Rapid endotoxin detection device based on recombinant C factor method

Abstract

The utility model discloses a rapid endotoxin detection device based on a recombinant C factor method, which comprises a shell with an opening on the front side; the kit is pushed into or pulled out of the shell through the opening by the bracket; the kit is placed on the bracket, a reaction cavity is formed in the kit, and a recombinant C factor reagent is stored in the reaction cavity; and the fluorescence analyzer is fixedly arranged above the kit in the shell. According to the endotoxin rapid detection device based on the recombinant C-factor method, recombinant C-factor reagents are added into the reagent storage area of the kit, the consistency of the recombinant C-factor reagents is high, batch difference does not exist, the accuracy is high when endotoxin detection is carried out, and the endotoxin rapid detection device is not limited by the yield of tachypleus amebocyte lysate. In the detection process, the recombinant C factor reagent is not exposed in the air, so that the influence of endotoxin in the air on the detection result is reduced.

Description

Technical Field

[0001] This invention relates to the field of biological detection technology, and more specifically, to a rapid endotoxin detection device based on the recombinant C factor method. Background Technology

[0002] Endotoxins are lipopolysaccharides found in the cell walls of Gram-negative bacteria and can cause severe inflammatory responses and sepsis. Traditional methods for detecting endotoxins mainly rely on horseshoe crab reagent (LAL), which is made using the lysate of horseshoe crab blood amoeboids. For example, patent CN113092767B discloses a lyophilized LAL microsphere, its preparation method, and its applications.

[0003] Patent document CN210442378U discloses an endotoxin detection device for the horseshoe crab reagent method, including an ampoule holder, a constant temperature water bath device, a camera device, and a display device electrically connected to the camera device; the ampoule holder includes a rotating rod for placing ampoules, a clamping mechanism for clamping ampoules is provided on the rotating rod, and a driving device for driving the rotating rod to rotate is connected to the rotating rod.

[0004] Existing endotoxin detection devices use horseshoe crab reagents. Since these reagents are made from horseshoe crab blood, the differences between individual horseshoe crabs can lead to variations between different batches of reagents, affecting the accuracy of the test results. Furthermore, during endotoxin testing, the reagents need to be added manually and mixed thoroughly, increasing the risk of airborne endotoxins entering the reagents.

[0005] Therefore, it is necessary to propose a rapid endotoxin detection device based on the recombinant C factor method to solve the problems existing in the prior art. Utility Model Content

[0006] The utility model description section introduces a series of simplified concepts, which will be further explained in detail in the detailed description section. This utility model description section is not intended to limit the key features and essential technical features of the claimed technical solution, nor is it intended to determine the scope of protection of the claimed technical solution.

[0007] To address the above problems, this invention provides a rapid endotoxin detection device based on the recombinant factor C method, comprising:

[0008] The outer casing has an opening on the front;

[0009] A tray allows the reagent kit to be pushed into or pulled out of the housing through an opening;

[0010] The reagent kit is placed on a tray. The reagent kit has a reaction chamber, which stores the recombinant factor C reagent.

[0011] The fluorescence analyzer is fixedly mounted above the reagent kit inside the casing.

[0012] Preferably, a guide rail is fixedly installed on the bottom plate of the outer casing, with the guide rail facing the opening, and a guide groove is opened on the side wall of the bracket, with the guide groove slidingly connected to the guide rail.

[0013] Preferably, two guide rails are arranged symmetrically.

[0014] Preferably, a baffle is fixedly installed at the end of the guide rail away from the opening, and a connecting plate is slidably installed between the two guide rails. The side of the baffle facing the opening is fixedly connected to one end of the spring, and the other end of the spring is fixedly connected to the end of the connecting plate that extends into the guide rail.

[0015] A magnet is fixedly installed in the middle of the side wall of the connecting plate away from the baffle. A magnet is fixedly installed on the outer side of the rear wall of the bracket at a position corresponding to the magnet. The polarity of the magnet is opposite to that of the magnet.

