CN109913352A - A microfluidic device and method for manipulating and capturing microparticles and cells based on non-contact dielectrophoresis - Google Patents
A microfluidic device and method for manipulating and capturing microparticles and cells based on non-contact dielectrophoresis Download PDFInfo
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Abstract
本发明提供一种基于非接触式介电电泳力操控捕获微颗粒和细胞的微流控装置，包括：依次连接的进样泵，微量注射器，微流控芯片，以及废液收集器；所述微流控芯片由玻璃基底层和聚二甲基硅氧烷芯片层贴合而成，其中，所述聚二甲基硅氧烷芯片层包括：样品进样口，样品进样通道；鞘流进样口，鞘流进样通道；分别独立成封闭环状的、布置于样品分离通道两侧的第一、第二液体电极沟道，以向其施加一个高频高压非均匀电场；目标产物收集通道和废液收集通道；以及出样口。本发明还提供一种基于非接触式介电电泳力操控捕获微颗粒和细胞的方法。根据本发明提供的装置和方法，具有分离效率高，操作简单，成本低，通量高的优点。
The invention provides a microfluidic device for controlling and capturing microparticles and cells based on non-contact dielectrophoresis force, comprising: a sampling pump, a microinjector, a microfluidic chip, and a waste liquid collector connected in sequence; the The microfluidic chip is formed by laminating a glass base layer and a polydimethylsiloxane chip layer, wherein the polydimethylsiloxane chip layer includes: a sample injection port, a sample injection channel; a sheath flow Injection port, sheath flow injection channel; independent first and second liquid electrode channels arranged in a closed ring and arranged on both sides of the sample separation channel to apply a high frequency and high voltage non-uniform electric field to it; target product Collection channel and waste collection channel; and sample outlet. The present invention also provides a method for manipulating and capturing microparticles and cells based on non-contact dielectrophoretic force. The device and method provided by the present invention have the advantages of high separation efficiency, simple operation, low cost and high throughput.
Description
Technical field
The present invention relates to field of biological detection, relate more specifically to a kind of based on the manipulation capture of contactless dielectrophoresis force
The micro fluidic device and method of microparticle and cell.
Background technique
Cancer is still high one of the disease of the global death rate at present.2008, the crowd for dying of cancer reached 7,600,000
(account for total toll 13%), and to the year two thousand thirty may be more than 13,000,000.The early diagnosis and therapy of primary cancer with
And effective treatment of metastatic cancer is the key that anticancer.
Circulating tumor cell (Circulating Tumor Cells, CTCs) is one kind from primary tumo(u)r, into blood
The tumour cell of fluid circulation.This kind of cell can be shifted by blood circulation system, and it is swollen to reach the realization of other organs
The transfer of tumor, so the occurrence and development and transfer of CTCs and tumour have close relationship.In view of CTCs in metastases
The important function played, we can monitor the effect and recurrence of oncotherapy in real time by the variation of detection CTCs.
Averagely contain 5 × 10 in normal person's 1mL peripheral blood10Red blood cell, 7 × 106Leucocyte and 2.95 × 106Blood is small
Plate.Analysis tumour cell is a huge challenge in the blood so complicated from ingredient.If be able to achieve CTCs separation and
Enrichment, so that it may the number and characterization of molecules of CTCs are analyzed, treated for diagnosing tumor parting and drug target provide according to
According to.
