CN114231389B - Rhesus monkey cochlear membrance labyrinth tissue cell nondestructive extraction device and use method - Google Patents
Rhesus monkey cochlear membrance labyrinth tissue cell nondestructive extraction device and use method Download PDFInfo
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- C12M33/00—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus
- C12M33/10—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus by centrifugation ; Cyclones
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- C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
- C12M41/12—Means for regulation, monitoring, measurement or control, e.g. flow regulation of temperature
- C12M41/18—Heat exchange systems, e.g. heat jackets or outer envelopes
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- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
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- C12N2509/00—Methods for the dissociation of cells, e.g. specific use of enzymes
- C12N2509/10—Mechanical dissociation
Abstract
The invention discloses a rhesus monkey cochlear membrane labyrinth tissue cell nondestructive extraction device and a use method, relates to the technical field of biological cell extraction, and solves the technical problem that cochlear tissue cells cannot be obtained through clinical human body material taking; the extraction method is used for clinically extracting the tissue cells, and has the advantage that the cochlear tissue cells are obtained by human body material drawing.
Description
Technical Field
The invention relates to the technical field of biological cell extraction, in particular to the technical field of a rhesus monkey cochlear membrane labyrinth tissue cell nondestructive extraction device and an extraction method.
Background
Compared with animals such as mice and pigs, the rhesus monkey which is the primate animal similar to human has higher gene homology similarity, so that the rhesus monkey is more advantageous to be used as a research object for researching human pathogenic genes. The cochlear tissue cells are deeply buried in the temporal bone to form a spiral structure, and the labyrinthic lumen is extremely narrow, so the cochlear tissue cells can hardly be obtained by clinical human body material drawing. Therefore, it is necessary to study the physiological basis of cochlea and the mechanism of deafness in human beings by means of the cochlear periosteum labyrinth tissue cells of rhesus monkey. At present, due to the particularity of the dissecting structure of the cochlear duct labyrinth tissue of the rhesus monkey, it is difficult to obtain complete and nondestructive cochlear duct labyrinth tissue cells of the rhesus monkey through a conventional dissecting experiment, so that the nondestructive extraction device for the cochlear duct labyrinth tissue cells of the rhesus monkey is urgently needed to be invented.
The prior art is as follows: because the cochlear tissue cells are deeply buried in the temporal bone to form a spiral structure and the labyrinth tube cavity is extremely narrow, the bone grinding method is usually adopted to extract the cochlear membrane labyrinth tissue cells of the rhesus monkey at present. The technical scheme of the bone grinding method includes that firstly, a cutting drill is used for cutting the cochlea structure of the rhesus monkey integrally. And then, grinding the cochlear structure of the rhesus monkey through a grinding drill to fully expose cochlear tissue cells of the cochlear membrane of the rhesus monkey. And finally, stripping the cochlear duct labyrinth tissue cells of the rhesus monkey by using a crochet hook.
The technical scheme of the bone grinding method has the following defects:
(1) Because the rhesus cochlear tissue cells are deeply buried in the temporal bone to form a spiral structure, the requirement of extracting the structure on the anatomical skill of test operators is very high, and few experimenters can complete the operation.
(2) The damage to the lumen of the membrane labyrinth is easily caused in the polishing process of the cochlear membrane of the rhesus monkey, the damage to the tissue cells of the membrane labyrinth is further caused, and the tissue cell waste is caused.
(3) During the polishing process of the rhesus monkey cochlear membrane, the abrasion drill rubs with sclerotin to generate high temperature, so that the activity of the tissue cells of the membrane labyrinth is further reduced, and the damage of the tissue cells of the membrane labyrinth is aggravated.
(4) The whole bone grinding experiment takes about 2 hours, the activity of the membranous labyrinth tissue cells is reduced due to the long-time in-vitro state, the protein is degraded, and the destruction of the membranous labyrinth tissue cells is further aggravated.
(5) The whole bone grinding experiment is carried out in an unsealed environment, membrane labyrinth tissue cells are easily polluted, and subsequent experiment data are influenced.
Disclosure of Invention
The invention aims to: in order to solve the problem that cochlear tissue cells cannot be obtained by clinical human body material drawing, the invention provides a rhesus monkey cochlear membrane labyrinth tissue cell nondestructive extraction device and an extraction method.