[0016] Preferably, a reaction chamber is formed on the upper surface of the reagent kit, and multiple reaction chambers are arranged in a rectangular array.

[0017] Preferably, the upper surface of the kit has sample loading holes and reaction buffer storage chambers on both sides of the reaction chamber. The reaction buffer storage chamber is connected to the reaction chamber through the liquid inlet, and the sample loading holes are connected to the reaction chamber through capillary pores.

[0018] Preferably, a glass cover is sealed above the reaction chamber and the reaction buffer storage chamber, and the glass cover is made of quartz glass or fused silica glass.

[0019] Preferably, a separator groove is formed between the reaction chamber and the reaction buffer storage chamber on the upper surface of the reagent kit, the separator groove is connected to the liquid inlet, and a separation membrane is provided inside the separator groove.

[0020] Preferably, the separator is double-folded, and an adhesive layer is provided on the side of the separator near the edge of the reaction buffer storage chamber. The adhesive layer is bonded to the side wall of the separator located on the reaction buffer storage chamber side.

[0021] Preferably, a display screen is installed on the top surface of the casing, and the display screen is electrically connected to the fluorescence analyzer.

[0022] Compared with the prior art, the present invention has at least the following beneficial effects:

[0023] The endotoxin rapid detection device based on the recombinant C factor method described in this invention adds recombinant C factor reagent to the reagent storage area of ​​the kit. The recombinant C factor reagent has high consistency and no batch differences, resulting in high accuracy in endotoxin detection. It is not limited by the production volume of horseshoe crab reagent. During the detection process, the recombinant C factor reagent is not exposed to the air, reducing the impact of airborne endotoxins on the detection results.

[0024] The rapid endotoxin detection device based on the recombinant C factor method described in this invention will be partly apparent from the following description, and partly understood by those skilled in the art through research and practice of this invention. Attached Figure Description

[0025] The accompanying drawings are provided to further illustrate the present invention and form part of the specification. They are used together with the embodiments of the present invention to explain the present invention, but do not constitute a limitation thereof. In the drawings:

[0026] Figure 1 This is a schematic diagram of the structure of the rapid endotoxin detection device based on the recombinant C factor method disclosed in this utility model;

[0027] Figure 2 This is a schematic diagram of the outer shell and guide rail disclosed in this utility model;

[0028] Figure 3 This is a cross-sectional structural schematic diagram of the detection device disclosed in this utility model;

[0029] Figure 4 This is a schematic diagram of the structure of the spring and connecting plate arranged on the guide rail disclosed in this utility model;

[0030] Figure 5 This is a schematic diagram of the bracket disclosed in this utility model;

[0031] Figure 6 This is a schematic diagram of the disassembled structure of the reagent kit disclosed in this utility model;

[0032] Figure 7 This is a cross-sectional structural schematic diagram of the reagent kit disclosed in this utility model;

[0033] Figure 8 This is a schematic diagram of the structure of the separator membrane being adhered in the separator groove as disclosed in this utility model;

[0034] Figure 9 This is a schematic diagram of the structure of the oscillation component disclosed in this utility model;

[0035] Figure 10 This is a schematic diagram of the camshaft disclosed in this utility model. Detailed Implementation

[0036] The present invention will now be described in further detail with reference to the accompanying drawings and embodiments, so that those skilled in the art can implement it based on the description.

[0037] It should be understood that terms such as “having,” “comprising,” and “including” as used herein do not exclude the presence or addition of one or more other elements or combinations thereof.

[0038] like Figures 1-8 As shown, the rapid endotoxin detection device based on the recombinant factor C method includes:

[0039] 1. Outer shell with an opening 2 on the front;

[0040] The tray 3 allows the reagent kit 4 to be pushed into or pulled out of the outer shell 1 through the opening 2;

[0041] Reagent kit 4 is placed on tray 3. Reaction chamber 5 is set on reagent kit 4, and recombinant C factor reagent is stored in reaction chamber 5.