What the enrichment of CTCs was mainly realized according to the special physicochemical properties of target cell.Traditional enrichment method
There are density-gradient centrifugation method, membrane filter method, micro-fluid chip method and separation method dependent on antibody etc..Wherein, U.S. FDA batch
Quasi- commercialized CellSearchTM system (Veridex LLC) just belongs to a kind of circulating tumor cell dependent on antibody point
From method.The system is to utilize the marker of magnetic bead and the surface epithelial cell expression of antibody modification, epithelial cell adhesion molecule
(Epithelial cell adhesion molecule, EpCAM) specifically binds to be enriched with CTCs.But this detection side
Method is only applied in breast cancer, colon cancer and prostate cancer [Miller MC, Doyle GV, Terstappen LW at present
Significance of CirculatingTumor Cells Detected by the CellSearch System in
Patients with MetastaticBreast Colorectal and Prostate Cancer.J Oncol,2010,
617421], the verification and measurement ratio in lung cancer is relatively low.Tanaka etc. passes through 125 patients with lung cancer of CellSearch system detection,
CTCs (30.6%) [Tanaka F, Yoneda K, et al.Circulating is only detected in 38 peripheral blood in patients
tumor cell as a diagnostic marker in primary lung cancer.J Clin Cancer Res,
2009,15(22):6980-6986].The capture rate of immunomagnetic bead detection method depends on CTCs marker EpCAM, can
It is that EpCAM antigen is expressed in different types of tumour with heterogeneity, or even is expressed in the tumour in some non-epidermis sources
It is missing from.The deficiencies of so that there are detection sensitivities is low for this detection method, application range is narrow, at high cost, and consuming time is long.
It is a kind of to be separated using circulating tumor cell with the difference of normal plasma cell physical characteristic in order to solve these deficiencies
The method of target cell is come into being.In recent years, constantly improve and develop with micro-fluidic and micromachining technology, realizes
The micro-fluidic chip channel of accurate dimension and aperture are possibly realized.The micro-fluidic chip method based on cell size can in this way
Higher capture rate is obtained, in addition to this, micro-fluidic chip also has many advantages, such as that small in size, analysis speed is fast, at low cost.This
All be conducive to micro-fluidic chip method being applied to clinical application a bit, to make up the deficiency of existing clinical testing procedure.Go out at present
There are many existing method based on micro-fluidic chip method detection circulating tumor cell, and patent document 1 (CN 103642756A) is announced
It is a kind of to be captured in conjunction with the antibody for being marked with complementary nucleotide in micro-fluidic chip microchannel surface modification single-stranded nucleotide
The method of circulating tumor cell.
It has disclosed in technology at present, for the Acquisition Scheme of circulating tumor cell, rough can be classified as passively sorting
With actively sort two major classes: passive sorting technology includes micro-structure filtering, field flow and waterpower sorting, certainty laterally offset, is used to
Property sorting, bionical sorting, compatibility sorting the methods of；Active sorting technology includes dielectrophoresis sorting, magnetic separation, sound sorting, light
The methods of sorting, we select dielectrophoresis sorting schemes herein, and dielectrophoresis is different from general electrophoresis, its scope of application
Larger, effect is uncharged neutrophil granule, thus can be used to manipulate the neutral microparticle of sorting and cell.Dielectrophoresis produces
Raw dielectrophoresis force is a kind of contactless force, therefore will not be to microparticle and cell during manipulating microparticle and cell
Mechanical damage is generated, while dielectrophoresis force is the electrical parameter by microparticle and cell itself, makes it in high-frequency and high-voltage electric field
Polarization motion, therefore do not need additional marking operation.The advantages of program includes: 1. effect neutrophil granules；2. non-contact
Power；3. without label, still select dielectrophoresis scheme realize the manipulation to microparticle and cell capture.
But traditional metal contact dielectrophoresis chip, electrode are contacted with sample, electric heating in contaminated samples, electrode
There is larger impact etc. to sample, while must first deposit metal electrode in chip base, then, then is directed at bonding, technique is multiple
Miscellaneous, chip cost is higher.
Summary of the invention
The object of the present invention is to provide it is a kind of based on contactless dielectrophoresis force manipulate capture microparticle and cell it is micro-
Flow control apparatus and method, to solve traditional metal contact dielectrophoresis chip contaminated samples, complex process and chip
The problem of higher cost.
