CN209098700U - A kind of intermittent streaming electrotransfection device - Google Patents
A kind of intermittent streaming electrotransfection device Download PDFInfo
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- CN209098700U CN209098700U CN201821508609.XU CN201821508609U CN209098700U CN 209098700 U CN209098700 U CN 209098700U CN 201821508609 U CN201821508609 U CN 201821508609U CN 209098700 U CN209098700 U CN 209098700U
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- tube body
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- liquid
- electrotransfection
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Abstract
The utility model discloses a kind of intermittent streaming electrotransfection devices, including tube body, first electrode, second electrode, piston, it is characterized in that, the first electrode, second electrode is placed in the inner wall of the tube body in parallel, sealed end is arranged in one end of the tube body, the piston that the other end setting of the tube body can be relatively moved along the tube body, the inside of the piston and tube body has the cavity for accommodating liquid sample, at least one liquid entrance is arranged in the tube body and at least one liquid passes in and out two way valve, the liquid entrance is located at the sealed end of the tube body, the liquid disengaging two way valve is located at the tube body liquid entrance.Streaming electrotransfection device disclosed by the utility model provides a kind of device of large volume electrotransfection sample solution, and by way of moveable piston pressurization, inhibit to generate bubble during electrotransfection, improve field uniformity, so as to improve the stability of electrotransfection device, electrotransfection efficiency is improved.
Description
Technical field
The utility model relates to electrotransfection fields, are related to a kind of intermittent streaming electrotransfection device, more specifically, this reality
It is related to a kind of intermittent streaming electrotransfection device for inhibiting electrotransfection to generate bubble by way of the mobile pressurization of piston with novel.
Background technique
Cell membrane is the thin film for being enclosed in pericellular, be cell with the external world carry out selective substances exchange it is penetrating
Property barrier.Cell membrane makes cell become an independent living unit, and possesses a metastable interior environment.Ambient enviroment
In some substances can be by cell membrane, other substances are then not all right.Cell can be absorbed by cell membrane from ambient enviroment
Nutriment is discharged metabolite, the transhipment of substance is made to reach equilibrium state.So the basic function of cell membrane is exactly to maintain cell
Interior microenvironment relatively stable simultaneously selectively carries out mass exchange with external environment.
The study found that if applying the electro photoluminescence of some strength to cell and continuing for some time, so that it may inducing cell
Some micropores are generated on film, enhance the permeability of cell, and so-called cell electroporation (Electroporation) just refers to cell
Under the action of adding impulse electric field outside, the biophysics process of instantaneous micropore is formed on cell membrane lipid bilayer.Electrotransfection
It (Electrotransfection) is to be imported the large biological molecule of external source, such as DNA, RNA or protein using electroporation technology
The technology of cell.When electroporation occurs for cell membrane, permeability and membrane conductance can instantaneously increase, and make hydrophilic molecules, DNA, egg
White matter, virion, drug granule etc. cannot be able to enter cell by micropore by the molecule of cell membrane under normal circumstances.?
After removing electro photoluminescence in short time, micropore disappears on cell membrane, and cell membrane becomes selective permeation barrier again.
Compared with traditional chemical transfection and virus transfection, due to electrotransfection with no chemical contamination, cell will not be made
At permanent damage, efficiency is higher the advantages that, have in fields such as biophysics, molecular biology, clinical medicine wide
Application prospect.
Although the mechanism of electrotransfection effect is not entirely clear that, cell electrotransfection herein is well known, including thin
The movement of after birth lipid bilayer causes to form temporary micropore on film, and exogenous molecule is allowed to enter cell by micropore.
In the prior art, three classes method mainly completes the process of cell electrotransfection:
Cell is placed in a pair of between several millimeters to several centimeters parallel pole.Make the electricity of cell between the electrodes
By electro photoluminescence in, to realize the purpose of electrotransfection.For example, United States Patent (USP) US5389069.
