A device for real-time measurement of cellular metabolism

Abstract

This invention relates to a real-time cell metabolism measurement device for measuring metabolic cells in a cell culture flask. It includes a pump mechanism and a first chamber and a second chamber sequentially connected to the outlet of the pump mechanism. The inlet of the pump mechanism is used to connect to the cell culture flask, and the second chamber has an outlet for connecting to the cell culture flask. It also includes necessary connecting pipes to form a circulation path between the pump mechanism, the first chamber, the second chamber, and the cell culture flask. The first chamber is equipped with probes for at least three of the following sensors: a pH sensor, a dissolved oxygen sensor, a dissolved carbon dioxide sensor, and an ion sensor. The second chamber is equipped with multiple electrochemical biosensors. This enables real-time metabolic detection.

Description

Technical Field

[0001] This invention belongs to the field of biomedical engineering and relates to a real-time cell metabolism measurement device that can measure cell metabolism in real time and improve temporal resolution. Background Technology

[0002] Metabolomics is a crucial tool in metabolic research, providing a global quantitative assessment of metabolites within biological systems. Cell culture metabolomics is an important branch of metabolomics research, reflecting cellular metabolic characteristics by studying changes in metabolites in the culture medium during cell culture. Cell metabolomics research can be applied to: optimizing cell lines and bioreactor culture conditions; cell classification based on metabolic phenotypes; simulating drug efficacy, target and toxicity analysis; identifying and modeling metabolic networks; and discovering biomarkers and drug targets.

[0003] Currently, the main research methods in metabolomics include mass spectrometry, chromatography, and nuclear magnetic resonance (NMR) (such as Chinese patent CN113874711A). While these methods can analyze a large number of metabolites simultaneously and have advantages in quantitative resolution, many times it is necessary to achieve real-time, non-destructive detection of metabolites, improve temporal resolution, and discover more details of metabolic process changes, which is something that current mainstream research methods cannot achieve. Summary of the Invention

[0004] The purpose of this invention is to provide a real-time cell metabolism measurement device that solves the problem of real-time detection of cell metabolites.

[0005] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0006] This invention provides a real-time cell metabolism measurement device for measuring metabolic cells in a cell culture flask. It includes a pump mechanism and a first chamber and a second chamber connected sequentially to the outlet of the pump mechanism. The inlet of the pump mechanism is used to connect to the cell culture flask, and the second chamber has an outlet for connecting to the cell culture flask. It also includes necessary connecting pipes so that the pump mechanism, the first chamber, the second chamber, and the cell culture flask form a circulation path. The first chamber is equipped with probes of at least three of the following sensors: a pH sensor, a dissolved oxygen sensor, a dissolved carbon dioxide sensor, and an ion sensor. The second chamber is equipped with multiple electrochemical biosensors.

[0007] Preferably, the first chamber includes a flow channel seat and a cover plate detachably connected to the flow channel seat. A flow channel is provided inside the flow channel seat. The pH sensor, dissolved oxygen sensor, dissolved carbon dioxide sensor, or ion sensor is combined with the cover plate and the probe extends into the flow channel.

[0008] Furthermore, the first chamber has placement slots corresponding to the sensors, and the flow channels between adjacent placement slots are elongated.

[0009] Furthermore, the flow channel in the flow channel seat has an inlet channel and an outlet channel at both ends, and the height of the inlet channel and the outlet channel is lower than the top edge of the flow channel seat.

[0010] Furthermore, a fixing plate is provided on both sides of the first chamber, and a pressure strip is connected between the two fixing plates. The pressure strip is connected to the fixing plate by bolts, and the pressure strip is pressed tightly against the upper surface of the cover plate.

[0011] Preferably, the second chamber includes an upper guide plate, a lower guide plate, and an electrode plate socket stacked sequentially from top to bottom. The upper surface of the lower guide plate is provided with a liquid collection groove extending from top to bottom. The bottom of the liquid collection groove is provided with a plurality of connecting grooves that communicate with the liquid collection groove and extend downward. A plurality of electrochemical biosensors facing the connecting grooves are placed in the electrode plate socket. The upper guide plate is provided with an inlet channel and an outlet channel that respectively connect to both ends of the liquid collection groove.

