A biomolecule affinity gravity column device with positive pressure drive and sterile protection

Abstract

This utility model discloses a biomolecular affinity gravity column device with positive pressure drive and aseptic protection, belonging to the technical field of bioseparation and purification equipment. It includes a column purification body comprising a gravity column, a gravity column support, and a waste liquid tank. The column purification body is housed within a biosafety cabinet. A small hole is provided on one side of the biosafety cabinet for a connecting pipe to pass through. A silicone sealing ring is provided at the edge of the small hole to achieve a sealed connection. One end of the connecting pipe is connected to the column purification body, and the other end passes through a pressurization component and connects to a sterile air supply component. A pressure gauge is provided at the end of the connecting pipe near the column purification body, and a pressure gauge is provided at the end near the sterile air supply component. This utility model, employing the above-described structure, provides a biomolecular affinity gravity column device with positive pressure drive and aseptic protection, achieving efficient, rapid, and high-purity affinity purification of biomolecules.

Description

Technical Field

[0001] This utility model relates to the field of biological separation and purification equipment technology, and in particular to a biomolecular affinity gravity column device with positive pressure drive and sterile protection. Background Technology

[0002] Affinity column chromatography is a technique that utilizes specific and reversible affinity interactions between biomolecules (such as antigen-antibody, receptor-ligand, enzyme-substrate, etc.) to achieve efficient separation and purification of target molecules. Its core principle is the selective binding of specific ligands immobilized on the column packing material to target molecules (such as proteins and nucleic acids), followed by altering the elution conditions to achieve the separation and recovery of the target molecules.

[0003] Existing gravity column apparatus mainly consists of a gravity column shell, affinity packing material, and support and collection components. The gravity column shell is a cylindrical body with an open top and a narrow bottom, serving as the primary site for the purification process. The affinity packing material, composed of immobilized ligands and an inert carrier, is packed within the gravity column. The immobilized ligands are uniformly loaded onto the surface of the inert carrier, providing specific binding sites for biomolecules. The support and collection components include a simple support for fixing the column and a waste collection container.

[0004] Its core working principle is based on gravity-driven liquid flow. In practice, a sample solution (or purification reagent) containing the target biomolecule (such as mRNA or protein) is poured into the column through the opening at the top. Under the influence of gravity, the liquid naturally permeates through the packing layer. The target molecule specifically binds to the immobilized ligands (such as antibodies, metal ions, and nucleic acid aptamers) on the surface of the packing, while non-target impurities flow out with the mobile phase. Subsequently, by changing different elution reagents, the target molecule dissociates from the packing and flows out with the eluent from the narrow opening at the bottom, thus completing the purification process.

[0005] Traditional gravity columns rely solely on gravity for fluid propagation, with flow rates typically ranging from 0.5 to 2 mL / min, influenced by packing particle size and liquid viscosity. A single column pass can take 30-60 minutes, and with multiple washes or gradient elutions, the total time can reach several hours. Furthermore, biomolecules are prone to degradation and denaturation due to prolonged environmental exposure, severely impacting the recovery rate and activity of the target product.

[0006] The samples used for purification often contain tiny solid particles, lipids, or salt crystals. These impurities tend to deposit in the gaps between the packing material at the top of the column or in the column head region, causing physical blockage. This is because low flow rates make it easier for impurities to accumulate locally. After blockage, backwashing is required, but frequent flushing is cumbersome and shortens the life of the packing material, leading to high costs.

[0007] Traditional gravity column experiments are often performed in open lab benches, where airborne microorganisms or particulate matter can easily be introduced during sample addition and liquid changes. Biomolecules are susceptible to contamination by bacterial metabolic products, leading to subsequent experimental failures.

[0008] Therefore, there is an urgent need for an improved gravity column device that can overcome the above problems and improve the efficiency, stability and safety of affinity chromatography by introducing a controllable positive pressure system, optimizing the structural design, improving the aseptic operating environment and anti-clogging performance. Utility Model Content

[0009] The purpose of this invention is to provide a biomolecular affinity gravity column device with positive pressure drive and sterile protection, so as to solve the problems of low liquid flow rate and easy blockage caused by traditional gravity columns relying solely on gravity drive. At the same time, the experimental body is in a sterile environment, avoiding the problems of bacterial contamination caused by the open experimental environment of traditional gravity columns.

