Pretreatment device and method for extracting nucleic acid from OCT (optical coherence tomography) embedded kidney biopsy tissue

Abstract

The invention provides a pretreatment method for extracting nucleic acid from OCT (Optical Coherence Tomography) embedded kidney biopsy tissue, which comprises the following steps: firstly, carrying out OCT embedding on a kidney tissue sample and then immediately carrying out liquid nitrogen quick-freezing by adopting a device for the pretreatment for extracting the nucleic acid from the OCT embedded kidney biopsy tissue; s03, soaking the OCT-embedded human kidney tissue stored at the temperature of 1-80 DEG C and tinfoil in liquid nitrogen in a liquid nitrogen tank for 10-20 minutes, wrapping the OCT-embedded human kidney tissue with the tinfoil, putting the human kidney tissue into a knocking tank with a low-temperature base, knocking the human kidney tissue by using a knocking hammer until the human kidney tissue is broken, clamping all tissue fragments by using tweezers, putting the tissue fragments into a test tube containing a lysis solution, and carrying out ultrasonic treatment on the human kidney tissue at the temperature of 1-80 DEG C for 10-20 minutes to obtain the OCT-embedded human kidney tissue. Immediately carrying out homogenization treatment; according to the method, the OCT embedding kidney biopsy tissue is quickly broken in the ultralow-temperature environment, the OCT embedding medium is separated out, and degradation of extracted nucleic acid caused by tissue melting is avoided. And tissue blocks are separated for subsequent nucleic acid extraction through rapid breaking in an ultralow-temperature environment, so that the quality of a nucleic acid sample is guaranteed.

Description

Technical Field

[0001] The invention belongs to the field of medical research technology, and in particular relates to a device and method for pre-processing of OCT-embedded renal biopsy tissue for nucleic acid extraction. Background Art

[0002] Percutaneous renal biopsy (RB) is an important technique for the diagnosis, treatment and research of kidney diseases. The renal tissue obtained by puncture of renal biopsy is further determined by light microscopy, electron microscopy and immunopathology. The renal biopsy tissue is embedded in paraffin and sliced ​​for light microscopy; fixed in glutaraldehyde fixative for electron microscopy; and embedded in OCT to make frozen sections for immunopathology. Usually, frozen sections of OCT-embedded renal biopsy tissue are used for clinical diagnosis, histopathology and molecular biology research. The remaining OCT-embedded renal biopsy tissue after frozen sectioning can be used for sample storage, providing sample resources for downstream scientific research, especially for genome sequencing analysis. OCT compound is an embedding medium widely used for long-term cryopreservation of different tissues. It is a viscous aqueous solution at room temperature and is mainly composed of polyethylene glycol, benzalkonium chloride, antifungal agent and resin polyvinyl alcohol. In order to make full use of the value of such stored sample resources and extract nucleic acids of good quality, appropriate pretreatment operations should be taken to remove the OCT embedding agent outside the tissue.

[0003] A method for re-making paraffin sections from the remaining frozen tissue of a renal biopsy by immunofluorescence, publication number CN 111879590A, provides a method for re-making paraffin sections from the remaining frozen tissue of a renal biopsy by immunofluorescence, which uses a PBS buffer solution to quickly thaw the remaining renal tissue after frozen sections and dissolve the OCT glue, and then re-makes paraffin sections, and uses the paraffin sections for special staining and light microscopy to make up for the situation where there are no glomeruli or too few glomeruli in the light microscopy. In this method, the frozen renal tissue stored in a -80°C ultra-low temperature refrigerator is taken out and placed in a PBS buffer solution at room temperature to thaw the frozen renal tissue and fully dissolve the OCT glue. This process is prone to degradation of nucleic acid extracted from tissues, which is not conducive to extracting RNA from such samples for gene sequencing analysis. To this end, we explored a more suitable method for pretreatment of nucleic acid extraction from OCT-embedded renal biopsy tissue through comparative experiments, and designed a device for pretreatment of nucleic acid extraction from OCT-embedded renal biopsy tissue based on this method, which avoids the melting of tissue samples during pretreatment of nucleic acid extraction from such samples and causes nucleic acid degradation, thereby facilitating the extraction of good-quality nucleic acids for downstream research. Summary of the invention

[0004] In order to solve the above problems, the present invention discloses a device and method for pre-processing of OCT-embedded renal biopsy tissue for nucleic acid extraction.

