CN113755318A - Composite pulse cell electrofusion instrument and control method - Google Patents
Composite pulse cell electrofusion instrument and control method Download PDFInfo
- Publication number
- CN113755318A CN113755318A CN202111030837.7A CN202111030837A CN113755318A CN 113755318 A CN113755318 A CN 113755318A CN 202111030837 A CN202111030837 A CN 202111030837A CN 113755318 A CN113755318 A CN 113755318A
- Authority
- CN
- China
- Prior art keywords
- pulse
- module
- nanosecond
- switch relay
- pulse module
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000000034 method Methods 0.000 title claims abstract description 21
- 239000002131 composite material Substances 0.000 title claims abstract description 20
- 230000004927 fusion Effects 0.000 claims abstract description 14
- 230000007246 mechanism Effects 0.000 claims abstract description 13
- 238000007599 discharging Methods 0.000 claims description 31
- 230000000149 penetrating effect Effects 0.000 claims description 14
- 230000033228 biological regulation Effects 0.000 claims description 9
- 238000003780 insertion Methods 0.000 claims description 7
- 230000037431 insertion Effects 0.000 claims description 7
- 239000011111 cardboard Substances 0.000 claims description 6
- 239000007788 liquid Substances 0.000 claims description 6
- 238000004806 packaging method and process Methods 0.000 claims description 4
- 230000004888 barrier function Effects 0.000 claims description 3
- 238000005538 encapsulation Methods 0.000 claims description 3
- 238000010257 thawing Methods 0.000 claims description 3
- 230000001681 protective effect Effects 0.000 claims 4
- 230000007910 cell fusion Effects 0.000 abstract description 10
- 230000002457 bidirectional effect Effects 0.000 abstract description 6
- 230000009471 action Effects 0.000 abstract description 2
- 230000009286 beneficial effect Effects 0.000 abstract description 2
- 230000008569 process Effects 0.000 description 9
- 230000000903 blocking effect Effects 0.000 description 3
- 238000010586 diagram Methods 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 108090000623 proteins and genes Proteins 0.000 description 3
- 102000004169 proteins and genes Human genes 0.000 description 2
- 230000008859 change Effects 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 230000000093 cytochemical effect Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000005684 electric field Effects 0.000 description 1
- 238000012407 engineering method Methods 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 238000009396 hybridization Methods 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 230000004807 localization Effects 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000001404 mediated effect Effects 0.000 description 1
- 238000002844 melting Methods 0.000 description 1
- 230000008018 melting Effects 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 231100000956 nontoxicity Toxicity 0.000 description 1
- 230000008672 reprogramming Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000014639 sexual reproduction Effects 0.000 description 1
- 230000000392 somatic effect Effects 0.000 description 1
- 230000003612 virological effect Effects 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M23/00—Constructional details, e.g. recesses, hinges
- C12M23/38—Caps; Covers; Plugs; Pouring means
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M23/00—Constructional details, e.g. recesses, hinges
- C12M23/46—Means for fastening
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M23/00—Constructional details, e.g. recesses, hinges
- C12M23/48—Holding appliances; Racks; Supports
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M35/00—Means for application of stress for stimulating the growth of microorganisms or the generation of fermentation or metabolic products; Means for electroporation or cell fusion
- C12M35/02—Electrical or electromagnetic means, e.g. for electroporation or for cell fusion
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N13/00—Treatment of microorganisms or enzymes with electrical or wave energy, e.g. magnetism, sonic waves
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q3/00—Condition responsive control processes
Abstract
The invention discloses a composite pulse cell electrofusion instrument and a control method, wherein the composite pulse cell electrofusion instrument comprises a protection box, a circuit control assembly and a positioning assembly, wherein the circuit control assembly comprises a power supply box, a high-frequency high-voltage sine module, a microsecond pulse module and a nanosecond pulse module; the positioning assembly is arranged at the bottom of the fixed bin and comprises two first fixed blocks and two second fixed blocks, the two positive middle parts of the two first fixed blocks and the two second fixed blocks are alternately connected with a bidirectional screw rod, the two sides of the two first fixed blocks are provided with control mechanisms, and the two sides of the two second fixed blocks are provided with clamping mechanisms. The invention has the beneficial effects that: the invention utilizes the characteristic that nanosecond pulse can perforate cells with different sizes, and can well fuse the cells with different sizes; the microsecond pulse is utilized to ensure that the perforation generated by the cells under the action of the nanosecond pulse is continuously and further expanded, thereby facilitating the fusion of the cells and effectively improving the success rate of the cell fusion.
Description
Technical Field
The invention relates to a cell capacitance instrument, in particular to a composite pulse cell electric fusion instrument and a control method, and belongs to the technical field of cell fusion instruments.
Background
Cell fusion, also known as cell hybridization, refers to the process of forming a cell with binuclear or polynuclear nuclei from two or more cells by means of mediated and cultured, artificial fusion in an ex vivo condition. The sexual reproduction of nature ensures the stability of genetic materials, various barriers are arranged among species, and the artificial induction cell fusion technology can fuse single cells from different kinds of organisms into a heterokaryocyte, and the new cell contains genetic materials of a plurality of parent cells, has new genetic or biological characteristics, and can be cultured into new species, strains or new cell engineering products.
Cell fusion, a rapidly developing new cell engineering method, has achieved pioneering research results in the fields of agriculture, medicine, and the like, and the application field is expanding, becoming a conventional means for studying somatic reprogramming and gene localization. The following four methods are mainly used for cell fusion: viral fusion, cytochemical fusion, cytoelectrofusion and laser fusion of cells.