[0042] The fluorescence analyzer 6 is fixedly mounted above the reagent kit 2 inside the outer casing 1.

[0043] Furthermore, a guide rail 7 is fixedly installed on the bottom plate of the outer casing 1, the guide rail 7 is positioned opposite to the opening 2, and a guide groove 8 is opened on the side wall of the bracket 3, the guide groove 8 is slidably connected to the guide rail 7.

[0044] Furthermore, two guide rails 7 are symmetrically arranged.

[0045] Furthermore, a baffle 9 is fixedly installed at the end of the guide rail 7 away from the opening 2, and a connecting plate 10 is slidably installed between the two guide rails 7. The side of the baffle 9 facing the opening 2 is fixedly connected to one end of the spring 11, and the other end of the spring 11 is fixedly connected to the end of the connecting plate 10 that extends into the guide rail 7.

[0046] A magnet is fixedly installed in the middle of the side wall of the connecting plate 10 away from the baffle 9. A magnet is fixedly installed on the outer side of the rear wall of the bracket 3 at a position corresponding to the magnet. The polarity of the magnet is opposite to that of the magnet.

[0047] Furthermore, the upper surface of the reagent kit 4 has a reaction chamber 5, and multiple reaction chambers 5 are arranged in a rectangular array.

[0048] Furthermore, sample application holes 12 and reaction buffer storage chambers 13 are provided on both sides of the reaction chamber 5 on the upper surface of the reagent kit 4. The reaction buffer storage chamber 13 is connected to the reaction chamber 5 through the liquid inlet hole 14, and the sample application hole 12 is connected to the reaction chamber 5 through the capillary pore 15.

[0049] Furthermore, a glass cover 16 is sealed on the reaction chamber 5 and the reaction buffer storage chamber 13. The glass cover 16 is made of quartz glass or fused silica glass.

[0050] Furthermore, a separator 17 is formed between the reaction chamber 5 and the reaction buffer storage chamber 13 on the upper surface of the reagent kit 4. The separator 17 is connected to the liquid inlet 14, and a separation membrane 18 is provided inside the separator 17.

[0051] Furthermore, the isolation membrane 18 is double-folded, and an adhesive layer is provided on the side of the isolation membrane 18 near the edge on one side of the reaction buffer storage cavity 13. The adhesive layer is bonded to the side wall of the separator 17 on the side of the reaction buffer storage cavity 13.

[0052] Furthermore, a display screen 19 is provided on the top surface of the outer casing 1, and the display screen 19 is electrically connected to the fluorescence analyzer 6.

[0053] The working principle of the above technical solution:

[0054] Before performing endotoxin testing, first take out the reagent kit 4. According to the number of samples to be tested, remove the isolation membrane in the separator 17 on the reagent kit 4, so that the buffer in the buffer storage chamber 13 enters the reaction chamber 5 and mixes with the recombinant C factor reagent. Then inject the sample into the sample well 12. The sample enters the reaction chamber 5 through the capillary 15. Then place the reagent kit into the placement slot 20 on the upper end face of the holder 3, push the holder 3 into the shell 1, so that the magnet 2 on the rear wall of the holder 3 is attracted to the magnet 1 on the connecting plate 10. At this time, the reagent kit 4 is located directly below the fluorescence analyzer 6.

[0055] Two guide rails 7 are symmetrically arranged with the vertical plane containing the center line of the outer shell along its length as the symmetrical plane. Needle roller bearings can be installed on the guide rails 7 and connected to the guide grooves 8 through the needle roller bearings, which can reduce the friction when the bracket 3 is pushed in or pulled out, making the pushing in or pulling out of the bracket 3 smoother.

