CN108070618B - Microinjection operation method - Google Patents
Microinjection operation method Download PDFInfo
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- CN108070618B CN108070618B CN201711315349.4A CN201711315349A CN108070618B CN 108070618 B CN108070618 B CN 108070618B CN 201711315349 A CN201711315349 A CN 201711315349A CN 108070618 B CN108070618 B CN 108070618B
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- needle
- operating arm
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- injection
- knob
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- 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
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/87—Introduction of foreign genetic material using processes not otherwise provided for, e.g. co-transformation
- C12N15/89—Introduction of foreign genetic material using processes not otherwise provided for, e.g. co-transformation using microinjection
Abstract
The invention discloses a microinjection operation method, which at least comprises a needle pulling instrument, a fixed needle, an injection needle, a fixed needle operating arm, an injection needle operating arm and a microscope, wherein the fixed needle is rotationally connected with the fixed needle operating arm, the injection needle is rotationally connected with the injection needle operating arm, the microscope comprises an object stage, a cell carrier is arranged on the object stage, a first knob is arranged on the fixed needle operating arm, a second knob is arranged on the injection needle operating arm, and the fixed needle has a bending angle of 30 degrees. Provides a convenient and rapid microinjection operation method which can conveniently and rapidly obtain a needle opening with a proper size when the needle is broken by adjusting the position of the injection needle and the angle of the fixed needle.
Description
Technical Field
The invention relates to microinjection operation, in particular to a microinjection operation method.
Background
The microinjection technology is a technology for injecting exogenous genes or other substances into cells by using a microinjection needle after the injected cells are adjusted and fixed in a microscope field, is one of important technical means of modern bioengineering, and particularly belongs to the field of cell engineering such as transgenosis, cloning and test-tube infants. Compared with other transgenic technologies, microinjection has the advantages of stable transgenic efficiency, unlimited size of injected genes, no need of vectors and no special requirements for operation objects and the like, and is widely applied to endangered animal protection, reproduction and health, species improvement and intracellular cytoplasm research.
During microinjection operation, the most critical step is the breaking of the injection needle. The needle point of the injection needle has too thick cut, so that the death rate of cells is increased during injection; the fracture is too thin, the injection amount is too small, and the needle is easy to block. The general needle breaking method of the injection needle is that the injection needle collides against the wall of a fixed needle tube to break the needle point of the injection needle, but the size of the needle opening after the needle breaking is random, and the inner diameter of the needle opening after the needle breaking needs to have proper size, so that the operation needs certain technical content, the operation time is generally long, and the operation time is accidental. The method also has high requirements on the injection needle, and the needle point of the injection needle needs to be very sharp, short and hard. When the needle pulling instrument pulls the injection needle, the pulled specification is different due to the difference of parameters and machines, most of the needle point of the pulled injection needle has certain length and toughness, and the method is not suitable for breaking the injection needle.
The invention discloses a Chinese patent No. CN105420099A with publication date of 2016, 3, 23, and a high-efficiency microinjection device, which comprises a quartz substrate and an injection body, wherein the injection body is fixed on the quartz substrate, the injection body is provided with an input micro-channel, an output micro-channel and a cell conveying micro-channel, a micro-injection channel and a cell movement control channel are also arranged between the input micro-channel and the output micro-channel on the injection body, the micro-injection channel is vertical and is vertically communicated with the cell conveying micro-channel, a micro-injection needle is fixed on the micro-injection channel, a gap is arranged between the micro-injection channel and the micro-injection needle, the size of the gap is smaller than the diameter of the cell, a limit bulge is arranged in the microinjection flow channel, the flow channel for controlling the movement of the cell is vertically communicated with the microinjection flow channel, and the flow channel for controlling the cell movement is positioned above the limiting bulge, and indirect control is provided by controlling the movement of the microfluid.
The cell moves to make the cell 'self' collide with the microinjection needle, thereby completing the microinjection operation of the cell. The defect is that the inner diameter of the needle opening of the broken needle can not be controlled corresponding to the needle point of the injection needle pulled by the needle pulling instrument.
Disclosure of Invention
The invention provides a convenient and quick microinjection operation method which can conveniently and quickly obtain a needle opening with a proper size when the needle is broken by adjusting the position of the injection needle and the angle of a fixed needle so as to solve the problem that the inner diameter of the broken needle opening cannot be controlled corresponding to the needle point of the injection needle pulled out by a needle pulling instrument.
The technical scheme adopted by the invention for solving the technical problem is as follows: a microinjection operation method at least comprises a needle pulling instrument, a fixed needle, an injection needle, a fixed needle operating arm, an injection needle operating arm and a microscope, wherein the fixed needle is rotatably connected with the fixed needle operating arm, the injection needle is rotatably connected with the injection needle operating arm, the microscope comprises an objective table, a cell carrier is arranged on the objective table, a first knob is arranged on the fixed needle operating arm, a second knob is arranged on the injection needle operating arm, and the fixed needle has a bending angle of 30 degrees.
