Flushing device capable of stably obtaining in-vitro fallopian tube cavity fluid

Abstract

The utility model relates to a flushing device capable of stably obtaining in-vitro fallopian tube cavity fluid. The flushing device comprises a support, a first flat plate, a second flat plate, a sterile flusher, a first haemostatic forceps, a second haemostatic forceps, a sterile centrifugal tube and an ice box. The support comprises a supporting rod and a base. The supporting rod is vertically fixed on the upper surface of the base; the first flat plate and the second flat plate are fixed on a supporting rod of the bracket; the first haemostatic forceps are fixedly supported on the upper surface of the first flat plate; the first haemostatic forceps are used for clamping the umbrella part of the fallopian tube; the second hemostatic forceps are fixedly supported on the upper surface of the second flat plate; the second hemostatic forceps are used for clamping the isthmus of the fallopian tube; the sterile flusher is used for injecting flushing fluid into the fallopian tube from the umbrella part of the fallopian tube; the sterile centrifugal tube is arranged under the isthmus of the fallopian tube; the bottom of the sterile centrifugal tube is inserted into a fixing groove of the ice box; the ice box is arranged on the upper surface of the base; the sterile centrifugal tube is matched with the ice box; the flushing fluid is sterile normal saline or sterile phosphate buffer.

Description

Technical Field

[0001] The utility model relates to the technical field of reproductive medicine, in particular to a flushing device for stably obtaining extracorporeal fallopian tube lumen fluid. Background Art

[0002] In the female mammalian reproductive tract, the fallopian tube, a tubular organ connecting the ovary and uterus, plays a crucial physiological role in gamete transport, fertilization, and pre-implantation embryonic development. Anatomically, the fallopian tube consists of three main sections: the fimbria, closest to the ovary, which is finger-like and responsible for capturing oocytes released from the ovary; the ampulla, the site of fertilization, which contains numerous ciliated epithelial cells. Cilia beating transports the pre-implantation embryo to the isthmus, closer to the uterus, where it then enters the uterus for implantation.

[0003] The fallopian tube provides a favorable developmental microenvironment for key reproductive processes such as gamete transport, fertilization, and embryonic division, as well as key reproductive developmental events such as zygotic genome activation (ZGA). This includes stable temperature, pH, and dynamic secretion of substances. Dynamic secretion of the fallopian tube consists of two components: exudation from the systemic circulation and active production by the fallopian tube secretory epithelial cells, which together form the fallopian tube fluid.

[0004] Fallopian tubal fluid (also called luminal fluid) plays a crucial role in gamete and embryo development. First, it plays a nutritional role. Pyruvate, lactate, lipids, amino acids, and numerous cytokines and growth factors are present in fallopian tubal fluid. Pyruvate and lactate serve as the primary energy sources for oxidative metabolism in the preimplantation embryo. Growth factors (such as epidermal growth factor, transforming growth factor, insulin-like growth factor, and fibroblast growth factor) can positively influence embryonic development. Second, it plays a protective role. Fallopian tubal fluid protects gametes and embryos from environmental stressors such as oxidative stress, heat stress, and immune responses. Fallopian tubal fluid also contains antioxidant enzymes (such as superoxide dismutases (SODS), glutathione peroxidases (GPXs), and catalase) and non-enzymatic antioxidants (such as glutathione, taurine, transferrin, and albumin) that can mitigate oxidative stress on the embryo caused by reactive oxygen species (ROS). Fallopian tubal fluid also contains heat shock factors (HSFs), which ensure proper protein folding and protect cells from heat stress. In addition, the fallopian tubes also contain estrogen and estrogen receptor α, which not only protect the embryo from attacks by the maternal immune system, but also coordinately regulate oviposit secretion, ciliated epithelial cell oscillation, and smooth muscle contraction.