[0056] The sample reacts with the recombinant C factor reagent, causing the fluorescent substrate to fluoresce. The fluorescence analyzer 6 analyzes the fluorescence data and displays the concentration of endotoxin in the sample on the display screen 16, thereby quickly and accurately detecting the concentration of endotoxin in the sample.

[0057] Reagent kit 4 is manufactured in a sterile environment and packaged aseptically to prevent contamination of reagent kit 4 and to avoid affecting the accuracy of endotoxin detection.

[0058] The use of a fluorescence analyzer 6 for quantitative analysis of endotoxins using recombinant factor C reagent is existing technology. Patent document CN116643047A discloses a method for detecting bacterial endotoxins from flucytosine injection based on the recombinant factor C method. The method involves gently mixing a fluorescent substrate solution equilibrated to room temperature, a assay buffer, and an rFC enzyme solution in a 5:4:1 ratio to prepare the substrate reagent. 100 μl of the substrate reagent is added to each well of a preheated 96-well plate, and the plate is incubated at 37°C for 1 hour. The microplate reader is set to excitation wavelength of 380 nm and emission wavelength of 440 nm to detect endotoxins before incubation. 0h ) and incubation for 1 hour (RFU) 1hThe fluorescence value of the instrument can be determined. In practice, a suitable model of fluorescence analyzer can be selected from the market.

[0059] The recombinant C factor reagent includes recombinant C factor and a fluorescent substrate. Patent document CN116449019A discloses a method for detecting endotoxins in a novel coronavirus vaccine using the recombinant C factor method. The method involves diluting the novel coronavirus vaccine with Tris buffer, adding it to a microplate, then adding a reaction mixture, incubating at 37°C, and reading the values ​​at 0h and 1h using a microplate reader. The reaction mixture includes a buffer, a fluorescent substrate, and recombinant C factor.

[0060] After the test is completed, the bracket 3 can be pulled out from the outer shell 1 to separate magnet 1 and magnet 2. After the bracket 3 is pulled out from the outer shell, the next test can be carried out.

[0061] The glass cover 16 is made of quartz glass or fused silica glass, which does not affect the transmission of fluorescence.

[0062] The isolation membrane 18 is made of polyethylene material. An adhesive layer is provided on the side edge of the isolation membrane 18 on one side of the reaction buffer storage chamber 13. The bottom surface of the separator 17 is lower than the bottom of the inlet hole 14. The width of the separator 17 is greater than the diameter of the inlet hole 14, and the vertical center line of the separator 17 passes through the axis of the inlet hole 14, so that the adhesive layer can completely seal the inlet hole 14 when it is bonded to the side wall of the separator 17. The isolation membrane 18 is double-folded, with one folded end located at the bottom of the separator 17. An adhesive layer is provided on the outer surface of the layer closest to the reaction buffer storage chamber 13. The adhesive layer is inserted into the separator 17 using a thin tool and then pressed towards the reaction buffer storage chamber 13 to firmly bond the adhesive layer to the side wall of the separator 17. The length of the layer of the separator 18 near the reaction chamber 5 is greater than the length of the layer near the reaction buffer storage chamber 13. A pull ring can be provided at the top of the layer near the reaction chamber 5 to facilitate pulling out the separator 18. When pulling out the separator 18, since the lower part is folded, pulling upwards at the end near the reaction chamber 5 will tear the separator 18 off the side wall of the separator 17 from below.

[0063] Then, add buffer solution to reaction buffer storage chamber 13, and add recombinant factor C reagent to reaction chamber 5. Next, seal the upper parts of reaction chamber 5 and reaction buffer storage chamber 13 with glass cover 16; the seal can be made with silicone. Before detection, pull out the isolation membrane 18.

[0064] The reagent kit 4 can be made of a material with lower adhesion to silicone than polyethylene, such as polytetrafluoroethylene. When the separator 18 is peeled off, the silicone layer on the separator 18 is completely adhered to the separator 18 and is peeled off without leaving any residue in the reagent kit. Due to the high stability of silicone, even if a small amount of silicone adheres to the separator 17, it will not affect the detection.