The invention specifically adopts the following technical scheme for realizing the purpose:
the utility model provides a harmless extraction element of rhesus monkey cochlea membrane labyrinth tissue cell, includes body, upper end cover, side tray, centrifuge module and lower end cover, the upper end of body have the upper end cover passes through internal thread pin joint with the body, and hydraulic interface passes through internal thread pin joint with the upper end cover to adopt the rubber circle sealed, the side tray is located the left end of body and through its inside guide rail and body gomphosis, and the lower extreme of body has the lower end cover, the lower end cover passes through internal thread pin joint with the body, and the right-hand member of body has cell extract filling opening and cell separation liquid discharge port, cell extract filling opening and cell separation liquid discharge port pass through internal thread pin joint with the body, and cell extract filling opening and cell separation liquid discharge port pass through internal thread pin joint with the body to adopt the rubber circle sealed, cell extract filling opening and cell separation liquid discharge port lead to the inside of body.
Further, the upper end cover is in contact connection with an I-shaped upper top shaft through a spring, the I-shaped upper top shaft and the body are sealed by a rubber ring, and the cell extracting solution injection port is finally connected to the cell separating solution discharge port through the I-shaped upper top shaft, the rhesus monkey cochlea, the pressure-bearing tray and the centrifugal groove.
Further, the lower end of the I-shaped upper jacking shaft is provided with a baffle plate, and the baffle plate presses the rhesus cochlea on the upper end of the pressure-bearing tray.
Further, the pressure-bearing tray is placed in a groove formed above the centrifugal groove, the centrifugal groove is connected with the centrifuge module through a key groove, and the centrifuge module is fixed at the upper end of the lower end cover through welding.
Further, the side tray is connected with the power supply wiring port through an internal thread pin and is sealed by a rubber ring.
Further, a constant-temperature heating module is embedded in the side tray, the constant-temperature heating module is communicated with the power supply wiring port through an electric wire, and the side tray is hermetically embedded with the pressure-bearing tray through a groove structure.
A use method of a rhesus monkey cochlear membrance labyrinth tissue cell nondestructive extraction device comprises the following steps:
the first step is as follows: the centrifugal groove is placed in a groove at the right end of the side tray, the pressure-bearing tray is placed in a groove at the upper end of the centrifugal groove, the pressure-bearing tray and the side tray are sealed through rubber rings, and the rhesus cochlea is placed at the upper end of the pressure-bearing tray and is in contact with the three groups of rubber sealing rings on the pressure-bearing tray.
The second step is that: the side tray is embedded into the left end of the body through the internal guide rail, the side tray is fixed to the body through the internal thread pin, and meanwhile, the power supply wiring port is connected with a power line to electrify the constant-temperature heating module, so that the temperature of a middle cavity of the body is maintained at 37 ℃.
The third step: the upper end cover is fixed above the body through an internal thread pin, hydraulic oil is injected through the hydraulic interface to enable the I-shaped upper jacking shaft to move downwards, the pressure of the hydraulic oil is maintained at 0.2-0.5MPa, and the fact that the baffle at the lower part of the I-shaped upper jacking shaft presses the rhesus monkey cochlea on the pressure-bearing tray to be firmly fixed is guaranteed.
The fourth step: the whole centrifuge module welded on the upper end of the lower end cover is migrated into the lower end of the body and fixed on the body through the internal thread pin, and the centrifugal groove is connected with the centrifuge module through the key groove.
The fifth step: injecting cell extract from the cell extract injection port, maintaining the injection pressure at 0.5Mpa, allowing the cell extract to enter through one end of the cochlear lumen tube of rhesus monkey, discharging the cells carrying cochlear tissue cells from the other end of the cochlear lumen tube, starting the centrifuge module to centrifuge the cell extract carrying cochlear tissue cells, and discharging the liquid without cochlear tissue cells through the cell separation liquid discharge port.
And a sixth step: and starting timing after liquid is returned from the cell separation liquid outlet, stopping injecting the cell extraction liquid from the cell extraction liquid injection port after 30min, stopping the work of the centrifuge module, relieving the pressure of the hydraulic interface, moving the I-shaped upper jacking shaft upwards under the action of the pulling force of the spring, and separating the cochlea of the rhesus monkey from the baffle plate at the lower end of the I-shaped upper jacking shaft.
The seventh step: and the lower end cover and the centrifuge module are integrally moved out from the lower part of the body, so that the upper end of the centrifuge module is separated from the key groove of the centrifugal groove.
Eighth step: and taking out the side tray, and quickly moving the rhesus cochlear membrane labyrinth tissue cells in the centrifugal groove into a cell culture dish to finish extraction of the rhesus cochlear membrane labyrinth tissue cells.