Compared with other cell fusion technologies, the cell electrofusion technology has the advantages of strong controllability, strong repeatability, high heterologous cell fusion efficiency, no toxicity to cells, wide application objects, low sample loss and the like, and gradually becomes the most important means for realizing cell fusion. The microsecond pulse electrofusion is adopted in the traditional cell electrofusion, but the microsecond pulse has certain defects, when cells with different sizes are fused, large cells can generate perforation earlier than small cells, so that under the condition of acting on an electric field with the same strength, when the small cells are perforated, the large cells can be in a death state of excessive perforation, and the problems of low success rate, high cost, low efficiency, poor repeatability and the like exist in the experimental process.
Disclosure of Invention
The present invention is directed to solving the above problems and providing a composite pulse cell thawing apparatus and a control method.
The invention achieves the above objects by the following technical scheme, a composite pulse cell electric melting instrument, comprising:
the top end of the protection box is provided with a packaging cover, and a fixed bin is arranged inside the protection box;
the circuit control assembly comprises a power box, a high-frequency high-voltage sine module, a microsecond pulse module and a nanosecond pulse module, wherein the power box, the high-frequency high-voltage sine module, the microsecond pulse module and the nanosecond pulse module are electrically connected in sequence;
locating component, locating component sets up the bottom in fixed storehouse, locating component includes two first fixed blocks and two second fixed blocks, two the second fixed block sets up between two first fixed blocks, two the positive middle part interlude of first fixed block and two second fixed blocks is connected with two-way screw rod, two the both sides of first fixed block all are provided with control mechanism, two the both sides of second fixed block all are provided with presss from both sides fixed establishment.
Preferably, the combination groove has been seted up on the top of protection box both sides wall, the both sides fixedly connected with compoboard, two of encapsulation lid bottom the combination board and two combination groove joints, two direction spouts have been seted up to the inner wall of fixed storehouse bottom both sides, one of them side of protection box, and run through fixed storehouse and seted up viewing aperture and a plurality of control hole, the opposite side of protection box just runs through fixed storehouse and has seted up a plurality of wiring mouths, the front of protection box just runs through fixed storehouse and has seted up the regulation hole.
Preferably, two equal fixedly connected with cardboard in one side that first fixed block carried on the back mutually, two the regulation screw has all been seted up at the positive middle part of first fixed block and two cardboards, two-way screw rod and four regulation screw thread interlude are connected, two the equal fixedly connected with barrier plate in top of cardboard.
Preferably, the two sides of the two second fixing blocks are provided with rotating ports, the two right middle parts of the second fixing blocks are provided with penetrating holes, the two-way screw rod and the two penetrating holes are connected in a sliding penetrating mode, one end of the two-way screw rod is provided with a rotating block, and the rotating block penetrates through the adjusting hole and is arranged on the right side of the protection box.
Preferably, the clamping mechanism comprises two supporting plates, two supporting plates and one of the guide sliding grooves are connected in a sliding and penetrating mode, guide holes are formed in the front faces of the supporting plates, swing rods are arranged between the supporting plates, the top of each supporting plate and the swing rods rotate to be connected with rotating rods in a penetrating mode, and one side of the top of each swing rod is fixedly connected with the clamping plate.
Preferably, the control mechanism comprises a control block, the control block is connected with one of the guide chutes in a sliding and penetrating manner, a control inserted rod is fixedly connected to the back of the control block, and the control inserted rod is connected with the guide hole in a sliding and penetrating manner.
Preferably, the back surfaces of the high-frequency high-voltage sine module, the microsecond pulse module and the nanosecond pulse module are provided with connecting poles, the front surfaces of the power box, the high-frequency high-voltage sine module and the microsecond pulse module are provided with two connecting holes, six connecting poles are correspondingly inserted into the six connecting holes, and one side of the power box is provided with a display screen;
a third switch relay is arranged on one side of the high-frequency high-voltage sine module and is set to be K3;
a second switching relay and a fifth switching relay are arranged on one side of the microsecond pulse module and are respectively set to be K2 and K5;
one side of nanosecond pulse module is provided with first switching relay and fourth switching relay, and sets up to K1 and K4 respectively.
