Filter for agarose bead elution solid-liquid separation in centrifuge tube
By designing a filter for eluting solid-liquid separation of agarose beads in centrifuge tubes, the problem of incomplete separation of agarose beads and solution in traditional methods is solved, and more efficient solid-liquid separation is achieved and solution residue is reduced. and sample loss.
Patent Information
- Application Number
- CN202421220319.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Utility models(China)
- Current Assignee / Owner
- Filing Date
- 2024-05-31
- Publication Date
- 2025-02-11
- Estimated Expiration
- 2034-05-31
AI Technical Summary
The traditional solid-liquid separation method causes incomplete separation of agarose beads from the solution after centrifugation, resulting in an increase in solution residue, affecting the data quality of subsequent experiments, and requires multiple PBS cleaning, which is time-consuming and easy to cause sample loss.
A filter for separating solid-liquid separation of agarose beads in centrifuge tubes is designed, including rubber rods, rubber seals and nylon filters. After the agarose beads are layered from the solution by standing, the filter is centrifuged. Complete separation of agarose beads from the upper solution is achieved in the tube.
The complete separation of agarose beads and solution was achieved, reducing solution residues, reducing the content of detergent, reducing the number of PBS cleanings, improving experimental efficiency and avoiding sample losses.
Smart Images
Figure CN222445557U_ABST
Abstract
Description
Technical Field
[0001] The utility model belongs to the field of immunocoprecipitation experimental supplies, and more specifically relates to a filter for eluting solid-liquid separation of agarose beads in a centrifugal tube. Background Art
[0002] Co-Immunoprecipitation (CoIP) is a classic method for studying protein interactions based on the specific interaction between antibodies and antigens. It is an effective method to determine the physiological interaction between two proteins in intact cells. The principle is that when cells are lysed under non-denaturing conditions, many protein-protein interactions existing in intact cells are preserved. If protein X is immunoprecipitated with an antibody against protein X, then protein Y that binds to X in vivo can also be precipitated.
[0003] At present, most refined protein A is pre-bound and solidified on agarose beads, so that after reacting with a solution containing an antigen and an antibody, the protein A on the agarose beads can adsorb the antigen to achieve the purpose of purification. The CoIP standardized operation process involves multiple processes of combining and separating agarose beads with lysis buffer, washing buffer, and elution buffer, which is a process of solid-liquid separation. The traditional solid-liquid separation method is to place a 1.5 mL centrifuge tube containing agarose bead suspension (see Figure 1 , also called 1.5mL transparent conical bottom microcentrifuge tube) is placed in a centrifuge at 1000×g to centrifuge, and after the agarose beads and the solution are separated, the solution is directly sucked out with a pipette, leaving the agarose beads at the bottom of the centrifuge tube to achieve the purpose of solid-liquid separation. However, when the solution is sucked with a pipette, since the agarose beads are light, after centrifugation, there will still be some agarose beads floating in the solution near the top of the large agarose beads at the bottom. The suction force of the pipette can also easily make the agarose beads at the bottom float up into the solution, so that the pipette will suck the agarose beads at the bottom, and it is impossible to achieve a good separation effect between the agarose beads and the solution during suction.
[0004] If the agarose beads at the bottom are not sucked, it is necessary to stop sucking the liquid when the liquid surface of the solution is a certain distance away from the agarose beads at the bottom, which will result in a lot of solution remaining and a lot of solution residue. Most researchers doing CO-IP experiments will choose "proteomics IP-MS" as the next step to explore the interaction between proteins, but the solution used in the last step of the CO-IP experiment contains detergents, and the mass spectrometer used in the subsequent "proteomics IP-MS" is not compatible with detergents. If the detergent enters the instrument, it will affect the detection signal of the mass spectrometer, resulting in poor data quality of the sample and contamination of the instrument. Therefore, researchers will add PBS (phosphate buffered saline) to wash the sample (i.e., agarose beads) between the CO-IP experiment and the proteomics IP-MS, dilute or even remove the detergent to avoid the detergent from causing adverse effects on subsequent IP-MS research.