[0010] To achieve the above objectives, this utility model provides a biomolecular affinity gravity column device with positive pressure drive and sterile protection, comprising a column purification body, the column purification body including a gravity column, a gravity column support for placing the gravity column, and a waste liquid tank disposed below the gravity column and detachably connected to the gravity column support. The column purification body is disposed inside a biosafety cabinet, and a small hole is provided on one side of the biosafety cabinet for a first connecting pipe to pass through. A silicone sealing ring is provided at the edge of the small hole to achieve a sealed connection. One end of the connecting pipe is connected to the column purification body, and the other end passes through a pressurization component and is connected to a sterile air supply component. A pressure gauge is provided at the end of the connecting pipe near the column purification body, and a pressure gauge is provided at the end near the sterile air supply component.

[0011] Preferably, the gravity column includes a gravity column body, which is cylindrical and has an upper cap connected to the top. The upper cap is fixed to the gravity column body by a closing clip. Inside the gravity column body, an upper screen plate, a packing material, and a lower screen plate are arranged sequentially from top to bottom. The lower end of the lower screen plate is detachably connected to a lower cap.

[0012] Preferably, the gravity column support includes two support plates and a column plate disposed between the two support plates. The support plates are provided with vertical stepped grooves, and the side of the column plate is provided with stepped buckles that are embedded in the grooves. The column plate is provided with column holes.

[0013] Preferably, the waste liquid tank is detachably connected to the bottom end of the support plate.

[0014] Preferably, the biosafety cabinet includes a cabinet body, a power switch is provided on the upper front side of the cabinet body, an operating cavity is provided inside the cabinet body, an ultraviolet lamp is provided on the inner wall of the top of the operating cavity, a socket is provided on the side wall of the operating cavity, a transparent observation window is provided at the front opening of the operating cavity, and support legs are provided at the bottom of the cabinet body.

[0015] Preferably, the column purification body is disposed within the operating chamber.

[0016] Preferably, the pressurizing component is a peristaltic pump equipped with an adjustment knob and a speed display screen.

[0017] Preferably, the sterile air supply component is a sterile air generator containing an air filter.

[0018] Therefore, the biomolecular affinity gravity column device with positive pressure drive and sterile protection using the above structure of this utility model has the following beneficial effects: (1) By using a peristaltic pump to provide positive pressure instead of traditional gravity, the clogging of the purification column is effectively prevented and the liquid flow rate is significantly improved. This not only greatly shortens the time required for the purification process, but also effectively avoids the sample degradation problem caused by the excessively long process. At the same time, the device achieves efficient and precise control of pressure by adjusting the speed of the peristaltic pump and cooperating with the pressure monitoring and adjustment device. (2) The integrated sterile air generator and biosafety cabinet provide a reliable sterile environment for the entire purification process; this design effectively isolates external contamination, thereby significantly improving the final purification purity of the target molecule.

[0019] The technical solutions in the embodiments of this utility model will now be clearly and completely described with reference to the accompanying drawings. Attached Figure Description

[0020] Figure 1 This is a schematic diagram of the overall structure of a biomolecular affinity gravity column device with positive pressure drive and sterile protection according to the present invention. Figure 2 This is a schematic diagram of the gravity column structure of a biomolecular affinity gravity column device with positive pressure drive and sterile protection according to this utility model; Figure 3 This is a schematic diagram of the structure of a gravity column support for a biomolecular affinity gravity column device with positive pressure drive and sterile protection according to this utility model. Figure reference numerals: 1. Column purification body; 11. Gravity column; 111. Gravity column body; 112. Packing material; 113. Upper cap; 114. Lower cap; 115. Sealing clip; 116. Upper sieve plate; 117. Lower sieve plate; 12. Gravity column support; 121. Support plate; 122. Column plate; 13. Waste liquid tank; 2. Biosafety cabinet; 21. Socket; 22. Ultraviolet lamp; 23. Power switch; 24. Cabinet body; 25. Operating chamber; 26. Transparent observation window; 27. Support leg; 3. Connecting pipeline; 31. Pressure gauge; 32. Pressure gauge; 4. Pressurization assembly; 5. Sterile air supply assembly. Detailed Implementation

[0021] The technical solution of this utility model will be further described below with reference to the accompanying drawings and embodiments.

[0022] Unless otherwise defined, the technical or scientific terms used in this utility model shall have the ordinary meaning understood by one of ordinary skill in the art to which this utility model pertains. The terms "first," "second," and similar terms used in this utility model do not indicate any order, quantity, or importance, but are merely used to distinguish different components. Terms such as "comprising" or "including" mean that the element or object preceding the word encompasses the elements or objects listed following the word and their equivalents, without excluding other elements or objects. Terms such as "connected" or "linked" are not limited to physical or mechanical connections, but can include electrical connections, whether direct or indirect. Terms such as "upper," "lower," "left," and "right" are used only to indicate relative positional relationships; when the absolute position of the described object changes, the relative positional relationship may also change accordingly.