A control method of a composite pulse cell electrofusion instrument comprises one of the following working modes;
the first working mode comprises the following steps:
s1: setting the number of output pulses of the microsecond pulse module, the number of output pulses of the nanosecond pulse module and the output time of the high-frequency high-voltage sine module;
s2: firstly, the controller controls the third switch relay K3 to be switched on, then controls the first switch relay K1, the second switch relay K2, the fourth switch relay K4 and the fifth switch relay K5 to be switched off, starts the high-frequency high-voltage sine module, applies sine alternating-current voltage to a culture dish containing cells and fusion liquid, and promotes the cells to form a queue and be tightly attached through the sine alternating-current voltage;
s3: when the output time of the sine alternating voltage reaches a set time value, the controller closes the third switch relay K3 to stop applying the sine alternating voltage to the culture dish;
s4: the first switch relay K1 is conducted through the controller, the nanosecond pulse module is electrified to start to operate, and nanosecond square pulse waves are applied to the culture dish;
s5: after the pulse number of the nanosecond pulse module reaches the set number, the controller controls to disconnect the first switch relay K1, and the nanosecond pulse module stops applying nanosecond square pulse waves to the culture dish;
s6: controlling a second switch relay K2 to be conducted, electrifying the microsecond pulse module to start to operate, and applying microsecond square pulse waves to the culture dish;
s: after the pulse number of the microsecond pulse module reaches the set number, the second switch relay K2 is controlled to be switched off by the controller, and the microsecond pulse module stops applying nanosecond square pulse waves to the culture dish;
s: controlling the fourth switching relay K4 to be switched on, electrifying a discharging loop connected with the nanosecond pulse module, and starting discharging the nanosecond pulse module through the first discharging resistor;
s: the fifth switch relay K5 is controlled to be switched on, a discharging loop connected with the microsecond pulse module is powered on, and the microsecond pulse module starts to discharge through a second discharging resistor R2;
the second working mode comprises the following steps:
the first step is as follows: setting the number of output pulses of the microsecond pulse module, the number of output pulses of the nanosecond pulse module 3 and the output time of the high-frequency high-voltage sine module 1;
the second step is that: firstly, the controller controls the third switch relay K3 to be switched on, then controls the first switch relay K1, the second switch relay K2, the fourth switch relay K4 and the fifth switch relay K5 to be switched off, starts the high-frequency high-voltage sine module, applies sine alternating-current voltage to a culture dish containing cells and fusion liquid, and promotes the cells to form a queue and be tightly attached through the sine alternating-current voltage;
the third step: when the output time of the sine alternating voltage reaches a set time value, the controller closes the third switch relay K3 to stop applying the sine alternating voltage to the culture dish;
the fourth step: controlling a second switch relay K2 to be conducted, electrifying the microsecond pulse module to start to operate, and applying microsecond square pulse waves to the culture dish;
the fifth step: after the pulse number of the microsecond pulse module reaches the set number, the second switch relay K2 is controlled to be switched off by the controller, and the microsecond pulse module stops applying nanosecond square pulse waves to the culture dish;
and a sixth step: the first switch relay K1 is conducted through the controller, the nanosecond pulse module is electrified to start to operate, and nanosecond square pulse waves are applied to the culture dish;
the seventh step: after the pulse number of the nanosecond pulse module reaches the set number, the controller controls to disconnect the first switch relay K1, and the nanosecond pulse module stops applying nanosecond square pulse waves to the culture dish;
eighth step: controlling the fourth switching relay K4 to be switched on, switching on a discharging loop connected with the nanosecond pulse module, and starting discharging the nanosecond pulse module through the first discharging resistor R1;
the ninth step: the fifth switch relay K5 is controlled to be turned on, the discharging loop connected with the microsecond pulse module is powered on, and the microsecond pulse module starts to discharge through the second discharging resistor R2
Preferably, the sinusoidal voltage output by the high-frequency high-voltage sinusoidal module ranges from 0V to 500V, the frequency ranges from 0 MHz to 3MHz, and the duration time ranges from 0s to 100 s.
Preferably, the nanosecond pulse module outputs pulses with the amplitude of 0-10kV, the pulse width of 50-1000 ns, the pulse frequency of 0.1-1000 Hz, the number of the pulses is adjustable, and the waveform is a pulse square wave;
the microsecond pulse module outputs pulses with the amplitude of 0-5kV, the pulse width of 1us-1000us, the pulse frequency of 0.1Hz-1000Hz, the number of the pulses is adjustable, and the waveform is pulse square waves.
The invention has the beneficial effects that:
firstly, the invention utilizes the characteristic that nanosecond pulse can perforate cells with different sizes, and can well fuse the cells with different sizes; the microsecond pulse is utilized to ensure that the perforation generated by the cells under the action of the nanosecond pulse is continuously and further expanded, thereby facilitating the fusion of the cells and effectively improving the success rate of the cell fusion.
Secondly, the clamping assembly is arranged in the protection box, and the circuit control assembly is clamped through the clamping assembly, so that the power supply box, the high-frequency high-voltage sine module, the microsecond pulse module and the nanosecond pulse module can be clamped step by step, if any one of the modules is damaged, disassembly, maintenance and replacement can be conveniently carried out, and the detection time and the production cost are saved.
The clamping assembly is controlled in two aspects, the two first fixing blocks are controlled by the bidirectional screw rod to horizontally and longitudinally clamp the two second fixing blocks, and then the control structure is matched with the two clamping mechanisms to horizontally and transversely clamp the two second fixing blocks, so that the whole clamping process is uniformly carried out, and the clamping is very firm.
Drawings
FIG. 1 is a schematic view of the overall structure of the present invention;
FIG. 2 is a schematic sectional view and an internal structure of the protection case according to the present invention;
FIG. 3 is a schematic diagram of the connection between the circuit control assembly and the positioning assembly of the present invention;
FIG. 4 is a schematic structural view of a positioning assembly of the present invention;
FIG. 5 is an enlarged view of the structure at A in FIG. 4 according to the present invention;
FIG. 6 is a schematic view of the construction of the sway bar of the present invention;
FIG. 7 is a circuit diagram of the circuit control assembly of the present invention;
fig. 8 is a circuit diagram of the present invention.