The invention has the following beneficial effects:
1. the method is used for extracting the cochlear tissue cells of the rhesus monkey, the operation is simpler and faster, the nondestructive extraction of the cochlear tissue cells of the rhesus monkey can be realized, the whole operation is carried out under the closed constant temperature condition, the activity of the cochlear tissue cells of the membrane labyrinth is ensured, the cochlear tissue cells are not polluted by the external environment, and the reliability of later-period experimental data is further ensured.
2. The operation is simpler and more convenient by using the invention, and common experimenters can operate the invention; the operation is quicker, only about 0.5h is needed for completing one experiment, and 1.5h is saved compared with a bone grinding method.
3. The invention relates to a constant-temperature and closed extraction structure for cochlear duct-labyrinthine tissue cells of rhesus monkeys.
4. The cochlea of the rhesus monkey is pressed between the baffle plate arranged at the lower end of the I-shaped upper top shaft and the upper end of the pressure-bearing tray to fixedly extract a membrane of the cochlea of the rhesus monkey, and the rotation of the centrifugal groove does not cause the damage of a lumen of a membranous labyrinth and the waste of tissue cells in the grinding process.
5. The centrifugal groove does not rub against bone of the cochlear membrane of the rhesus monkey to generate high temperature in the rotating process, so that the activity of the tissue cells of the membrane labyrinth is further improved.
Drawings
Fig. 1 is a schematic structural view of the present invention.
Reference numerals are as follows: the cell separation device comprises a body 1, an upper end cover 2, a hydraulic interface 3, an I-shaped upper top shaft 4, a side tray 5, a rhesus monkey cochlea 6, a cell extract liquid injection port 7, a cell separation liquid discharge port 8, a power connection port 9, a constant temperature heating module 10, a centrifuge module 11, a lower end cover 12, a centrifugal groove 13 and a pressure-bearing tray 14.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, 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 some, but not all, embodiments of the present invention. The components of embodiments of the present invention generally described and illustrated in the figures herein may be arranged and designed in a wide variety of different configurations.
Example 1
As shown in fig. 1, the present embodiment provides a rhesus monkey cochlear duct tissue cell nondestructive extraction device, which includes a body 1, an upper end cap 2, a side tray 5, a centrifuge module 11 (connected to a power supply from the front end or the rear end of the body 1) and a lower end cap 12, wherein the upper end cap 2 provided at the upper end of the body 1 is connected to the body 1 through an internal thread pin, a hydraulic interface 3 is connected to the upper end cap 2 through an internal thread pin and sealed by a rubber ring, the side tray 5 is located at the left end of the body 1 and is embedded to the body 1 through an internal guide rail thereof and is fixedly connected by a pin, the lower end of the body 1 is provided with a lower end cap 12, the lower end cap 12 is connected to the body 1 through an internal thread pin, the right end of the body 1 is provided with a cell extract injection port 7 and a cell separation liquid discharge port 8, the cell extract port 7 and the cell separation liquid discharge port 8 are connected to the body 1 through an internal thread pin and sealed by a rubber ring, and the cell extract port 7 and the cell separation liquid discharge port 8 are led to the inside of the body 1.
Specifically, the upper end cover 2 is in contact connection with an I-shaped upper top shaft 4 through a spring, the I-shaped upper top shaft 4 and the body 1 are sealed through a rubber ring, the cell extracting solution injection port 7 is finally communicated with the cell separating solution discharge port 8 through the I-shaped upper top shaft 4, the rhesus monkey cochlea 6, the pressure-bearing tray 14 and the centrifugal groove 13, the lower end of the I-shaped upper top shaft 4 is provided with a baffle, and the rhesus monkey cochlea 6 is pressed on the upper end of the pressure-bearing tray 14 through the baffle.
The pressure-bearing tray 14 is placed in a groove formed above the centrifugal groove 13, the centrifugal groove 13 is connected with the centrifuge module 11 through a key groove, and the centrifuge module 11 is fixed at the upper end of the lower end cover 12 through welding.
The side tray 5 is connected with the power supply wiring port 9 through an internal thread pin and sealed through a rubber ring, a constant-temperature heating module 10 is embedded in the side tray 5, the constant-temperature heating module 10 is communicated with the power supply wiring port 9 through an electric wire, and the side tray 5 is hermetically embedded with the pressure-bearing tray 14 through a groove structure.