In order to solve the above-mentioned technical problem, the invention adopts the following technical scheme:
According to the first aspect of the invention, it provides a kind of based on contactless dielectrophoresis force manipulation capture microparticle and thin
The micro fluidic device of born of the same parents, comprising: sequentially connected sampling pump, micro syringe, micro-fluidic chip and liquid waste collector；Institute
It states micro-fluidic chip to be bonded by glass substrate layers and polydimethylsiloxanechip chip layer, wherein the polydimethylsiloxanes
Alkane chip layer includes: sample feeding mouth, the sample feeding channel connecting with the sample feeding mouth；Sheath stream injection port, described in
Sheath stream injection port is drawn, and the two sheath stream sample intake passages to be formed that are divided into two, the end point of two sheath stream sample intake passages
Do not converge in the two sides in sample feeding channel with the sample feeding channel, forms a sample split tunnel；Independently at
Closed circular, the first, second liquid electrode channel for being arranged in sample split tunnel two sides, first liquid electrode
Channel includes: first electrode liquid injection port and channel tip wave structure, and the second liquid electrode channel includes: second electrode
Liquid injection port and the gentle structure in channel tip, wherein channel tip wave structure and the gentle structure in channel tip point
It is not arranged in the opposite sides thereof of the sample split tunnel, it is non-homogeneous to apply a high-frequency and high-voltage to the sample split tunnel
Electric field；It is drawn from the end of the sample split tunnel, and the target product collection channel and waste liquid being respectively formed that be divided into two
Collection channel；And respectively with the target product collection channel and the channel attached outlet of waste collection.
The micro-fluidic chip in micro fluidic device provided according to the present invention, the first liquid electrode channel and second liquid electricity
Pole channel is respectively arranged in the opposite sides thereof of sample split tunnel, on the one hand, eletrode tip and sample split tunnel border
Certain distance is separated by with cured PDMS, therefore referred to as a kind of non-contact liquid channel electrode；On the other hand, due to first,
The two-part main structure of second liquid electrode channel is almost the same, but is that wave is convex in channel tip portion side shape
It rises, side shape is gently extension, therefore also known as local asymmetry electrode.Its working principle is that: according to point effect, electrode
Surface curvature radius is smaller, and surface charge density is bigger, and power line is more intensive, and electric field strength is higher；Conversely, then electric field strength is got over
It is low, apply a high-frequency and high-voltage inhomogeneous field to realize to sample split tunnel, and then in induced samples split tunnel
Sample polarization, then sorting deflects sample.
Unlike using contact metal electrode from dielectrophoresis chip disclosed in present technology, the invention
Using contactless local asymmetry liquid electrode channel replace traditional metal electrode, while reaching effect substantially
The manufacturing process for simplifying chip, reduces the manufacturing cost and bonding required precision of chip, while avoiding metal electrode
The pollution of sample and electric heating are influenced.
The high unity of the entire micro-fluidic chip, preferably height are 30 μm -50 μm, most preferably 40 μm.
Preferably, it in the micro-fluidic chip, is provided in the biggish liquid storage channel of electrode section and is used to support channel
Cylindrical columns, diameter are about 100 μm.
Preferably, the sample feeding channel is centrally arranged with serpentine channels, and the sheath stream sample intake passage is respectively separate
The two sides in sample feeding channel are arranged, then gradually close to converge with the sample feeding channel, form sheath flow structure.
Before converging, the sample feeding channel and the width of sheath stream sample intake passage are become narrow gradually.
Preferably, it described 150-200 μm of sample feeding channel width, is become narrow gradually in sheath flow structure part, final width becomes
At 30-40 μm, the sheath stream sample intake passage is 100-120 μm wide, becomes narrow gradually in sheath flow structure part, final width becomes 20-
30μm。
Most preferably, 200 μm of the sample feeding channel width becomes narrow gradually in sheath flow structure part, and final width becomes
40 μm, the sheath stream sample intake passage is 100 μm wide, becomes narrow gradually in sheath flow structure part, final width becomes 30 μm.
The first liquid electrode channel further include: first electrode liquid liquid storage area and first electrode liquid buffer area, described
Two liquid electrode channels further include: second electrode liquid liquid storage area and second electrode liquid buffer area.
