CN106124388A - Capillary sample inlet system and sample injection method, unicellular electrology characteristic detecting system - Google Patents
Capillary sample inlet system and sample injection method, unicellular electrology characteristic detecting system Download PDFInfo
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- CN106124388A CN106124388A CN201610407657.9A CN201610407657A CN106124388A CN 106124388 A CN106124388 A CN 106124388A CN 201610407657 A CN201610407657 A CN 201610407657A CN 106124388 A CN106124388 A CN 106124388A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
- G01N15/10—Investigating individual particles
- G01N15/1031—Investigating individual particles by measuring electrical or magnetic effects
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
- G01N15/10—Investigating individual particles
- G01N2015/1006—Investigating individual particles for cytology
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Abstract
The invention provides a kind of capillary sample inlet system and sample injection method, unicellular electrology characteristic detecting system.This capillary sample inlet system includes: syringe pump, capillary tube, micro-fluidic chip and negative pressure module, sets gradually pressure channel, cell recovery approach and negative pressure channel in micro-fluidic chip, and three forms the microchannel of micro-fluidic chip；The front end of pressure channel is connected to the liquid outlet of microchannel as the inlet of microchannel, its rear end by cell recovery approach and negative pressure channel, and negative pressure module is connected to the liquid outlet of this microchannel；The rear end of capillary tube is connected to syringe pump, controlled to carry out suction and the discharge of cell suspension by syringe pump, the cell suspension that capillary tube instills in inlet side, microchannel, under the suction function that negative pressure module provides, cell suspension sequentially enters pressure channel and cell recovery approach.The present invention can effectively reduce the loss rate of sample, it is achieved high-recovery, may be used for the electrology characteristic detection of the cell sample of cell concentration and rareness thereof.
Description
Technical field
The present invention relates to microfluidic art, particularly relate to a kind of capillary sample inlet system and sample injection method, unicellular
Electrology characteristic detecting system.
Background technology
Unicellular electrology characteristic, as cell membrane is than electric capacity and cytoplasm electric conductivity, for understanding that cell function and state have
Very important meaning and potential value.As a kind of cell characteristic without mark, it can be used for defining cell
And classification.
In the research of unicellular electrology characteristic, cell is equivalent to a capacitance resistance network, wherein phospholipid bilayer knot
The cell membrane of structure is equivalent to electric capacity, and the predominantly Cytoplasm of electrolyte is equivalent to resistance.Differ in view of cell size, make
Intercellular comparison just can be carried out by unit are cell membrane capacitance and the cytoplasm electric conductivity independent of cell size.
Tradition studies the method for unicellular electrology characteristic mainly patch-clamp etc., can obtain cell membrane than electric capacity, but
Testing efficiency is the lowest, thus flux is relatively low.
Microflow control technique is one important science and technology in this century, due to its channel size and the linear chi of cells of mamma animals
Very little comparable, can conveniently carry out the manipulation of cell, have been widely used for the research of cell biophysics.At micro-fluidic chip
On, generally use microoperation method and drive cell clamp between electrode or flow, by inter-electrode impedance in test process
Change the anti-electrology characteristic pushing away cell.
But, when using micro-fluidic chip to carry out the research of unicellular electrology characteristic in the prior art, generally use and suck
Cell is clamped in interelectrode method, although relatively traditional method flux improves, but the lowest.Between electrode, flowing is micro-
The method of impedance flow cytometry, becomes by allowing cell measure impedance when flowing through in the passage of electrode is placed in one or both sides
Change.In this kind of method, the method using runner to be uncompressed passage (i.e. channel cross-sectional area is less than cell sectional area size), although
Flux has the biggest lifting, but it is very big to be because electrical leak between electrodes stream in test system, and lacks electrical model, it is impossible to obtain solely
Stand on the unicellular electrology characteristic of cell size.