[0005] To achieve the above object, the technical solution of the present invention is as follows:

[0006] The present invention provides a method for pre-treatment of OCT-embedded renal biopsy tissue for nucleic acid extraction, comprising the following steps:

[0007] S01. Immediately freeze the renal tissue sample in liquid nitrogen after embedding in OCT, freeze-section at -20°C, cover the remaining surface of the OCT-embedded renal tissue sample with OCT droplets, and store at -80°C.

[0008] S02. Soak the OCT-embedded human kidney tissue and tin foil stored at -80°C in step S01 in liquid nitrogen in a liquid nitrogen tank for 10-20 minutes, then wrap the OCT-embedded human kidney tissue with tin foil, place it in a knocking tank, and knock it with a knocker until it breaks into pieces. Use tweezers to pick up all tissue fragments, put them into a test tube containing lysis solution, and immediately perform homogenization.

[0009] Furthermore, the number of knocking is 3-10 times, preferably, the number of knocking is 5-8 times, and the size of the tissue fragments is 0.5-1.5 mm.

[0010] The present invention also provides a device for pre-processing nucleic acid extraction from OCT-embedded renal biopsy tissue, comprising an ultra-low temperature base, on which a liquid nitrogen area and a pre-processing area are arranged; the liquid nitrogen area comprises a liquid nitrogen tank, a tweezers rack, and tweezers; the pre-processing area comprises a splash-proof enclosure, a light strip, a temperature display, and a knocker, and the bottom of the splash-proof enclosure is threadedly connected to the ultra-low temperature base; the knocker comprises a C-shaped support base, a vertical support rod, and a support cross arm, the vertical support rod is placed on the C-shaped support base, a support cross arm is arranged on the top of the vertical support rod, a cylindrical carrier shell is arranged at the suspended end of the support cross arm, a first guide hole is arranged at the upper end of the carrier shell, a second guide hole is arranged at the lower end surface of the carrier shell, a knocking rod is arranged in the first guide hole and the second guide hole, a knocking hammer is arranged at the bottom end of the knocking rod, a first limit plate is arranged at the top of the knocking rod, and a splash-proof cover is also arranged on the knocking rod, and the splash-proof cover is arranged above the knocking hammer and below the carrier shell.

[0011] Furthermore, the ultra-low temperature base includes a first ultra-low temperature base and a second ultra-low temperature base, the first ultra-low temperature base and the second ultra-low temperature base are connected through a connecting channel, and an insulation layer is arranged outside the first ultra-low temperature base, the second ultra-low temperature base and the connecting channel.

[0012] Furthermore, the first ultra-low temperature base is provided with a liquid nitrogen tank, a tweezers rack, and tweezers, the liquid nitrogen tank is matched with a liquid nitrogen tank cover, a hollow partition is provided at the bottom of the liquid nitrogen tank, a baffle is provided at the edge of the hollow partition, the liquid nitrogen tank cover is connected to the upper edge of the baffle through a connecting rod, a liquid nitrogen inlet is provided on the side of the liquid nitrogen tank, liquid nitrogen is injected into the liquid nitrogen tank through the liquid nitrogen inlet, a sample is placed on the hollow partition, an insulation layer is provided on the outer side of the bottom of the liquid nitrogen tank, the hollow partition is connected to the first liquid nitrogen channel in the first ultra-low temperature base, and the first liquid nitrogen channel is connected to the connecting channel. Liquid nitrogen overflows the hollow partition and enters the first liquid nitrogen channel. The connection arrangement of the liquid nitrogen tank cover and the hollow partition can be lifted out of the liquid nitrogen together with the partition as a whole, which is convenient for sample removal and addition.