[0005] Assisted reproductive technology (ART) (including in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI)) is currently an effective means of solving the problem of infertility for couples. However, compared with the natural pregnancy rate of 45%-85%, the IVF / ICSI pregnancy success rate is only 20%-30%, and the live birth rate is only 43%. The biggest difference between IVF and natural conception is the difference in fertilization and the early embryonic development environment. Compared with the dynamic fallopian tube environment in the body of natural conception, IVF culture medium is static and predetermined, and cannot respond in time to harmful substances such as ROS produced by the embryo. Therefore, the key to improving the IVF blastocyst development rate is to design a culture medium that simulates normal female fallopian tube fluid. Accurately obtaining fallopian tube fluid and in-depth understanding of its material composition and subtle changes are prerequisites for the development of the above-mentioned culture medium. Carrying out such work is of great significance.

[0006] The existing technology does not have a complete set of stable flushing devices for obtaining extracorporeal fallopian tube fluid. It usually requires two or more people to clamp the fimbria, isthmus and other parts of the fallopian tube and use a syringe to collaboratively obtain extracorporeal fallopian tube fluid (also called fallopian tube lavage fluid).

[0007] The existing method for obtaining in vitro fallopian tube lavage fluid is to use a syringe to draw air and a certain volume of PBS, saline, or culture medium. This is then injected into the isthmus of the in vitro fallopian tube tissue obtained from a patient who has given informed consent and has a clear surgical indication. The fallopian tube outflow fluid is then collected from the fimbria, which is then centrifuged to remove cells and debris. The supernatant, the fallopian tube lavage fluid, is then stored for subsequent testing. However, this existing method suffers from the following two major deficiencies:

[0008] First, accurately obtaining tubal lavage fluid is difficult. 1) Because the lumen of the isthmus is narrow, typically only 1mm in diameter, using a syringe with a needle to insert the needle through the isthmus can easily puncture the tubal tissue, making it difficult to inject fluid into the lumen and recover tubal lavage fluid from the fimbria. 2) Even if the needle is removed, the diameter of the tip of a conventional syringe is still larger than the diameter of the isthmus, and the tip of a conventional syringe is short, making it difficult to reliably inject fluid from the isthmus. Consequently, multiple people are often required to perform the procedure, which takes a long time and results in a small volume of tubal fluid recovered. 3) Even if a syringe without the needle is used to inject fluid from the fimbria, although the diameter of the fimbria is slightly larger than the syringe, the short tip of the syringe prevents it from being firmly positioned within the fimbria for lavage. Consequently, the fimbria requires prolonged handholding, which is time-consuming and laborious, resulting in inefficient lavage. Furthermore, even the slightest carelessness can cause lavage fluid to flow from the outside of the tubal lumen, resulting in the possibility of not obtaining the intended lavage fluid and even the presence of external contaminants.

[0009] Secondly, the obtained fallopian tube lavage fluid is not very reliable for scientific research. 1) The existing technology flushing fluid flushes directly through the fallopian tube cavity, and the time it flows through the fallopian tube is short, and the material is not fully washed out; and the fallopian tube is not filled, and the fallopian tube fluid in some folds may not be flushed down; these two factors together lead to a large difference in the type and content of the substances in the fallopian tube cavity itself and the actual fallopian tube lavage fluid obtained. 2) The slightest carelessness during flushing can easily cause the flushing fluid to splash, and the fallopian tube is difficult to ensure that it is placed vertically, resulting in some liquid remaining in the fallopian tube cavity. These factors lead to large errors in the weight / volume and other aspects of the fallopian tube lavage fluid obtained between different samples. The scientificity of continuing to use these lavage fluids for subsequent comparative studies between samples is questionable.

[0010] Therefore, it is necessary to develop a flushing device and its supporting method for stably obtaining fallopian tube fluid in vitro. Summary of the Invention

[0011] The utility model aims to provide a flushing device for stably obtaining extracorporeal fallopian tube cavity fluid, and the technical problems to be solved include at least the difficulty in accurately obtaining extracorporeal fallopian tube cavity fluid and the low reliability of the obtained extracorporeal fallopian tube cavity fluid caused by the lack of a flushing device for stably obtaining extracorporeal fallopian tube cavity fluid.

