CN112586491A - Device and method for preparing mouse intestine volume label book - Google Patents
Device and method for preparing mouse intestine volume label book Download PDFInfo
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- CN112586491A CN112586491A CN202011571509.3A CN202011571509A CN112586491A CN 112586491 A CN112586491 A CN 112586491A CN 202011571509 A CN202011571509 A CN 202011571509A CN 112586491 A CN112586491 A CN 112586491A
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Abstract
A device and a method for preparing a mouse intestine volume label book relate to the technical field of animal specimen preparation. The preparation device of the mouse intestine volume label comprises a winder and a wax strip; the winder comprises a handle and a clamping seat fixedly connected to one end of the handle, and the end part of the clamping seat far away from the handle is provided with a jack; one end of the wax strip is inserted in the insertion hole of the clamping seat, and the other end of the wax strip extends out of the insertion hole of the clamping seat. The preparation method of the mouse intestine volume label book is applied to the preparation device of the mouse intestine volume label book. The invention has the advantages that: 1. the Bouins liquid is used for pre-hardening the intestinal skin, which is beneficial to keeping the smoothness of the intestinal skin. The pulling and pressing of the intestine skin operation on the intestine skin can be reduced, the operation difficulty in the process of winding the intestine skin is reduced on one hand, and the integrity of the intestinal tract morphological structure is ensured on the other hand. 2. The preparation method of the sausage roll is simple and easy to operate, has short time consumption, can block endogenous or exogenous enzyme reaction in time, and is favorable for ensuring the stability of protein in small intestine tissues.
Description
Technical Field
The invention relates to the technical field of preparation of animal specimens, in particular to a device and a method for preparing a mouse intestine volume label book.
Background
The intestinal tract is an important organ for conversion and absorption of organic matters, water, poisons and medicaments, and is different from other parenchymal organ tissues, and the intestinal tract tissue has a tubular cavity-like structure. The pathophysiological verification of intestinal tissue is an indispensable part of medical experimental research, in which small animals such as rats, mice, rabbits, etc. are generally used as the collection objects of intestinal tract samples. How to keep the integrity of the intestinal morphological structure and the stability of the detected protein in the acquisition and fixation part of the sample is a technical problem which troubles researchers in laboratories.
The existing experimental animal intestinal tissue fixing method comprises a transverse cutting fixing method and a longitudinal cutting fixing method.
The cross-cut fixation method operates as follows: 1. sampling intestinal tract tissues and transversely cutting the intestinal tract tissues at a proper position to form a circular incision; 2. soaking the cut intestinal tissue into formalin solution for dehydration and preservative treatment; 3. the dehydrated and preserved intestinal tissue was embedded in paraffin and the incision was revealed intact. Intestinal structures such as intestinal villi, intestinal muscular layer, intestinal mucosa and the like can be shown through the incision. But the transverse cutting fixing method has the following defects: the intestinal structure that can only be revealed by incision is very limited and does not fully present the full picture of the intestinal tissue.
The slitting fixing method is operated as follows: 1. sampling intestinal tract tissues, and cutting the intestinal tract along the length direction (longitudinal direction) of the intestinal tract tissues to form a rectangular intestinal tract skin, wherein one surface of the intestinal tract skin is the outer wall of the intestinal tract, and the other surface of the intestinal tract skin is the inner wall of the intestinal tract; 2. soaking the intestinal skin into formalin solution for dehydration and preservative treatment; 3. and (4) spreading and flattening the dehydrated and antiseptic intestinal skin, and embedding the intestinal skin into paraffin. Intestinal structures such as intestinal villi, intestinal muscular layers, intestinal mucosa and the like can be displayed through the unfolded intestinal skin. But the longitudinal cutting fixing method has the following defects: 1. when the intestine skin is soaked in formalin solution, the intestine skin is twisted, overturned and bent due to dehydration, so that the subsequent leveling operation is not difficult; 2. when the intestine skin is flattened, the operations such as pressing, stretching and the like are needed, which is difficult to avoid causing the intestinal injury and is not beneficial to keeping the integrity of the intestinal morphological structure.
