CN208921647U - For adherent single celled electrical impedance detecting electrode chip and its electrode array chip - Google Patents
For adherent single celled electrical impedance detecting electrode chip and its electrode array chip Download PDFInfo
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- CN208921647U CN208921647U CN201820258483.9U CN201820258483U CN208921647U CN 208921647 U CN208921647 U CN 208921647U CN 201820258483 U CN201820258483 U CN 201820258483U CN 208921647 U CN208921647 U CN 208921647U
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Abstract
The utility model discloses one kind to be used for adherent single celled electrical impedance detecting electrode chip and its electrode chip array, the contre electrode (1) including being less than or equal to projected area when individual cells suspend；The combined type circle electrode around the contre electrode (1) being arranged on the same base；The contre electrode and combined type circle electrode constitute a pair of of measuring electrode；Combined type circle electrode is constituted with the adherent unicellular required measurement area being gradually increased under growing state.Electrode chip array composed by the measuring electrode that multipair contre electrode and combined type circle electrode are constituted constitutes the measurement area that is gradually increased required in growth, division growth with adherent unicellular array.As cell division is proliferated, the adherent area of cell is gradually expanded, and the combination of electrode changes correspondingly, it can be achieved that the impedance variations of the monoclonal cell group of unicellular formation are detected, from behaviors such as period, the proliferation of analysis of variance procedure cell of electrical impedance.
Description
Technical field
The utility model relates to multiple fields such as biological cell impedance detection, the designing techniques of microelectrode, more particularly to
A kind of anti-detecting electrode chip design of cell resistance.
Background technique
Since there is the knots that heterogeneity, many biological studies are obtained from the average effect of group's cell between cell
Fruit can not provide information useful enough, and therefore, unicellular research becomes hot spot in recent years.
In the diagnosis of many cancers, we it can be found that " key cells " in pathological tissues proliferation, differentiation capability
It is relatively strong, and some can also be shifted, this be cause the one of the major reasons that sb.'s illness took a turn for the worse, so, from individual cell level to cell
The proliferation of state the biological properties such as breaks up, migrates and carry out research and is of great significance.There is researcher to prove, using cell electricity
Resistance technique can be monitored to cell state and study (Kurz et al.) from individual cell level.Due to technology itself
Not damaged, non-marked, and prolonged status monitoring can be carried out, thus unicellular research is carried out using the anti-technology of cell resistance
With unique advantage.
Realize that unicellular electrical impedance detection generally will be to individual cells precise manipulation, efficiently nondestructively single living cell
Capture on the electrode, allows cell to adhere to growth (i.e. adherent) on the electrode, could be realized by detecting to cell resistance is anti-
The monitoring of cell state.This not only needs efficiently to capture cell, it is often more important that, adherent single celled electrical impedance is detected,
The design of electrode is crucial.Existing electrode design can be realized the detection of equal acts and efforts for expediency adherent to cell, but be difficult to realize grow
Time supervision.
Adherent single celled state change can be detected by how allowing electrode to have enough sensitivity, and further be monitored
The biological behaviours such as single celled growth, division growth, cell cycle in long-time, while it being able to satisfy general micro fabrication again
Requirement to glass electrode is laid out enough electrode numbers in limited single level, realizes array type single cell detection,
This is that the emphasis of electrode design is also difficult point, is that the utility model solves the problems, such as.
Utility model content
The utility model aim is to propose one kind for adherent single celled electrical impedance detecting electrode chip and its array, benefit
With combined type circle electrode constitute with it is adherent it is unicellular under growing state needed for the measurement area being gradually increased, and can be with
Electrode array chip is constructed using the electrode system, realizes the electrical impedance detection in attached cell and its growth course.
One kind of the utility model is used for adherent single celled electrical impedance detecting electrode chip, including is less than or equal to single
Projected area contre electrode 1 when cell suspends；The combined type circle electricity around the contre electrode 1 being arranged on the same base
Pole；The contre electrode and combined type circle electrode constitute a pair of of measuring electrode；On the contre electrode, combined type circle electrode respectively
Contact conductor is arranged to connect the pad of electrode and chip edge；A layer insulating is covered on contact conductor；Combined type
It encloses electrode and constitutes the measurement area that is gradually increased required under growing state with adherent individual cells, the combined type circle
Electrode includes at least inner ring electrode 2 and outer ring electrode 3.