[0012] In order to achieve the above-mentioned purpose, the utility model provides a flushing device for stably obtaining in vitro fallopian tube cavity fluid, comprising a bracket, a first plate, a second plate, a sterile flusher, a first hemostatic forceps, a second hemostatic forceps, a sterile centrifuge tube and an ice box; the bracket comprises a support rod and a base; the support rod is vertically fixed on the upper surface of the base; the first plate and the second plate are fixed on the support rod of the bracket; the first hemostatic forceps is fixedly supported on the upper surface of the first plate; the first hemostatic forceps is used to clamp the fallopian tube fimbria; the second hemostatic forceps is fixedly supported on the upper surface of the second plate; the second hemostatic forceps is used to clamp the fallopian tube isthmus; the sterile flusher is used to inject flushing fluid from the fallopian tube fimbria into the inside of the fallopian tube; the sterile centrifuge tube is arranged directly below the fallopian tube isthmus; the bottom of the sterile centrifuge tube is inserted into the fixing groove of the ice box; the ice box is arranged on the upper surface of the base; the sterile centrifuge tube and the ice box match each other; the flushing fluid is sterile saline or sterile phosphate buffer.

[0013] Preferably, the first flat plate and the second flat plate are adjustably fixed to the support rod of the bracket by means of clips or threads; and the distance between the first flat plate and the second flat plate is 8-12 cm.

[0014] Preferably, the first flat plate and the second flat plate are made of iron or stainless steel, preferably stainless steel.

[0015] Preferably, the annular handles on one side of the first hemostatic forceps and the second hemostatic forceps are respectively inserted into the support rod, or the first hemostatic forceps is fixedly supported on the upper surface of the first flat plate by a first locking component, and the second hemostatic forceps is fixedly supported on the upper surface of the second flat plate by a second locking component; the first locking component and the second locking component adopt a clamping or snapping method to achieve a fixed connection between the first hemostatic forceps and the first flat plate, and the second hemostatic forceps and the second flat plate.

[0016] Preferably, the first hemostatic forceps and the second hemostatic forceps are stainless steel straight full-tooth hemostatic forceps specially used for surgery.

[0017] Preferably, the clamp ends of the first hemostatic forceps and the second hemostatic forceps extend out of the edge of the upper surface of the first plate and the second plate respectively; the two clamp ends extend out of the edge by at least 0.5 cm, and the front ends of the clamp ends correspond vertically.

[0018] Preferably, the sterile centrifuge tube and the ice box match each other, and the specifications are one of 15 mL and 50 mL.

[0019] Preferably, the sterile flusher is disposable; the flushing fluid is sterile physiological saline or sterile phosphate buffer.

[0020] Compared with the prior art, the beneficial effects of the present invention are:

[0021] The key innovation of the present invention is that it proposes for the first time a flushing device for stably obtaining fallopian tube lumen fluid in vitro. This flushing device effectively solves the problem of the difficulty in accurately obtaining fallopian tube lavage fluid in existing / previous technical methods. The present application only requires one person to suspend and fix the fallopian tube on the invented flushing device for fallopian tube lavage, which is significantly more advantageous than the existing technology that requires close cooperation of multiple people (i.e., it saves manpower). The present application also achieves time-saving and convenient effects by replacing an ordinary syringe with a medical flushing device, and changing the method of injecting the flushing fluid from the isthmus of the fallopian tube to the fimbria.

[0022] At the same time, the flushing device described in this application is more scientific than the prior art for stably obtaining fallopian tube lumen fluid in vitro. By replacing the prior art's rapid outflow of the flushing fluid with a system that allows the fluid to remain within the fallopian tube for a period of time before flowing out, and by replacing the fallopian tube's horizontal position with a naturally stretched vertical position, the present invention allows the fluid within the fallopian tube to fully contact the flushing fluid and be washed out, thereby increasing the content of substances. It also reduces the dead volume remaining in the fallopian tube, resulting in a higher volume of the obtained fallopian tube lumen fluid. The flushing fluid is slowly injected, thus avoiding the risk of splashing and spillage common in the prior art, making quantitative determination more accurate and significantly improving the reliability of comparisons between samples. Furthermore, the use of the device described in this application significantly reduces the need for repeated manual handling of the fallopian tube, thereby reducing the risk of introducing external contaminants. The ice box quickly and cold preserves the fallopian tube effluent, preserving more active substances in the fluid. The distance between the first and second clamping plates of the flushing device can be adjusted according to the length of different fallopian tubes.