[0013] Furthermore, the second ultra-low temperature base is provided with an upwardly protruding connecting flange, the outside of the connecting flange is provided with an external thread, the splash-proof enclosure is threadedly connected to the outside of the connecting flange, an insulation layer is provided on the outside of the bottom of the splash-proof enclosure, a light strip is provided above the insulation layer, a bottom plate is provided in the connecting flange, a plurality of supporting columns are provided at the bottom of the bottom plate, the bottom of the supporting columns penetrates the insulation layer and is connected to the steel plate, a liquid nitrogen distribution plate is designed in the second ultra-low temperature insulation base, one end of the liquid nitrogen distribution plate is connected to the connecting channel, a plurality of liquid nitrogen outlets are provided on the liquid nitrogen distribution plate, and the second ultra-low temperature insulation base is also provided with a liquid nitrogen discharge outlet, which is convenient for the discharge and recycling of the liquid nitrogen after use.

[0014] Furthermore, a limiting cylinder is arranged outside the knocking rod in the carrier shell, the limiting cylinder opens downward, the knocking rod passes through the top of the limiting cylinder, a first limiting ring is arranged outwardly on the bottom edge of the limiting cylinder, a first limiting block is arranged in the middle part of the inner wall of the carrier shell, the first limiting block is placed above the first limiting ring, a second limiting block is also arranged above the splashproof cover of the knocking rod, the second limiting block is quickly placed in the carrier shell, a spring is arranged between the limiting cylinder, the knocking rod and the second limiting block ring, and the spring is sleeved on the knocking rod; a first adjusting hole is axially arranged on the side surface of the carrier shell, a first push rod is arranged on the first limiting ring, and the first push rod is placed in the first adjusting hole.

[0015] Furthermore, the first adjustment hole comprises a vertical strip hole, and a plurality of strip branch holes are arranged at intervals on both sides of the strip hole.

[0016] Furthermore, a second annular limit plate is provided at the inner bottom of the carrier shell, a ball is provided between the second limit plate and the inner bottom surface of the carrier shell, and an annular groove matching the sliding of the ball is provided on the lower end surface of the second limit plate and the inner bottom surface of the carrier shell; a plurality of radially outward first limit recesses and a plurality of radially inward limit protrusions are provided at the inner edge of the second limit plate, the limit protrusions are symmetrically arranged, and the first limit recesses are symmetrically arranged, a second adjustment hole is provided on the carrier shell contacted by the outer edge of the second limit plate, a second push rod is provided at the outer edge of the second limit plate, and the second push rod is placed in the second adjustment hole at the bottom of the carrier shell.

[0017] Furthermore, a second limiting recess corresponding to the first limiting recess is arranged at the bottom of the carrier shell, and the second limiting recess is larger than the first limiting recess.

[0018] The beneficial effects of the present invention are:

[0019] The present invention utilizes the rapid crushing of OCT-embedded renal biopsy tissue in an ultra-low temperature environment to separate the OCT embedding agent, thereby avoiding the degradation of extracted nucleic acids caused by tissue melting. By rapidly crushing in an ultra-low temperature environment, the tissue block is separated for subsequent nucleic acid extraction, thereby ensuring the quality of the nucleic acid sample. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 It is a three-dimensional schematic diagram of the device for pre-treatment of OCT-embedded renal biopsy tissue for nucleic acid extraction according to the present invention;

[0021] Figure 2 Extract RNA integrity RIN values ​​for Examples and Comparative Examples;

[0022] Figure 3 This is a schematic diagram of the internal structure of the device used for pre-treatment of OCT-embedded renal biopsy tissue for nucleic acid extraction according to the present invention;

[0023] Figure 4 is a top view of the second limiting plate of the present invention;

[0024] Figure 5 for Figure 3 Enlarged view of part A in the figure.