[0005] Therefore, if a large amount of solution remains in the sample (i.e., agarose beads) after solid-liquid separation, the content of detergent in the sample (i.e., agarose beads) increases. The traditional method of solid-liquid separation after centrifugation will result in more solution residues and a higher detergent content. The sample (i.e., agarose beads) needs to be washed with PBS three times or more, which is time-consuming. In addition, repeated washing will take away more samples (agarose beads), resulting in sample loss. Utility Model Content
[0006] In view of the above defects or improvement needs of the prior art, the utility model provides a filter for solid-liquid separation of agarose beads eluted in a centrifuge tube. After the agarose bead suspension is allowed to stand to separate the agarose beads and the solution, the filter can achieve a relatively thorough solid-liquid separation of the agarose beads and the upper solution in a 1.5 mL centrifuge tube, making it easy for a pipette to absorb the solution, and the pipette will not absorb the agarose beads.
[0007] To achieve the above object, according to the utility model, a filter for solid-liquid separation of agarose beads eluted in a centrifuge tube is provided, wherein the centrifuge tube is a 1.5 mL centrifuge tube, and the centrifuge tube comprises an upper cylindrical structure and a lower conical structure connected to the upper cylindrical structure, characterized in that the filter comprises a rubber rod, a rubber sealing ring and a nylon filter, wherein:
[0008] The center line of the rubber sealing ring is arranged vertically, and the rubber sealing ring is in the shape of a truncated cone with a larger top and a smaller bottom;
[0009] The rubber rod is vertically arranged, and the rubber rod is connected to the top surface of the rubber sealing ring and is integrally formed with the rubber sealing ring;
[0010] The nylon filter is installed on the rubber sealing ring and covers the inner hole of the rubber sealing ring;
[0011] The outer diameter of the top surface and the outer diameter of the bottom surface of the rubber sealing ring are D1 and D2 respectively, the distance between the top surface and the bottom surface of the rubber sealing ring is h, and So that the outer wall of the rubber sealing ring is in close contact with the inner wall of the lower conical structure of the centrifuge tube.
[0012] Preferably, D1≤0.8cm, D2≥0.6cm.
[0013] Preferably, D1 = 0.8 cm, D2 = 0.6 cm.
[0014] Preferably, the diameter of the inner hole of the rubber sealing ring is 0.4 cm to 0.5 cm.
[0015] Preferably, the bottom surface of the nylon filter is flush with the bottom surface of the rubber sealing ring.
[0016] Preferably, the pore size of the nylon filter is less than 50 μm.
[0017] Preferably, the diameter D of the filter holes of the nylon filter is in the range of 10 μm≤D<50 μm.
[0018] Preferably, the rubber rod is arranged at a position close to the outer edge of the top surface of the rubber sealing ring.
[0019] Preferably, the length of the rubber rod is 1.8 cm to 2 cm.
[0020] Preferably, the thickness of the nylon filter is 0.1 cm to 0.12 cm.
[0021] In general, the above technical solutions conceived by the utility model can achieve the following beneficial effects compared with the prior art:
[0022] 1) The filter for eluting agarose beads in a centrifuge tube for solid-liquid separation of the utility model is truncated cone-shaped as a whole. When in use, it is directly placed in a 1.5 mL centrifuge tube containing agarose beads and a liquid in which agarose beads and a solution are layered after the agarose bead suspension is allowed to stand. The rubber sealing ring can be completely fitted with the inner wall of the lower conical structure of the 1.5 mL centrifuge tube to prevent the agarose beads from passing through, and it is not easy to tip over, so it has a better separation effect and can also avoid the adverse effects of liquid residue on the experimental effect. The nylon filter makes the agarose beads below the nylon filter screen and the upper solution (supernatant) above the nylon filter screen, so that the solid-liquid separation (separation of agarose beads and the upper solution) operation can be achieved. In addition, the nylon filter screen is made of nylon material, which is a material with stable chemical properties and has no adverse effects on the chemical properties of the agarose beads.