[0023] The applicant has completed metabolomics and proteomics testing of fallopian tube fluid in more than 100 patients undergoing bilateral hysterectomy and salpingo-oophorectomy using the device invented in this application, confirming the feasibility of the technical solution of this application.

[0024] The flushing device for stably obtaining in vitro fallopian tube fluid described in the present application is time-saving and labor-saving, convenient and efficient, easy to carry, and highly flexible. It can meet the needs of clinical research and has great application value in clinical practice. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] The accompanying drawings are used to provide a further understanding of the technical solution of the present invention and constitute a part of the specification. Together with the specific implementation methods of this application, they are used to explain the technical solution of the present invention and do not constitute a limitation on the technical solution of the present invention.

[0026] Figure 1 It is a structural schematic diagram of the flushing device for obtaining extracorporeal fallopian tube cavity fluid according to the utility model.

[0027] Figure 2 1 is a comparison chart of the flushing time using the conventional method and the method of the present application in Examples 1, 2, and 3.

[0028] Figure 3 1 is a comparison chart of the volumes of the flushing fluids obtained by the conventional method and the method of the present application in Examples 1, 2, and 3.

[0029] Figure 4 This is a BCA method total protein detection chart of the flushing liquid obtained by the previous method and the method of the present application in Examples 1, 2, and 3. DETAILED DESCRIPTION

[0030] The present invention is described in more detail below to facilitate understanding of the present invention.

[0031] like Figure 1 As shown, the flushing device for stably obtaining in vitro fallopian tube cavity fluid of the present invention comprises a bracket 1, a first plate 2, a second plate 4, a sterile flusher 6, a first hemostatic forceps 7, a second hemostatic forceps 8, a sterile centrifuge tube 9 and an ice box 10; the bracket 1 comprises a support rod 13 and a base 14; the support rod 13 is vertically fixed to the upper surface of the base 14; the first plate and the second plate are fixed to the support rod of the bracket; the first hemostatic forceps is fixedly supported on the upper surface of the first plate; the first hemostatic forceps is used to clamp The fallopian tube fimbria 11; a second hemostatic forceps is fixedly supported on the upper surface of the second flat plate; the second hemostatic forceps is used to clamp the isthmus of the fallopian tube; the sterile flusher 6 is used to inject flushing fluid into the interior of the fallopian tube from the fimbria 11 of the fallopian tube; a sterile centrifuge tube 9 is arranged directly below the isthmus 12 of the fallopian tube; the bottom of the sterile centrifuge tube 9 is inserted into the fixing groove of the ice box 10; the ice box 10 is arranged on the upper surface of the base 14; the sterile centrifuge tube and the ice box match each other; the flushing fluid is sterile saline or sterile phosphate buffer.

[0032] Preferably, the first flat plate and the second flat plate are adjustably fixed to the support rod of the bracket by means of clips or threads; and the distance between the first flat plate and the second flat plate is 8-12 cm.

[0033] Preferably, the first flat plate and the second flat plate are made of iron or stainless steel, preferably stainless steel.

[0034] Preferably, the annular handles on one side of the first hemostatic forceps 7 and the second hemostatic forceps 8 are respectively inserted into the support rod 13, or the first hemostatic forceps 7 is fixedly supported on the upper surface of the first flat plate 2 by a first locking component, and the second hemostatic forceps 8 is fixedly supported on the upper surface of the second flat plate 4 by a second locking component; the first locking component and the second locking component adopt a clamping or snapping method to achieve a fixed connection between the first hemostatic forceps and the first flat plate, and the second hemostatic forceps and the second flat plate.

[0035] Preferably, the clamp ends of the first hemostatic forceps and the second hemostatic forceps extend out of the edge of the upper surface of the first plate and the second plate respectively; the two clamp ends extend out of the edge by at least 0.5 cm, and the front ends of the clamp ends correspond vertically.