[0065] The beneficial effects of the above technical solution are as follows:

[0066] The endotoxin rapid detection device based on the recombinant C factor method described in this invention adds recombinant C factor reagent to the reagent storage area of ​​the kit. The recombinant C factor reagent has high consistency and no batch differences, resulting in high accuracy in endotoxin detection. It is not limited by the production volume of horseshoe crab reagent. During the detection process, the recombinant C factor reagent is not exposed to the air, reducing the impact of airborne endotoxins on the detection results.

[0067] In one embodiment, such as Figure 4 , Figures 9-10 As shown, the rapid endotoxin detection device based on the recombinant C factor method also includes an oscillation assembly. The oscillation assembly includes an L-shaped bracket 21 fixedly mounted on the outer shell 1. A camshaft 22 is rotatably mounted between the bracket 21 and the bottom plate of the outer shell 1. A cam 23 on the camshaft 22 contacts the connecting plate 10. A first pulley 24 is spaced apart from the cam 23 on the outer circumference of the camshaft 22. A reduction motor 25 is fixedly mounted on the bottom plate of the outer shell 1. A second pulley 26 is fixedly mounted on the power output shaft of the reduction motor 25. The first pulley 24 and the second pulley 26 are connected in a transmission manner.

[0068] The working principle of the above technical solution:

[0069] To improve the reaction speed between the sample and the recombinant factor C reagent and increase detection efficiency, an oscillation assembly is installed on the bottom plate inside the outer shell 1. After the reagent kit is pushed into the outer shell, the reduction motor 25 is turned on. The reduction motor 25 drives the first pulley 24 to rotate through the second pulley 26, which in turn drives the camshaft 22 to rotate. The cam 23 squeezes the connecting plate 10 once every one rotation, causing the connecting plate 10 to slide away from the camshaft 22. When the cam 23 is not squeezing the connecting plate 10, the connecting plate 10 slides towards the camshaft 22 under the action of the spring 11. The camshaft 22 rotates continuously, and the connecting plate 10 reciprocates under the alternating action of the cam 23 and the spring 11, which in turn drives the reagent kit 4 to reciprocate through the bracket 3, so that the recombinant factor C reagent in the reagent kit and the sample are mixed more quickly, thereby improving detection efficiency.

[0070] Bearing holes are provided on the bottom plate of the housing 1 and the horizontal plate of the bracket 21. The two ends of the camshaft 23 are respectively interference-fitted with the inner ring of the bearing in the bearing hole. A flange can be set on the upper part of the horizontal plate of the bracket 21 to fix the bearing and prevent the camshaft 22 from moving up and down. The installation of the camshaft 22 is existing technology and will not be described in detail here.

[0071] In the description of this utility model, it should be understood that the terms "center", "longitudinal", "transverse", "length", "width", "thickness", "upper", "lower", "front", "rear", "left", "right", "vertical", "horizontal", "top", "bottom", "inner", "outer", "clockwise", "counterclockwise", "axial", "radial", "circumferential", etc., indicating the orientation or positional relationship are based on the orientation or positional relationship shown in the accompanying drawings, and are only for the convenience of describing this utility model and simplifying the description, and are not intended to indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation, and therefore should not be construed as a limitation of this utility model.

[0072] In this utility model, unless otherwise explicitly specified and limited, the terms "installation," "connection," "joining," and "fixing," etc., should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral part; they can refer to a mechanical connection, an electrical connection, or a connection that allows communication between them; they can refer to a direct connection or an indirect connection through an intermediate medium; they can refer to the internal communication of two components or the interaction between two components, unless otherwise explicitly limited. Those skilled in the art can understand the specific meaning of the above terms in this utility model according to the specific circumstances.

