving electricity
Distribution both sides, pole；Positive drive electrode with bipolar electrode negative electrode by supporting that microchannel connects, feminine gender drive electrode and bipolar electrode
Anode is by reporting that microchannel connects；The surrounding of described microchannel is hydrophobic wax dam；
Described microchannel is hydrophilic.
Cloth chip the most according to claim 1, it is characterised in that: two described microchannels are not communicated with, and bipolar electrode is horizontal
Across two microchannels.
Cloth chip the most according to claim 1, it is characterised in that: described bipolar electrode is divided into three parts, its two ends
Being placed in and support that in microchannel and report microchannel be negative electrode and anode respectively, clip by wax dam in the middle of it is the 3rd portion
Point.
Cloth chip the most according to claim 1, it is characterised in that: described support microchannel and report microchannel, it is long
Spending equal, width is equal.
Cloth chip the most according to claim 1, it is characterised in that: described electrode is made up of conductive carbon paste.
Cloth chip the most according to claim 1, it is characterised in that: its shape of described electrode is rectangle；Bipolar electrode arrives
The distance of drive electrode positive, negative is equal.
7. the preparation method of the cloth chip described in any one of claim 1-6, it is characterised in that comprise the following steps:
(1) design the pattern of microchannel and electrode, then make microchannel web plate and electrode web plate；
(2) take pieces of cloth and be close to below electrode web plate, electrode web plate is brushed conductive carbon paste, by electrode print on pieces of cloth；
Brush complete, pieces of cloth are toasted 20-40 minute at 80-120 DEG C, makes electrode resistance tend towards stability, the coldest after baking
But；
(3) take and be printed on the pieces of cloth of electrode, microchannel web plate is close on the pieces of cloth back side, with solid wax, smooth utensil micro-logical
Successively smear on road web plate, mill；Then pieces of cloth are put on hot plate together with the web plate of microchannel, at 70-130 DEG C, heat number
Second, wax infiltration forms microchannel in pieces of cloth；
(4) pieces of cloth are cooled to room temperature, prepare closed type bipolar electrode electrogenerated chemiluminescence cloth chip.
8. the cloth chip described in any one of claim 1-6 is at detection tripropyl amine (TPA), H2O2With the application in glucose.
Cloth chip the most according to claim 8 is at detection tripropyl amine (TPA), H2O2With the application in glucose, it is characterised in that bag
Include following steps:
(1) cloth chip is fixed on a support, below support, pads last layer pad；
Described pad is emptied corresponding to the area requirement of cloth chip microchannel；
(2) use conducting adhesive cloth and alligator clamp to be connected with power supply by the drive electrode of cloth chip on support, and put into dark together
On object stage in case；Regulation cloth chip position and CCD focal length make to can show that on computer screen cloth chip figure clearly
Picture；CCD relevant parameter is set so that the ECL image needed for Huo get ing；
(3) on chip, support microchannel adds buffer, in report microchannel, add test fluid, wait that the several seconds makes molten
Liquid is full of whole microchannel, starts CCD automatic camera pattern；
(4) connect DC source to drive electrode applying appropriate voltage to trigger C-BPE-ECL, use CCD realtime imaging ECL mistake
Journey；The image of captured in real-time processes by analysis, obtains data and curves.
Application the most according to claim 9, it is characterised in that: for detection H2O2And glucose, chip bending angle θ is
180 °, solution dropping mode is: first solution drop in formation drop on clean disposable PE thin film；Then by hydrophilic
Microchannel contact droplet solution, is drawn to solution support in microchannel or report microchannel under capillary force effect.
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