Disclosure of Invention
The invention aims to overcome the defects of the prior art, and provides a device and a method for preparing a mouse intestinal volume label book, which solve the problems that the existing mouse intestinal tissue fixing method cannot completely show the full appearance of the intestinal tissue by a transverse cutting fixing method, has high operation difficulty by a longitudinal cutting fixing method, and is easy to cause intestinal injury.
The technical scheme of the invention is as follows: the preparation device of the mouse intestine volume label comprises a winder and a wax strip; the winder comprises a handle and a clamping seat fixedly connected to one end of the handle, and the end part of the clamping seat far away from the handle is provided with a jack; one end of the wax strip is inserted in the insertion hole of the clamping seat, and the other end of the wax strip extends out of the insertion hole of the clamping seat.
The further technical scheme of the invention is as follows: the insertion hole of the clamping seat is a regular hexagonal prism-shaped hole, a chamfer is arranged at the hole opening of the insertion hole, and correspondingly, the wax strip is in a regular hexagonal prism shape and is adaptive to the size of the insertion hole.
The invention further adopts the technical scheme that: the diameter of the largest inscribed circle of the cross section of the wax strip is 10mm, and the depth of the jack of the clamping seat is 20 mm.
The further technical scheme of the invention is as follows: the handle is a regular quadrangular prism-shaped rod, and the axis of the wax strip is coincided with the axis of the handle.
The further technical scheme of the invention is as follows: the centering swing mechanism comprises an insert sleeve, a bearing seat and an end cover; an inner cavity for mounting a bearing is arranged in the bearing seat, and an annular step surface is arranged in the inner cavity; the plug bush is cylindrical, the center of the plug bush is provided with a through hole A which runs through the end faces of the two sides of the plug bush, and the through hole A is matched with the appearance size of the handle; the bearing is sleeved on the insertion sleeve and is arranged in the inner cavity of the bearing seat, one end of the bearing is positioned through an annular step surface in the inner cavity of the bearing seat, and the other end of the bearing is positioned through the end cover; the end cover is arranged at one end of the bearing seat and is fixedly connected with the bearing seat and the plug bush, and the center of the end cover is provided with a through hole B for the handle to pass through; the centering and rotating device is inserted into the handle of the winder through a through hole A on the plug bush.
The technical scheme of the invention is as follows: the preparation method of the mouse intestine volume label book comprises the following steps:
s01, sampling mouse small intestine:
a. anaesthetizing the mouse, fixing the mouse on an operation table, exposing the chest wall, and performing cardiac puncture bleeding until the mouse loses hemorrhagic shock;
b. after the mouse dies, sterilizing the abdomen, cutting the abdominal wall, exposing the abdominal cavity, dissociating and cutting off the cardia part of the upper section of the small intestine, ligating the opening of the upper section of the small intestine, dissociating and cutting off the ileocaecal part of the lower section of the small intestine;
c. physiological saline is injected into the small intestine through the injector to wash the inner cavity of the small intestine, so that the content in the inner cavity of the small intestine is discharged from the lower section incision of the intestinal tract;
d. ligating an opening at the lower section of the small intestine, dissociating the whole small intestine, and injecting 2ml of physiological saline into the dissociated small intestine by using a syringe so as to avoid cavity wall adhesion caused by the cavity collapse in the small intestine;
in the step, the adopted anesthetic is isoflurane, and the dosage is lethal dose;
in the step, the distance between the ligation point of the upper segment of the small intestine and the incision of the upper segment of the small intestine is 1-2 cm;
in the step, the injection point of the injector is positioned at the rear end of the ligation point of the upper segment of the small intestine by 1-2 cm;
s02, preparing a rectangular sausage skin:
placing the small intestine injected with the physiological saline into a glass dish, cutting a transverse cut at a position close to a ligation opening at the upper section of the small intestine, extending a cutting edge of a scissors into the inner cavity of the small intestine from the transverse cut, cutting the small intestine from top to bottom along the longitudinal direction of the small intestine, cutting the small intestine at the upper section of the transverse cut, and cutting a ligation line at the lower section of the small intestine to finally form a rectangular intestine skin;