It is penetrated in tissue or cell liquid using miniature needle electrode and is shocked by electricity to cell, achieve the purpose that electrotransfection.Example
Such as, United States Patent (USP) US5389069.
One chamber is placed between a pair of of parallel pole, while so that the aaerosol solution of cell flows in the chamber
It is shocked by electricity.For example, United States Patent (USP) US6773669.
Chinese patent CN201010242144 discloses a kind of streaming electrotransfection apparatus and system, which includes: streaming
Electrotransfection device, including: substrate, and the electrode being produced on substrate, the electrode are parallel and place in pairs
, each pair of electrode includes the anode and cathode being oppositely arranged；It is placed in the channel of the limitation fluid flowing on electrode；The channel
Starting point has multiple inlet branch channels, and pools a main channel, and end has multiple outlet branch channels, described
Setting has the top cover of multiple fluid inlets and outlet above channel；Syringe pump is connected to the streaming electrotransfection by pipeline and fills
Set the entrance of middle top cover and the flow velocity of control of export fluid；Voltage source is set by electrical connector connection electrode and generates pulse electricity
Pressure.Streaming electrotransfection system realizes various suspension in the fluid passage using fluid channel and the syringe pump being connected
Continuous flowing, so that cell be enable to be continued to carry out by the process of electrotransfection, realization quickly handles a large amount of samples.
Chinese patent CN201610806987 discloses the disposable product of the electrotransfection for cell comprising: in institute
State the fluid compartment of the inside of disposable product；First fluid port is used to provide cell suspension to the fluid compartment；With
And second fluid port, be used for will include to by the fluid delivery of electrotransfection at least one of cell compound extremely
The fluid compartment；First electrode and second electrode in the fluid compartment are set；At least one outlet port, delivering
The fluid from the fluid compartment, wherein the first fluid port and the second fluid port and hybrid channel are flowed
Body connection, the hybrid channel and the fluid compartment are in fluid communication.
But streaming electrotransfection device disclosed above can generate a large amount of bubbles during practical electrotransfection, generation
Bubble, which is attached on electrode plate, influences field uniformity, unstable so as to cause electrotransfection effect.Inventor passes through retrieval discovery,
The technical issues of electrotransfection process generates bubble is solved not yet in streaming electrotransfection field.
Summary of the invention
The utility model is provided a kind of by movably living for above-mentioned existing technology
The mode of pressurization is filled in, the intermittent streaming electrotransfection device of generation bubble during electrotransfection is inhibited.The utility model it is another
Purpose is to improve the stability of electrotransfection device, improves the efficiency of electrotransfection.
In order to achieve the above object, the technical solution of intermittent streaming electrotransfection device provided by the utility model is summarized
It is as follows: a kind of intermittent streaming electrotransfection device, including tube body, first electrode, second electrode, piston, which is characterized in that described
First electrode, second electrode are placed in the inner wall of the tube body in parallel, and one end of the tube body is arranged sealed end, the tube body it is another
The inside of the piston that one end setting can be relatively moved along the tube body, the piston and tube body has the chamber for accommodating liquid sample
Body, at least one liquid entrance is arranged in the tube body and at least one liquid passes in and out two way valve, the liquid entrance position
In the sealed end of the tube body, the liquid disengaging two way valve is located at the tube body liquid entrance.The liquid disengaging
The opening and closing of two way valve control liquid entrance.
It is preferred that at least two liquid entrances are arranged in the tube body, the liquid entrance either liquid-inlet,
It can be liquid outlet, corresponding at least two liquid that are arranged pass in and out two way valve；The liquid disengaging two way valve is for controlling
Liquid sample introduction and liquid discharge sample processed；At least one liquid-inlet and a liquid outlet is arranged in the further preferred tube body, until
Few two liquid pass in and out two way valve；The further preferred liquid entrance is located at the tube body sealed end on the piston opposite.