One kind of the utility model is used for adherent single celled electrical impedance detecting electrode array chip, including multipair center electricity
Electrod-array composed by the measuring electrode that pole and combined type circle electrode are constituted, is constituted as adherent individual cells array is in life
Long, required in the case of the division growth measurement area being gradually increased；What is be arranged on the same base surrounds the contre electrode
(1) the combined type circle electrode；The contre electrode and combined type circle electrode constitute a pair of of measuring electrode；The center electricity
Contact conductor connection electrode and chip edge pad is respectively set on pole, combined type circle electrode；One layer is covered on contact conductor absolutely
Edge layer；Combined type circle electrode constitutes the measurement area that is gradually increased required under growing state with adherent individual cells,
The combined type circle electrode includes at least inner ring electrode (2) and outer ring electrode (3).
One kind of the utility model is used for adherent single celled electrical impedance detecting electrode chip, including is less than or equal to single
The contre electrode 1 of projected area when cell suspends；The combined type circle electricity around the contre electrode 1 being arranged on the same base
Pole；Contact conductor is respectively set on the contre electrode, combined type circle electrode；The layer insulating covered on contact conductor；Group
Box-like circle electrode is constituted with adherent individual cells after the acquisition, required in the case of growth, division growth to be gradually increased
Measurement area；The combined type circle electrode includes at least inner ring electrode 2 and outer ring electrode 3, the contre electrode 1 and combined type
The AC field applied between circle electrode captures the single thin of suspension for attracting the individual cells to suspend to contre electrode 1
Born of the same parents.
One kind of the utility model is used for adherent single celled electrical impedance detecting electrode array chip, including multipair center electricity
Electrod-array, each contre electrode 1 and each combined type composed by the measuring electrode that pole and combined type circle electrode are constituted
The AC field applied between circle electrode captures multiple suspensions for attracting the individual cells to suspend to each contre electrode 1
Individual cells；It constitutes with adherent individual cells after the acquisition, it is required in the case of growth, division growth to be gradually increased
Measurement area；The combined type circle electrode around the contre electrode 1 being arranged on the same base；The contre electrode
A pair of of measuring electrode is constituted with combined type circle electrode；Contact conductor company is respectively set on the contre electrode, combined type circle electrode
Receiving electrode and chip edge pad；A layer insulating is covered on contact conductor；Combined type circle electrode is constituted with adherent single
The cell measurement area being gradually increased required under growing state, the combined type circle electrode include at least 2 He of inner ring electrode
Outer ring electrode 3.
Compared with prior art, the advantage of the utility model design scheme be when individual cells capture contre electrode simultaneously
When adherent growth, inner ring electrode and contre electrode work, and detect the impedance of individual cells, with cell division, the adherent face of cell
Product is gradually expanded, when to close to inner ring electrode area, outer ring electrode and the work of inner ring electrode, and the Dan Ke that detection individual cells are formed
Grand impedance variations, from behaviors such as period, the proliferation of analysis of variance procedure cell of electrical impedance；
By setting extend outer ring electrode, can be realized when a cell division be proliferated out it is thin under more many cells situation
Born of the same parents' impedance detection.
Using electrod-array can be realized simultaneously the array in region to be measured arrangement it is multiple it is unicellular it is adherent after, growth
And the electrical impedance detection under division growth situation.
Detailed description of the invention
Fig. 1 is a kind of electrical impedance detector electrode structure schematic diagram for attached cell；
Fig. 2 is a kind of insulation layer structure schematic diagram of electrical impedance detecting electrode for attached cell；
Fig. 3 is a kind of electrical impedance detecting electrode chip structure sectional view for attached cell；
Fig. 4 is the electrode array chip overall scheme of multiple Single cell analysis units of the utility model embodiment；Side
Part in frame is structure corresponding with Fig. 1；
Appended drawing reference:
1, contre electrode, 2, inner ring electrode, 3, outer ring electrode, 4, contact conductor；5, insulating layer (corresponding with Fig. 1), 6, nothing
Insulation layer structure, 7, chip base, 8, pad.
Specific embodiment
The embodiments of the present invention is described in further detail below in conjunction with attached drawing.
Face will be described in further detail the embodiments of the present invention in conjunction with attached drawing.