[0036] Preferably, the sterile centrifuge tube and the ice box match each other, and the specifications are one of 15 mL and 50 mL.

[0037] Preferably, the sterile flusher is disposable; the flushing fluid is sterile physiological saline or sterile phosphate buffer.

[0038] Preferably, the ice box 10 is a low-temperature ice box, and the low-temperature temperature is about 4°C.

[0039] The flushing method using the flushing device for stably obtaining in vitro fallopian tube lumen fluid of the utility model comprises the following steps:

[0040] S1. Place the stent flushing device in a standard Class 10,000 laminar flow operating room.

[0041] A Class 10,000 laminar flow operating room is one with a cleanliness level of Class 10,000 and a dust particle count of 0.5 μm or larger exceeding 35,000 particles / m 3 (35 particles / L) to less than or equal to 350,000 particles / m 3 (350 particles / L); the number of dust particles larger than or equal to 5μm is greater than 300 particles / m 3 (0.3 particles / L) to less than or equal to 3000 particles / m 3 (3 grains / L).

[0042] S2. Fill a sterile irrigator (which can be disposable) with a fixed amount of irrigating fluid. Gently insert the blunt end of the sterile irrigator into the fimbria of the fallopian tube, ensuring that the front end of the sterile irrigator is placed inside the fallopian tube lumen. Then, clamp the fallopian tube with the first hemostatic forceps, enclosing the fimbria of the sterile irrigator. Allow the fallopian tube to droop naturally, with the lowest point of the isthmus of the fallopian tube 1-2 cm from the opening of a sterile centrifuge tube directly below the ice box.

[0043] S3. The rinsing fluid is slowly injected or flows naturally into the fallopian tube, and when the fallopian tube is filled and expanded and there is watery outflow below the isthmus, immediately clamp the isthmus of the fallopian tube with the second hemostatic forceps below and stop the injection of the rinsing fluid;

[0044] S4. After keeping the fallopian tube filled for an appropriate amount of time (eg, 5-60 min), release the second hemostatic forceps and collect the effluent from the isthmus of the fallopian tube with the sterile centrifuge tube below; and continue to use the sterile irrigator above to slowly inject the irrigating fluid into the fallopian tube lumen for lavage until the irrigating fluid in the sterile irrigator is completely used up;

[0045] S5. The liquid in the sterile centrifuge tube is centrifuged to obtain the supernatant, which is the oviduct cavity fluid, and then the supernatant is packaged and stored.

[0046] The key innovation of this application is that it proposes for the first time a flushing device for stably obtaining fallopian tube cavity fluid in vitro. Compared with the existing technology, this application has a shorter flushing time, a larger volume of fallopian tube cavity flushing fluid obtained, and is more labor-saving. The application of the device described in this application reduces human touch and the risk of introducing external contaminants. The low-temperature treatment retains more active substances in the fallopian tube fluid, which is more mobile and can be adjusted according to the length of different fallopian tubes. It is operational and convenient in clinical applications. At present, more than ten metabolomics and proteomics tests on fallopian tube fluid of patients undergoing bilateral hysterectomy have been completed, confirming the feasibility of this technology.

[0047] The following three specific examples verify the use effect of the flushing device for obtaining extracorporeal fallopian tube fluid described in the present invention.

[0048] Example 1:

[0049] A patient underwent a hysterectomy with bilateral salpingo-oophorectomy for ovarian teratoma and multiple uterine fibroids. Both fallopian tubes were cut at the isthmus. The length and width of the left fallopian tube were 2.75 cm × 0.5 cm, and the right fallopian tube was 3.5 cm × 0.5 cm.

[0050] The left fallopian tube was flushed using conventional methods. A 5mL disposable sterile syringe was used to draw up 5mL of sterile saline. One person held the isthmus of the fallopian tube and the syringe, inserting the syringe through the isthmus into the tubal lumen while maintaining stability. The other person held a sterile centrifuge tube to collect the fluid flowing from the fimbria of the fallopian tube.

[0051] The right fallopian tube is flushed using the device described in this application to obtain flushing fluid. The flushing method specifically includes the following steps:

[0052] S1. Place the stent flushing device in a standard Class 10,000 laminar flow operating room.