[0025] List of Figure Symbols:

[0026] 101 liquid nitrogen tank; 102 tweezers rack; 103 tweezers; 104 first ultra-low temperature base; 105, liquid nitrogen tank cover; 106, hollow partition; 107, baffle plate; 201 second ultra-low temperature base; 202 splash-proof baffle; 203 light strip; 204 temperature display; 3 smasher; 301 C-type support base; 302 vertical support rod; 303 support cross arm; 304 knocking rod; 305 carrier shell; 306 splash-proof cover; 307 first adjustment hole; 308 second limit plate; 309 second push rod; 310 second limit block; 311 spring; 312 first limit recess; 313 limit protrusion; 314 second adjustment hole. DETAILED DESCRIPTION

[0027] The present invention will be further explained below in conjunction with the accompanying drawings and specific embodiments. It should be understood that the following specific embodiments are only used to illustrate the present invention and are not used to limit the scope of the present invention.

[0028] Example 1-Example 5

[0029] Taking 5 cases of OCT-embedded human renal biopsy tissues stored at -80℃ for 5 years as an example, the specific operation steps are as follows:

[0030] Tissue sample storage: Human kidney tissue samples were obtained by puncture during renal biopsy. After sampling, they were embedded in OCT, immediately frozen in liquid nitrogen, and frozen and sectioned at -20℃. The remaining OCT-embedded kidney tissue sample cut surface was covered with OCT droplets and stored at -80℃.

[0031] OCT crushing treatment: Soak the OCT-embedded human kidney tissue and tin foil in liquid nitrogen for about 20 minutes, then wrap the tissue with tin foil and quickly knock 5-8 times on the pre-treatment area with a hammer until the tissue fragments are broken into pieces with a size of 0.5-1.5 mm. Use tweezers to grab all tissue fragments, put them into a test tube containing lysis solution, and immediately perform homogenization treatment. Labeled as Example 1, Example 2, Example 3, Example 4, Example 5.

[0032] The knocking is performed using the device shown in Example 6.

[0033] Comparative Example 1-Comparative Example 5

[0034] Taking 5 cases of OCT-embedded human renal biopsy tissues stored at -80°C for 5 years in Example 1-5 as an example, the specific operation steps are as follows:

[0035] Tissue sample storage: Human kidney tissue samples were obtained by puncture during renal biopsy. After sampling, they were embedded in OCT, immediately frozen in liquid nitrogen, and frozen and sectioned at -20℃. The remaining OCT-embedded kidney tissue sample cut surface was covered with OCT droplets and stored at -80℃.

[0036] PBS washing treatment: Place the OCT-embedded human kidney tissue in a 4°C PBS solution, wash each sample 3 times, replace the 4°C PBS solution each time, and each washing time is about 15 seconds. After washing, the OCT outside the tissue is completely dissolved, and then the tissue is taken out of the PBS solution, placed on a dust-free paper to gently wipe off the surface liquid, placed in a test tube containing a lysis solution, and immediately homogenized. Marked as Comparative Example 1, Comparative Example 2, Comparative Example 3, Comparative Example 4, Comparative Example 5.

[0037] RNA extraction and quality analysis

[0038] use Kit (Omega Bio-tek, EZNATotal RNAkit I, Cat No: R6834-01) was used to extract RNA from tissue samples of the implementation group and the control group. The quality of RNA was evaluated by yield, purity, and integrity. The RNA concentration and purity were analyzed using a Nanodrop2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA). The integrity was evaluated using an Agilent 2100 bioanalyzer (Agilent Technologies, Palo Alto, California). The standard for good integrity was RIN ≥ 6.0; the standard for good yield was yield ≥ 0.5 μg; the purity A 260 / 280 Good in the range of 1.9 to 2.1; 260 / 230 Increase, indicating that the amount of pollutants such as salt ions decreased.