s03, hardening treatment of the rectangular intestine skin:
spreading the rectangular sausage skin on an operation table top with the fluff surface facing upwards, spreading the rectangular sausage skin on the operation table top, then spreading Bouins liquid on the rectangular sausage skin, and hardening the rectangular sausage skin for 5-10min to fix the shape of the rectangular sausage skin, so that subsequent sausage roll manufacturing is facilitated;
s04, preparing an intestine roll:
a. taking one end of the rectangular sausage skin by forceps pincers, placing the end on a wax strip, ensuring that the fluff surface of the rectangular sausage skin faces the surface of the wax strip, then rotating a handle, and driving the wax strip to rotate through the handle, so that the rectangular sausage skin is wound around the wax strip one by taking the wax strip as an axis to form a cylindrical sausage roll;
b. after the connection between the wax strips and the clamping seat is disconnected, the wax strips and the intestine rolls are placed in the Bouins solution for soaking, and the intestine rolls are hardened to fix the shape of the intestine rolls and prevent the intestine rolls from being scattered or deformed when the intestine rolls are subsequently soaked in the formalin solution;
c. taking out the wax strips and the intestinal rolls from the Bouins solution, transferring the wax strips and the intestinal rolls into a formalin solution, soaking for 6-10 hours, further fixing tissues of small intestines to stop endogenous or exogenous enzyme reaction, preventing autolysis or autolysis of the tissues, keeping the original shape of the tissues and avoiding antigen inactivation or dispersion;
d. cutting along the radial surface of the intestine roll by a sharp cutting edge, cutting tissues at two ends of the intestine roll and wax strip parts beyond the two ends of the intestine roll, only keeping the axial width of the intestine roll of 2-3cm, finally obtaining a cylindrical intestine roll according with the size of an embedding box, and placing the cylindrical intestine roll in the embedding box;
s05, dehydrating the intestinal roll:
a. washing the embedding box in flowing water for 10min to fully remove the formalin solution on the cylindrical intestine roll;
b. soaking the embedding box in a dehydration box A filled with 75% alcohol for 80 min;
c. taking out the embedding box from the dehydration box A, and soaking in a dehydration box B filled with 85% alcohol for 80 min;
d. taking out the embedding box from the dehydrating box B, and soaking in the dehydrating box C filled with 95% alcohol for 80 min;
e. taking out the embedding box from the dehydrating box C, and soaking in the dehydrating box D filled with pure alcohol for 80 min;
f. taking out the embedding box from the dewatering box D, and soaking in the dewatering box E filled with pure alcohol for 80 min;
s06, waxing the intestinal roll:
a. taking out the embedding box from the dehydrating box E, and soaking in the solution box A filled with the xylene stock solution for 20 min;
b. taking out the embedding box from the wax dipping box A, and soaking in a solution box B filled with a xylene stock solution for 30 min;
c. placing the wax dipping box A filled with the soft wax in an oven for baking to melt the soft wax, taking out the embedding box from the solution box B, and placing the embedding box in the wax dipping box A to soak the liquid soft wax for 60 min;
d. placing the wax dipping box B filled with the soft wax in an oven to be baked to be softened into a liquid state, taking the embedding box out of the wax dipping box A, and placing the embedding box in the wax dipping box B to be dipped in the liquid soft wax for 60 min;
e. placing the wax dipping box C filled with the hard wax in an oven to be baked to be softened into a liquid state, taking the embedding box out of the wax dipping box B, and placing the embedding box in the wax dipping box C to be dipped in the liquid hard wax for 120 min;
in the step, in the process of soaking the soft wax, setting the temperature of an oven to be 58-66 ℃ to maintain the melting state of the soft wax;
in the step, in the process of soaking the hard wax, setting the temperature of an oven to be 58-66 ℃ to maintain the melting state of the hard wax;
s07, embedding the steamed roll:
a. rolling the dehydrated and waxed cylindrical sausage out, and placing the rolled cylindrical sausage in the center of the bottom of the groove of the embedding mold;
b. filling the whole embedding mold groove with liquid hard wax, and covering the cover of the embedding mold;
c. placing the whole embedding mould in a refrigerated cabinet with the temperature of-18 to-22 ℃ for standing for 20 minutes;
d. and taking down the cover of the embedding mold, inverting the embedding mold, and tapping the embedding mold to completely remove the wax block containing the intestinal roll from the embedding mold.