Using the electrical impedance detector electrode structure for attached cell as shown in Figure 1 to Figure 2 as embodiment.Wherein, including
It is contre electrode 1 positioned at most intermediate small electrode, is in addition provided with inner ring electrode 2 and outer ring electrode 3, surrounds center electricity from periphery
The area of pole 1, inner ring electrode 2 and outer ring electrode 3 is gradually increased；In contre electrode 1, the first peripheral electrode 2 and the second periphery electricity
Contact conductor is respectively set in pole 3；Suspension cell is closely sized to contre electrode 1, therefore contre electrode 1 is matched with electrode 2 or electrode 3
Conjunction can capture individual cells on electrode 1, be closely sized to after cell is adherent with electrode 2, at this time electrode 1 and inner ring electrode 2
For detecting；The area to outer ring electrode 3 is gradually expanded in the further division growth of attached cell, area, at this time electrode 1 and electrode
2 are shorted as an electrode with electrode 3 together for detecting.
Specifically, the shape of contre electrode 1 is circle, and diameter is 5~15 microns, and the inner ring electrode 2 is in
The annulus of electrode 1 is entreated, the inner and outer diameter of annulus is 7~50 microns, and wherein internal diameter is bigger in the outer diameter of electrode 1 than center；It is described outer
The shape of electrode 3 is enclosed to be 15~200 microns around the inner and outer diameter of the annulus of inner ring electrode 2, annulus, wherein internal diameter compares inner ring
The outer diameter of electrode 2 is big, the 3 also settable more extension outer rings electrode in periphery of outer ring electrode.
Contre electrode 1, inner ring electrode 2, outer ring electrode 3 are respectively independent, are not connected to mutually；All electrodes are connected by lead
It is connect to chip edge and with back-end impedance detection circuit.
The combination of electrode is adjusted as needed.As a pair of electrodes needed for impedance bioelectrical measurement, Different electrodes
Between can combine to carry out impedance detection, pair-wise combination mode has contre electrode 1 and inner ring electrode 2, contre electrode 1 and outer ring
Electrode 3, inner ring electrode 2 are combined with outer ring electrode 3, inner ring electrode 2 and outer ring electrode 3 are combined with contre electrode 1 again after being shorted, in
Centre electrode 1 and inner ring electrode 2 be shorted after combine again with outer ring electrode 3, contre electrode 1 and outer ring electrode 3 short circuit after again with inner ring
Electrode 2 combines, and this 6 kinds, if expansion electrode can increase accordingly more combinations again for outer ring.It is, adjacent electrode or
Being shorted for Different electrodes the electrode adjacent with its outside or inside can form compound electrode again.
As shown in figure 4, the electrode array chip overall picture of the Single cell analysis unit for the utility model embodiment.
The chip preparation method of the utility model, concrete operations are as follows:
Optimization layout on the glass sheet, design electrode, lead and insulating layer domain:
Contact conductor layout design: in glass electrode edge designs pad, lead；In one electricity of glass-chip central design
Pole array；Electrode domain is designed, the diameter of detection position electrode area is 20~40 μm, in order to guarantee precision and the reduction of processing
Electrode direct impedance, the extra fine wire width of electrode are 5 μm, and lead extra fine wire width is 10 μm.According to electrode, pad, lead pattern
Design insulating layer domain.
Gold electrode, insulating layer are processed on glass-chip according to domain:
1, one layer of titanium (30nm) and gold (300nm) are successively sputtered on clean glass first；
2, using electrode domain (Fig. 1) with photoetching process to metal layer patterning, by gluing, exposure, elution, etching,
Remove photoresist and etc., the electrode pattern needed；
3, SiO is processed by pecvd process in electrode surface2/Si3N4/SiO2Film, then (schemed with insulating layer domain
2), photoetching process patterns the film layer, also pass through gluing, exposure, elution, etch, remove photoresist and etc. obtain figure
The insulating layer of case, so that electrode section is exposed, lead is covered by insulating layer, to improve detection sensitivity, reduces signal interference；
PCB interface board matched with glass-chip is designed, electrode chip is connected on pcb board by scolding tin；
Pad is covered using epoxy AB glue, guarantees to cover after insulation, glue are dried one layer of PDMS mixture and solid on glue
Change, prevents glue to the toxic effect of cell；
One fence is fixed on chip pcb board with PDMS, forms a culture cavity, and mix a transparent plastic
Lid；
After the completion of device fabrication, first stayed overnight with distilled water immersion, after PBS is rinsed well, ultraviolet disinfection；
Pcb board is connected after drawing contact conductor with Impedance Analysis instrument, and realization powers on interpolar impedance detection to chip.