[0012] Furthermore, a first sealing ring is provided around the liquid collection tank, which abuts against the upper guide plate and the lower guide plate. A second sealing ring is provided around the sensing area of ​​the electrochemical biosensor, which abuts against the lower guide plate and the electrode plate socket. The second sealing ring abuts against the outer periphery of the connecting groove.

[0013] Furthermore, the second chamber is provided with fixing plates on both sides, and pressure strips are connected between the two fixing plates. The pressure strips are fixed to the fixing plates by bolts.

[0014] Furthermore, the pressure strip has a first end and a second end, both of which are provided with through holes for bolts to pass through, and the second end is also provided with a slip groove extending to the edge of the pressure strip on the side of the through hole. The width of the slip groove is greater than the outer diameter of the bolt thread portion, and the slip groove penetrates the pressure strip in the thickness direction.

[0015] Furthermore, the signal collection end of the electrochemical biosensor extends forward from between the lower guide plate and the electrode plate socket, and a notch is provided in the fixing plate on the front side of the second chamber, the position of the notch corresponding to the position of the signal collection end of the electrochemical biosensor.

[0016] Due to the application of the above technical solution, the present invention has the following advantages compared with the prior art:

[0017] The real-time cell metabolism measurement device of the present invention extracts the culture medium from the cell culture flask via a pump mechanism, pumps it to a first chamber for pH, dissolved oxygen, dissolved carbon dioxide, or ion detection, and then the culture medium flows to a second chamber for real-time monitoring of indicators such as glucose, lactate, glutamate, glutamine, and hydrogen peroxide using an electrochemical biosensor. Finally, the culture medium flows back to the cell culture flask. This achieves real-time metabolic detection. Attached Figure Description

[0018] The following sections will describe some specific embodiments of the invention in detail by way of example and not limitation, with reference to the accompanying drawings. The same reference numerals in the drawings denote the same or similar parts or portions. Those skilled in the art should understand that these drawings are not necessarily drawn to scale. In the drawings:

[0019] Figure 1 This is a schematic diagram showing the connection between the real-time cell metabolism measurement device of the present invention and the cell culture flask;

[0020] Figure 2 This is a schematic diagram of the real-time cell metabolism measurement device of the present invention, excluding the pump mechanism and necessary connecting pipes;

[0021] Figure 3 yes Figure 2 A schematic diagram showing the breakdown of the first chamber;

[0022] Figure 4 yes Figure 3 Another viewpoint;

[0023] Figure 5 This is the bottom view of the base plate;

[0024] Figure 6 yes Figure 5 Enlarged view of point A in the image;

[0025] Figure 7 This is a schematic diagram of the internal structure of the flow channel seat in the first chamber;

[0026] Figure 8 yes Figure 7 Enlarged view at point B in the middle;

[0027] Figure 9 A cross-sectional view of the first chamber is shown;

[0028] Figure 10 This is a schematic diagram of the cover plate structure in the first chamber;

[0029] Figure 11 yes Figure 10 Enlarged view at point D;

[0030] Figure 12 This is a schematic diagram of the second chamber split;

[0031] Figure 13 This is a schematic diagram of the upper guide vane structure in the second chamber;

[0032] Figure 14 This is a schematic diagram of the lower guide vane structure in the second chamber;

[0033] Figure 15 This is a cross-sectional view of the second chamber;

[0034] Figure 16 yes Figure 15 Enlarged view at point F;

[0035] Figure 17 This is a 3D schematic diagram of the molding strip;

[0036] Figure 18 yes Figure 2 The left view;

[0037] Figure 19 This is a schematic diagram showing the fit between the pressure strip and the fixing plate;

[0038] Figure 20 yes Figure 2 A cross-sectional view of the first chamber in the middle;