[0039] The quality of RNA extracted from tissues of the embodiment and comparative example after different pre-treatments is shown in Table 1, Table 2, Figure 2 :

[0040] Table 1 RNA quality of tissues extracted from examples and comparative examples after different pre-treatments

[0041]

[0042]

[0043] Table 2 Comparison of RNA extraction quality between the embodiment and the comparative example

[0044]

[0045] According to statistical analysis, the OCT crushing treatment in the embodiment is not significantly different from the PBS washing treatment in the control example, and the RNA yield and purity (A260 / 280, A260 / 230) extracted from the tissue in the embodiment are not significantly different from those in the control example. However, the RNA integrity RIN value of the embodiment is significantly higher than that of the control example, that is, the RNA extracted in the embodiment is of better quality and more conducive to the use of downstream research. Therefore, a device and method for pre-treatment of nucleic acid extraction from OCT-embedded renal biopsy tissue has substantial significance for the pre-treatment of nucleic acid extraction from such sample resources.

[0046] The present invention also provides a device for the pretreatment of nucleic acid extraction from OCT-embedded biopsy tissues, which includes a cryogenic base 201. A liquid nitrogen area 1 and a pretreatment area 2 are arranged on the cryogenic base 201; the liquid nitrogen area 1 includes a liquid nitrogen tank 101, a forceps rack 102, and forceps 103; the pretreatment area 2 includes a splash-proof enclosure 202, a light strip 203, a temperature display 204, and a crusher 3. The bottom of the splash-proof enclosure 202 is threadedly connected to the cryogenic base; the crusher includes a C-shaped support base 301, a vertical support rod 302, and a support cross-arm 303. The vertical support rod 302 is placed on the C-shaped support base 301. A support cross-arm 303 is arranged at the top of the vertical support rod 302. A cylindrical carrier shell 305 is arranged at the suspended end of the support cross-arm 303. A first guiding hole is arranged at the upper end of the carrier shell 305, and a second guiding hole is arranged at the lower end face of the carrier shell 305. A striking rod 304 is arranged in the first guiding hole and the second guiding hole. A striking hammer is arranged at the bottom end of the striking rod 304, and a first limiting plate is arranged at the top of the striking rod 304. A splash-proof cover 306 is also arranged on the striking rod 304, and the splash-proof cover 306 is arranged above the striking hammer and below the carrier shell 305.

[0047] The cryogenic base includes a first cryogenic base 104 and a second cryogenic base 201. The first cryogenic base 104 and the second cryogenic base 201 are connected through a communication channel. Heat insulation layers are arranged outside the first cryogenic base 104, the second cryogenic base 201, and the communication channel.

[0048] A liquid nitrogen tank 101, a forceps rack 102, and forceps 103 are arranged on the first cryogenic base 104. A liquid nitrogen tank cover 105 is arranged in a matching manner on the liquid nitrogen tank 101. A hollow partition plate 106 is arranged at the inner bottom of the liquid nitrogen tank 101. A baffle 107 is arranged at the edge of the hollow partition plate 106. The liquid nitrogen tank cover 105 and the upper edge of the baffle 107 are connected through a connecting rod. A liquid nitrogen inlet is arranged on the side of the liquid nitrogen tank 101. Liquid nitrogen is injected into the liquid nitrogen tank 101 through the liquid nitrogen inlet. Samples are placed on the hollow partition plate 106. A heat insulation layer is arranged outside the bottom of the liquid nitrogen tank 101. The hollow partition plate 106 is connected to a first liquid nitrogen channel inside the first cryogenic base 104, and the first liquid nitrogen channel is connected to the communication channel. Liquid nitrogen overflows the hollow partition plate 106 and enters the first liquid nitrogen channel. The connection between the liquid nitrogen tank cover 105 and the hollow partition plate 106 enables the whole partition plate to be lifted out of the liquid nitrogen, facilitating the removal and addition of samples.

