A multi-well plate sperm screening device based on the upstream method

Abstract

The utility model relates to a porous plate sperm screening device based on upstream method, including porous plate base and sperm screening spare, the body middle part recessed and recessed bottom interval of porous plate base is provided with two or more than two culture holes, and each culture hole is provided with a sperm screening spare in a matched mode, the main part of sperm screening spare is embedded in the culture hole, and the culture hole wall top is sleeved in through the sleeve setting of sperm screening spare top, the main part bottom of sperm screening spare sets up the annular buffer ring groove that forms relative to the main part of sperm screening spare and extends inwards, and multiple channels are arranged on the inner side groove wall of buffer ring groove, and the main part bottom of sperm screening spare forms the communicating port in the inner side of buffer ring groove, and the filter membrane that seals the communicating port and is used for sperm upstream is arranged at the communicating port, at least one concave sampling groove is arranged at the bottom of buffer ring groove.

Description

Technical Field

[0001] This utility model relates to the technical field of devices for sperm selection using the sperm upstream method in human assisted reproduction, and particularly to a multi-well plate sperm screening device based on the upstream method. Background Technology

[0002] In the field of assisted reproductive technology, sperm optimization is a key step in improving the success rate of male infertility treatment. Currently, the main technologies in this area include sperm swim-up, density gradient centrifugation, membrane filtration, and microfluidic technology.

[0003] The sperm swim-up method utilizes the upstream characteristics of sperm to screen for highly motile sperm. While traditional sperm swim-up methods are simple to perform, surface agitation can easily cause low-motile sperm to flow back, reducing the recovery rate of highly motile sperm in the final extracted sperm sample. This can also affect sperm quality, consequently impacting fertilization success rates. The problem lies in surface agitation leading to the backflow of low-motile sperm and low recovery rates. While density gradient centrifugation can screen for sperm with good morphology and motility, the centrifugation process can cause sperm DNA damage, reducing the success rate of assisted reproduction and potentially having long-term adverse effects on fetal development.

[0004] In recent years, membrane filtration technology has been increasingly applied to sperm screening. For example, Chinese utility model patent CN213570453U, entitled "A Sperm Concentrator," uses membrane filtration to concentrate sperm instead of centrifugal concentration, reducing operational steps and the generation of oxygen free radicals. However, its function is to concentrate semen that has already undergone upstream screening, and it cannot achieve high-throughput screening of highly motile sperm. Additionally, Chinese utility model patent CN213085967U, entitled "An Assisted Reproductive Operating Device for Direct Upstream Sperm Screening," while capable of screening highly motile sperm, requires adding culture medium directly above the sperm screening membrane. This can easily disrupt the surface of the semen sample below the membrane, causing inactive or weakly motile sperm to pass through the membrane and rise above it, affecting the quality of sperm screening.

[0005] In addition, although microfluidic technology can achieve high-throughput sperm screening, its high equipment cost and complex operation limit its widespread clinical application.

[0006] The limitations of existing sperm screening technologies have restricted the development and application of sperm optimization techniques to some extent, failing to meet clinical needs for high-throughput, high-recovery, and non-destructive screening of high-quality sperm. Utility Model Content

[0007] To address the aforementioned problems, this invention provides a multi-well plate sperm screening device based on the upstream method.

[0008] To achieve the above objectives, the present invention adopts the following technical solution:

[0009] A multi-well plate sperm screening device based on the upstream method includes a multi-well plate base and a sperm screening component. The multi-well plate base has a concave center with two or more culture wells spaced apart at the bottom of the concave area. Each culture well is equipped with a sperm screening component. The main body of the sperm screening component is embedded in the culture well and is fitted onto the top of the culture well wall by a sleeve provided at the top of the sperm screening component. The bottom of the main body of the sperm screening component has an annular buffer groove extending inward relative to the main body of the sperm screening component. Multiple channels are opened at intervals on the inner wall of the buffer groove. The bottom of the main body of the sperm screening component forms a connecting port inside the buffer groove. A filter membrane is provided at the connecting port to seal the connecting port and allow sperm to pass upstream. At least one concave sampling groove is provided at the bottom of the buffer groove. The sampling groove is used to hold hyaluronic acid to induce high-quality sperm to enter the sampling groove. The added height of the hyaluronic acid is lower than the top surface of the sampling groove.