Additionally, pure microflow control technique based on pressure channel, driven the cell stream in micro-fluidic sample intake passage by negative pressure
Cross sectional area less than during pressure channel, measure cell tensile length and pressure channel two terminal impedance, in conjunction with electrical model
Obtain unicellular electrology characteristic parameter, although the unicellular electrology characteristic independent of cell size that can obtain certain flux is joined
Number, but be because fluid dead band in micro-fluidic sample intake passage and exist, cause cell loss rate the highest, need input ratio measurement to arrive
The number of cells of number of cells two to three orders of magnitude big, and the cell kinds such as a lot of cells, such as circulating tumor cell (CTCs)
Class is rare, it is impossible to provide a large amount of cell for test.
Therefore, extremely low unicellular independent of cell size that can obtain again having statistical significance of a kind of Loss Rate is sought
Electrology characteristic detecting system is the most with practical value and practical significance.
Summary of the invention
(1) to solve the technical problem that
In view of above-mentioned technical problem, the invention provides a kind of capillary sample inlet system and sample injection method, unicellular electricity
Characteristics Detection system, to improve detection flux, reduces Loss Rate.
(2) technical scheme
One aspect of the embodiment of the present invention provides a kind of capillary sample inlet system.This capillary sample inlet system includes:
Syringe pump, capillary tube, micro-fluidic chip and negative pressure module, wherein: set gradually pressure channel in micro-fluidic chip, cell reclaims
Passage and negative pressure channel, three forms the microchannel of micro-fluidic chip；The front end of pressure channel as the inlet of microchannel, its
Rear end is connected to the liquid outlet of microchannel by cell recovery approach and negative pressure channel, and negative pressure module is connected to going out of this microchannel
Liquid mouth；The rear end of capillary tube is connected to syringe pump, syringe pump control to carry out suction and the discharge of cell suspension, and capillary tube is micro-
The cell suspension that passage inlet side instills, under the suction function that negative pressure module provides, cell suspension sequentially enters compression
Passage and cell recovery approach.
Another aspect of the embodiment of the present invention additionally provides the sample injection method of a kind of capillary sample inlet system.This sample introduction side
Method includes: sucked in capillary tube by cell suspending liquid；And capillary tube is moved near the inlet of pressure channel discharge liquid
Body.
Another aspect of the embodiment of the present invention additionally provides a kind of unicellular electrology characteristic detecting system.This unicellular electricity
Learning Characteristics Detection system to include: above-mentioned capillary sample inlet system, wherein, micro-fluidic chip includes: transparent substrates and tight with it
The supporting body of close combination, forms pressure channel, cell recovery approach and negative pressure channel in supporting body；Image detection module, is used for
In the bottom surface of micro-fluidic chip, through the situation by pressure channel of the cell in transparent substrates observation of cell suspension；Impedance is surveyed
Amount module, its inlet in microchannel and liquid outlet are connected to electrode, for carrying out the impedance operator of cell suspension
Measure.
(3) beneficial effect
From technique scheme it can be seen that capillary sample inlet system of the present invention and sample injection method, unicellular electrology characteristic
Detecting system has the advantages that
(1) microflow control technique is combined with capillary tube microoperation technology, by capillary tube, sample is directly injected into use
Pressure channel mouth in the micro-fluidic chip detecting unicellular electrology characteristic, it is achieved that the detection of unicellular electrology characteristic.In inspection
Survey flux and detection data validity on it has been reported that suitable based on the pure micro-fluidic detection technique comprising pressure channel.
(2) with it has been reported that based on compared with comprising the pure detection method of microfluidic chip of pressure channel, based on capillary tube
The method of sample introduction can effectively reduce the loss rate of sample, it is achieved and high-recovery (or claim extremely low Loss Rate, i.e. think that measurement arrives
The number of cells of number of cells and input is a magnitude), the electricity of the cell sample that may be used for cell concentration and rareness thereof is special
Property detection, this be it has been reported that can not accomplish based on the pure micro-fluidic detection technique comprising pressure channel.