[0073] Although the embodiments of this utility model have been disclosed above, they are not limited to the applications listed in the specification and embodiments. They can be applied to various fields suitable for this utility model. For those skilled in the art, other modifications can be easily made. Therefore, without departing from the general concept defined by the claims and their equivalents, this utility model is not limited to the specific details and the illustrations shown and described herein.

Claims

1. A rapid endotoxin detection device based on the recombinant factor C method, characterized in that, include: The outer shell (1) has an opening (2) on the front; The tray (3) allows the reagent kit (4) to be pushed into or pulled out of the outer shell (1) through the opening (2); The reagent kit (4) is placed on the tray (3). A reaction chamber (5) is set on the reagent kit (4). The reaction chamber (5) stores the recombinant C factor reagent. The fluorescence analyzer (6) is fixedly installed above the reagent kit (4) inside the outer casing (1).

2. The rapid endotoxin detection device based on the recombinant C factor method according to claim 1, characterized in that, A guide rail (7) is fixedly installed on the bottom plate of the outer shell (1). The guide rail (7) is positioned opposite to the opening (2). A guide groove (8) is opened on the side wall of the bracket (3). The guide groove (8) is slidably connected to the guide rail (7).

3. The rapid endotoxin detection device based on the recombinant C factor method according to claim 2, characterized in that, Two guide rails (7) are symmetrically arranged.

4. The rapid endotoxin detection device based on the recombinant C factor method according to claim 3, characterized in that, A baffle (9) is fixedly installed at the end of the guide rail (7) away from the opening (2), and a connecting plate (10) is slidably installed between the two guide rails (7). The side of the baffle (9) facing the opening (2) is fixedly connected to one end of the spring (11), and the other end of the spring (11) is fixedly connected to the end of the connecting plate (10) that extends into the guide rail (7). A magnet is fixedly installed in the middle of the side wall of the connecting plate (10) away from the baffle (9), and a magnet is fixedly installed on the outer side of the rear wall of the bracket (3) at a position corresponding to the magnet. The polarity of the magnet is opposite to that of the magnet.

5. The rapid endotoxin detection device based on the recombinant C factor method according to claim 1, characterized in that, A reaction chamber (5) is opened on the upper end face of the reagent kit (4), and multiple reaction chambers (5) are arranged in a rectangular array.

6. The rapid endotoxin detection device based on the recombinant C factor method according to claim 5, characterized in that, On the upper surface of the reagent kit (4), sample application holes (12) and reaction buffer storage chambers (13) are opened on both sides of the reaction chamber (5). The reaction buffer storage chamber (13) is connected to the reaction chamber (5) through the liquid inlet hole (14), and the sample application hole (12) is connected to the reaction chamber (5) through the capillary pore (15).

7. The rapid endotoxin detection device based on the recombinant C factor method according to claim 6, characterized in that, The upper part of the reaction chamber (5) and the reaction buffer storage chamber (13) is sealed with a glass cover (16), which is made of quartz glass or fused silica glass.

8. The rapid endotoxin detection device based on the recombinant C factor method according to claim 7, characterized in that, A separator (17) is formed between the reaction chamber (5) and the reaction buffer storage chamber (13) on the upper surface of the reagent kit (4). The separator (17) extends to the upper surface of the glass cover plate (16). The separator (17) is connected to the liquid inlet (14). An isolation membrane (18) is set inside the separator (17).

9. The rapid endotoxin detection device based on the recombinant C factor method according to claim 8, characterized in that, The isolation membrane (18) is double-folded. An adhesive layer is provided on the side of the isolation membrane (18) near the edge of the reaction buffer storage cavity (13). The adhesive layer is bonded to the side wall of the separator (17) on the side of the reaction buffer storage cavity (13).

10. The rapid endotoxin detection device based on the recombinant C factor method according to claim 1, characterized in that, A display screen (19) is provided on the top surface of the outer casing (1), and the display screen (19) is electrically connected to the fluorescence analyzer (6).

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