[0010] Furthermore, the body of the porous plate base is square or cylindrical, the top surface of the culture well is lower than or flush with the top surface of the body of the porous plate base, and the main body of the sperm screening component is cylindrical to match the shape of the culture well.

[0011] Furthermore, the porous plate base is a four-well culture dish.

[0012] Furthermore, the space between the body of the sperm screening component and the culture well allows the pipette tip to extend into it.

[0013] Furthermore, the sleeve is an annular or semi-annular piece with an inverted L-shaped cross-section, integrally formed with the main body of the sperm screening component and extending outward relative to the main body of the sperm screening component, or a plurality of pendants with an inverted L-shaped cross-section arranged at intervals on the top of the main body of the sperm screening component by integral forming or by adhesive bonding.

[0014] Furthermore, the inner wall of the buffer ring groove has multiple channels evenly spaced, and the bottom surface of the channels is higher than the bottom surface of the buffer ring groove.

[0015] Furthermore, the filter membrane is a porous polycarbonate membrane, which is bonded to the bottom of the sperm screening element body by a biocompatible epoxy resin adhesive.

[0016] Furthermore, the shape of the sampling groove matches the shape of the pipette tip.

[0017] Furthermore, the sperm screening component is directly formed by 3D printing from biocompatible materials.

[0018] This utility model has the following beneficial effects:

[0019] The design employs a buffer ring groove with channels, which reduces the impact on the semen sample during culture medium addition, preventing low-motility sperm, poorly developed sperm, and fragmented impurities from entering the culture medium within the buffer ring groove, while also ensuring sufficient contact area between the two. A sampling groove for hyaluronic acid, which induces sperm entry, is designed at the bottom of the buffer ring groove. After passing through the filter membrane, sperm are induced into the sampling groove, improving the purity of the sampled sperm and facilitating the sampling of high-quality sperm.

[0020] This device avoids the cumbersome and time-consuming nature of traditional operations and the potential damage to sperm caused by centrifugation, significantly improving the recovery rate of high-motility sperm, enhancing the efficiency of upstream sperm preparation, and reducing operational difficulty, thus enabling the acquisition of more high-quality sperm for in vitro fertilization.

[0021] In addition, the device has a simple structure and is easy to prepare. It can be directly formed by 3D printing, which meets the clinical requirements for single use. The biocompatible materials used are widely available, easy to obtain and cost-controllable, which facilitates its promotion and application and has high clinical practical value. Attached Figure Description

[0022] Figure 1 This is a three-dimensional structural diagram of the present invention;

[0023] Figure 2 This is a top-view structural schematic diagram of the present invention;

[0024] Figure 3 This is a three-dimensional structural diagram of the sperm screening component of this utility model;

[0025] Figure 4 This is another three-dimensional structural diagram of the sperm screening component of this utility model. Detailed Implementation

[0026] The present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments:

[0027] See Figures 1 to 4 A multi-well plate sperm screening device based on the upstream method includes a multi-well plate base 1 and a sperm screening component 2.

[0028] The body 11 of the multi-well plate base 1 is square or cylindrical. The middle of the body 11 is concave, and two or more culture wells 13 are spaced apart at the bottom of the concave part 12. The top surface of the culture wells 13 is lower than or flush with the top surface of the body 11 of the multi-well plate base 1. In this embodiment, a four-well culture dish is specifically selected as the multi-well plate base 1.

[0029] Each culture well 13 is equipped with a sperm screening component 2. The main body 21 of the sperm screening component 2 is cylindrical, matching the shape of the culture well 13, and is embedded within the culture well 13. Preferably, the space between the main body 21 of the sperm screening component 2 and the culture well 13 is sufficient for a pipette tip to extend into. In this embodiment, the sperm screening component 2 is directly formed by 3D printing from a biocompatible material.