Accompanying drawing explanation
Fig. 1 is the structural representation according to the embodiment of the present invention unicellular electrology characteristic detecting system based on capillary sample inlet
Figure；
Fig. 2 is the flow chart of facture of microchip process in unicellular electrology characteristic detecting system shown in Fig. 1；
Fig. 3 is to perform the schematic diagram of device cross-section after each step during facture of microchip shown in Fig. 2；
Fig. 4 is the flow chart of unicellular electrology characteristic detecting system operational approach shown in Fig. 1；
Fig. 5 is the flow chart in unicellular electrology characteristic detecting system operational approach shown in Fig. 4 carried out micro-fluidic chip.
Detailed description of the invention
Microfluidic chip technology is combined by the present invention with capillary tube microoperation technology, by using capillary tube directly to draw
The pressure channel injection port of cell sample introduction to be measured to micro-fluidic chip is directly carried out electrology characteristic by the few cells suspension concentrated
Detection, devising one can be high-throughout, and high-recovery is applicable to the unicellular electrology characteristic detection of very small amount cell and is
System.
For making the object, technical solutions and advantages of the present invention clearer, below in conjunction with specific embodiment, and reference
Accompanying drawing, the present invention is described in more detail.
One aspect of the embodiment of the present invention provides the detection of a kind of unicellular electrology characteristic based on capillary sample inlet
System.Refer to Fig. 1, this unicellular electrology characteristic detecting system includes: syringe pump, capillary tube, micro-fluidic chip, negative pressure module,
Impedance measurement module and image detection module.
Wherein, pressure channel in the horizontal direction and cell recovery approach are set in micro-fluidic chip, and along vertically
The negative pressure channel in direction.Pressure channel, cell recovery approach and the microchannel of negative pressure channel composition micro-fluidic chip.Wherein, pressure
The front end of contracting passage is connected to microchannel as the inlet of microchannel, its rear end by cell recovery approach and negative pressure channel
Liquid outlet.Negative pressure module provides the driving force of flowing for the cell suspension flowing through microchannel.
The cell suspension that capillary tube instills in inlet side, microchannel, provides in the negative pressure module being positioned at negative pressure channel side
Suction function under, cell suspension sequentially enters pressure channel and cell recovery approach.Image detection module is at micro-fluidic chip
Bottom surface observe the situation by pressure channel of the cell in cell suspension.The impedance measurement module inlet in microchannel and going out
Liquid mouth is connected to electrode, measures the impedance operator of cell suspension.
Each ingredient of electrology characteristic detecting system unicellular to the present embodiment is described in detail below.
In the present embodiment, capillary tube refers to the pipeline of diameter very thin (generally less than 1mm), its material generally glass or have
Organic polymeric material, the internal diameter in thick portion, its upper end is between 0.5mm～1mm, and the internal diameter in thin portion, lower end is between 20 μm～60 μm
Between (ensure more than cell dia)；The appearance liquid measure in thin portion, front end is between 10 μ L-100 μ L, and it has can manipulate μ L level liquid
The feature of amount.Wherein, capillary tube is mounted in three-dimensional movement platform, and rear end is connected to syringe pump.In real work,
Capillary exit can be moved near the inlet of microchannel by three-dimensional movement platform, and the syringe pump connected by rear end is carried out carefully
The suction of born of the same parents' suspension and discharge.
Refer to Fig. 1, micro-fluidic chip includes: transparent substrates and with the supporting body combined closely on substrate.At supporting body
Pressure channel, cell recovery approach and the negative pressure channel that interior formation is above-mentioned.
Transparent substrates can be that (polydimethylsiloxane is called for short for sheet glass, microscope slide or polydimethylsiloxane
PDMS) the transparent sheet-like material such as sheet, above-mentioned material ensure that in the lower section of pressure channel and cell recovery approach be transparent, from
And utilize microscope or the video camera can the mobility status of observation of cell easily.Certainly, if only needing to measure unicellular electricity
Learning characteristic, without concern for the mobility status of cell, this substrate can also use other non-transparent material to make.