Preferably, the present invention comprises the steps of:
the method comprises the following steps: placing the cells in a cell carrier on a microscope stage;
step two: adjusting an injection needle operating arm through a first knob to enable the needle point of the injection needle to be wholly tiled at the bottom of the cell carrier;
step three: rotating the fixed needle to enable the bent back of the rotating needle to face upwards;
step four: the fixed needle operating arm is adjusted through the second knob, so that the needle head of the fixed needle is slowly contacted with the bottom of the vessel;
step five: the needle operating arm is adjusted through the first knob, and the needle fixing operating arm is adjusted through the second knob, so that the needle point of the injection needle and the needle head of the fixing needle are on the same horizontal plane;
step six: selecting the position of the needle point of the injection needle, and finely adjusting the fixed needle operating arm through a second knob;
step seven: and breaking the needle point of the injection needle by using the fixed needle head to finish the needle breaking.
Preferably, the inner diameter of the needle opening after needle breakage is 3-5 μm. The inner diameter of the needle opening is 3-5 μm after the needle is broken, the needle point opening of the injection needle is too thick, and the death rate of cells is increased during injection; the fracture is too thin, the injection amount is too small, and the needle is easy to block.
Preferably, the length of the needle tip of the injection needle is 5-8 mm. The needle point of the injection needle has stronger hardness and high sharpness.
Preferably, the length of the needle point of the injection needle pulled out by the needle pulling instrument is 10-20 mm.
The practical significance of the invention is as follows:
(1) the operation is convenient, quick and easy;
(2) by adjusting the position of the injection needle and the angle of the fixed needle, the needle opening with proper size can be conveniently and quickly obtained when the injection needle is broken.
(3) The device can adapt to the different specifications of the injection needles pulled out by the needle pulling instrument due to the difference of parameters and machines when the injection needles are pulled.
Detailed Description
During microinjection operation, the most critical step is the breaking of the injection needle. The needle point of the injection needle has too thick cut, so that the death rate of cells is increased during injection; the fracture is too thin, the injection amount is too small, and the needle is easy to block. The common needle breaking method of the injection needle is that the injection needle collides against the wall of a fixed needle tube to break the needle point of the injection needle, but the size of the needle opening after the needle breaking is random, and the size of the inner diameter of the needle opening after the needle breaking is 3-5 mu m is most suitable, so the operation needs a certain technical content, the operation time is generally longer and the operation time is accidental. The method also has high requirements on the injection needle, and the needle point of the injection needle needs to be sharp, short (5-8 mm) and certain hardness. When the needle pulling instrument pulls the injection needle, the pulled specifications are different due to the difference of parameters and machines, most of the needle points of the pulled injection needle have certain length (10-20 mm) and are flexible, and the needle opening with the proper size can be conveniently and quickly obtained when the injection needle is broken by adjusting the position of the injection needle and the angle of the fixed needle.
This embodiment includes a fixed needle, an injection needle, an operating arm for the fixed needle and the injection needle, and a vessel for loading cells on a microscope stage. The needle point of the injection needle has certain length and toughness, and is slowly regulated by the operating arm of the injection needle to be completely tiled at the bottom of the vessel for loading cells. When the fixed needle is arranged on the operating arm, the angle of the fixed needle is adjusted (the fixed needle has a bending angle of about 30 degrees) so that the bent back of the fixed needle is upward, and the fixed needle operating arm is adjusted so that the needle head of the fixed needle is slowly contacted with the bottom of the vessel. And then adjusting the two operating arms to enable the needle point of the injection needle and the needle point of the fixed needle to be on the same horizontal plane, selecting the position with the proper size of the needle point of the injection needle, finely adjusting the operating arm of the fixed needle, and breaking the needle point of the injection needle by using the needle point of the fixed needle to finish the needle breaking.
The above-described embodiment is only a preferred embodiment of the present invention, and is not intended to limit the invention in any way, and other variations and modifications may be made without departing from the spirit of the invention as set forth in the claims.