[0030] See Figure 3 and Figure 4 The sperm screening component 2 has a retainer 22 fitted onto the top of the culture well 13. Specifically, the retainer 22 can be an annular or semi-annular component with an inverted L-shaped cross-section, integrally formed with the main body 21 of the sperm screening component 2 and extending outward relative to the main body 21. Alternatively, it can be a plurality of inverted L-shaped hanging components spaced apart on the top of the main body 21 of the sperm screening component 2, either integrally formed or bonded. In this embodiment, the retainer 22 is specifically a semi-annular component with an inverted L-shaped cross-section, integrally formed with the main body 21 of the sperm screening component 2 and extending outward relative to the main body 21. The pipette tip can be inserted from the other side of the semi-annular component into the space between the main body 21 of the sperm screening component 2 and the culture well 13 to add semen samples into the culture well 13.

[0031] The sperm screening element 2 has a ring-shaped buffer groove 23 extending inward from the main body 21 at its bottom. Multiple channels 232 are spaced apart on the inner wall 231 of the buffer groove 23, with the bottom surface of each channel 232 higher than the bottom surface of the buffer groove 23. In this embodiment, six channels 232 are evenly spaced on the inner wall 231 of the buffer groove 23. At least one recessed sampling groove 24 is provided at the bottom of the buffer groove 23. The sampling groove 24 is used to hold hyaluronic acid that induces high-quality sperm to enter the sampling groove 24. The added height of the hyaluronic acid is lower than the top surface of the sampling groove 24, and the shape of the sampling groove 24 matches the shape of the pipette tip, facilitating the pipette tip to aspirate high-quality sperm from the sampling groove 24.

[0032] The bottom of the main body 21 of the sperm screening element 2 forms a communication port inside the buffer ring groove 23. A filter membrane 25, which seals the communication port but is permeable from top to bottom, is provided to allow sperm to pass through upstream. In this embodiment, the filter membrane 25 is specifically a porous polycarbonate membrane, which is connected to the bottom of the main body 21 of the sperm screening element 2 by means of biocompatible epoxy resin adhesive. The filter membrane and the sperm screening element are bonded together with biocompatible epoxy resin adhesive to ensure airtightness, and the sperm survival rate can reach more than 80% after being stored in it for 48 hours.

[0033] The sperm screening component 2 is fitted onto the culture well 13 of the multi-well plate base 1 using a sleeve 22, with the main body 21 of the sperm screening component 2 embedded within the culture well 13 of the multi-well plate base 1. Simultaneously or before installing the sperm screening component 2 onto the multi-well plate base 1, a semen sample is added to the culture well 13 of the multi-well plate base 1. Culture medium is slowly added along the inner sidewall of the main body 21 of the sperm screening component 2, flowing along the inner sidewall to the buffer ring groove 23. The buffer ring groove 23 reduces the direct impact on the semen sample during culture medium addition. Multiple channels 232 on the inner wall 231 of the buffer ring groove 23 ensure sufficient contact area between the culture medium and the semen sample, facilitating faster screening of high-quality sperm. After sperm screening is complete, the sperm sample is removed from the buffer ring groove 23 and the sampling groove 24.

[0034] The method for screening highly motile sperm using the above-mentioned upstream-based multi-well plate sperm screening device includes the following steps:

[0035] S100. Add 2-4 mL of semen sample to the culture well 13 of the multi-well plate base 1 and let it stand on the test tube rack for 1 min until impurities precipitate.

[0036] S200. Attach the filter membrane 25 to the bottom of the main body 21 of the sperm screening element 2 using biocompatible epoxy resin adhesive. Add 100-400 μL of 0.5 mg / ml hyaluronic acid to the sampling groove 24 (ensuring the hyaluronic acid level is below the top surface of the sampling groove 24). Then, embed the sperm screening element 2 entirely into the culture well 13 and attach it to the top of the side wall of the culture well 13 of the multi-well plate base 1 using a clip 22.

[0037] S300. Slowly add 2-4 mL of culture medium into the buffer ring trough 23 along the inner sidewall of the sperm selection component 2 main body 21. Due to the upstream characteristics of sperm, morphologically normal and highly motile sperm enter the culture medium of the buffer ring trough 23 through the channel 232 on the inner sidewall 231 of the buffer ring trough 23 via the porous polycarbonate membrane.