Compared with the prior art, the invention has the following advantages:
1. in the method for making the sausage roll, the Bouins liquid is used for pre-hardening the sausage skin, so that the hardness of the sausage skin can be improved to a certain extent, and the smoothness of the sausage skin can be kept. Reducible follow-up coiling intestines skin operation is to dragging and pressing of intestines skin, (dragging and pressing are in order to flatten the intestines skin arrangement, but drag and press and can destroy the small intestine fine hair, enlarge the small intestine and hold the clearance, destroy the integrality of intestinal form structure), the operation degree of difficulty when having reduced coiling intestines skin on the one hand, it is long to be favorable to shortening the preparation of intestines skin, on the other hand is owing to reduced dragging and pressing to the intestines skin, and then has guaranteed the integrality of intestinal form structure.
2. The small intestine is subjected to autolysis caused by endogenous or exogenous enzyme reaction along with the lapse of time after being separated from the body, so that the observation of protein tissues in a specimen is influenced, and therefore, the simplification and optimization of the preparation steps of the intestinal roll are the key points for improving the quality of the tissue sample of the intestinal roll. The preparation method of the sausage roll is simple and easy to operate, consumes short time, can block endogenous or exogenous enzyme reaction in time, and is favorable for ensuring the stability of protein in small intestine tissues.
3. The slice prepared based on the cylindrical sausage can completely show the full appearance of the intestinal tissue, and is convenient for observation and experiment.
4. The preparation device of the mouse intestine volume label sample can make the preparation process of the intestine volume sample more convenient and faster. On the other hand, the wax strip at the center of the sausage roll is not required to be removed before the step of waxing the sausage roll, and can be perfectly fused with the wax used in the step of waxing the sausage roll into a whole. The clamping seat in the winder is used for fixing the wax strips, and the handle is used for driving the wax strips to rotate. The centering and rotating mechanism can drive the handle to rotate around the axis line of the handle, so that the wax strip can only rotate around the axis line of the handle, and when the wax strip rotates around the axis line of the wax strip, a cylindrical sausage roll with a regular shape can be formed.
The invention is further described below with reference to the figures and examples.
Drawings
FIG. 1 is a schematic structural view of the present invention;
fig. 2 is a sectional view a-a of fig. 1.
Detailed Description
Example 1:
as shown in figures 1-2, the device for preparing the mouse intestine volume label comprises a winder, a wax strip 3 and a centering rotary mechanism.
The winder comprises a handle 1 and a clamping seat 2 fixedly connected to one end of the handle 1, the handle 1 is a regular quadrangular prism-shaped rod, one end of the clamping seat 2 is fixedly welded to one end of the handle 1, and the other end of the clamping seat is provided with a jack. One end of the wax strip 3 is inserted into the jack of the clamping seat 2, the other end of the wax strip extends out of the jack of the clamping seat 2, and the axis of the wax strip 3 is coincided with the axis of the handle 1.
The centering swing mechanism includes a bearing seat 41, a plug bush 42, a bearing 43, and an end cap 44. The bearing seat 41 is internally provided with an inner cavity for mounting a bearing, and an annular step surface is arranged in the inner cavity. The plug bush 42 is cylindrical, a through hole A penetrating through end faces of two sides of the plug bush is formed in the center of the plug bush, and the through hole A is matched with the overall dimension of the handle 1. The bearing 43 is fitted over the sleeve 42 and mounted in the interior of the bearing housing 41 with one end positioned by an annular step surface in the interior of the bearing housing 41 and the other end positioned by an end cap 44. The end cap 44 is disposed at one end of the bearing seat 41, and is fixedly connected to the bearing seat 41 and the insert 42, and a through hole B for the handle 1 to pass through is disposed at the center thereof. The centering and rotating device is inserted into the handle 1 of the winder through a through hole A on the plug bush 42, and the through hole A of the plug bush 42 is in transition fit with the outer wall of the handle 1.