Claims (2)
1. A microinjection operation method is suitable for a microinjection device and is characterized in that the microinjection device at least comprises a needle pulling instrument, a fixed needle, an injection needle, a fixed needle operating arm, an injection needle operating arm and a microscope, wherein the fixed needle is rotatably connected with the fixed needle operating arm, the injection needle is rotatably connected with the injection needle operating arm, the microscope comprises a stage, a cell carrier is arranged on the stage, a first knob is arranged on the fixed needle operating arm, a second knob is arranged on the injection needle operating arm, and the fixed needle has a bending angle of 30 degrees;
the method of microinjection manipulation includes the steps of:
the method comprises the following steps: placing the cells in a cell carrier on a microscope stage;
step two: adjusting an injection needle operating arm through a second knob to enable the needle point of the injection needle to be wholly tiled at the bottom of the cell carrier;
step three: rotating the fixing needle to enable the bent back of the fixing needle to face upwards;
step four: the fixed needle operating arm is adjusted through the first knob, so that the needle head of the fixed needle is slowly contacted with the bottom of the vessel;
step five: the needle operating arm is adjusted through the second knob, and the fixed needle operating arm is adjusted through the first knob at the same time, so that the needle point of the injection needle and the needle head of the fixed needle are on the same horizontal plane;
step six: selecting the position of the needle point of the injection needle, and finely adjusting the fixed needle operating arm through a first knob;
step seven: breaking the needle point of the injection needle by using a fixed needle head, and completing needle breaking; the method is used for non-disease diagnostic and therapeutic purposes.
2. A method of microinjection manipulation according to claim 1, wherein the inside diameter of the needle opening after the needle break is 3 to 5 μm.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
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| CN201711315349.4A CN108070618B (en) | 2017-12-12 | 2017-12-12 | Microinjection operation method |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201711315349.4A CN108070618B (en) | 2017-12-12 | 2017-12-12 | Microinjection operation method |
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| CN108070618A CN108070618A (en) | 2018-05-25 |
| CN108070618B true CN108070618B (en) | 2021-03-05 |
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| CN201711315349.4A Active CN108070618B (en) | 2017-12-12 | 2017-12-12 | Microinjection operation method |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN109402220A (en) * | 2018-11-28 | 2019-03-01 | 安徽农业大学 | A method of collecting blastocoele liquid |
| US11402331B2 (en) * | 2019-05-08 | 2022-08-02 | City University Of Hong Kong | Imaging and manipulation of biological sample |
| CN112779119A (en) * | 2019-11-08 | 2021-05-11 | 中国科学院大连化学物理研究所 | Small manual five-dimensional needle tip adjusting frame for cell experiment |
| CN111693196A (en) * | 2020-06-16 | 2020-09-22 | 梁诗豪 | Live embryo surface zona pellucida tension detection device |
| NL2029490B1 (en) * | 2021-10-22 | 2023-05-19 | Life Science Methods B V | Microinjection system and method |
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| JPH06343478A (en) * | 1993-06-08 | 1994-12-20 | Hitachi Ltd | Micro-injection method and apparatus |
| CN1763170A (en) * | 2004-10-19 | 2006-04-26 | 章维一 | Cell microinjection instrument |
| CN201067425Y (en) * | 2007-08-17 | 2008-06-04 | 周淑梅 | Oocyte micro-injection puncture needle |
| CN101481653A (en) * | 2008-01-11 | 2009-07-15 | 南京理工大学 | Single cell microoperation apparatus for microscopic injection |
| CN103255049A (en) * | 2013-05-20 | 2013-08-21 | 苏州大学 | Composite piezoelectric injection system and injection method |
| CN203938668U (en) * | 2014-06-18 | 2014-11-12 | 华南理工大学 | A kind of automatization microinjection device |
| CN104493825A (en) * | 2014-10-29 | 2015-04-08 | 北京理工大学 | Cooperating device for micro-nano operating robot and operating method for microinjection |
| CN107245431A (en) * | 2017-08-04 | 2017-10-13 | 重庆三峡医药高等专科学校 | A kind of microinjection device precisely injected for cell drug and its operating method |
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2017
- 2017-12-12 CN CN201711315349.4A patent/CN108070618B/en active Active
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH06343478A (en) * | 1993-06-08 | 1994-12-20 | Hitachi Ltd | Micro-injection method and apparatus |
| CN1763170A (en) * | 2004-10-19 | 2006-04-26 | 章维一 | Cell microinjection instrument |
| CN201067425Y (en) * | 2007-08-17 | 2008-06-04 | 周淑梅 | Oocyte micro-injection puncture needle |
| CN101481653A (en) * | 2008-01-11 | 2009-07-15 | 南京理工大学 | Single cell microoperation apparatus for microscopic injection |
| CN103255049A (en) * | 2013-05-20 | 2013-08-21 | 苏州大学 | Composite piezoelectric injection system and injection method |
| CN203938668U (en) * | 2014-06-18 | 2014-11-12 | 华南理工大学 | A kind of automatization microinjection device |
| CN104493825A (en) * | 2014-10-29 | 2015-04-08 | 北京理工大学 | Cooperating device for micro-nano operating robot and operating method for microinjection |
| CN107245431A (en) * | 2017-08-04 | 2017-10-13 | 重庆三峡医药高等专科学校 | A kind of microinjection device precisely injected for cell drug and its operating method |
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| CN108070618A (en) | 2018-05-25 |
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