In the figure: 1. a protection box; 101. a package cover; 102. a composition board; 103. a guide chute; 2. a first fixed block; 201. clamping a plate; 202. a blocking plate; 3. a control block; 301. a control inserted link; 4. a second fixed block; 401. a support plate; 402. a guide hole; 5. a swing lever; 501. clamping a plate; 6. a bidirectional screw; 601. rotating the block; 7. a nanosecond pulse module; 8. a microsecond pulse module; 9. a high-frequency high-voltage sine module; 10. a power supply box; 1001. a display screen; 11. connecting the pole; 12. and connecting the holes.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Referring to fig. 1-8, a composite pulse cell electrofusion apparatus includes:
the protection box comprises a protection box 1, wherein a packaging cover 101 is arranged at the top end of the protection box 1, and a fixed bin is arranged inside the protection box 1;
the combination groove has been seted up on the top of protection box 1 both sides wall, the both sides fixedly connected with compoboard 102 of encapsulation lid 101 bottom, two compoboards 102 and two combination groove joints, two direction spouts 103 have been seted up to the inner wall of fixed storehouse bottom both sides, one of them side of protection box 1, and run through the fixed storehouse and seted up viewing aperture and a plurality of control hole, protection box 1's opposite side, and run through the fixed storehouse and seted up a plurality of wiring mouths, protection box 1's front, and run through the fixed storehouse and seted up the regulation hole.
The circuit control assembly comprises a power box 10, a high-frequency high-voltage sine module 9, a microsecond pulse module 8 and a nanosecond pulse module 7, wherein the power box 10, the high-frequency high-voltage sine module 9, the microsecond pulse module 8 and the nanosecond pulse module 7 are electrically connected in sequence;
the back sides of the high-frequency high-voltage sine module 9, the microsecond pulse module 8 and the nanosecond pulse module 7 are respectively provided with a connecting pole post 11, the front sides of the power box 10, the high-frequency high-voltage sine module 9 and the microsecond pulse module 8 are respectively provided with two connecting holes 12, the six connecting pole posts 11 and the six connecting holes 12 are correspondingly inserted, and one side of the power box 10 is provided with a display screen 1001;
a third switch relay is arranged on one side of the high-frequency high-voltage sine module 9 and is set to be K3;
a second switching relay and a fifth switching relay are arranged on one side of the microsecond pulse module 8 and are respectively set to be K2 and K5;
one side of the nanosecond pulse module 7 is provided with a first switching relay and a fourth switching relay, which are respectively set to K1 and K4
The positioning assembly is arranged at the bottom of the fixed bin and comprises two first fixed blocks 2 and two second fixed blocks 4, the two second fixed blocks 4 are arranged between the two first fixed blocks 2, the middle parts of the front surfaces of the two first fixed blocks 2 and the two second fixed blocks 4 are connected with two-way screws 6 in an inserting mode, two sides of each of the two first fixed blocks 2 are provided with control mechanisms, and two sides of each of the two second fixed blocks 4 are provided with clamping mechanisms;
the clamping mechanism comprises two supporting plates 401, the two supporting plates 401 are connected with one of the guide sliding grooves 103 in a sliding and penetrating manner, guide holes 402 are formed in the front faces of the two supporting plates 401, a swing rod 5 is arranged between the two supporting plates 401, a rotating rod is connected between the tops of the two supporting plates 401 and the swing rod 5 in a rotating and penetrating manner, and one side of the top of the swing rod 5 is fixedly connected with a clamping plate 501;
the control mechanism comprises a control block 3, the control block 3 is connected with one of the guide chutes 103 in a sliding and penetrating way, the back of the control block 3 is fixedly connected with a control inserted rod 301, and the control inserted rod 301 is connected with a guide hole 402 in a sliding and penetrating way
Clamping plates 201 are fixedly connected to the opposite sides of the two first fixing blocks 2, adjusting screw holes are formed in the middles of the front sides of the two first fixing blocks 2 and the front sides of the two clamping plates 201, the two-way screw 6 and the four adjusting screw holes are connected in a threaded penetrating mode, and blocking plates 202 are fixedly connected to the top ends of the two clamping plates 201;
the rotation mouth has all been seted up to the both sides of two second fixed blocks 4, and the interlude hole has all been seted up at the positive middle part of two second fixed blocks 4, and two-way screw rod 6 and two interlude hole slip are connected, and wherein one end of two-way screw rod 6 is provided with turning block 601, and turning block 601 runs through the regulation hole and sets up in the front of protection box 1.