Preferably, the insertion hole of the clamping seat 2 is a regular hexagonal prism-shaped hole, a chamfer is arranged at the orifice of the insertion hole, and correspondingly, the wax strip is in a regular hexagonal prism shape and is adaptive to the size of the insertion hole.
Preferably, the maximum inscribed circle of the cross section of the wax strip 3 has a diameter of 10mm, and the depth of the insertion hole of the clamping seat 2 is 20 mm.
The preparation method of the mouse intestine volume label book is applied to the preparation device of the mouse intestine volume label book, and comprises the following steps:
s01, sampling mouse small intestine:
a. anaesthetizing the mouse, fixing the mouse on an operation table, exposing the chest wall, and performing cardiac puncture bleeding until the mouse loses hemorrhagic shock;
b. after the mouse dies, sterilizing the abdomen, cutting the abdominal wall, exposing the abdominal cavity, dissociating and cutting off the cardia part of the upper section of the small intestine, ligating the opening of the upper section of the small intestine, dissociating and cutting off the ileocaecal part of the lower section of the small intestine;
c. physiological saline is injected into the small intestine through the injector to wash the inner cavity of the small intestine, so that the content in the inner cavity of the small intestine is discharged from the lower section incision of the intestinal tract;
d. the lower section opening of the small intestine is ligated to free the whole small intestine, and 2ml of physiological saline is injected into the free small intestine by using a syringe to avoid the cavity of the small intestine from collapsing and causing the adhesion of the cavity wall.
In this step, the purpose of cardiac puncture exsanguination is to remove erythrocytes in the intestinal wall vessel as much as possible so as to optimize the subsequent staining effect.
In this step, the purpose of the injection of saline is to facilitate the subsequent longitudinal cutting of the small intestine.
In the step, the adopted anesthetic is isoflurane, and the dosage is lethal dose.
In the step, the distance between the ligation point of the upper segment of the small intestine and the incision of the upper segment of the small intestine is 1-2 cm.
In the step, the injection point of the injector is positioned at the rear end of the ligation point of the upper segment of the small intestine by 1-2 cm.
S02, preparing a rectangular sausage skin:
the small intestine injected with the physiological saline is placed in a glass dish, a transverse cut is cut at the position, close to a ligation opening, of the upper section of the small intestine, then the cutting edge of the scissors stretches into the inner cavity of the small intestine from the transverse cut, the small intestine is cut from top to bottom along the longitudinal direction of the small intestine, the small intestine at the upper section of the transverse cut is cut, a ligation line at the lower section of the small intestine is cut, and finally a rectangular intestine skin is formed.
S03, hardening treatment of the rectangular intestine skin:
the method comprises the steps of enabling the fluff surface of a rectangular intestine skin to face upwards, spreading the rectangular intestine skin on an operation table, then scattering Bouins liquid on the rectangular intestine skin (the Bouins can enable proteins in the intestine skin to be solidified), and hardening the rectangular intestine skin for 5-10min to fix the shape of the rectangular intestine skin, so that subsequent intestine roll manufacturing is facilitated.
S04, preparing an intestine roll:
a. taking one end of the rectangular sausage skin by forceps pincers, placing the end on a wax strip, ensuring that the fluff surface of the rectangular sausage skin faces the surface of the wax strip, then rotating a handle, and driving the wax strip to rotate through the handle, so that the rectangular sausage skin is wound around the wax strip one by taking the wax strip as an axis to form a cylindrical sausage roll;
b. after the connection between the wax strips and the clamping seat is broken, the wax strips and the intestine rolls are placed in the Bouins solution to be soaked together (the Bouins can solidify protein in the intestine rolls), and the intestine rolls are hardened to fix the shape of the intestine rolls and prevent the intestine rolls from being scattered or deformed when being subsequently soaked in formalin solution;
c. taking out the wax strips and the intestinal rolls from the Bouins solution, transferring the wax strips and the intestinal rolls into a formalin solution, soaking for 6-10 hours, further fixing tissues of small intestines to stop endogenous or exogenous enzyme reaction, preventing autolysis or autolysis of the tissues, keeping the original shape of the tissues and avoiding antigen inactivation or dispersion;
d. cutting along the radial surface of the intestine roll by a sharp blade, cutting off tissues at two ends of the intestine roll and the wax strip part beyond the two ends of the intestine roll, only keeping the axial width of the intestine roll of 2-3cm, finally obtaining a cylindrical intestine roll according with the size of an embedding box, and placing the cylindrical intestine roll in the embedding box.