In the present embodiment, the material of supporting body is PDMS.It will be apparent to those skilled in the art that in addition to PDMS, also
Can use lucite, the transparent plastic material such as SU-8 is molded the above-mentioned supporting body of formation.
Supporting body is by PDMS material injection mo(u)lding, and pressure channel, cell recovery approach and negative pressure channel are in injection mo(u)lding
During formed.Wherein, pressure channel, cell recovery approach and the circular in cross-section of negative pressure channel or ellipse.
Single celled diameter is generally 10 μm～30 μm, and correspondingly, its cross sectional dimensions is between 80 μm2～700 μm2Between.
From the point of view of pressure channel, its cross-sectional area is about long-pending (the about 40-600 μm of cell cross section to be measured2) 40%-90%, and
For cell recovery approach and negative pressure channel, its cross sectional dimensions is more than single celled cross sectional dimensions.Wherein, compression is logical
Road and cell recovery approach are horizontally disposed, and the negative pressure channel that the end of cell recovery approach is by near vertical direction
25 connect up the liquid outlet to micro-fluidic chip.
In the present embodiment, micro-fluidic chip is double-deck pressure channel based on PDMS, uses fine process to make.With
Under to this cell electrology characteristic detection micro-fluidic chip manufacture method be described in detail, refer to Fig. 1 and Fig. 2, this making
Method includes:
Step S202: photoresist SU-8 5 makes Seed Layer；
Microscope slide cleans in acetone, ethanol and deionized water successively, dries rear surface Rotating with Uniform one layer of SU-8 of coating
5, exposure forms Seed Layer.
Step S204: photoresist SU-8 5 makes pressure channel layer；
One layer of SU-8 5 of even application, puts mask exposure the most again, as shown in (A) in Fig. 3.
Step S206: photoresist SU-8 25 makes cell recovery approach layer；
One layer of SU-8 25 of even application on SU 8-5, puts mask plate exposure, as shown in (B) in Fig. 3；
Step S208: development, forms pressure channel and the SU-8 formpiston of cell recovery approach；
Use SU-8 developer solution that photoresist is developed, form the double-layer structure of pressure channel and cell recovery approach
SU-8 formpiston, in its structure such as Fig. 3 shown in (C).
The cast of step S210:PDMS, overmolded, punching；
The double-deck SU-8 formpiston of use as die casting PDMS, solidification after overmolded obtain pressure channel and
The PDMS overmolded of cell recovery approach, in one end punching that cell recovery approach is not connected with pressure channel, forms negative pressure channel,
In its structure such as Fig. 3 shown in (D).
The punching of step S212:PDMS is cut with pressure channel mouth；
PDMS device is cut along pressure channel mouth, by pressure channel one end open, in later experiments
Suck cell and carry out unicellular electrology characteristic detection, in its structure such as Fig. 3 shown in (E).
Step S214:PDMS is bonded with glass, forms micro-fluidic chip；
After the PDMS device clean of cutting, it is bonded with sheet glass, forms device, in its structure such as Fig. 3 shown in (F).
Refer to Fig. 1, in the present embodiment, negative pressure module includes: airtight flexible pipe and negative pressure source.Wherein, airtight flexible pipe is
One T-shaped flexible pipe, its first end inserts in negative pressure channel, and the second end is connected to negative pressure source, and three-terminal link is to fluid outlet side.?
Under the suction function that negative pressure source provides, described pressure channel is passed through in cytomorphosis to be measured, then enters cell recovery approach.