[0023] Example like Figures 1-3As shown, a biomolecular affinity gravity column device with positive pressure drive and sterile protection includes a column purification body 1. The column purification body 1 includes a gravity column 11 filled with affinity packing material 112, a gravity column support 12 for placing the gravity column 11, and a waste liquid tank 13 disposed below the gravity column 11 and detachably connected to the gravity column support 12. The gravity column 11 includes a gravity column body 111, which is cylindrical and made of polypropylene (PP). The upper end cap 113 is connected to the gravity column body 111. A gas inlet is located in the center of the upper end cap 113. A rubber gasket made of silicone rubber is sandwiched between the upper end cap 113 and the opening of the gravity column body 111 to ensure airtightness. The upper end cap 113 is fixed to the gravity column body 111 by a sealing clip 115. Inside the gravity column body 111, from top to bottom, are arranged an upper sieve plate 116, a packing material 112, and a lower sieve plate 117. The holes in the upper sieve plate 116 and the lower sieve plate 117... The diameter is ≤ 1 / 5 of the particle size of filler 112, and its function is to prevent the filler 112 from leaking. The type of filler 112 can be selected according to the target molecule (such as Ni-NTA resin, Oligo(dT) cellulose). The lower end of the lower sieve plate 117 is detachably connected to the lower end cap 114, which can be removed during the experiment to allow liquid to flow out. The gravity column support 12 includes two support plates 121 and a column plate 122 set between the two support plates 121. The support plates 121 are provided with vertical stepped grooves, and the side of the column plate 122 is provided with stepped buckles. The buckles can be inserted into the grooves to adjust the height of the column plate 122. The column plate 122 is provided with 12 column holes for the gravity column 11 to be inserted. A maximum of 12 gravity columns 11 can be placed in one column plate 122. The waste liquid tank 13 is detachably connected to the bottom end of the support plate 121. The waste liquid tank 13 has a capacity of 1000mL and is used to collect waste liquid, such as unbound samples and washing liquid.

[0024] The column purification unit 1 is installed inside the biosafety cabinet 2, which has a cleanliness level of Class II A2, providing a sterile operating environment. A small hole is provided on one side of the biosafety cabinet 2 for the first connecting pipe 3 to pass through. A silicone sealing ring is provided at the edge of the small hole to achieve a sealed connection, ensuring that outside air does not contaminate the interior of the biosafety cabinet 2. The biosafety cabinet 2 includes a cabinet body 24. A power switch 23 is located on the upper front side of the cabinet body 24. An operating chamber 25 is located inside the cabinet body 24. An ultraviolet lamp 22 is installed on the inner top wall of the operating chamber 25 for regular ultraviolet disinfection before and after experiments to ensure that the biosafety cabinet 2 remains sterile. A socket 21 is located on the side wall of the operating chamber 25. A transparent observation window 26 is located at the front opening of the operating chamber 25. Support legs 27 are located at the bottom of the cabinet body 24. The column purification unit 1 is installed inside the operating chamber 25.

[0025] The connecting pipe 3 is made of medical-grade silicone tubing, which is resistant to chemical corrosion and has high cleanliness. It serves as the passage for pressurized gas transmission. One end of the connecting pipe 3 is connected to the sterile air supply component 5, passes through the pressurization component 4, and the other end is connected to the gas inlet of the cap 113 on the upper end of the gravity column 111. A pressure gauge 31 is installed on the end of the connecting pipe 3 near the gravity column 11 to monitor the pressure in front of the column in real time and prevent overpressure damage to the column. A pressure reducer 32 is installed on the end of the connecting pipe 3 near the sterile air supply component 5. The pressure reducer 32 integrates an automatic pressure reduction mechanism, which can monitor the pressure in front of the pump in real time and maintain the pressure within a safe range by changing the cross-sectional area of ​​the flow channel or releasing excess medium when the pressure exceeds a preset threshold.

[0026] The pressurization component 4 is a peristaltic pump with adjustment function and display screen. It generates positive pressure by mechanically squeezing air to drive the liquid to flow through the gravity column 11. The peristaltic pump is equipped with an adjustment knob and a speed display screen to display the pump speed in real time, ranging from 10 to 300 rpm. The output pressure can be precisely controlled by adjusting the speed.

[0027] The sterile air supply component 5 is a sterile air generator containing an air filter. The compressed air is filtered through a 0.22μm sterilization filter to ensure that the gas entering the connecting pipe 3 is clean.