[0023] 2) The filter used for solid-liquid separation of eluting agarose beads in a centrifuge tube of the utility model can leave little or no solution residue in the sample (i.e., agarose beads) after solid-liquid separation, and the sample only needs to be washed once with PBS (phosphate buffered saline); therefore, the filter reduces the solution residue on the agarose beads, which can not only avoid the adverse effects of high-content detergent on the data quality and instrument of subsequent "proteomics IP-MS", but also reduce the cleaning steps after the CO-IP experiment, and can avoid sample loss caused by multiple PBS washings. BRIEF DESCRIPTION OF THE DRAWINGS
[0024] Figure 1 It is a schematic diagram of an existing 1.5mL centrifuge tube;
[0025] Figure 2 It is a schematic diagram of the utility model;
[0026] Figure 3 It is a schematic diagram of the utility model placed in a 1.5mL centrifuge tube. DETAILED DESCRIPTION
[0027] In order to make the purpose, technical solution and advantages of the utility model more clear, the utility model is further described in detail below in conjunction with the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain the utility model and are not used to limit the utility model. In addition, the technical features involved in each embodiment of the utility model described below can be combined with each other as long as they do not conflict with each other.
[0028] See also Figure 2 , Figure 3 , a filter 100 for eluting agarose beads in a centrifuge tube 200 for solid-liquid separation, wherein the centrifuge tube 200 is a 1.5 mL centrifuge tube (a conventionally used 1.5 mL centrifuge tube with a conical bottom, also called a 1.5 mL transparent conical bottom microcentrifuge tube), wherein the centrifuge tube 200 comprises an upper cylindrical structure and a lower conical structure connected to the upper cylindrical structure, and the side wall of the 1.5 mL centrifuge tube generally has scale lines at the volumes of 0.1 mL, 0.5 mL, 1 mL and 1.5 mL, and the scale line at the junction of the upper cylindrical structure and the lower conical structure is 0.5 mL, and the inner diameter corresponding to the 0.5 mL scale line is about 0.8 cm, and the inner diameter corresponding to the 0.1 mL scale line is about 0.6 cm, and the filter 100 comprises a rubber rod 1, a rubber sealing ring 2 and a nylon filter 3, wherein:
[0029] The center line of the rubber sealing ring 2 is arranged vertically. The rubber sealing ring 2 is in the shape of a truncated cone with a larger upper part and a smaller lower part. It has an inner hole. The diameter of the inner hole of the rubber sealing ring 2 is 0.4 cm to 0.5 cm, preferably 0.5 cm. The diameter design of the inner hole can ensure that the rubber sealing ring 2 has sufficient strength to maintain its shape, and can also ensure the filtering effect of the nylon filter 3 and facilitate the pipette to absorb the upper solution.
[0030] The rubber rod 1 is arranged vertically, connected to the top surface of the rubber sealing ring 2 and integrally formed with the rubber sealing ring 2; the rubber rod 1 is arranged at a position close to the outer edge of the top surface of the rubber sealing ring 2, the length of the rubber rod 1 is 1.8 cm to 2 cm, preferably 2 cm, for easy taking and placing, and the rubber rod 1 is located as a whole inside the 1.5 mL centrifuge tube.
[0031] The nylon filter 3 is installed on the rubber sealing ring 2 and covers the inner hole of the rubber sealing ring. The thickness of the nylon filter 3 is 0.1 cm to 0.12 cm, preferably 0.1 cm. The thickness design of the nylon filter 3 can ensure the overall strength of the nylon filter 3 and allow as much solution as possible to be sucked away by the pipette.
[0032] The outer diameter of the top surface and the outer diameter of the bottom surface of the rubber sealing ring 2 are D1 and D2 respectively, the distance between the top surface and the bottom surface of the rubber sealing ring 2 is h, and That is, the cone angle of the truncated cone-shaped rubber sealing ring 2 is 20°, so that the outer wall of the rubber sealing ring 2 is in close contact with the inner wall of the lower cone structure of the centrifuge tube 200 (the cone angle of the lower cone structure of the centrifuge tube 200 is also 20°).