It is preferred that the cross section of the piston is identical or close as the cross section of the tubular body, it is further preferably described
The shape of the cross section of tubular body be it is rectangular, round or other shapes also can be set into；
It is preferred that the piston and tube body contact position are arranged sealing device, the preferred seal washer of the sealing device, sealant
Item or other devices etc. with sealing performance；
It is preferred that when the liquid passes in and out two way valve and gas valve and closes, the piston and tubular body form close
Enclosed cavity；
It is preferred that the piston is after tube body movement, the closed chamber when liquid disengaging two way valve is closed
The pressure value in internal portion is 1.0-2.5Mpa；The pressure value of the further preferred inside cavity is 1.1-2.0Mpa；Further
It is preferred that the pressure value of the inside cavity is 1.2-1.5Mpa；
It is preferred that described device further includes that conduit is several, it is divided into inlet tube and outlet tube, conduit and tube body sealed set；
It is preferred that described device further includes the push-pull device at fixed for enabling the piston to relatively move along the tube body, it is described to push away
The type of drawing device includes but is not limited to hydraulic stem, atmospheric pressure pole, spiral push-pull device at fixed, electromagnetic force push-pull device at fixed etc.；
It is preferred that the piston setting operating position and off-position, the piston are moved to work from off-position
Position, can be by liquid from liquid entrance inspiration cavity, and the piston is mobile from operating position to off-position, Neng Gouwei
Inside cavity provides pressure；
The preparation method of device described in the utility model: tube body, selection cured plastics work after capable of heating are prepared first
For the material of tube body, tube body is prepared using the mold for preparing tube body, Liquid sample introduction mouth and liquid discharge sample are respectively set on tube body
Mouthful, the electrode of two panels same shape, size is arranged in parallel in the opposite sides of inboard wall of tube body.One end of tube body is set as sealed end,
One piston parallel relative to tube body seal face is set in the other end opening of tube body, is adopted in piston and tube body contact position
It is sealed with sealing joint strip, is connect in the side that piston does not contact cavity with push-pull device at fixed.In the sealed end of tube body, one liquid is set
Body import and a liquid outlet.It is tightly connected in liquid-inlet and liquid outlet setting conduit and tube body, is set on conduit
Set two way valve.
Technical solutions of the utility model bring the utility model has the advantages that
The intermittent streaming electrotransfection device of the utility model is capable of handling a large amount of fluid samples.
The intermittent streaming electrotransfection device of the utility model, to liquid pressing, inhibits bubble to produce during electrotransfection
Raw, electric field strength inhomogeneities caused by reducing because of bubble attachment improves the stability of electrotransfection device, to improve electrotransfection
Efficiency.
The intermittent streaming electrotransfection apparatus structure of the utility model is novel in design, has initiative.
Detailed description of the invention
The intermittent streaming electrotransfection device exemplary diagram of Fig. 1 the utility model；
Attached drawing annotation: 1- first electrode, 2- second electrode, 3- tube body, 4- sealed end, 5- inlet tube, 6- outlet tube, 7- the
One two way valve, the second two way valve of 8-, 9- push-pull device at fixed, 10- piston off-position, the first operating position of 11- piston,
The second operating position of 12- piston.
Specific embodiment
Embodiment one
Overall construction design (one)
Intermittent streaming electrotransfection device provided by the utility model is as shown in Figure 1, the first electrode (1) and the second electricity
Pole (2) is arranged in parallel in the inner wall of tube body (3) respectively, and one end nozzle of tube body (3) is arranged to sealed end (4), the feed liquor
It manages (5) and outlet tube (6) is arranged on tube body sealed end (4), first two way valve (7) is placed in the inlet tube (5)
Interface close to inlet tube (5) and sealed end (4), second two way valve (8) is placed in close in the outlet tube (6)
The interface of outlet tube (6) and sealed end (4).The piston is set at the other end nozzle of the tube body (3), the piston
It is fixedly connected with push-pull device at fixed (9), piston can be moved to piston first from piston off-position (10) by push-pull device at fixed (9)
Operating position (11) or the second operating position of piston (12), the preferred hydraulic stem of the push-pull device at fixed (9) push piston to make in cavity
The liquid of portion's carrying bears certain pressure.Apply electric pulse in first electrode (1) and second electrode (2), electric pulse makes chamber
Electric field is formed in vivo, so that cell membrane forms micropore, so that the target substance in liquid sample be made to be able to enter in cell.Electricity turns
After the completion of dye, piston can be moved to piston off-position (10) from the second operating position of piston (12) by push-pull device at fixed (9),
Tube body (3) interior liquid is squeezed out into tube body.It repeats aforesaid operations and carries out intermittent streaming electrotransfection.