A control method of a composite pulse cell electrofusion instrument comprises one of the following working modes;
the first working mode comprises the following steps:
s1: setting the number of output pulses of the microsecond pulse module 8, the number of output pulses of the nanosecond pulse module 7 and the output time of the high-frequency high-voltage sine module 9;
s2: firstly, the controller controls the third switch relay K3 to be switched on, then controls the first switch relay K1, the second switch relay K2, the fourth switch relay K4 and the fifth switch relay K5 to be switched off, starts the high-frequency high-voltage sine module 9, applies sine alternating-current voltage to a culture dish containing cells and fusion liquid, and promotes the cells to form a queue and be tightly attached through the sine alternating-current voltage;
s3: when the output time of the sine alternating voltage reaches a set time value, the controller closes the third switch relay K3 to stop applying the sine alternating voltage to the culture dish;
s4: the first switch relay K1 is conducted through the controller, the nanosecond pulse module 7 is electrified to start to operate, and nanosecond square pulse waves are applied to the culture dish;
s5: after the number of pulses of the nanosecond pulse module 7 reaches the set number, the controller controls to turn off the first switch relay K1, and the nanosecond pulse module 7 stops applying nanosecond square pulse waves to the culture dish;
s6: controlling a second switch relay K2 to be conducted, electrifying the microsecond pulse module 8 to start to operate, and applying microsecond square pulse waves to the culture dish;
s7: after the number of pulses of the microsecond pulse module 8 reaches the set number, the controller controls to turn off the second switch relay K2, and the microsecond pulse module 8 stops applying nanosecond square pulse waves to the culture dish;
s8: controlling the fourth switching relay K4 to be switched on, electrifying a discharging loop connected with the nanosecond pulse module 7, and starting discharging through the first discharging resistor by the nanosecond pulse module 7;
s9: the fifth switch relay K5 is controlled to be turned on, a discharging loop connected with the microsecond pulse module 8 is powered on, and the microsecond pulse module 8 starts to discharge through a second discharging resistor R2;
the second working mode comprises the following steps:
the first step is as follows: setting the number of output pulses of the microsecond pulse module 8, the number of output pulses of the nanosecond pulse module 7 and the output time of the high-frequency high-voltage sine module 9;
the second step is that: firstly, the controller controls the third switch relay K3 to be switched on, then controls the first switch relay K1, the second switch relay K2, the fourth switch relay K4 and the fifth switch relay K5 to be switched off, starts the high-frequency high-voltage sine module 9, applies sine alternating-current voltage to a culture dish containing cells and fusion liquid, and promotes the cells to form a queue and be tightly attached through the sine alternating-current voltage;
the third step: when the output time of the sine alternating voltage reaches a set time value, the controller closes the third switch relay K3 to stop applying the sine alternating voltage to the culture dish;
the fourth step: controlling a second switch relay K2 to be conducted, electrifying the microsecond pulse module 8 to start to operate, and applying microsecond square pulse waves to the culture dish;
the fifth step: after the number of pulses of the microsecond pulse module 8 reaches the set number, the controller controls to turn off the second switch relay K2, and the microsecond pulse module 8 stops applying nanosecond square pulse waves to the culture dish;
and a sixth step: the first switch relay K1 is conducted through the controller, the nanosecond pulse module 7 is electrified to start to operate, and nanosecond square pulse waves are applied to the culture dish;
the seventh step: after the number of pulses of the nanosecond pulse module 7 reaches the set number, the controller controls to turn off the first switch relay K1, and the nanosecond pulse module 7 stops applying nanosecond square pulse waves to the culture dish;
eighth step: controlling the fourth switching relay K4 to be switched on, electrifying a discharging loop connected with the nanosecond pulse module 7, and starting discharging the nanosecond pulse module 7 through the first discharging resistor R1;
the ninth step: and the fifth switching relay K5 is controlled to be switched on, the discharging loop connected with the microsecond pulse module 8 is switched on, and the microsecond pulse module 8 starts to discharge through the second discharging resistor R2.
The sine voltage output by the high-frequency high-voltage sine module 9 ranges from 0V to 500V, the frequency ranges from 0 MHz to 3MHz, and the duration ranges from 0s to 100 s.
The nanosecond pulse module 7 outputs pulses with the amplitude of 0-10kV, the pulse width of 50-1000 ns, the pulse frequency of 0.1-1000 Hz, the number of the pulses is adjustable, and the waveform is a pulse square wave;
the microsecond pulse module 8 outputs pulses with the amplitude of 0-5kV, the pulse width of 1us-1000us, the pulse frequency of 0.1Hz-1000Hz, the number of the pulses is adjustable, and the waveform is pulse square waves.
When the invention is used, all components are firstly installed, and refer to fig. 1-6;
the first step of operation is implemented, the power supply box 10 and the nanosecond pulse module 7 are respectively clamped at the top ends of the two first fixed blocks 2, and the two side walls of the top of the first fixed block 2 and the blocking plate 202 are used for attaching and clamping the power supply box 10 and the nanosecond pulse module 7.
And implementing the second step of operation, and respectively clamping the microsecond pulse module 8 and the high-frequency high-voltage sine module 9 on the tops of the two second fixed blocks 4.
Implementing the third step of operation, pinching the rotating block 601 to rotate the bidirectional screw 6 clockwise, and performing the following processes;
the first process is as follows: when the bidirectional screw 6 rotates clockwise, the bidirectional screw is matched with the adjusting screw holes of the two first fixing blocks 2, and the two first fixing blocks 2 respectively drive the two power supply boxes 10 and the nanosecond pulse module 7 to move towards opposite directions under the guiding support of the control block 3 and the guiding chute 103;
and a second process: the two first fixed blocks 2 move and simultaneously extrude the two second fixed blocks 4 to move oppositely;
the third process: the two first fixing blocks 2 drive the connected control blocks 3 to move synchronously, the control blocks 3 drive the connected control insertion rods 301 to move synchronously, and meanwhile, the power supply box 10, the high-frequency high-voltage sine module 9, the microsecond pulse module 8 and the connecting pole 11 and the connecting hole 12 arranged in the nanosecond pulse module 7 are electrically inserted;
the process four is as follows: when the two control insertion rods 301 are close to the support plate 401, the two control insertion rods 301 are slowly inserted into the guide holes 402, under the interaction of the structures of the control insertion rods 301 and the swinging rods 5, the swinging rods 5 are extruded by the control insertion rods 301, the tops of the swinging rods swing towards the direction close to the center of the second fixed block 4, at the moment, the clamping plate 501 swings along with the swinging rods 5, and the microsecond pulse module 8 and the high-frequency high-voltage sine module 9 are clamped from two sides through the clamping plates 501;
and a fifth process: when the two first fixing blocks 2 move oppositely to surround and clamp the two second fixing blocks 4, the power supply box 10, the high-frequency high-voltage sine module, the microsecond pulse module 8 and the nanosecond pulse module 7 are inserted into the connecting pole 11 and the connecting hole 12 to complete clamping of the electric conduction circuit control assembly.