[0038] S400. Maintain the entire device in a 37°C water bath environment for 20 minutes. Remove the required sperm samples from the buffer ring groove 23 and the sampling groove 24 to complete the collection of highly motile sperm. Alternatively, depending on the actual operation, the water bath temperature can be selected as 36 or 38°C, and the water bath time can be adjusted to 15 minutes, 25 minutes, or 30 minutes.

[0039] The above description is only a specific embodiment of this utility model and does not limit the patent scope of this utility model. Any equivalent structural transformations made based on the contents of this utility model specification and drawings, or direct or indirect applications in other related technical fields, are similarly included within the patent protection scope of this utility model.

Claims

1. A multi-well plate sperm screening device based on the upstream method, characterized in that: The system includes a multi-well plate base (1) and a sperm screening component (2). The multi-well plate base (1) has a concave (12) center in the body (11) and two or more culture wells (13) spaced apart at the bottom of the concave (12). Each culture well (13) is equipped with a sperm screening component (2). The main body (21) of the sperm screening component (2) is embedded in the culture well (13) and is fitted onto the top of the culture well (13) via a sleeve (22) provided at the top of the sperm screening component (2). The bottom of the main body (21) of the sperm screening component (2) is provided with an inward extension relative to the main body (21) of the sperm screening component (2). A ring-shaped buffer groove (23) is formed by stretching. Multiple channels (232) are opened at intervals on the inner wall (231) of the buffer groove (23). The bottom of the main body (21) of the sperm screening component (2) forms a communication port on the inner side of the buffer groove (23). A filter membrane (25) is set at the communication port to seal the communication port and allow sperm to pass upstream. At least one recessed sampling groove (24) is set at the bottom of the buffer groove (23). The sampling groove (24) is used to hold hyaluronic acid that induces high-quality sperm to enter the sampling groove (24). The height of the added hyaluronic acid is lower than the top surface of the sampling groove (24).

2. The multi-well plate sperm screening device based on the upstream method according to claim 1, characterized in that: The body (11) of the porous plate base (1) is square or cylindrical. The top surface of the culture well (13) is lower than or flush with the top surface of the body (11) of the porous plate base (1). The body (21) of the sperm screening component (2) is cylindrical to match the shape of the culture well (13).

3. The multi-well plate sperm screening device based on the upstream method according to claim 1, characterized in that: The multi-well plate base (1) is a four-well culture dish.

4. The multi-well plate sperm screening device based on the upstream method according to claim 1, characterized in that: The sleeve (22) is integrally formed with the main body (21) of the sperm screening component (2) and extends outward relative to the main body (21) of the sperm screening component (2) to form a semi-circular part with an inverted L-shaped cross section. The space between the main body (21) of the sperm screening component (2) and the culture well (13) is available for the pipette tip to be inserted.

5. The multi-well plate sperm screening device based on the upstream method according to claim 1, characterized in that: The sleeve (22) is an annular or semi-annular piece with an inverted L-shaped cross section that is integrally formed with the main body (21) of the sperm screening component (2) and extends outward relative to the main body (21) of the sperm screening component (2). Alternatively, it is a plurality of pendants with an inverted L-shaped cross section that are spaced apart and connected to the top of the main body (21) of the sperm screening component (2) by integral forming or by adhesive bonding.

6. The multi-well plate sperm screening device based on the upstream method according to claim 1, characterized in that: The inner wall (231) of the buffer ring groove (23) has multiple channels (232) evenly opened, and the bottom surface of the channel (232) is higher than the bottom surface of the buffer ring groove (23).

7. The multi-well plate sperm screening device based on the upstream method according to claim 1, characterized in that: The filter membrane (25) is a porous polycarbonate membrane, which is bonded to the bottom of the main body (21) of the sperm screening component (2) by a biocompatible epoxy resin adhesive.

8. The multi-well plate sperm screening device based on the upstream method according to claim 1, characterized in that: The shape of the sampling groove (24) matches the shape of the pipette tip.

9. The multi-well plate sperm screening device based on the upstream method according to claim 1, characterized in that: The sperm screening component (2) is directly formed by 3D printing from biocompatible materials.

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