In the present embodiment, image detection module is used for observing cell to be measured in shooting testing sample and passes through under suction function
The process of described pressure channel.Refer to Fig. 1, this image detection module includes: biological inverted microscope and photographic head.Wherein,
The image-forming component of biological inverted microscope is directed at described pressure channel from the back side of transparent glass substrate, for by small passage
Image and cell image etc. are converted to can the image size of camera-shot.The mesh of the biological inverted microscope of photographic head alignment
Mirror, the image during being deformed by described pressure channel under suction function by biological inverted microscope record cell
Information.
In the present embodiment, refer to Fig. 1, the first measurement electrode of impedance measurement module stretches at the inlet of microchannel
In testing sample, second measures electrode is stretched in the testing sample in negative pressure channel by airtight flexible pipe.Measured by the two
Electrode, impedance measurement module records corresponding low frequency and high frequency during cell to be measured deforms by pressure channel under suction function
The time dependent waveform of impedance of two Frequency point pressure channel both sides.
In the present embodiment, the amplification of biological inverted microscope is 400 times, and the scanning speed of photographic head is 25 frames/second.
The sample frequency of electric impedance analyzer is 25 sampled points per second, and measuring frequency is 1kHz and 100kHz.
In based on pressure channel the unicellular electrology characteristic detection method of prior art, generally comprise: Preparatory work of experiment, former
Beginning impedance and image data acquiring and data process three steps.Wherein, sample preparation mainly about 1 milliliter concentration of preparation is
106The cell suspension of individual/ml, on-line measurement mainly includes that suspension directly uses liquid-transfering gun to instill the hand-hole of micro-fluidic chip
In, it being acquired the associative operations such as cycle afterwards, it is that the initial data of on-line measurement is combined data processing model that data process
Process with software platform, obtain single celled cell membrane than electric capacity and cytoplasm electric conductivity.
And utilize the unicellular electrology characteristic detecting system of the present embodiment, have only to prepare 10 microlitres in the suspension preparatory stage
The concentration of left and right is 104-106The cell suspension of individual/ml, sucks in capillary tube, then in test process by controlling syringe pump
Synchronize sample introduction.Specifically, refer to Fig. 4, the operational approach of this unicellular electrology characteristic detecting system is as follows:
Step S402: prepare cell suspending liquid；
It is suspended in cell culture fluid or phosphate buffer (phosphate after cell is broken up in the cell sample of acquirement
Buffered saline, is called for short PBS) in.Suspension is loaded in centrifuge tube, use centrifuge to be centrifuged making cell aggregation
Bottom centrifuge tube, carrying out concentration allotment according to general number of cells after removing the supernatant, concentration is arranged in 104Individual/ml
To 106Between individual/ml.
Step S404: the drop of cell suspension is dropped to the inlet of microchannel；
Refer to Fig. 5, this step specifically includes:
Sub-step S404a: use liquid-transfering gun to be dropped on hydrophobic surface by cell suspension drop；
This sub-step is particularly as follows: the cell suspension drop after using liquid-transfering gun to take 10 microlitres allotments drops in and facilitates capillary tube to inhale
On the surface taken.In order to reduce cell loss in transfer process as far as possible, select hydrophobic surface to be advisable as far as possible, as
Parafilm etc..
Sub-step S404b: the front end of capillary tube is inserted in cell suspension drop；
This sub-step is particularly as follows: on the syringe that is connected on by capillary tube thick tube part on device and syringe pump, thin head inserts
In cell suspension drop.
Sub-step S404c: be adjusted to by syringe pump as decimation pattern, sucks cell suspension drop in capillary tube；
This sub-step specifically includes: after being adjusted to by syringe pump as " extraction (refill) " pattern, that is taken out by liquid-transfering gun is thin
Born of the same parents' suspension drop all sucks in capillary tube.In order to eliminate the capillary tube time decalage in extraction and push, in suction
Before, capillary tube and the solution such as rear end pipeline and syringe use pure water can be filled full, suck fluid afterwards by filling liquid
Isolate with cell suspension, form filling liquid-fluid-cell suspension three-decker.