[0033] The filter 100 used in traditional production and food manufacturing industries is mostly made of stainless steel or plastic, etc. These materials are characterized by high hardness and difficulty in deformation. The cylindrical filter 100 placed in the centrifuge tube 200 will inevitably have a gap with the inner wall of the centrifuge tube 200 with a slope, and it is not easy to fix, and there is a risk of tipping over, which will result in poor filtering effect. The rubber seal ring 2 of the filter 100 is made of rubber material and is designed to be truncated. When placed in the centrifuge tube 200, it can better fit the inner wall of the lower conical structure of the centrifuge tube 200. It can be well stuck on the inner wall of the centrifuge tube 200 with a little force to prevent agarose beads from passing through, and it is not easy to tip over, which has a better separation effect and can also avoid the adverse effects of liquid residue between each step on the experimental effect. The rubber rod 1 and the rubber seal ring 2 are two parts connected together, both of which are made of rubber material, which is convenient for the manufacture and one-piece molding of the two parts.
[0034] Furthermore, D1≤0.8cm, D2≥0.6cm, and the rubber sealing ring 2 is entirely located in the lower conical structure of the centrifuge tube 200 , which facilitates the pipette to absorb the supernatant in the upper cylindrical structure of the centrifuge tube 200 . As a further preference, D1=0.8cm, D2=0.6cm. At present, the amount of agarose beads used in immunoprecipitation experiments is generally about 0.1mL. Therefore, the outer diameter of the bottom surface of the rubber sealing ring 2 (0.6cm) is designed to be the same as the inner diameter of the 0.1mL scale of the 1.5mL centrifuge tube (0.6cm). The cone angle of the cross section of the filter 100 and the cone angle of the lower conical structure of the centrifuge tube 200 are both 20°. In this way, when the rubber sealing ring 2 of the filter 100 is placed in the 1.5mL centrifuge tube after the agarose beads and the solution are layered, the rubber sealing ring 2 of the filter 100 can fit the inner wall of the lower conical structure of the 1.5mL centrifuge tube without a gap, the agarose beads are below the nylon filter 3 of the filter 100, and the supernatant is above the nylon filter 3 of the filter 100, so as to achieve solid-liquid separation. Preferably, the bottom surface of the nylon filter 3 is flush with the bottom surface of the rubber sealing ring 2, so that the area below the filter 3 is entirely filled with agarose beads without any solution, and the separation is more thorough.
[0035] Furthermore, the pore size of the filter holes of the nylon filter 3 is less than 50μm. Compared with the large-sized filters 100 used in the fields of traditional production and food manufacturing, the filter 100 of the utility model is more delicate and compact. Since the diameter of agarose beads is generally 50μm-150μm, the pore size of the filter hole must also be less than 50μm to separate the agarose beads. As a further preference, the range of the diameter D of the filter holes of the nylon filter 3 is: 10μm≤D<50μm, and more preferably D=40μm. When the pore size of the nylon filter 3 is too small (less than 10μm), the speed of aspirating the liquid on the nylon filter 3 with a pipette will be too slow. Therefore, the filter 100 can use a nylon filter 3 with a pore diameter D of 10μm≤D<50μm, which is smaller than the diameter of the agarose beads and has a better separation effect.