Example 2
A use method of a rhesus monkey cochlear membrance labyrinth tissue cell nondestructive extraction device comprises the following steps:
the first step is as follows: the centrifugal groove 13 is placed in a groove at the right end of the side face tray 5, the pressure bearing tray 14 is placed in a groove at the upper end of the centrifugal groove 13, the pressure bearing tray 14 and the side face tray 5 are sealed through rubber rings, and the rhesus monkey cochlea 6 is placed at the upper end of the pressure bearing tray 14 and is in contact with the three groups of rubber sealing rings on the pressure bearing tray 14.
The second step is that: the side tray 5 is embedded into the left end of the body 1 through an internal guide rail, is fixed on the body 1 through an internal thread pin, and is connected with an electric wire through a power supply wiring port 9 to electrify the constant-temperature heating module 10, so that the temperature of the middle chamber of the body 1 is maintained at 37 ℃.
The third step: the upper end cover 2 is fixed above the body 1 through an internal thread pin, hydraulic oil is injected through the hydraulic interface 3 to enable the I-shaped upper jacking shaft 4 to move downwards, the hydraulic oil pressure is maintained at 0.2-0.5MPa, and the fact that the rhesus monkey cochlea 6 is pressed on the pressure bearing tray 14 to be firmly fixed by a baffle plate at the lower part of the I-shaped upper jacking shaft 4 is guaranteed.
The fourth step: the whole centrifuge module 11 welded on the upper end of the lower end cover 12 is migrated into the lower end of the body 1 and fixed on the body 1 through the internal thread pin, and the centrifuge groove 13 is connected with the centrifuge module 11 through the key groove.
The fifth step: injecting cell extract from the cell extract injection port 7, maintaining the injection pressure at 0.5Mpa, allowing the cell extract to enter through one end of the 6-lumen tube of the cochlea of the rhesus monkey, discharging cells carrying cochlear tissue cells from the other end of the lumen tube, starting the centrifuge module 11 to centrifuge the cell extract carrying cochlear tissue cells, and discharging the liquid without containing cochlear tissue cells through the cell separation liquid discharge port 8.
And a sixth step: and starting timing after liquid is returned from the cell separation liquid outlet 8, stopping injecting the cell extraction liquid from the cell extraction liquid injection port 7 after 30min, stopping the work of the centrifuge module 11, unloading the pressure of the hydraulic interface 3, moving the I-shaped upper jacking shaft 4 upwards under the action of the spring tension, and separating the rhesus cochlea 6 from the baffle plate at the lower end of the I-shaped upper jacking shaft 4.
The seventh step: the lower end cap 12 and the centrifuge module 11 are integrally removed from below the body 1, so that the upper end of the centrifuge module 11 is separated from the key groove of the centrifugal groove 13.
Eighth step: and (3) taking out the side tray 5, and quickly moving the rhesus cochlear membrane labyrinth tissue cells in the centrifugal groove 13 into a cell culture dish, thereby finishing the extraction of the rhesus cochlear membrane labyrinth tissue cells.
The foregoing is merely a preferred embodiment of the invention, which is intended to be illustrative and not limiting. It will be understood by those skilled in the art that various changes, modifications and equivalents may be made therein without departing from the spirit and scope of the invention as defined in the appended claims.
Claims (2)
1. The utility model provides a harmless extraction element of rhesus monkey cochlea membrane labyrinth tissue cell, includes body (1), upper end cover (2), side tray (5), centrifuge module (11) and lower end cover (12), its characterized in that: the upper end cover (2) arranged at the upper end of the body (1) is connected with the body (1) through an internal thread pin, the hydraulic interface (3) is connected with the upper end cover (2) through the internal thread pin and sealed by a rubber ring, the side tray (5) is positioned at the left end of the body (1) and is embedded with the body (1) through an internal guide rail, the lower end of the body (1) is provided with a lower end cover (12), the lower end cover (12) is connected with the body (1) through the internal thread pin, the right end of the body (1) is provided with a cell extracting solution injection port (7) and a cell separating solution discharge port (8), the cell extracting solution injection port (7) and the cell separating solution discharge port (8) are connected with the body (1) through the internal thread pin and sealed by the rubber ring, and the cell extracting solution injection port (7) and the cell separating solution discharge port (8) lead to the inside of the body (1);
the upper end cover (2) is in contact connection with an I-shaped upper top shaft (4) through a spring, the I-shaped upper top shaft (4) and the body (1) are sealed by a rubber ring, and the cell extracting solution injection port (7) finally reaches a cell separating solution discharge port (8) through the I-shaped upper top shaft (4), a rhesus cochlea (6), a pressure-bearing tray (14) and a centrifugal groove (13);
the lower end of the I-shaped upper jacking shaft (4) is provided with a baffle plate, and the baffle plate presses the rhesus cochlea (6) on the upper end of the pressure-bearing tray (14);
the pressure-bearing tray (14) is placed in a groove formed above the centrifugal groove (13), the centrifugal groove (13) is connected with the centrifuge module (11) through a key groove, and the centrifuge module (11) is fixed at the upper end of the lower end cover (12) through welding;
the side tray (5) is connected with the power supply wiring port (9) through an internal thread pin and is sealed by a rubber ring;
a constant-temperature heating module (10) is embedded in the side tray (5), the constant-temperature heating module (10) is communicated with the power supply wiring port (9) through an electric wire, and the side tray (5) is hermetically embedded with the pressure-bearing tray (14) through a groove structure;
the cochlea of the rhesus monkey is pressed between the baffle plate arranged at the lower end of the I-shaped upper top shaft and the upper end of the pressure-bearing tray so as to fix and extract the cochlea membrane of the rhesus monkey.