Wherein, the first, second electrode solution liquid storage area works as a buffer, and prevents subsequent in the first, second electrode solution injection port
It is shaken when being inserted into metal electrode, leads to the cutout of electrode solution in electrode channel, influence liquid electrode conduction.
Channel tip wave structure is one section of channel with wave-like, and the gentle structure in channel tip is one
Section has the channel of rectilinear form.
The micro fluidic device further includes the multiple target product liquid storage chambers connecting with the target product collection channel.It answers
When understanding, different numbers or different size of target product liquid storage chamber can be set according to actual needs, for example, three, four
It is a or six.
Sequentially connected sampling pump and micro syringe are adopted as the dynamical system in micro fluidic device provided by the invention
With the mode of positive pressure sample introduction, sample introduction flow velocity can be accurately controlled.
Flow dividing structure step by step is additionally provided between the target product collection channel and the target product liquid storage chamber.
The micro fluidic device further includes the electrode for being inserted into the first, second electrode solution injection port respectively, is believed with the electrode
The high-voltage amplifier of number connection, function generator and computer.
According to the second aspect of the invention, it provides a kind of based on contactless dielectrophoresis force manipulation capture microparticle and thin
The method of born of the same parents, comprising the following steps: 1) provide a kind of as described above based on the manipulation of contactless dielectrophoresis force capture micro-
The micro fluidic device of grain and cell；2) it imports washing lotion to be rinsed the micro-fluidic chip, then simultaneously that sheath flow liquid, sample is same
When import the micro-fluidic chip, after sheath flow liquid stable in the micro-fluidic chip is formed, i.e., openable external circuit is opened
Row is put into capture the manipulation of microparticle or cell；And 3) observe the fortune of microparticle and cell in electric field in real time under the microscope
Emotionally condition, optimal physical parameter of the analysis for different microparticles or cell.
Specifically, step 1) further include: suck the sample into micro syringe, micro syringe is then connected to sampling pump
On, then by tubule micro syringe is connected with micro-fluidic chip.
Step 2) further include: micro-fluidic chip is placed in vacuum kettle and is vacuumized, then draws DEP buffer with pipette tips
It is injected separately into the first, second electrode solution injection port.
Step 2) further include: rinse the chip with DEP buffer, i.e., by sample feeding mouth and sheath fluid injection port, first will
DEP buffer carries out a sample introduction process, so that DEP buffer is full of all channels, plays a flushing and immersional wetting, convenient
Subsequent sample sample introduction.
Step 2) further includes that sample introduction flow rate pump is adjusted to 10mL/h-30mL/h, make sample and buffer start together into
Sample.
According to sample dielectric parameter and physical size difference, current field condition, optimization buffer Jie can be applied by changing
Electrical parameter, adjustment channel flow velocity, main channel dimensions, liquid channel size and at a distance from tap drain road, it is inclined to reach differentiation
Turn the purpose of sorted sample.
According to the present invention, microparticle refers to that the electroneutral particle of micron level, such as polystyrene microsphere etc., cell can wrap
Include circulating tumor cell etc..
When method provided by the invention is used for the capture of polystyrene microsphere, its working principle is that: the polyphenyl of electroneutral
Dielectric polarization, i.e. free charge a small amount of inside polystyrene microsphere first occur in high frequency inhomogeneous field for ethylene microballoon outside
Charge rearrangement under boundary's electric field action, formation dipole moment, can be with two identical carried charges but opposite polarity charge is come table
Show, when this dipole moment is located in inhomogeneous field, makes a resulting net force in the difference of the local electric field strength on particle both sides
It generates, this resulting net force is referred to as dielectrophoresis force, polystyrene microsphere can be made to translate fortune under the resulting net force by optimal conditions
It is dynamic, it is collected into target product collection channel.