[0028] Working Principle: The entire purification process is conducted within the aseptic operating environment provided by the biosafety cabinet 2. At the start of the experiment, the operator, inside the operating chamber 25 of the biosafety cabinet 2, installs the gravity column 11, filled with specific affinity packing material 112, onto the gravity column support 12, and seals it with the upper cap 113 and its silicone rubber gasket to ensure airtightness. Sterile compressed air generated by a sterile air generator is delivered via connecting pipe 3, and a peristaltic pump provides precise and controllable positive pressure through compression. This positive pressure replaces traditional gravity, serving as the core driving force for the liquid (such as sample, washing solution, eluent) to flow through the packing material 112 within the gravity column 11. Driven by pressure, the liquid flows from top to bottom, and the target molecules specifically bind to the ligands on the packing material 112, while impurities flow out with the waste liquid to the waste liquid tank 13 below. By adjusting the speed of the peristaltic pump, the flow rate and pressure can be precisely controlled. The pressure gauge 31 monitors the pressure at the column inlet in real time, while the pressure reducer 32 ensures that the system pressure is always within a safe range, jointly preventing overpressure damage to the gravity column 111. When it is necessary to collect the purified target molecules, the waste liquid tank 13 is removed and replaced with a collection tube. The target molecules flow out with the eluent, thus completing an efficient and rapid affinity purification process.

[0029] Therefore, this utility model provides a biomolecular affinity gravity column device with positive pressure drive and aseptic protection, which effectively solves the problems of easy clogging, slow flow rate, long time consumption and easy contamination of traditional gravity columns by combining controllable positive pressure drive with comprehensive aseptic protection, and achieves efficient, rapid and high-purity affinity purification of biomolecules.

[0030] Finally, it should be noted that the above embodiments are only preferred embodiments of the present utility model, but the protection scope of the present utility model is not limited thereto. Any equivalent substitutions or changes made by those skilled in the art within the technical scope disclosed in the present utility model, based on the technical solution and the inventive concept of the present utility model, should be covered within the protection scope of the present utility model.

Claims

1. A biomolecular affinity gravity column device with positive pressure drive and sterile protection, characterized in that: The device includes a column purification unit, comprising a gravity column, a gravity column support for placing the gravity column, and a waste liquid tank disposed below the gravity column and detachably connected to the gravity column support. The column purification unit is housed inside a biosafety cabinet. A small hole is provided on one side of the biosafety cabinet for a connecting pipe to pass through. A silicone sealing ring is provided at the edge of the small hole to achieve a sealed connection. One end of the connecting pipe is connected to the column purification unit, and the other end passes through a pressurization component and is connected to a sterile air supply component. A pressure gauge is provided at the end of the connecting pipe near the column purification unit, and a pressure gauge is provided at the end near the sterile air supply component.

2. The biomolecular affinity gravity column device with positive pressure drive and sterile protection according to claim 1, characterized in that: The gravity column includes a gravity column body, which is cylindrical and has an upper cap connected to the top. The upper cap is fixed to the gravity column body by a closing clip. Inside the gravity column body, from top to bottom, there are an upper screen plate, a packing material, and a lower screen plate. The lower end of the lower screen plate is detachably connected to a lower cap.

3. The biomolecular affinity gravity column device with positive pressure drive and sterile protection according to claim 1, characterized in that: The gravity column support includes two support plates and a column plate disposed between the two support plates. The support plates are provided with vertical stepped grooves, and the side of the column plate is provided with stepped buckles that are embedded in the grooves. The column plate is provided with column holes.

4. The biomolecular affinity gravity column device with positive pressure drive and sterile protection according to claim 3, characterized in that: The waste liquid tank is detachably connected to the bottom end of the support plate.

5. The biomolecular affinity gravity column device with positive pressure drive and sterile protection according to claim 1, characterized in that: The biosafety cabinet includes a cabinet body, a power switch located on the upper front side of the cabinet body, an operating chamber located inside the cabinet body, an ultraviolet lamp located on the inner top wall of the operating chamber, a socket located on the side wall of the operating chamber, a transparent observation window located at the front opening of the operating chamber, and support legs located at the bottom of the cabinet body.

6. A biomolecular affinity gravity column device with positive pressure drive and sterile protection according to claim 5, characterized in that: The column purification body is disposed within the operating chamber.

7. The biomolecular affinity gravity column device with positive pressure drive and sterile protection according to claim 1, characterized in that: The pressurization component is a peristaltic pump equipped with an adjustment knob and a speed display screen.

8. A biomolecular affinity gravity column device with positive pressure drive and sterile protection according to claim 1, characterized in that: The sterile air supply component is a sterile air generator containing an air filter.

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