[0049] The second ultra-low temperature base 201 is provided with an upwardly protruding connecting flange, the outside of the connecting flange is provided with an external thread, the splash-proof enclosure 202 is threadedly connected to the outside of the connecting flange, an insulation layer is provided on the outside of the bottom of the splash-proof enclosure 202, a light strip 203 is provided above the insulation layer, a bottom plate is provided in the connecting flange, a plurality of supporting columns are provided at the bottom of the bottom plate, the bottom of the supporting columns penetrates the insulation layer and is connected to the steel plate, a liquid nitrogen distribution plate is designed in the second ultra-low temperature insulation base, one end of the liquid nitrogen distribution plate is connected to the connecting channel, a plurality of liquid nitrogen outlets are provided on the liquid nitrogen distribution plate, and the second ultra-low temperature insulation base 201 is also provided with a liquid nitrogen discharge outlet, which is convenient for the discharge and recycling of liquid nitrogen after use.

[0050] A limiting cylinder is arranged outside the knocking rod 304 in the carrier shell 305, and the limiting cylinder opens downward, and the knocking rod 304 passes through the top of the limiting cylinder, and a first limiting ring is arranged outward from the bottom edge of the limiting cylinder, and a first limiting block is arranged in the middle part of the inner wall of the carrier shell 305, and the first limiting block is placed above the first limiting ring, and a second limiting block 310 is also arranged above the splash-proof cover 306 of the knocking rod 304, and the second limiting block is quickly placed in the carrier shell 305, and a spring 311 is arranged between the limiting cylinder, the knocking rod 304 and the second limiting block ring, and the spring 311 is sleeved on the knocking rod 304; a first adjusting hole 307 is axially arranged on the side of the carrier shell 305, and a first push rod is arranged on the first limiting ring, and the first push rod is placed in the first adjusting hole 307.

[0051] The first adjustment hole 307 includes a vertical strip hole, and a plurality of strip branch holes are arranged at intervals on both sides of the strip hole.

[0052] A second annular limit plate 308 is provided at the inner bottom of the carrier shell 305, and a ball is provided between the second limit plate 308 and the inner bottom surface of the carrier shell 305. The lower end surface of the second limit plate 308 and the inner bottom surface of the carrier shell 305 are both provided with annular grooves matching the sliding of the ball; the inner edge of the second limit plate 308 is provided with a plurality of radially outward first limit recesses 312 and a plurality of radially inward limit protrusions 313, the limit protrusions 313 are symmetrically arranged, and the first limit recesses 312 are symmetrically arranged. A second adjustment hole 314 is provided on the carrier shell 305 contacted by the outer edge of the second limit plate 308, and a second push rod 309 is provided at the outer edge of the second limit plate 308, and the second push rod 309 is placed in the second adjustment hole 314 at the bottom of the carrier shell 305.

[0053] A second limiting recess corresponding to the first limiting recess 312 is disposed at the bottom of the carrier shell 305 , and the second limiting recess is larger than the first limiting recess 312 .

[0054] When in use, liquid nitrogen is injected into the liquid nitrogen tank 101. After the temperature display 204 on the ultra-low temperature base shows that the temperature is stable, the forceps 103 on the forceps rack 102 are removed, and the OCT-embedded renal biopsy tissue is clamped and placed on the hollow partition 106 of the liquid nitrogen tank 101, and then placed in the liquid nitrogen tank 101 for 20 minutes. Turn on the light strip 203 for illumination, use the forceps 103 to clamp the OCT-embedded renal biopsy tissue sample on the hollow partition 106 of the liquid nitrogen tank 101, place it in the middle area of ​​the splash-proof enclosure 202, start the smasher 3 to uniformly smash the sample and the embedded OCT compound, and use the forceps 103 to quickly clamp the tissue fragments and place them in the lysate for the next step of nucleic acid extraction.