A kind of detection method of electrical impedance detector electrode structure for attached cell of the utility model, this method are specific
Steps are as follows:
It after individual cells are captured electrode surface, is partnered electrode by contre electrode 1 and inner ring electrode 2, carries out electricity
Impedance detection, to detect adherent, the growing state of this individual cells；
When contre electrode 1 and inner ring electrode 2 detect, cell detection has reached threshold value to impedance or microscope is observed carefully
After born of the same parents are completely adherent, after contre electrode 1 and inner ring electrode 2 are become an electrode by respective contact conductor short circuit,
Cell measurement electrode is constituted with outer ring electrode 3 again to be formed to progress impedance detection, or only with inner ring electrode 2 and outer ring electrode 3
Cell measurement electrode is to progress impedance detection；
During cell progress division growth, adherent again, when cell quantity is more and more, the resistance of cell measurement formation
Resist increasing.
When detecting under more cell proliferative conditions that a cell division is proliferated out, outer ring electricity is further set
Pole, and be shorted with contre electrode 1, inner ring electrode 2 and 3 three of outer ring electrode and constitute an electrode, together with extension outer ring electrode
It constitutes a pair of electrodes and carries out impedance detection, inner ring electrode 2, independent 3 electrode of outer ring electrode 3 or outer ring can also be used as one
Electrode constitutes cell measurement electrode pair with outer ring electrode, and so on.
Individual cells are fixed to the embodiment of the method at electrode detection position:
(1) additional chip auxiliary law
PDMS (dimethyl silicone polymer) pipeline with unicellular capturing function is fitted in chip electrode surface, and
It is aligned under microscope, the structure positioning of cell will be captured on electrode (1) surface；
Attached cell is digested and is suspended with cell culture fluid as the cell suspension of certain density；
The PDMS chip processed is attached on clean culture dish bottom surface, after plasma cleaning processing, is first instilled
Not celliferous a small amount of culture solution carefully aspirates from exit with syringe, culture medium is allowed to take pipeline；
Cell is captured, a small amount of cell suspension is instilled from inlet, is aspirated from exit with syringe, microscopically observation is caught
When obtaining effect and reaching requirement, stop suction, be further dropped into cell culture fluid, cleans unwanted cell in entrance and pipeline；
Entrance surplus liquid is sucked out, a large amount of cell culture fluids are added, culture dish is transferred to incubator and carries out cell culture；
To cell it is adherent after, PDMS layer is opened, is added more culture mediums, the culture of individual cells array is carried out and simultaneously examines
Its impedance variations is surveyed, and then analyzes the physiological status situation of change of individual cells level.
(2) electrode dielectric power prize law
Certain density cell suspension is added dropwise on the electrode, then applies certain frequency, voltage between electrode 1 and 2,3
AC field, to attract on individual cells to electrode 1；
Apply certain mobilization force, the unwanted cell not being attracted is washed away；
Stablize and stand chip, remove AC field, allows and cell adherent growth and start impedance bioelectrical measurement.
Claims (6)
1. one kind is used for adherent single celled electrical impedance detecting electrode chip, which is characterized in that single thin including being less than or equal to
The contre electrode (1) of projected area when born of the same parents suspend；The combined type circle around the contre electrode (1) being arranged on the same base
Electrode；The contre electrode and combined type circle electrode constitute a pair of of measuring electrode；Divide on the contre electrode, combined type circle electrode
It She Zhi not contact conductor connection electrode and chip edge pad；A layer insulating is covered on contact conductor；Combined type circle electrode structure
At the measurement area that is gradually increased required under growing state with adherent individual cells, the combined type circle electrode is at least
Including inner ring electrode (2) and outer ring electrode (3).