[0053] S2. Fill a sterile irrigator (which can be disposable) with a fixed amount of irrigating fluid. Gently insert the blunt end of the sterile irrigator into the fimbria of the fallopian tube, ensuring that the front end of the sterile irrigator is placed inside the fallopian tube lumen. Then, clamp the fallopian tube with the first hemostatic forceps, enclosing the fimbria of the sterile irrigator. Allow the fallopian tube to droop naturally, with the lowest point of the isthmus of the fallopian tube 1-2 cm from the opening of a sterile centrifuge tube directly below the ice box.

[0054] S3. The rinsing fluid is slowly injected or flows naturally into the fallopian tube, and when the fallopian tube is filled and expanded and there is watery outflow below the isthmus, immediately clamp the isthmus of the fallopian tube with the second hemostatic forceps below and stop the injection of the rinsing fluid;

[0055] S4. After keeping the fallopian tube filled for an appropriate amount of time (e.g., 5 min), release the second hemostatic forceps and collect the effluent from the isthmus of the fallopian tube with the sterile centrifuge tube below; and continue to use the sterile irrigator above to slowly inject the irrigating fluid into the fallopian tube lumen for irrigation until the irrigating fluid in the sterile irrigator is completely used up;

[0056] S5. The liquid in the sterile centrifuge tube is centrifuged to obtain the supernatant, which is the oviduct cavity fluid, and then the supernatant is packaged and stored.

[0057] Example 2:

[0058] A patient underwent a hysterectomy with bilateral salpingo-oophorectomy for multiple uterine fibroids and abnormal uterine bleeding. Both fallopian tubes were sectioned at the isthmus. Measurements of the length and width of the tubal bodies revealed the left tubal size to be 2 cm × 0.5 cm, and the right tubal size to be 2.75 cm × 0.5 cm.

[0059] The left fallopian tube was flushed using the conventional method.

[0060] The right fallopian tube is flushed using the flushing device of the present application to obtain flushing fluid. The flushing method specifically includes the following steps:

[0061] S1. Place the stent flushing device in a standard Class 10,000 laminar flow operating room.

[0062] S2. Fill a sterile irrigator (which can be disposable) with a fixed amount of irrigating fluid. Gently insert the blunt end of the sterile irrigator into the fimbria of the fallopian tube, ensuring that the front end of the sterile irrigator is placed inside the fallopian tube lumen. Then, clamp the fallopian tube with the first hemostatic forceps, enclosing the fimbria of the sterile irrigator. Allow the fallopian tube to droop naturally, with the lowest point of the isthmus of the fallopian tube 1-2 cm from the opening of a sterile centrifuge tube directly below the ice box.

[0063] S3. The rinsing fluid is slowly injected or flows naturally into the fallopian tube, and when the fallopian tube is filled and expanded and there is watery outflow below the isthmus, immediately clamp the isthmus of the fallopian tube with the second hemostatic forceps below and stop the injection of the rinsing fluid;

[0064] S4. After keeping the fallopian tube filled for an appropriate amount of time (eg, 5-60 min), release the second hemostatic forceps and collect the effluent from the isthmus of the fallopian tube with the sterile centrifuge tube below; and continue to use the sterile irrigator above to slowly inject the irrigating fluid into the fallopian tube lumen for lavage until the irrigating fluid in the sterile irrigator is completely used up;

[0065] S5. The liquid in the sterile centrifuge tube is centrifuged to obtain the supernatant, which is the oviduct cavity fluid, and then the supernatant is packaged and stored.

[0066] Example 3:

[0067] A patient underwent hysterectomy and bilateral adnexectomy due to ovarian chocolate cyst, adenomyosis and other diseases. The fallopian tubes on both sides were cut at the isthmus, and the length and width of the fallopian tubes were measured: the left fallopian tube was 3cm×0.5cm, and the right fallopian tube was 3cm×0.5cm.

[0068] The left fallopian tube was flushed using the conventional method.

[0069] The right fallopian tube is flushed with the device of the present application to obtain flushing fluid.