[0055] As a specific implementation of the present invention, it is possible to adjust which strip-shaped branch hole in the first adjustment hole 307 the first push rod is placed in according to the requirements.

[0056] It should be noted that the above content only illustrates the technical idea of ​​the present invention and cannot be used to limit the protection scope of the present invention. For ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications all fall within the protection scope of the claims of the present invention.

Claims

1. A method for pre-treatment of OCT-embedded renal biopsy tissue for nucleic acid extraction, characterized in that: The following steps are involved: S01. Immediately freeze the renal tissue sample in liquid nitrogen after embedding in OCT, freeze-section at -20°C, cover the remaining surface of the OCT-embedded renal tissue sample with OCT droplets, and store at -80°C. S02. Soak the OCT-embedded human kidney tissue and tin foil stored at -80°C in step S01 in liquid nitrogen in a liquid nitrogen tank for 10-20 minutes, then wrap the OCT-embedded human kidney tissue with tin foil, place it in a percussion tank, and use a percussion hammer to knock it into pieces. Use tweezers to pick up all tissue fragments, put them into a test tube containing lysis solution, and immediately perform homogenization.

2. A method for pre-treatment of nucleic acid extraction from OCT-embedded renal biopsy tissue according to claim 1, characterized in that: The number of knocking is 3-10 times, and the size of the tissue fragments is 0.5-1.5 mm.

3. A device for pre-treatment of OCT-embedded renal biopsy tissue for nucleic acid extraction, characterized in that: The invention comprises an ultra-low temperature base (201), on which a liquid nitrogen area (1) and a pre-treatment area (2) are arranged; the liquid nitrogen area (1) comprises a liquid nitrogen tank (101), a tweezers rack (102), and tweezers (103); the pre-treatment area (2) comprises an anti-splash enclosure (202), a light strip (203), a temperature display (204), and a knocker (3); the bottom of the anti-splash enclosure (202) is threadedly connected to the ultra-low temperature base; the knocker comprises a C-shaped support base (301), a vertical support rod (302), and a support cross arm (303); the vertical support rod (302) is arranged on the C-shaped On the support base (301), a support cross arm (303) is arranged at the top of the vertical support rod (302), a cylindrical carrier shell (305) is arranged at the suspended end of the support cross arm (303), a first guide hole is arranged at the upper end of the carrier shell (305), a second guide hole is arranged at the lower end surface of the carrier shell (305), a knocking rod (304) is arranged in the first guide hole and the second guide hole, a knocking hammer is arranged at the bottom end of the knocking rod (304), a first limit plate is arranged at the top of the knocking rod (304), and a splash-proof cover (306) is also arranged on the knocking rod (304), and the splash-proof cover (306) is arranged above the knocking hammer and below the carrier shell (305).

4. The device for pre-treatment of nucleic acid extraction from OCT-embedded renal biopsy tissue according to claim 3, characterized in that: The ultra-low temperature base comprises a first ultra-low temperature base (104) and a second ultra-low temperature base (201); the first ultra-low temperature base (104) and the second ultra-low temperature base (201) are connected via a connecting channel; and a thermal insulation layer is arranged outside the first ultra-low temperature base (104), the second ultra-low temperature base (201) and the connecting channel.

5. The device for pre-treatment of OCT-embedded renal biopsy tissue for nucleic acid extraction according to claim 4, characterized in that: The first ultra-low temperature base (104) is provided with a liquid nitrogen tank (101), a tweezers rack (102), and tweezers (103); the liquid nitrogen tank (101) is matched with a liquid nitrogen tank cover (105); a hollow partition (106) is provided at the bottom of the liquid nitrogen tank (101); a baffle plate (107) is provided at the edge of the hollow partition plate (106); the liquid nitrogen tank cover (105) is connected to the upper edge of the baffle plate (107) through a connecting rod; a liquid nitrogen inlet is provided on the side of the liquid nitrogen tank (101); an insulation layer is provided on the outer side of the bottom of the liquid nitrogen tank (101); the hollow partition plate (106) is connected to a first liquid nitrogen channel in the first ultra-low temperature base (104); and the first liquid nitrogen channel is connected to the connecting channel.