And performing the fourth step, namely clamping the packaging cover 101 on the top end of the protection box 1 to complete the assembly of the whole device, and then completing the fusion operation of the cells according to the using method in the specification.
If one of them module of circuit control assembly damages in the use of whole device, then can anticlockwise rotation turning block 601 for two-way screw rod 6 follows turning block 601 and carries out anticlockwise rotation, can overhaul and change the module that damages afterwards.
Furthermore, it should be understood that although the present description refers to embodiments, not every embodiment may contain only a single embodiment, and such description is for clarity only, and those skilled in the art should integrate the description, and the embodiments may be combined as appropriate to form other embodiments understood by those skilled in the art.
Claims (10)
1. A composite pulse cell electrofusion apparatus, comprising:
the protective box comprises a protective box (1), wherein a packaging cover (101) is arranged at the top end of the protective box (1), and a fixed bin is arranged inside the protective box (1);
the circuit control assembly comprises a power box (10), a high-frequency high-voltage sine module (9), a microsecond pulse module (8) and a nanosecond pulse module (7), wherein the power box (10), the high-frequency high-voltage sine module (9), the microsecond pulse module (8) and the nanosecond pulse module (7) are electrically connected in sequence;
locating component, locating component sets up the bottom in fixed storehouse, locating component includes two first fixed blocks (2) and two second fixed blocks (4), two second fixed block (4) set up between two first fixed blocks (2), two first fixed block (2) and two positive middle parts of second fixed block (4) alternate and are connected with two-way screw rod (6), two the both sides of first fixed block (2) all are provided with control mechanism, two the both sides of second fixed block (4) all are provided with presss from both sides fixed establishment.
2. The composite pulse cell electrofusion apparatus according to claim 1, wherein: the combined groove has been seted up on the top of protection box (1) both sides wall, both sides fixedly connected with compoboard (102) of encapsulation lid (101) bottom, two compoboard (102) and two combined groove joints, two direction spout (103) have been seted up to the inner wall of fixed storehouse bottom both sides, wherein one side of protection box (1), and run through the fixed storehouse and seted up viewing aperture and a plurality of control hole, the opposite side of protection box (1), and run through the fixed storehouse and seted up a plurality of wiring mouths, the front of protection box (1), and run through the fixed storehouse and seted up the regulation hole.
3. The composite pulse cell electrofusion apparatus according to claim 1, wherein: two equal fixedly connected with cardboard (201) in one side that first fixed block (2) carried on the back mutually, two the regulation screw has all been seted up at the positive middle part of first fixed block (2) and two cardboard (201), two-way screw rod (6) and four regulation screw thread alternate connection, two the equal fixedly connected with in top of cardboard (201) barrier plate (202).
4. The composite pulse cell electrofusion apparatus according to claim 1, wherein: two the rotation mouth has all been seted up to the both sides of second fixed block (4), two the interlude hole has all been seted up at the positive middle part of second fixed block (4), two-way screw rod (6) and two interlude hole sliding connection, one of them one end of two-way screw rod (6) is provided with turning block (601), just turning block (601) run through the regulation hole and set up in the front of protection box (1).
5. The composite pulse cell electrofusion apparatus according to claim 1, wherein: clamping mechanism includes two backup pads (401), two backup pad (401) and one of them direction spout (103) slip interlude are connected, two bullport (402) have all been seted up in the front of backup pad (401), two be provided with swinging arms (5) between backup pad (401), two it has the rotary rod to rotate to alternate between the top of backup pad (401) and swinging arms (5), one side fixedly connected with clamping plate (501) at swinging arms (5) top.
6. The composite pulse cell electrofusion apparatus according to claim 1, wherein: the control mechanism comprises a control block (3), the control block (3) is connected with one of the guide sliding grooves (103) in a sliding and penetrating mode, a control insertion rod (301) is fixedly connected to the back face of the control block (3), and the control insertion rod (301) is connected with a guide hole (402) in a sliding and penetrating mode.
7. The composite pulse cell electrofusion apparatus according to claim 1, wherein: the back surfaces of the high-frequency high-voltage sine module (9), the microsecond pulse module (8) and the nanosecond pulse module (7) are respectively provided with a connecting pole column (11), the front surfaces of the power box (10), the high-frequency high-voltage sine module (9) and the microsecond pulse module (8) are respectively provided with two connecting holes (12), six connecting pole columns (11) and six connecting holes (12) are correspondingly inserted, and one side of the power box (10) is provided with a display screen (1001);
a third switch relay is arranged on one side of the high-frequency high-voltage sine module (9) and is set to be K3;
a second switching relay and a fifth switching relay are arranged on one side of the microsecond pulse module (8) and are respectively set to be K2 and K5;
one side of the nanosecond pulse module (7) is provided with a first switch relay and a fourth switch relay which are respectively set to K1 and K4.