2. as described in claim 1 a kind of for adherent single celled electrical impedance detecting electrode chip, which is characterized in that electrode
Between constitute a pair of electrodes combination and carry out impedance detection, contre electrode (1), inner ring electrode (2), between outer ring electrode (3) in total
It include following 6 kinds of combinations: contre electrode (1) and inner ring electrode (2) combination, contre electrode (1) and outer ring electrode (3) group
Close, inner ring electrode (2) and outer ring electrode (3) combine, contre electrode (1), inner ring electrode (2) short circuit again with outer ring electrode (3) group
It closes, is combined again with contre electrode (1) after inner ring electrode (2) and outer ring electrode (3) short circuit, contre electrode (1) and outer ring electrode (3)
It is shorted and is combined again with inner ring electrode (2).
3. one kind is used for adherent single celled electrical impedance detecting electrode array chip, which is characterized in that including multipair contre electrode
With combined type circle electrode constitute measuring electrode composed by electrod-array, constitute with adherent individual cells array growth,
The measurement area being gradually increased needed in the case of division growth；What is be arranged on the same base surrounds the contre electrode (1)
The combined type circle electrode；The contre electrode and combined type circle electrode constitute a pair of of measuring electrode；The contre electrode, group
Contact conductor connection electrode and chip edge pad is respectively set on box-like circle electrode；A layer insulating is covered on contact conductor；
The combined type circle electrode composition measurement area that is gradually increased required under growing state with adherent individual cells, described group
Box-like circle electrode includes at least inner ring electrode (2) and outer ring electrode (3).
4. one kind is used for adherent single celled electrical impedance detecting electrode chip, which is characterized in that single thin including being less than or equal to
The contre electrode (1) of projected area when born of the same parents suspend；The combined type circle around the contre electrode (1) being arranged on the same base
Electrode；The pad of contact conductor connection electrode and chip edge is respectively set on the contre electrode, combined type circle electrode；Electrode
A layer insulating is covered on lead；Combined type circle electrode constitutes with adherent individual cells growth, division growth after the acquisition
In the case of needed for the measurement area being gradually increased；The combined type circle electrode includes at least inner ring electrode (2) and outer ring electrode
(3), the AC field applied between the contre electrode (1) and combined type circle electrode, for attracting the individual cells to suspend to arrive
On contre electrode (1), the individual cells of suspension are captured.
5. as claimed in claim 4 a kind of for adherent single celled electrical impedance detecting electrode chip, which is characterized in that electrode
Between constitute a pair of electrodes combination and carry out impedance detection, contre electrode (1), inner ring electrode (2), between outer ring electrode (3) in total
It include following 6 kinds of combinations: contre electrode (1) and inner ring electrode (2) combination, contre electrode (1) and outer ring electrode (3) group
Close, inner ring electrode (2) and outer ring electrode (3) combine, contre electrode (1), inner ring electrode (2) short circuit again with outer ring electrode (3) group
It closes, is combined again with contre electrode (1) after inner ring electrode (2) and outer ring electrode (3) short circuit, contre electrode (1) and outer ring electrode (3)
It is shorted and is combined again with inner ring electrode (2).
6. one kind is used for adherent single celled electrical impedance detecting electrode array chip, which is characterized in that including multipair contre electrode
Electrod-array, each contre electrode (1) and each combined type composed by the measuring electrode constituted with combined type circle electrode
The AC field applied between circle electrode captures multiple outstanding for attracting the individual cells to suspend to each contre electrode (1)
Floating individual cells；It constitutes with adherent individual cells after the acquisition, required gradually increasing in the case of growth, division growth
Big measurement area；The combined type circle electrode around the contre electrode (1) being arranged on the same base；The center
Electrode and combined type circle electrode constitute a pair of of measuring electrode；Electrode is respectively set on the contre electrode, combined type circle electrode to draw
Line connection electrode and chip edge pad；A layer insulating is covered on contact conductor；Combined type circle electrode is constituted with adherent
The individual cells measurement area being gradually increased required under growing state, the combined type circle electrode include at least inner ring electrode
(2) and outer ring electrode (3).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201820258483.9U CN208921647U (en) | 2018-02-14 | 2018-02-14 | For adherent single celled electrical impedance detecting electrode chip and its electrode array chip |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201820258483.9U CN208921647U (en) | 2018-02-14 | 2018-02-14 | For adherent single celled electrical impedance detecting electrode chip and its electrode array chip |
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| CN208921647U true CN208921647U (en) | 2019-05-31 |
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| CN201820258483.9U Active CN208921647U (en) | 2018-02-14 | 2018-02-14 | For adherent single celled electrical impedance detecting electrode chip and its electrode array chip |
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2018
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