Embodiment two
Liquid batch formula feed liquor mode and pressuring method
When feed liquor, the first two way valve (7) is opened, and the second two way valve (8) is closed, and push-pull device at fixed (9) is by piston from work
It fills in off-position (10) drawing and arrives the first operating position of piston (11), target liq sample enters inside cavity from inlet tube (5)；
It closes the first two way valve (7), push-pull device at fixed (9) shifts piston onto piston the second working position from the first operating position of piston (11)
(12) are set, to liquid pressing in cavity；The electrotransfection of liquid is carried out in pressurized state.After the completion of electrotransfection, it is two-way to open second
Valve (8), push-pull device at fixed (9) shifts piston piston off-position (10) from the second operating position of piston (12) onto, in tube body
Liquid is pushed out tube body.The movement of the above liquid in-out is repeated, realizes the electrotransfection to big quantity of fluid.
Embodiment three
Biological test experiment
The CHO-S cell (Chinese hamster ovary cell) for being in logarithmic growth phase is collected, 1000 revs/min of revolving speed are centrifuged 5 points
Clock abandons supernatant, and cell is resuspended with electrotransfection buffer, so that the density of cell is 1X107A/milliliter, addition need electrotransfection
It is transferred to the plasmid pCDNA3.1-GFP of cell, makes 20 mcg/ml of concentration of plasmid, it is soft to be uniformly mixed.
The electrotransfection device being prepared using the utility model embodiment one is tested, and controls piston using hydraulic stem
Mobile in operating position and off-position, cooperation electrotransfection instrument uses, and carries out electro photoluminescence: voltage 180 using following condition
Volt, 6 milliseconds of pulse width, pulse number 2 times, the pulse spacing 1 second.
Comparative test is not apply pressure to liquid in cavity, and piston is maintained at the first operating position of piston (11),
Liquid filling is had one's bosom filled with after body, directly progress electrotransfection；
The test of technical solutions of the utility model is to apply pressure to liquid in cavity, after body is had one's bosom filled in liquid filling, is led to
It crosses hydraulic stem and piston is moved to the second operating position of piston (12) from the first operating position of piston (11), liquid is added
Pressure carries out electrotransfection.
After electrotransfection, the cell suspension after transfection is placed in centrifuge tube, 1000 revs/min, is centrifuged 5 minutes.It abandons
Supernatant is added CD-OptiCHO culture medium and cell is resuspended, is inoculated in triangular pyramidal culture in glassware, culture density 2 × 106A/
Then triangular pyramidal bottle is placed on shaking table and cultivates by ml, 125 revs/min of shaking speed, condition of culture: temperature is 37 DEG C, dioxy
Change concentration of carbon 5%.After 24 hours fluorescence microscopy under the microscope, and with flow cytomery electrotransfection efficiency and cell survival
Rate.
By the observation to electrotransfection process, comparative test can generate a small amount of gas during electrotransfection near electrode
Bubble, influences electrotransfection effect；And the test of technical solutions of the utility model is during electrotransfection and bubble-free generates.
By the way that electrotransfection Comparative result, the electrotransfection efficiency of comparative test is 60%-64%, and cell survival rate is
70%-83%；The electrotransfection efficiency of technical solutions of the utility model test is 84%-86%, cell survival rate 82%-
85%.The experimental results showed that technical solutions of the utility model test provide electrotransfection device have higher electrotransfection efficiency and
Cell survival rate.