When method provided by the invention is used for the capture of circulating tumor cell, needs before sample introduction, first use sample
PBS rinses concussion centrifugation twice, and is resuspended in DEP buffer (8.5% sucrose [wt/vol], 0.3% glucose of our preparations
[wt/vol]) in.Then sample introduction in 100 μ L micro syringes is sucked again.The process of sample introduction needs to use sampling pump two in parallel
Channel.Sample introduction starts that subsequent dielectric electrophoretic procedures can be carried out after sheath flows out and stablizes.
The present invention is based on contactless dielectrophoresis capture circulating tumor cell micro-fluidic chip can and fluorescent staining
Identification combines the detection method for separating for constructing a kind of circulating tumor cell, dynamically detects during flowing to realize
With capture circulating tumor cell.Miniflow provided by the invention based on contactless dielectrophoresis technology capture circulating tumor cell
Control device greatly improves the capture ability to circulating tumor cell, and cost is relatively low for the micro-fluidic chip, on the one hand, silicon
After sheet mold is carried out, it can reuse；On the other hand, electrode section is replaced with channel in chip, is eliminated in substrate
On the step of depositing metal electrode in advance, therefore carved on mold simultaneously because electrode channel and sample channel are in same layer
While losing sample channel, electrode channel can etch together to be come, simple process, and eliminates pair of later period chip bonding
Quasi- process.
In short, easy to operate, at low cost, flux is high, during flowing the present invention provides a kind of separative efficiency height
Apply the micro fluidic device and method of active force manipulation capture circulating tumor cell.
Detailed description of the invention
Shown in FIG. 1 is the overall structure signal of the micro fluidic device provided according to a preferred embodiment of the present invention
Figure；
Shown in Fig. 2 is the structural schematic diagram of the micro-fluidic chip in micro fluidic device；
Fig. 3 is the detail enlarged diagram of sheath flow structure as shown in Figure 2；
Fig. 4 be liquid electrode channel portion as shown in Figure 2 be not filled by conduction liquid and fill conduction liquid after structure show
It is intended to；
Fig. 5 is to apply different electric pulse field parameters in embodiment 2 in microballoon sample introduction, different sized microspheres deflection situation comparisons
Figure；
Fig. 6 is when implementing to set circulating tumor cell H446 for sample introduction sample in 3, to start circulating tumor cell after electrode
The captured result figure of H446；
Fig. 7 is that Fig. 6 dielectric electrophoresis region potential electric field distribution situation such as emulates schematic diagram.
Specific embodiment
Below in conjunction with specific embodiment, the present invention will be further described.It should be understood that following embodiment is merely to illustrate this
The range of invention and is not intended to limit the present invention.
According to a preferred embodiment of the present invention, it provides a kind of based on the manipulation of contactless dielectrophoresis force capture micro-
The micro fluidic device of grain and cell, as shown in Figure 1, comprising: sequentially connected sampling pump 100, micro syringe 200 are micro-fluidic
Chip 300, liquid waste collector 400, high-voltage amplifier 500, function generator 600 and computer 700.
Specifically, micro-fluidic chip 300 structure as shown in Fig. 2, the micro-fluidic chip 300 by dimethyl silicone polymer
(PDMS) chip layer 301 and glass substrate layers 302 are bonded.
In conjunction with shown in Fig. 2, Fig. 3, Fig. 4, polydimethylsiloxanechip chip layer 301 includes: sample feeding mouth 1, with sample into
The sample feeding channel 2 that sample mouth 1 connects；Sheath stream injection port 3 is drawn from sheath stream injection port 3, and two sheaths to be formed that are divided into two
Sample intake passage 4 is flowed, the end of two sheath stream sample intake passages 4 is converged in the two sides in sample feeding channel 2 and sample feeding channel 2 respectively
It closes, forms a sample split tunnel 5；Independently at closed circular, be arranged in the first of 5 two sides of sample split tunnel,
Second liquid electrode channel, the first, second liquid electrode channel general structure is identical, include: sequentially connected electrode solution into
Sample mouth 6, electrode solution buffer area 7 and electrode solution liquid storage area 8, unlike, the first liquid electrode channel further includes a channel
Tip wave structure 9, second liquid electrode channel further include the gentle structure 10 in a channel tip.Wherein, channel tip wave
Structure 9 and the gentle structure 10 in channel tip are respectively arranged in the opposite sides thereof of sample split tunnel 5, with to sample split tunnel 5
Apply a high-frequency and high-voltage inhomogeneous field.The polydimethylsiloxanechip chip layer 301 further include: from sample split tunnel 5
End is drawn, and be divided into two the target product collection channel 11 being respectively formed and waste collection channel 12；And respectively with mesh
Mark the outlet 13,13 ' that collection of products channel 11 and waste collection channel 12 connect.