Sub-step S404d: capillary tube is moved discharge liquid near the inlet of pressure channel；
This sub-step specifically includes: be fixed in three-dimensional movement platform by capillary tube, utilizes this three-dimensional movement platform by hair
Tubule moves near pressure channel mouth discharge liquid；
So far, it is achieved the drop of cell suspension is dropped to the inlet of microchannel.
Step S406: debug image detection module, micro-fluidic chip, impedance measurement module and negative pressure module；
First, the micro-fluidic chip of preparation is loaded on the object stage of biological inverted microscope, makes in micro-fluidic chip
Pressure channel be partially in the middle of field of microscope；
Then, the cell culture fluid (or PBS) that cell suspension is identical is made the inlet from microchannel injects capillary tube
Fill microfluidic channel, remove the bubble in passage.And drip a dropping liquid in the outside of inlet and drip, flood inlet, be used for hindering
Electrode is inserted during anti-measurement.
After again, build in the measuring instrument in impedance measurement module, the video camera in image detection module and negative pressure module
The equipment such as pressure calibration instrument, this part is with consistent based on micro-fluidic unicellular electricity characteristic measuring mode before.
Finally, an electrode in impedance measurement module is contained in the T-shaped pipeline of negative pressure module insertion micro-fluidic chip
Liquid outlet, it is ensured that electrode effectively contacts with solution；Another root electrode of impedance measurement module inserts micro-fluidic chip inlet
In the drop of side, the liquid in microfluidic channel and electrode is made to form measuring circuit path.
So far, Preparatory work of experiment completes.
Step S408: carry out the electrology characteristic test of cell to be measured；
First, the pattern adjusting the syringe pump connecting capillary tube is " injection (inject) ", starts to release in capillary tube
Cell suspension, examines under a microscope simultaneously and whether begins with cell outflow in capillary tube.
When capillary tube begins with cell flow out, click on unicellular electrology characteristic test platform software start measure by
Button, proceeds by impedance data and image data acquiring.After cell suspension all flows out in capillary tube, stop test.
Repeat step S408, until all cells test sample completes or the number of cells measured reaches required
Number.
Step S410: the initial data obtained based on data processing module processes, obtains unicellular cells film than electricity
Hold and cytoplasm electric conductivity.
The concrete grammar processed about data, the pertinent literature being referred in this area, the most no longer describe in detail.
Another aspect of the embodiment of the present invention also provides for a kind of capillary sample inlet system.This capillary sample inlet system bag
Include: syringe pump, three-dimensional movement platform, capillary tube, micro-fluidic chip and negative pressure module.Wherein, set gradually in micro-fluidic chip
Pressure channel, cell recovery approach and negative pressure channel, three forms the microchannel of micro-fluidic chip；The front end of described pressure channel
As the inlet of microchannel, its rear end is connected to the liquid outlet of microchannel by cell recovery approach and negative pressure channel, described
Negative pressure module is connected to the liquid outlet of this microchannel.The rear end of capillary tube is connected to described syringe pump, described syringe pump control
Carrying out suction and the discharge of cell suspension, the cell suspension that described capillary tube instills in inlet side, microchannel, at negative pressure mould
Under the suction function that block provides, cell suspension sequentially enters pressure channel and cell recovery approach.Capillary tube is fixed in described
In three-dimensional movement platform.
In the present embodiment, micro-fluidic chip includes: transparent substrates and the supporting body combined closely with it, in described supporting body
Form described pressure channel, cell recovery approach and negative pressure channel.Pressure channel and cell recovery approach are in the horizontal direction, described
Negative pressure channel is vertically.Further, the cross section of pressure channel, cell recovery approach and negative pressure channel is circular or oval
Shape；The cross-sectional area of pressure channel is the 40%-90% of the cross-sectional area of cell to be measured, and described cell recovery approach and negative pressure are led to
The cross-sectional area in road is more than the cross-sectional area of cell to be measured.