2. The use method of the rhesus monkey cochlear membrance labyrinth tissue cell nondestructive extraction device according to claim 1 is characterized in that: the method comprises the following steps:
the first step is as follows: placing a centrifugal groove (13) in a groove at the right end of a side surface tray (5), placing a pressure-bearing tray (14) in a groove at the upper end of the centrifugal groove (13), sealing the pressure-bearing tray (14) and the side surface tray (5) through rubber rings, and placing a rhesus cochlea (6) at the upper end of the pressure-bearing tray (14) to be contacted with three groups of rubber sealing rings on the pressure-bearing tray (14);
the second step: embedding a side tray (5) into the left end of the body (1) through an internal guide rail, fixing the side tray to the body (1) through an internal thread pin, and simultaneously connecting a power supply wiring port (9) with a wire to electrify the constant-temperature heating module (10) so as to maintain the temperature of a middle cavity of the body (1) at 37 ℃;
the third step: the upper end cover (2) is fixed above the body (1) through an internal thread pin, hydraulic oil is injected through the hydraulic interface (3) to enable the I-shaped upper jacking shaft (4) to move downwards, the hydraulic oil pressure is maintained at 0.2-0.5MPa, and the fact that the rhesus cochlea (6) is pressed on the pressure-bearing tray (14) and is firmly fixed by a baffle at the lower part of the I-shaped upper jacking shaft (4) is guaranteed;
the fourth step: integrally moving a centrifuge module (11) welded at the upper end of a lower end cover (12) into the lower end of the body (1), fixing the centrifuge module to the body (1) through an internal thread pin, and connecting a centrifuge groove (13) with the centrifuge module (11) through a key groove;
the fifth step: injecting cell extracting solution from a cell extracting solution injection port (7), wherein the injection pressure is maintained at 0.5Mpa, the cell extracting solution enters through one end of a cavity tube of a rhesus monkey cochlea (6) and is discharged from the other end of the cavity tube with cochlear tissue cells, a centrifuge module (11) is started to carry out centrifugal treatment on the cell extracting solution with cochlear tissue cells, and liquid without cochlear tissue cells is discharged through a cell separation solution discharge port (8);
and a sixth step: starting timing after liquid begins to return from the cell separation liquid outlet (8), stopping injecting the cell extraction liquid from the cell extraction liquid injection port (7) after 30min, stopping the work of the centrifuge module (11), unloading the pressure of the hydraulic interface (3), moving the I-shaped upper jacking shaft (4) upwards under the action of spring tension, and separating the rhesus cochlea (6) from a baffle plate at the lower end of the I-shaped upper jacking shaft (4);
the seventh step: integrally removing the lower end cover (12) and the centrifuge module (11) from the lower part of the body (1) to separate the upper end of the centrifuge module (11) from the key groove of the centrifugal groove (13);
eighth step: and (4) taking out the side tray (5), and quickly moving the cochlear labyrinth tissue cells in the centrifugal groove (13) into a cell culture dish to finish extraction of the cochlear labyrinth tissue cells of the rhesus.
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| CN113005087B (en) * | 2021-03-22 | 2022-05-03 | 澎立生物医药技术(上海)股份有限公司 | Adherent culture method of cochlear organ of mouse |
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| CN213951215U (en) * | 2020-11-10 | 2021-08-13 | 浙江大学医学院附属第一医院 | In vitro type liver cell extraction device |
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