8. A control method of a composite pulse cell electrofusion instrument is characterized in that: comprises one of the following working modes;
the first working mode comprises the following steps:
s1: setting the number of output pulses of the microsecond pulse module (8), the number of output pulses of the nanosecond pulse module (7) and the output time of the high-frequency high-voltage sine module (9);
s2: firstly, the controller controls the third switch relay K3 to be switched on, then controls the first switch relay K1, the second switch relay K2, the fourth switch relay K4 and the fifth switch relay K5 to be switched off, starts the high-frequency high-voltage sine module (9), applies sine alternating-current voltage to a culture dish containing cells and fusion liquid, and promotes the cells to form a queue and be tightly attached through the sine alternating-current voltage;
s3: when the output time of the sine alternating voltage reaches a set time value, the controller closes the third switch relay K3 to stop applying the sine alternating voltage to the culture dish;
s4: the first switch relay K1 is conducted through the controller, the nanosecond pulse module (7) is electrified to start to operate, and nanosecond square pulse waves are applied to the culture dish;
s5: after the pulse number of the nanosecond pulse module (7) reaches the set number, the controller controls to disconnect the first switch relay K1, and the nanosecond pulse module (7) stops applying nanosecond square pulse waves to the culture dish;
s6: controlling a second switch relay K2 to be conducted, electrifying a microsecond pulse module (8) to start to operate, and applying microsecond square pulse waves to the culture dish;
s7: after the pulse number of the microsecond pulse module (8) reaches the set number, the controller controls to turn off the second switch relay K2, and the microsecond pulse module (8) stops applying nanosecond square pulse waves to the culture dish;
s8: controlling the fourth switching relay K4 to be switched on, electrifying a discharging loop connected with the nanosecond pulse module (7), and starting discharging the nanosecond pulse module (7) through the first discharging resistor;
s9: the fifth switch relay K5 is controlled to be switched on, a discharging loop connected with the microsecond pulse module (8) is electrified, and the microsecond pulse module (8) starts to discharge through a second discharging resistor R2;
the second working mode comprises the following steps:
the first step is as follows: setting the number of output pulses of the microsecond pulse module (8), the number of output pulses of the nanosecond pulse module (7) and the output time of the high-frequency high-voltage sine module (9);
the second step is that: firstly, the controller controls the third switch relay K3 to be switched on, then controls the first switch relay K1, the second switch relay K2, the fourth switch relay K4 and the fifth switch relay K5 to be switched off, starts the high-frequency high-voltage sine module (9), applies sine alternating-current voltage to a culture dish containing cells and fusion liquid, and promotes the cells to form a queue and be tightly attached through the sine alternating-current voltage;
the third step: when the output time of the sine alternating voltage reaches a set time value, the controller closes the third switch relay K3 to stop applying the sine alternating voltage to the culture dish;
the fourth step: controlling a second switch relay K2 to be conducted, electrifying a microsecond pulse module (8) to start to operate, and applying microsecond square pulse waves to the culture dish;
the fifth step: after the pulse number of the microsecond pulse module (8) reaches the set number, the controller controls to turn off the second switch relay K2, and the microsecond pulse module (8) stops applying nanosecond square pulse waves to the culture dish;
and a sixth step: the first switch relay K1 is conducted through the controller, the nanosecond pulse module (7) is electrified to start to operate, and nanosecond square pulse waves are applied to the culture dish;
the seventh step: after the pulse number of the nanosecond pulse module (7) reaches the set number, the controller controls to disconnect the first switch relay K1, and the nanosecond pulse module (7) stops applying nanosecond square pulse waves to the culture dish;
eighth step: controlling the fourth switching relay K4 to be switched on, electrifying a discharging loop connected with the nanosecond pulse module (7), and starting discharging the nanosecond pulse module (7) through the first discharging resistor R1;
the ninth step: and controlling the fifth switch relay K5 to be switched on, electrifying a discharging loop connected with the microsecond pulse module (8), and starting discharging the microsecond pulse module (8) through a second discharging resistor R2.
9. The method for controlling a composite pulse cell thawing apparatus according to claim 8, wherein: the sine voltage output by the high-frequency high-voltage sine module (9) ranges from 0V to 500V, the frequency ranges from 0 MHz to 3MHz, and the duration time ranges from 0s to 100 s.
10. The method for controlling a composite pulse cell thawing apparatus according to claim 8, wherein: the nanosecond pulse module (7) outputs pulses with the amplitude of 0-10kV, the pulse width of 50-1000 ns, the pulse frequency of 0.1-1000 Hz, the number of the pulses is adjustable, and the waveform is a pulse square wave;
the microsecond pulse module (8) outputs pulses with the amplitude of 0-5kV, the pulse width of 1us-1000us, the pulse frequency of 0.1Hz-1000Hz, the number of the pulses is adjustable, and the waveform is pulse square waves.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111030837.7A CN113755318A (en) | 2021-09-03 | 2021-09-03 | Composite pulse cell electrofusion instrument and control method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111030837.7A CN113755318A (en) | 2021-09-03 | 2021-09-03 | Composite pulse cell electrofusion instrument and control method |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN113755318A true CN113755318A (en) | 2021-12-07 |