S05, dehydrating the intestinal roll:
a. washing the embedding box in flowing water for 10min to fully remove the formalin solution on the cylindrical intestine roll;
b. soaking the embedding box in a dehydration box A filled with 75% alcohol for 80 min;
c. taking out the embedding box from the dehydration box A, and soaking in a dehydration box B filled with 85% alcohol for 80 min;
d. taking out the embedding box from the dehydrating box B, and soaking in the dehydrating box C filled with 95% alcohol for 80 min;
e. taking out the embedding box from the dehydrating box C, and soaking in the dehydrating box D filled with pure alcohol for 80 min;
f. taking out the embedding box from the dewatering box D, and soaking in the dewatering box E filled with pure alcohol for 80 min.
In the step, four dewatering boxes are prepared in advance and named as a dewatering box A, a dewatering box B, a dewatering box C and a dewatering box D respectively, 75% volume fraction alcohol is filled in the dewatering box A, 85% volume fraction alcohol is filled in the dewatering box B, 95% volume fraction alcohol is filled in the dewatering box C, and pure alcohol is filled in the dewatering box D.
S06, waxing the intestinal roll:
a. taking out the embedding box from the dehydrating box E, and soaking in the solution box A filled with the xylene stock solution for 20 min;
b. taking out the embedding box from the wax dipping box A, and soaking in a solution box B filled with a xylene stock solution for 30 min;
c. placing the wax dipping box A filled with the soft wax in an oven for baking to melt the soft wax, taking out the embedding box from the solution box B, and placing the embedding box in the wax dipping box A to soak the liquid soft wax for 60 min;
d. placing the wax dipping box B filled with the soft wax in an oven to be baked to be softened into a liquid state, taking the embedding box out of the wax dipping box A, and placing the embedding box in the wax dipping box B to be dipped in the liquid soft wax for 60 min;
e. and (3) placing the wax dipping box C filled with the hard wax in an oven to bake so as to soften the wax into liquid, taking the embedding box out of the wax dipping box B, and placing the embedding box in the wax dipping box C to soak the liquid hard wax for 120 min.
In the step, two solution boxes are prepared in advance and named as a solution box A and a solution box B respectively, xylene stock solution is filled in the solution box A and the solution box B, three wax dipping boxes are prepared in advance and named as a wax dipping box A, a wax dipping box B and a wax dipping box C respectively, soft wax is filled in the wax dipping box A and the wax dipping box B, and hard wax is filled in the wax dipping box C.
In the step, in the process of soaking the soft wax, the temperature of the oven is set to be 58-66 ℃, so that the soft wax is maintained in a melting state.
In the step, in the process of soaking the hard wax, the temperature of the oven is set to be 58-66 ℃, so that the hard wax is maintained in a melting state.
S07, embedding the steamed roll:
a. rolling the dehydrated and waxed cylindrical sausage out, and placing the rolled cylindrical sausage in the center of the bottom of the groove of the embedding mold;
b. filling the whole embedding mold groove with liquid hard wax, and covering the cover of the embedding mold;
c. placing the whole embedding mould in a refrigerated cabinet with the temperature of-18 to-22 ℃ for standing for 20 minutes;
d. and taking down the cover of the embedding mold, inverting the embedding mold, and tapping the embedding mold to completely remove the wax block containing the intestinal roll from the embedding mold.
Claims (8)
1. This preparation facilities of mouse intestines volume label, characterized by: comprises a winder and a wax strip; the winder comprises a handle and a clamping seat fixedly connected to one end of the handle, and the end part of the clamping seat far away from the handle is provided with a jack; one end of the wax strip is inserted in the insertion hole of the clamping seat, and the other end of the wax strip extends out of the insertion hole of the clamping seat.