It can be seen that the capillary sample inlet system of the present embodiment is a upper embodiment unicellular electrology characteristic detection in fact it is
For a part for sample introduction in system.About the details of this capillary sample inlet system structure, it is referred to an embodiment
Explanation.
The sample injection method of application given below above-mentioned capillary sample inlet system.This sample injection method includes:
Liquid-transfering gun is used to be dropped on hydrophobic surface by cell suspension drop；
The front end of described capillary tube is inserted in cell suspension drop；
Described syringe pump is adjusted to into decimation pattern, cell suspension drop is sucked in capillary tube；And
Capillary tube is moved discharge liquid near the inlet of pressure channel.
Preferably, also included before cell suspension drop is sucked the step in capillary tube: suck in capillary tube and fill out
Topping up；And in capillary tube, suck fluid；Wherein, after just cell suspension drop sucks in capillary tube, described capillary tube
Middle formation filling liquid-fluid-cell suspension three-decker.
Equally, about the detailed content of this sample injection method, it is referred to the explanation of an embodiment, the most no longer describes in detail.
So far, already in connection with accompanying drawing, the present embodiment has been described in detail.According to above description, those skilled in the art
Capillary sample inlet system of the present invention and sample injection method, unicellular electrology characteristic detecting system should be had and clearly recognize.
Additionally, the above-mentioned definition to each element and method is not limited in various concrete structures, the shape mentioned in embodiment
Shape or mode, it can be changed or replace, such as by those of ordinary skill in the art simply:
(1) capillary drive part is not limited to syringe pump, it is possible to use the modes such as air pressure driving.
(2) capillary tube selects use or do not use filling liquid to be filled with.If carried out filling, can choose whether
Oil droplet or other hydrophobic liquids is used to isolate.
(3) droplet size of capillary sample inlet is not limited to 10 microlitres, can the most suitably regulate.
(4) structures such as the pressure channel cross section in micro-fluidic chip is not limited to rectangle, could alternatively be trapezoidal, circular；
Negative pressure channel mouth is also not limited to circle, could alternatively be square, triangle etc..
(5) impedance measurement module is not limited to lock-in amplifier and functional generator, could alternatively be based on data acquisition
The virtual instrument etc. of card.
Additionally, it will be appreciated by persons skilled in the art that the demonstration that can provide the parameter comprising particular value herein, but this
A little parameters are worth equal to corresponding without definite, but can be similar to analog value in acceptable error margin or design constraint.
Further, the direction term mentioned in embodiment, such as " on ", D score, "front", "rear", "left", "right" etc., be only with reference to accompanying drawing
Direction, not be used for limit the scope of the invention.In embodiment of the method, unless specifically described or must sequentially occur
Step, the order of above-mentioned steps there is no and is limited to listed above, and can change according to required design or rearrange.Above-mentioned
Embodiment can be based on design and the consideration of reliability, and the collocation that is mixed with each other uses or uses with other embodiment mix and match, i.e.
Technical characteristic in different embodiments can freely form more embodiment.
In sum, microfluidic chip technology is combined by the present invention with capillary tube microoperation technology, it is provided that one can
High-throughout, high-recovery, it is applicable to unicellular electrology characteristic detecting system and the operational approach thereof of very small amount cell, tool
There are stronger practical value and application prospect.
Particular embodiments described above, has been carried out the purpose of the present invention, technical scheme and beneficial effect the most in detail
Describe in detail bright, be it should be understood that the specific embodiment that the foregoing is only the present invention, be not limited to the present invention, all
Within the spirit and principles in the present invention, any modification, equivalent substitution and improvement etc. done, should be included in the guarantor of the present invention
Within the scope of protecting.