[0070] The flushing method specifically includes the following steps:

[0071] S1. Place the stent flushing device in a standard Class 10,000 laminar flow operating room.

[0072] S2. Fill a sterile irrigator (which can be disposable) with a fixed amount of irrigating fluid. Gently insert the blunt end of the sterile irrigator into the fimbria of the fallopian tube, ensuring that the front end of the sterile irrigator is placed inside the fallopian tube lumen. Then, clamp the fallopian tube with the first hemostatic forceps, enclosing the fimbria of the sterile irrigator. Allow the fallopian tube to droop naturally, with the lowest point of the isthmus of the fallopian tube 1-2 cm from the opening of a sterile centrifuge tube directly below the ice box.

[0073] S3. The rinsing fluid is slowly injected or flows naturally into the fallopian tube, and when the fallopian tube is filled and expanded and there is watery outflow below the isthmus, immediately clamp the isthmus of the fallopian tube with the second hemostatic forceps below and stop the injection of the rinsing fluid;

[0074] S4. After keeping the fallopian tube filled for an appropriate amount of time (e.g., 5 min), release the second hemostatic forceps and collect the effluent from the isthmus of the fallopian tube with the sterile centrifuge tube below; and continue to use the sterile irrigator above to slowly inject the irrigating fluid into the fallopian tube lumen for irrigation until the irrigating fluid in the sterile irrigator is completely used up;

[0075] S5. The liquid in the sterile centrifuge tube is centrifuged to obtain the supernatant, which is the oviduct cavity fluid, and then the supernatant is packaged and stored.

[0076] The key innovation of this application is that it proposes for the first time a flushing device for stably obtaining fallopian tube lumen fluid in vitro.

[0077] 1. The fallopian tube flushing fluid obtained by the flushing device of the present application takes a shorter average flushing time than the previous method. Figure 2In the above three examples, the time taken by the previous method was 803±164.2s (n=3), while the time taken by the device of the present application to flush was 442.3±57.7s (n=3). The flushing time of the present application was shorter, and the difference was significant.

[0078] 2. Compared with the previous method, the volume of the oviduct flushing fluid obtained by the flushing of the present application is larger under the premise of the same flushing volume (all 5 mL), as shown in the attached Figure 3 In the above three examples, the volume of the rinsing fluid collected by the previous method was 2.88±0.31mL (n=3), and the volume of the rinsing fluid collected by the present application was 4.63±0.23mL (n=3). The volume of the rinsing fluid obtained by the present application was larger, and the difference was significant.

[0079] 3. Compared with the previous method, the oviduct rinsing fluid obtained by the present invention has a higher total protein content in the rinsing fluid collected from the isthmus under the premise of the same rinsing volume (all 5 mL). Figure 4 As shown. In Example 1, the total protein concentration in the fallopian tube lavage fluid obtained by the previous method is 322.33±46.6μg / mL, and the total protein concentration in the fallopian tube lavage fluid obtained by the lavage of the present application is 425.44±28.12μg / mL, and the difference is significant. In Example 2, the total protein concentration in the fallopian tube lavage fluid obtained by the previous method is 197.22±17.02μg / mL, and the total protein concentration in the fallopian tube lavage fluid obtained by the lavage of the present application is 681±49.19μg / mL, and the difference is highly significant. In Example 3, the total protein concentration in the fallopian tube lavage fluid obtained by the previous method is 282±32.34μg / mL, and the total protein concentration in the fallopian tube lavage fluid obtained by the lavage of the present application is 340.56±18.50μg / mL, and the difference is significant.