2. The apparatus for preparing a mouse intestine volume label according to claim 1, wherein: the insertion hole of the clamping seat is a regular hexagonal prism-shaped hole, a chamfer is arranged at the hole opening of the insertion hole, and correspondingly, the wax strip is in a regular hexagonal prism shape and is adaptive to the size of the insertion hole.
3. The apparatus for preparing a mouse intestine volume label according to claim 2, wherein: the diameter of the largest inscribed circle of the cross section of the wax strip is 10mm, and the depth of the jack of the clamping seat is 20 mm.
4. The apparatus for preparing a mouse intestinal volume label according to any one of claims 1 to 3, wherein: the handle is a regular quadrangular prism-shaped rod, and the axis of the wax strip is coincided with the axis of the handle.
5. The apparatus for preparing a mouse intestine volume label according to claim 4, wherein: the centering swing mechanism comprises an insert sleeve, a bearing seat and an end cover; an inner cavity for mounting a bearing is arranged in the bearing seat, and an annular step surface is arranged in the inner cavity; the plug bush is cylindrical, the center of the plug bush is provided with a through hole A which runs through the end faces of the two sides of the plug bush, and the through hole A is matched with the appearance size of the handle; the bearing is sleeved on the insertion sleeve and is arranged in the inner cavity of the bearing seat, one end of the bearing is positioned through an annular step surface in the inner cavity of the bearing seat, and the other end of the bearing is positioned through the end cover; the end cover is arranged at one end of the bearing seat and is fixedly connected with the bearing seat and the plug bush, and the center of the end cover is provided with a through hole B for the handle to pass through; the centering and rotating device is inserted into the handle of the winder through a through hole A on the plug bush.
6. A method for preparing a mouse intestine volume label book, which is applied to the mouse intestine volume label book preparation device according to any one of claims 1 to 5, and which is characterized by comprising the following steps:
s01, sampling mouse small intestine:
a. anaesthetizing the mouse, fixing the mouse on an operation table, exposing the chest wall, and performing cardiac puncture bleeding until the mouse loses hemorrhagic shock;
b. after the mouse dies, sterilizing the abdomen, cutting the abdominal wall, exposing the abdominal cavity, dissociating and cutting off the cardia part of the upper section of the small intestine, ligating the opening of the upper section of the small intestine, dissociating and cutting off the ileocaecal part of the lower section of the small intestine;
c. physiological saline is injected into the small intestine through the injector to wash the inner cavity of the small intestine, so that the content in the inner cavity of the small intestine is discharged from the lower section incision of the intestinal tract;
d. ligating an opening at the lower section of the small intestine, dissociating the whole small intestine, and injecting 2ml of physiological saline into the dissociated small intestine by using a syringe so as to avoid cavity wall adhesion caused by the cavity collapse in the small intestine;
s02, preparing a rectangular sausage skin:
placing the small intestine injected with the physiological saline into a glass dish, cutting a transverse cut at a position close to a ligation opening at the upper section of the small intestine, extending a cutting edge of a scissors into the inner cavity of the small intestine from the transverse cut, cutting the small intestine from top to bottom along the longitudinal direction of the small intestine, cutting the small intestine at the upper section of the transverse cut, and cutting a ligation line at the lower section of the small intestine to finally form a rectangular intestine skin;
s03, hardening treatment of the rectangular intestine skin:
spreading the rectangular sausage skin with the fluff surface facing upwards on an operation table, spreading the Bouins liquid on the rectangular sausage skin, and hardening the rectangular sausage skin for 5-10min to fix the shape of the rectangular sausage skin, so as to facilitate the subsequent production of sausage rolls;
s04, preparing an intestine roll:
a. taking one end of the rectangular sausage skin by forceps pincers, placing the end on a wax strip, ensuring that the fluff surface of the rectangular sausage skin faces the surface of the wax strip, then rotating a handle, and driving the wax strip to rotate through the handle, so that the rectangular sausage skin is wound around the wax strip one by taking the wax strip as an axis to form a cylindrical sausage roll;