Claims (10)
1. a capillary sample inlet system, it is characterised in that including: syringe pump, capillary tube, micro-fluidic chip and negative pressure module,
Wherein:
Setting gradually pressure channel, cell recovery approach and negative pressure channel in described micro-fluidic chip, three forms micro-fluidic core
The microchannel of sheet；Cell recovery approach and negative pressure are passed through as the inlet of microchannel, its rear end in the front end of described pressure channel
Passage is connected to the liquid outlet of microchannel, and described negative pressure module is connected to the liquid outlet of this microchannel；
The rear end of described capillary tube is connected to described syringe pump, described syringe pump control carry out the suction of cell suspension and tell
Go out, the cell suspension that described capillary tube instills in inlet side, microchannel, under the suction function that negative pressure module provides, cell
Suspension sequentially enters pressure channel and cell recovery approach.
Capillary sample inlet system the most according to claim 1, it is characterised in that the internal diameter in thick portion, described capillary tube upper end is situated between
Between 0.5mm～1mm, the internal diameter in thin portion, lower end is between 20 μm～60 μm, and the appearance liquid measure in thin portion, lower end is between 10 μ L-100 μ L
Between.
Capillary sample inlet system the most according to claim 1, it is characterised in that also include: three-dimensional movement platform；
Wherein, described capillary tube is fixed in described three-dimensional movement platform.
Capillary sample inlet system the most according to any one of claim 1 to 3, it is characterised in that described pressure channel and
In the horizontal direction, described negative pressure channel is vertically for cell recovery approach.
Capillary sample inlet system the most according to any one of claim 1 to 3, it is characterised in that described pressure channel, thin
The cross section of born of the same parents' recovery approach and negative pressure channel is circular or oval；
The cross-sectional area of described pressure channel is the 40%-90% of the cross-sectional area of cell to be measured, described cell recovery approach and
The cross-sectional area of negative pressure channel is more than the cross-sectional area of cell to be measured.
6. apply the sample injection method of capillary sample inlet system according to any one of claim 1 to 5 for one kind, it is characterised in that bag
Include:
Cell suspending liquid is sucked in capillary tube；And
Capillary tube is moved discharge liquid near the inlet of pressure channel.
Sample injection method the most according to claim 6, it is characterised in that described cell suspending liquid is sucked the step in capillary tube
Suddenly include:
Liquid-transfering gun is used to be dropped on hydrophobic surface by cell suspension drop；
The front end of described capillary tube is inserted in cell suspension drop；And
Described syringe pump is adjusted to into decimation pattern, cell suspension drop is sucked in capillary tube.
Sample injection method the most according to claim 7, it is characterised in that described by cell suspension drop suction capillary tube
Also include before step:
Filling liquid is sucked in capillary tube；And
Fluid is sucked in capillary tube；
Wherein, after just cell suspension drop sucks in capillary tube, described capillary tube is formed filling liquid-fluid-cell and hangs
Liquid three-decker.
9. a unicellular electrology characteristic detecting system, it is characterised in that including:
Capillary sample inlet system according to any one of claim 1 to 5, wherein, described micro-fluidic chip includes: transparent substrates
And in the supporting body combined closely with it, described supporting body, form described pressure channel, cell recovery approach and negative pressure channel；
Image detection module, in the bottom surface of described micro-fluidic chip, through in described transparent substrates observation of cell suspension
The cell situation by pressure channel；
Impedance measurement module, its inlet in microchannel and liquid outlet are connected to electrode, for the resistance to cell suspension
Anti-characteristic measures.
Unicellular electrology characteristic detecting system the most according to claim 9, it is characterised in that described capillary sample inlet system
In system, described negative pressure module includes: airtight flexible pipe and negative pressure source；
Wherein, described airtight flexible pipe is a T-shaped flexible pipe, and its first end inserts in the negative pressure channel of described micro-fluidic chip, and it is the years old
Two ends are connected to negative pressure source, and its 3rd end for inserting the liquid of cell recovery approach by the second electrode of impedance measurement module
In；
Wherein, during the first electrode of described impedance measurement module inserts the cell suspension of inlet side, microchannel.
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