[0080] 4. The fallopian tube lavage fluid obtained in this application contains more protein types than those obtained by previous methods. When the same mass of protein was loaded in each channel (2 μg total protein), the relative abundance of proteins was quantitatively analyzed using polyacrylamide gel electrophoresis combined with silver staining. From left to right, the sample channels are: Channel ①: 14kDa-250kDa protein molecular weight marker, channel; ②: total protein of the lavage fluid obtained by the previous method in Example 1, channel; ③: total protein of the lavage fluid obtained in this application in Example 1, channel; ④: total protein of the lavage fluid obtained by the previous method in Example 2, channel; ⑤: total protein of the lavage fluid obtained in this application in Example 2, channel; ⑥: total protein of the lavage fluid obtained by the previous method in Example 3, channel; ⑦: total protein of the lavage fluid obtained in this application in Example 3, channel; ⑧: total protein of the 293T cell line. As shown in Table 1, grayscale analysis of the bands using imageJ software revealed that the total band intensity values ​​of channels ③, ⑤, and ⑦ were stronger than those of channels ②, ④, and ⑥, respectively. Taking the protein band of <14 kDa as an example, the corresponding band intensities of channels ③, ⑤, and ⑦ are all stronger than those of channels ②, ④, and ⑥, respectively. This indicates that the oviduct fluid obtained in Examples 1, 2, and 3 using the present invention has more protein bands than the oviduct fluid obtained by the previous method, i.e., the relative abundance is higher and the protein types are more diverse.

[0081] 5. The previous method required two people to assist in completing the flushing and collection of fallopian tube fluid each time, but this application can be completed by one person.

[0082] Table 1 Analysis of the intensity of silver staining bands by Image J software

[0083]

[0084] The above describes the preferred embodiments of the present invention, but it is not intended to limit the present invention. Those skilled in the art may make improvements and changes to the embodiments disclosed herein without departing from the scope and spirit of the present invention.

Claims

1. A flushing device for stably obtaining extracorporeal fallopian tube fluid, characterized in that: The flushing device for stably obtaining in vitro fallopian tube cavity fluid includes a bracket, a first plate, a second plate, a sterile flusher, a first hemostatic forceps, a second hemostatic forceps, a sterile centrifuge tube and an ice box; the bracket includes a support rod and a base; the support rod is vertically fixed to the upper surface of the base; the first plate and the second plate are fixed to the support rod of the bracket; the first hemostatic forceps is fixedly supported on the upper surface of the first plate; the first hemostatic forceps is used to clamp the fallopian tube fimbria; the second hemostatic forceps is fixedly supported on the upper surface of the second plate; the second hemostatic forceps is used to clamp the fallopian tube isthmus; the sterile flusher is used to inject flushing fluid from the fallopian tube fimbria into the interior of the fallopian tube; the sterile centrifuge tube is arranged directly below the fallopian tube isthmus; the bottom of the sterile centrifuge tube is inserted into the fixing groove of the ice box; the ice box is arranged on the upper surface of the base; the sterile centrifuge tube and the ice box match each other; the flushing fluid is sterile saline or sterile phosphate buffer.

2. The flushing device for stably obtaining in vitro fallopian tube lumen fluid according to claim 1, characterized in that: The first and second flat plates are adjustably fixed to the support rods of the bracket by means of clips or threads; the distance between the first and second flat plates is 8-12 cm.

3. The flushing device for stably obtaining in vitro fallopian tube lumen fluid according to claim 1, characterized in that: The first flat plate and the second flat plate are made of iron or stainless steel.

4. The flushing device for stably obtaining in vitro fallopian tube lumen fluid according to claim 1, characterized in that: The annular handles on one side of the first hemostatic forceps and the second hemostatic forceps are respectively inserted into the support rod, or the first hemostatic forceps is fixedly supported on the upper surface of the first flat plate by the first locking component, and the second hemostatic forceps is fixedly supported on the upper surface of the second flat plate by the second locking component; the first locking component and the second locking component adopt a clamping or snapping method to achieve a fixed connection between the first hemostatic forceps and the first flat plate, and the second hemostatic forceps and the second flat plate.

5. The flushing device for stably obtaining in vitro fallopian tube lumen fluid according to claim 1, characterized in that: The first hemostatic forceps and the second hemostatic forceps are stainless steel straight full-tooth hemostatic forceps specially used for surgery.

6. The flushing device for stably obtaining in vitro fallopian tube lumen fluid according to claim 1, characterized in that: The clamp ends of the first hemostatic forceps and the second hemostatic forceps extend out of the edges of the upper surfaces of the first flat plate and the second flat plate respectively.

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