According to the preferred embodiment, flow dividing structure step by step is also devised between target product collection channel 11 and outlet 13
14 and four target product liquid storage chambers 15, as shown in Figure 2.It should be understood that target product liquid storage chamber 15 can be according to reality
Border Demand Design is different numbers or different size, for example, three, four or six, etc..
According to the preferred embodiment, as shown in figure 3, before converging, the width in sample feeding channel 2 and sheath stream sample intake passage 4
Degree becomes narrow gradually.Preferably, the main part in sample feeding channel 2 is 150-200 μm wide, gradually becomes in sheath flow structure part
Narrow, final width becomes 30-40 μm, and the main part of the sheath stream sample intake passage 4 is 100-120 μm wide, in sheath flow structure part
It becomes narrow gradually, final width becomes 20-30 μm.
Preferably, sample feeding channel 2, electrode solution buffer area 7, target product collection channel 11 and waste collection channel
12 are arranged with serpentine channels, as shown in Figure 2.
The preparation of 1 micro-fluidic chip of embodiment
1.1 firstly, be directed to micro-fluidic chip structure division, draw out required structure with AutoCAD drawing software
Figure makes mask, carries out gluing photoetching, reactive ion etching, cleaning of removing photoresist, gluing light by substrate of four cun of monocrystalline silicon pieces
It carves, deep reaction ion etching, etches the height of chip channel, obtain the silicon wafer mold with micro-structure.
1.2 are placed in a vacuum drying oven silicon wafer mold and open centrifuge tube equipped with 10 μ L silicon fluorides, are evacuated to negative
Pressure, vaporizes silicon fluoride, mold stands 5-6 hour in silicon fluoride steam.In ventilation, drying box is opened, ventilation 1
After a hour, by silicon chip extracting.The purpose of this step is to deposit one layer of organic matter in silicon chip surface, convenient for subsequent PDMS chip
Production.
1.3 weigh PDMS prepolymer and curing agent according to weight ratio 10:1 respectively, are then mixed and stirred for uniformly, being placed in true
It is vacuumized in empty drying box, stands 30min under negative pressure.After PDMS substantially without bubble after, be cast on silicon wafer mold, it is quiet
30min is set, then, puts it into 65 DEG C of baking oven heating 1h.Finally the PDMS chip layer being cured is stripped down from mold,
It is punched according to the inlet and outlet on figure, and the redundance other than structure is cut away.Finally by PDMS chip layer 301
Structure is put into plasma cleaner with glass substrate layers 302 up and cleans 1min, fits together rapidly after taking-up, i.e., complete
At the encapsulation of micro-fluidic chip 300.
Embodiment 2 carries out the deflection sorting of different size polystyrene microspheres using micro-fluidic chip prepared by embodiment 1
The packaged micro-fluidic chip 300 of embodiment 1 is put into vacuum kettle and vacuumizes 30min, is taken out later micro-fluidic
Chip 300, with liquid-transfering gun draw 100 μ L DEP conducting solutions, then by pipette tips be inserted into micro-fluidic chip 300 on electrode solution into
Sample mouth 6 stands 10min by the negative pressure in enclosed-electrode channel and DEP conduction liquid is sucked the first, second liquid electrode respectively
In channel.Liquid electrode channel portion is not filled by conduction liquid and the effect after filling conduction liquid as shown in A, B in Fig. 4.