Although the utility model has been described in detail, the modification in the spirit and scope of the utility model is for ability
Field technique personnel will be apparent.It should be understood that sheet described above and/or in the appended claims is practical
The part of novel aspect and various embodiments and various features can be combined or completely or partially be interchangeable.Such as this
Field technical staff will be realized that, in the description of aforementioned various embodiments, refers to those of another embodiment and implements
Scheme can suitably with other combination of embodiment.In addition, it will be recognized by those of ordinary skill in the art that foregoing description is only to make
To illustrate, and it is not intended to limit the utility model.
Claims (9)
1. a kind of intermittent streaming electrotransfection device, including tube body, first electrode, second electrode, piston, which is characterized in that institute
The inner wall that first electrode, second electrode are placed in the tube body in parallel is stated, sealed end is arranged in one end of the tube body, the tube body
The inside of the piston that other end setting can be relatively moved along the tube body, the piston and tube body, which has, accommodates liquid sample
Cavity, at least one liquid entrance is arranged in the tube body and at least one liquid passes in and out two way valve, the liquid entrance
Positioned at the sealed end of the tube body, the liquid disengaging two way valve is located at the tube body liquid entrance.
2. intermittent streaming electrotransfection device according to claim 1, which is characterized in that the tube body setting at least two
Liquid entrance and at least two liquid pass in and out two way valve.
3. intermittent streaming electrotransfection device according to claim 1, which is characterized in that operating position is arranged in the piston
And off-position.
4. intermittent streaming electrotransfection device according to claim 1, which is characterized in that the piston and tube body contact position
Sealing device is set.
5. intermittent streaming electrotransfection device according to claim 1, which is characterized in that when the liquid passes in and out two-way valve
When door is closed, the inside of the piston and tube body forms closed cavity.
6. intermittent streaming electrotransfection device according to claim 5, which is characterized in that when the liquid passes in and out two-way valve
When door is closed, for the piston after tube body relative movement, the pressure value of the closed inside cavity is 1.0-
2.50Mpa。
7. intermittent streaming electrotransfection device according to claim 1, which is characterized in that described device further include make it is described
The push-pull device at fixed that piston can be relatively moved along the tube body.
8. intermittent streaming electrotransfection device according to claim 7, which is characterized in that the push-pull device at fixed includes but not
It is limited to hydraulic stem, atmospheric pressure pole, spiral push-pull device at fixed, electromagnetic force push-pull device at fixed.
9. intermittent streaming electrotransfection device according to claim 1, which is characterized in that described device further includes conduit,
The conduit and liquid entrance are tightly connected.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201821508609.XU CN209098700U (en) | 2018-09-17 | 2018-09-17 | A kind of intermittent streaming electrotransfection device |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201821508609.XU CN209098700U (en) | 2018-09-17 | 2018-09-17 | A kind of intermittent streaming electrotransfection device |
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| Publication Number | Publication Date |
|---|---|
| CN209098700U true CN209098700U (en) | 2019-07-12 |
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Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201821508609.XU Active CN209098700U (en) | 2018-09-17 | 2018-09-17 | A kind of intermittent streaming electrotransfection device |
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| Country | Link |
|---|---|
| CN (1) | CN209098700U (en) |
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2018
- 2018-09-17 CN CN201821508609.XU patent/CN209098700U/en active Active
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| Date | Code | Title | Description |
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| GR01 | Patent grant | ||
| CP02 | Change in the address of a patent holder |
Address after: No.2 501, No.200, Shengpu Xingpu Road, Suzhou Industrial Park, Jiangsu Province Patentee after: SUZHOU ETTA BIOTECH CO., LTD. Address before: 215123 Ruoshui Road 388 E1504, Suzhou Industrial Park, Jiangsu Province Patentee before: SUZHOU ETTA BIOTECH CO., LTD. |
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