[0036] Compared with the large-sized filters 100 used in the traditional production and food manufacturing industries, the filter 100 of the utility model is more exquisite and compact. Since the inner diameter of a 1.5 mL centrifuge tube is relatively small, if a cylindrical filter is used, it is necessary to clamp the filter with tweezers to put it into the 1.5 mL centrifuge tube, and the filter is in line contact with the lower conical structure of the 1.5 mL centrifuge tube, which is easy to tip over and affect the sealing and filtering effect, and the position of the filter needs to be adjusted repeatedly. When the filter 100 of the utility model is used, the rubber rod 1 of the filter 100 is clamped by hand or tweezers, and then released to let the filter 100 fall into a 1.5mL centrifuge tube (the 1.5mL centrifuge tube contains a liquid with agarose beads having a volume of 0.5mL to 1mL after standing (standing for about 1 minute), and the volume of the agarose beads is about 0.1mL. The filter is directly used in the liquid after standing, and there is no need to centrifuge the agarose bead suspension). The rubber sealing ring 2 can contact the inner wall of the lower conical structure of the 1.5mL centrifuge tube under its own weight, and the nylon filter 3 can float the agarose beads above the large piece of agarose beads at the bottom in the solution. The sugar beads are pressed down to gather together with the large agarose beads at the bottom, and then the rubber seal ring 2 is slightly pressed downward with tweezers or other objects to make the rubber seal ring 2 close to the lower conical structure of the 1.5mL centrifuge tube, without the need to use tweezers to operate repeatedly, and the rubber seal ring 2 and the inner wall of the lower conical structure of the 1.5mL centrifuge tube are in surface contact, not easy to tip over, and after being close, the rubber seal ring 2 plays a good sealing role, the agarose beads are under the nylon filter 3 and will not float into the supernatant, and the supernatant is above the nylon filter 3. The supernatant can be sucked by a pipette close to the nylon filter 3 to achieve solid-liquid separation operation, and the pipette will not suck away the agarose beads. Since there is less solution in the agarose beads, it is only necessary to use PBS (phosphate buffered saline) to wash the sample once.
[0037] It is easy for those skilled in the art to understand that the above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the protection scope of the present invention.
Claims
1. A filter for solid-liquid separation of agarose beads eluted in a centrifuge tube, wherein the centrifuge tube is a 1.5 mL centrifuge tube, and the centrifuge tube comprises an upper cylindrical structure and a lower conical structure connected to the upper cylindrical structure, characterized in that: The filter comprises a rubber rod, a rubber sealing ring and a nylon filter screen, wherein: The center line of the rubber sealing ring is arranged vertically, and the rubber sealing ring is in the shape of a truncated cone with a larger top and a smaller bottom; The rubber rod is vertically arranged, and the rubber rod is connected to the top surface of the rubber sealing ring and is integrally formed with the rubber sealing ring; The nylon filter is installed on the rubber sealing ring and covers the inner hole of the rubber sealing ring; The outer diameter of the top surface and the outer diameter of the bottom surface of the rubber sealing ring are D1 and D2 respectively, the distance between the top surface and the bottom surface of the rubber sealing ring is h, and So that the outer wall of the rubber sealing ring is in close contact with the inner wall of the lower conical structure of the centrifuge tube.
2. The filter for solid-liquid separation of agarose beads eluted in a centrifuge tube according to claim 1, characterized in that: D1≤0.8cm, D2≥0.6cm.
3. The filter for solid-liquid separation of agarose beads eluted in a centrifuge tube according to claim 2, characterized in that: D1=0.8cm, D2=0.6cm.
4. The filter for solid-liquid separation of agarose beads eluted in a centrifuge tube according to claim 3, characterized in that: The diameter of the inner hole of the rubber sealing ring is 0.4 cm to 0.5 cm.
5. The filter for solid-liquid separation of agarose beads eluted in a centrifuge tube according to claim 1, characterized in that: The bottom surface of the nylon filter is flush with the bottom surface of the rubber sealing ring.
6. The filter for solid-liquid separation of agarose beads eluted in a centrifuge tube according to claim 1, characterized in that: The pore size of the nylon filter is less than 50 μm.
7. The filter for solid-liquid separation of agarose beads eluted in a centrifuge tube according to claim 1, characterized in that: The range of the diameter D of the filter holes of the nylon filter is: 10 μm≤D<50 μm.
8. The filter for solid-liquid separation of agarose beads eluted in a centrifuge tube according to claim 1, characterized in that: The rubber rod is arranged at a position close to the outer edge of the top surface of the rubber sealing ring.
9. The filter for solid-liquid separation of agarose beads eluted in a centrifuge tube according to claim 1, characterized in that: The length of the rubber rod is 1.8 cm to 2 cm.
10. The filter for solid-liquid separation of agarose beads eluted in a centrifuge tube according to claim 1, characterized in that: The thickness of the nylon filter is 0.1 cm to 0.12 cm.