b. after the connection between the wax strips and the clamping seat is disconnected, the wax strips and the intestine rolls are placed in the Bouins solution for soaking, and the intestine rolls are hardened to fix the shape of the intestine rolls and prevent the intestine rolls from being scattered or deformed when the intestine rolls are subsequently soaked in the formalin solution;
c. then taking out the wax strips and the intestinal rolls from the Bouins solution, transferring the wax strips and the intestinal rolls into a formalin solution to be soaked for 6-10h, and further fixing tissues of small intestines to stop endogenous or exogenous enzyme reaction, prevent autolysis or exsolution of the tissues, keep the original shapes of the tissues and avoid antigen inactivation or dispersion;
d. cutting along the radial surface of the intestine roll by a sharp cutting edge, cutting tissues at two ends of the intestine roll and wax strip parts beyond the two ends of the intestine roll, only keeping the axial width of the intestine roll of 2-3cm, finally obtaining a cylindrical intestine roll according with the size of an embedding box, and placing the cylindrical intestine roll in the embedding box;
s05, dehydrating the intestinal roll:
a. washing the embedding box in flowing water for 10min to fully remove the formalin solution on the cylindrical intestine roll;
b. soaking the embedding box in a dehydration box A filled with 75% alcohol for 80 min;
c. taking out the embedding box from the dehydration box A, and soaking in a dehydration box B filled with 85% alcohol for 80 min;
d. taking out the embedding box from the dehydrating box B, and soaking in the dehydrating box C filled with 95% alcohol for 80 min;
e. taking out the embedding box from the dehydrating box C, and soaking in the dehydrating box D filled with pure alcohol for 80 min;
f. taking out the embedding box from the dewatering box D, and soaking in the dewatering box E filled with pure alcohol for 80 min;
s06, waxing the intestinal roll:
a. taking out the embedding box from the dehydrating box E, and soaking in the solution box A filled with the xylene stock solution for 20 min;
b. taking out the embedding box from the wax dipping box A, and soaking in a solution box B filled with a xylene stock solution for 30 min;
c. placing the wax dipping box A filled with the soft wax in an oven for baking to melt the soft wax, taking out the embedding box from the solution box B, and placing the embedding box in the wax dipping box A to soak the liquid soft wax for 60 min;
d. placing the wax dipping box B filled with the soft wax in an oven to be baked to be softened into a liquid state, taking the embedding box out of the wax dipping box A, and placing the embedding box in the wax dipping box B to be dipped in the liquid soft wax for 60 min;
e. placing the wax dipping box C filled with the hard wax in an oven to be baked to be softened into a liquid state, taking the embedding box out of the wax dipping box B, and placing the embedding box in the wax dipping box C to be dipped in the liquid hard wax for 120 min;
s07, embedding the steamed roll:
a. rolling the dehydrated and waxed cylindrical sausage out, and placing the rolled cylindrical sausage in the center of the bottom of the groove of the embedding mold;
b. filling the whole embedding mold groove with liquid hard wax, and covering the cover of the embedding mold;
c. placing the whole embedding mould in a refrigerated cabinet with the temperature of-18 to-22 ℃ for standing for 20 minutes;
d. and taking down the cover of the embedding mold, inverting the embedding mold, and tapping the embedding mold to completely remove the wax block containing the intestinal roll from the embedding mold.
7. The method for preparing a mouse intestine volume label according to claim 6, wherein: in the step S01, isoflurane is used as anesthetic, and the dosage is lethal dose; in the step S01, the distance between the ligation point of the upper small intestine and the incision of the upper small intestine is 1-2 cm; in the step S01, the injection point of the injector is located 1-2 cm behind the ligation point of the upper segment of the small intestine.
8. The method for preparing a mouse intestine volume label according to claim 7, wherein: s06, setting the temperature of an oven to be 58-66 ℃ in the process of soaking the soft wax to maintain the melting state of the soft wax; and S06, setting the temperature of the oven to be 58-66 ℃ in the process of soaking the hard wax, and keeping the hard wax in a melting state.
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| CN115356182A (en) * | 2022-08-11 | 2022-11-18 | 蚌埠医学院第一附属医院（蚌埠医学院附属肿瘤医院） | Preparation method of mouse intestinal tissue wax block for slicing |
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