6. The device for pre-treatment of nucleic acid extraction from OCT-embedded renal biopsy tissue according to claim 4, characterized in that: The second ultra-low temperature base (201) is provided with an upwardly protruding connecting flange, the connecting flange is provided with an external thread, the splash-proof enclosure (202) is threadedly connected to the outside of the connecting flange, an insulation layer is provided on the outside of the bottom of the splash-proof enclosure (202), a light strip (203) is provided above the insulation layer, a bottom plate is provided inside the connecting flange, a plurality of supporting columns are provided at the bottom of the bottom plate, the bottom of the supporting columns penetrates the insulation layer and is connected to the steel plate, a liquid nitrogen distribution plate is designed inside the second ultra-low temperature insulation base, one end of the liquid nitrogen distribution plate is connected to the connecting channel, and a plurality of liquid nitrogen outlets are provided on the liquid nitrogen distribution plate.

7. The device for pre-treatment of nucleic acid extraction from OCT-embedded renal biopsy tissue according to claim 3, characterized in that: A limiting cylinder is arranged outside the knocking rod (304) in the carrier shell (305), the limiting cylinder opening is downward, the knocking rod (304) passes through the top of the limiting cylinder, a first limiting ring is arranged outwardly at the bottom edge of the limiting cylinder, a first limiting block is arranged in the middle part of the inner wall of the carrier shell (305), the first limiting block is arranged above the first limiting ring, a second limiting block (310) is arranged above the splash-proof cover (306) of the knocking rod (304), the second limiting block is arranged in the carrier shell (305), a spring (311) is arranged between the limiting cylinder, the knocking rod (304) and the second limiting block ring, and the spring (311) is sleeved on the knocking rod (304); a first adjusting hole (307) is axially arranged on the side surface of the carrier shell (305), a first push rod is arranged on the first limiting ring, and the first push rod is arranged in the first adjusting hole (307).

8. The device for pre-treatment of nucleic acid extraction from OCT-embedded renal biopsy tissue according to claim 3, characterized in that: The first adjustment hole (307) comprises a vertical strip hole, and a plurality of strip branch holes are arranged at intervals on both sides of the strip hole.

9. The device for pre-treatment of OCT-embedded renal biopsy tissue for nucleic acid extraction according to claim 7, characterized in that: The inner bottom of the carrier shell (305) is provided with an annular second limiting plate (308), a ball bearing is provided between the second limiting plate (308) and the inner bottom surface of the carrier shell (305), and an annular groove matching the sliding of the ball bearing is provided on the lower end surface of the second limiting plate (308) and the inner bottom surface of the carrier shell (305); the inner edge of the second limiting plate (308) is provided with a plurality of radially outward first limiting recesses (312) and a plurality of radially inward limiting protrusions (313), the limiting protrusions (313) are symmetrically arranged, and the first limiting recesses (312) are symmetrically arranged; a second adjustment hole (314) is provided on the carrier shell (305) contacted by the outer edge of the second limiting plate (308); a second push rod (309) is provided at the outer edge of the second limiting plate (308), and the second push rod (309) is placed in the second adjustment hole (314) at the bottom of the carrier shell (305).

10. The device for pre-treatment of OCT-embedded renal biopsy tissue for nucleic acid extraction according to claim 9, characterized in that: The bottom of the carrier shell (305) is provided with a second limiting recess corresponding to the first limiting recess (312), and the second limiting recess is larger than the first limiting recess (312).

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