Family
ID=78792791
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202111030837.7A Pending CN113755318A (en) | 2021-09-03 | 2021-09-03 | Composite pulse cell electrofusion instrument and control method |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113755318A (en) |
Citations (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4578168A (en) * | 1984-07-27 | 1986-03-25 | Biotronics | Apparatus for fusing live cells with electric fields |
| US4663292A (en) * | 1984-12-21 | 1987-05-05 | Wong Daniel T | High-voltage biological macromolecule transfer and cell fusion system |
| CN86202617U (en) * | 1986-10-23 | 1987-12-16 | 北京师范大学 | Special electric instrument for cell fusion |
| US5650305A (en) * | 1996-05-15 | 1997-07-22 | Health Research Inc. | Method to produce hybrid cells by electrofusion |
| CN2633892Y (en) * | 2003-09-05 | 2004-08-18 | 中国科学院上海技术物理研究所 | Space cell electra-fusion cultivation system |
| CN106834272A (en) * | 2017-03-08 | 2017-06-13 | 重庆大学 | Method based on the fusion of composite pulse electric field inducing cell |
| CN109576152A (en) * | 2019-01-17 | 2019-04-05 | 重庆大学 | A kind of composite pulse cell electricity melts instrument and control method |
| CN109628438A (en) * | 2019-01-17 | 2019-04-16 | 重庆大学 | A method of it is merged based on bipolar pulse electric field inducing cell |
| CN109777732A (en) * | 2019-01-17 | 2019-05-21 | 重庆大学 | A kind of bipolar cells fusion instrument and its control method |
| CN211300301U (en) * | 2019-11-25 | 2020-08-21 | 杭州睿笛生物科技有限公司 | High-voltage composite electric pulse modulation circuit and ablation equipment |
| CN112384810A (en) * | 2018-05-24 | 2021-02-19 | 齐默尔根公司 | Scalable, movable and reconfigurable modules for processing biological and chemical materials |
-
2021
- 2021-09-03 CN CN202111030837.7A patent/CN113755318A/en active Pending
Patent Citations (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4578168A (en) * | 1984-07-27 | 1986-03-25 | Biotronics | Apparatus for fusing live cells with electric fields |
| US4663292A (en) * | 1984-12-21 | 1987-05-05 | Wong Daniel T | High-voltage biological macromolecule transfer and cell fusion system |
| CN86202617U (en) * | 1986-10-23 | 1987-12-16 | 北京师范大学 | Special electric instrument for cell fusion |
| US5650305A (en) * | 1996-05-15 | 1997-07-22 | Health Research Inc. | Method to produce hybrid cells by electrofusion |
| CN2633892Y (en) * | 2003-09-05 | 2004-08-18 | 中国科学院上海技术物理研究所 | Space cell electra-fusion cultivation system |
| CN106834272A (en) * | 2017-03-08 | 2017-06-13 | 重庆大学 | Method based on the fusion of composite pulse electric field inducing cell |
| CN112384810A (en) * | 2018-05-24 | 2021-02-19 | 齐默尔根公司 | Scalable, movable and reconfigurable modules for processing biological and chemical materials |
| CN109576152A (en) * | 2019-01-17 | 2019-04-05 | 重庆大学 | A kind of composite pulse cell electricity melts instrument and control method |
| CN109628438A (en) * | 2019-01-17 | 2019-04-16 | 重庆大学 | A method of it is merged based on bipolar pulse electric field inducing cell |
| CN109777732A (en) * | 2019-01-17 | 2019-05-21 | 重庆大学 | A kind of bipolar cells fusion instrument and its control method |
| CN211300301U (en) * | 2019-11-25 | 2020-08-21 | 杭州睿笛生物科技有限公司 | High-voltage composite electric pulse modulation circuit and ablation equipment |
Non-Patent Citations (2)
| Title |
|---|
| CHENGXIANG LI等: "Cell electrofusion based on nanosecond/microsecond pulsed electric fields", 《PLOS ONE》 * |
| 吴梦等: "利用微秒和微纳秒脉冲电场诱导细胞融合", 《生物技术》 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| US6117660A (en) | Method and apparatus for treating materials with electrical fields having varying orientations | |
| US6078490A (en) | Method of treating materials with pulsed electrical fields | |
| JP2609690B2 (en) | Driving circuit and driving method for liquid crystal display device | |
| CN107598442B (en) | A kind of electric power distributing cabinet partition automation rapid welding equipment | |
| JP5885144B2 (en) | Method and apparatus for uniformly treating adherent cells | |
| US20160298074A1 (en) | Flow electroporation device | |
| WO1993002178A1 (en) | Apparatus and methods for electroporation and electrofusion | |
| CA2333394A1 (en) | Flow-through electroporation system for ex vivo gene therapy | |
| CN104617808A (en) | Novel adjustable polarity square wave high-voltage pulse power circuit and control strategy for generating double-polarity pulse | |
| Zimmermann et al. | Fusion of Avena sativa mesophyll cell protoplasts by electrical breakdown | |
| EP0968275B1 (en) | Method and apparatus for treating materials with electrical fields having varying orientations | |
| CN113755318A (en) | Composite pulse cell electrofusion instrument and control method | |
| JPH0676992B2 (en) | Electrophoresis method and apparatus for separating particles in a separation medium | |
| CN203535876U (en) | Electrical control experimental cabinet used for teaching | |
| Eynard et al. | Electrooptics studies of Escherichia coli electropulsation: orientation, permeabilization, and gene transfer | |
| CN109777732A (en) | A kind of bipolar cells fusion instrument and its control method | |
| CN206337280U (en) | Three-dimensional cell cultivation automates generating means | |
| CA2371880A1 (en) | Method and sample support system for separating and processing substances in situ | |
| CN109576152A (en) | A kind of composite pulse cell electricity melts instrument and control method | |
| EP1329502A2 (en) | Method and apparatus for treating materials with electrical fields having varying orientations | |
| CN208300832U (en) | A kind of power Transmission pylon scarer | |
| Gimsa et al. | Electroporation in rotating electric fields | |
| CN206710374U (en) | A kind of automatic glue instrument of new Ago-Gel | |
| CN206517265U (en) | A kind of Microsecond grade pulsed torsional load motor | |
| CN104242887B (en) | Plasma electro-optical switch driving circuit based on pulsed hydrogen thyratron and low-impedance cable Blumlein pulse-forming line |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20211207 |
|
| RJ01 | Rejection of invention patent application after publication |