By diameter be 5 μm and 10 μm of polystyrene microspheres clean concussion with deionized water and are centrifuged twice, is then resuspended in me
In the DEP buffer for preparing (deionized water adds PBS containing 0.05%Tween, and conductivity is adjusted to 10ms/m).When sorting, use
Micro syringe draws 100 μ L samples, slides into the tubule of connection micro syringe 200, allows sample solution full of whole
Then tubule pulls up micro syringe 200, pipette samples are spare to 100 μ L again.It is molten that 100 μ LDEP buffering is drawn with liquid-transfering gun
Liquid injection micro-fluidic chip 300 is cleaned, and is drawn DEP buffer with another micro syringe later and is repeated above-mentioned be full of carefully
The flow velocity of sampling pump 100 is set 15mL/h by the step of pipe, and diameter is set as 1.6mm, connects the thin of two micro syringes
Pipe, the other end are inserted into sample feeding mouth 1 and sheath fluid injection port 3 respectively.Micro-fluidic chip 300 is fixed on microscope in the following, pulling out
The DEP conduction liquid sample introduction pipette tips of power down pole channel, toward each piece platinum electrodes of insertion of two electrode solution injection ports 6, platinum electrode it is another
One end connects ATA2161 high-voltage amplifier 500, and amplifier is connected with MHS-5200A signal generator 600, signal generator 600
It is controlled by computer 700 with software kit.
It should be understood that electrode material used herein can also use other metal material insteads with good conductivity,
Such as: copper etc. selects platinum electrode herein, and main cause is that platinum electrode is inert electrode, not oxidizable.
100 start button of sampling pump is opened, after waiting two minutes, liquid stream is stablized, and main channel mesotheca stream forms stabilization, polyphenyl
Ethylene microballoon, which is stablized, flows into waste collection channel 12, is closed the switch of function generator 600 at this time, output parameter is adjusted to
5V, frequency 100KHz reclose ATA2161 switch, and parameter is adjusted to 200 times of amplification, and output mode is difference output.It can see
It observes, after a brief delay, the microballoon for flowing through sample split tunnel 5 starts to deflect to target product collection channel 11, polyphenyl
Ethylene microballoon flows into main channel end sample collection channel, most flows into four sample collection chambers through serpentine channels afterwards.Principle is benefit
After closure electrode switch, electrode is served as with liquid, the main channel region folded by electrode applies the non-homogeneous electricity of a high frequency
, the neutral microparticle for flowing through the region polarizes rapidly, then in inhomogeneous field, not due to both sides electric-field intensity distribution
The phenomenon that uniformly generating deflection.The electric pulse field parameter of function generator output is respectively 0V；2.5V, 100KHz；5V, 100KHz；
And 0V；3V, 100KHZ；6V, 100KHz, waveform are unified for square wave.Experimental result is as shown in A, B, C, D, E, F in Fig. 5.
Embodiment 3
The method that the micro-fluidic chip 300 and embodiment 2 provided using embodiment 1 is provided, by polystyrene microsphere sample
It is substituted for circulating tumor cell H446, circulating tumor cell is cleaned into concussion centrifugation twice with PBS (containing 0.05%Tween), so
It is resuspended in the DEP buffer (8.5% sucrose [wt/vol], 0.3% glucose [wt/vol]) of our preparations afterwards, conductivity tune
For 10ms/m, the electric pulse field parameter of function generator output is 6V, 100KHz, and waveform is unified for square wave.Circulation is swollen after starting electrode
The captured effect of oncocyte H446 is as shown in fig. 6, dielectrophoresis region potential electric field distribution situation simulated effect such as Fig. 7 institute
Show.
Above-described, only presently preferred embodiments of the present invention, the range being not intended to limit the invention, of the invention is upper
Stating embodiment can also make a variety of changes.Letter made by all claims applied according to the present invention and description
Single, equivalent changes and modifications, fall within the claims of the invention patent.The not detailed description of the present invention is normal
Advise technology contents.
Claims (10)
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