[0039] Figure 21 yes Figure 20 Enlarged view of point G in the image;

[0040] Figure 22 This is a schematic diagram after the lower guide plate and electrode plate socket are separated;

[0041] Figure 23 This is a schematic diagram of the electrochemical biosensor mating with the second sealing ring;

[0042] Figure 24 yes Figure 23 Another viewpoint;

[0043] The reference numerals in the attached figures are explained as follows:

[0044] 1. Pump mechanism; 2. First chamber; 21. Flow channel seat; 211. Flow channel; 2111. Placement groove; 212. Settlement platform; 22. Cover plate; 221. Boss; 222. Threaded mounting hole; 223. Smooth mounting hole; 224. Small screw hole; 23. Pressure plate; 231. Small through hole; 232. Perforation; 24. Washer ring; 25. Outlet channel; 26. Inlet channel; 3. Second chamber; 31. Upper guide vane; 311. Liquid inlet channel; 312. Liquid outlet channel; 313. First slot; 32. Lower guide vane; 321. Liquid collection tank; 322. Connecting groove; 323. Second slot; 324. Third slot; 33. Electrode plate socket; 331. Embedded groove; 3311. Extension; 34. First sealing ring; 35. Second sealing ring; 351. Liquid placement space; 36. Pad block 361. Connecting plate; 362. Support column; 4. Base plate; 41. Threaded hole; 42. Through hole; 5. pH sensor; 51. First threaded part; 52. First probe; 53. First enclosure plate; 531. First notch; 6. Dissolved oxygen sensor; 61. Second threaded part; 62. Second probe; 63. Second enclosure plate; 631. Second notch; 7. Dissolved carbon dioxide sensor; 71. Fastening surface; 8. Electrochemical biosensor; 81. Contact area; 9. Sleeve tube; 10. Pressure strip; 101. First end; 102. Second end; 103. Through hole; 104. Slip groove; 11. Fixing plate; 111. Notch; 112. Base; 113. Rod; 12. Bolt; 13. Connecting tube; 14. Rubber pad; 141. Bolt mounting hole; 15. Cell culture flask. Detailed Implementation

[0045] The technical solution of the present invention will now be clearly and completely described with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0046] In the description of this invention, it should be noted that the terms "center," "upper," "lower," "left," "right," "vertical," "horizontal," "inner," and "outer," etc., indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings. They are used only for the convenience of describing the invention and for simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation. Therefore, they should not be construed as limitations on the invention. Furthermore, the terms "first," "second," and "third" are used for descriptive purposes only and should not be construed as indicating or implying relative importance.

[0047] Furthermore, the technical features involved in the different embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0048] like Figure 1 As shown, the real-time cell metabolism measurement device includes a pump mechanism 1, a first chamber 2, and a second chamber 3, which are connected sequentially via multiple connecting pipes 13. The inlet of the pump mechanism 1 is connected to the outlet of the cell culture flask 15, and the outlet of the second chamber 3 is connected to the inlet of the cell culture flask 15. The pump mechanism 1 draws the culture medium from the cell culture flask 15 and then passes it sequentially through the first chamber 2 and the second chamber 3. Each of the first chamber 2 and the second chamber 3 is equipped with a corresponding sensor to perform real-time measurement of the culture medium in the cell culture flask 15.

[0049] like Figure 2 The first chamber 2 and the second chamber 3 are placed on a base plate 4, making the structure compact. The first chamber 2 contains a pH sensor 5, a dissolved oxygen sensor 6, and a dissolved carbon dioxide sensor 7, used to measure the pH, dissolved oxygen, and dissolved carbon dioxide of the culture medium, respectively. One of the sensors can be replaced with an ion sensor to measure parameters such as potassium ions. The second chamber 3 contains multiple electrochemical biosensors 8, used to measure indicators such as glucose, lactic acid, glutamate, glutamine, or hydrogen peroxide. Each end of the first chamber 2 has a retaining tube 9 for inlet and outlet of liquid (left side for inlet, right side for outlet), which connects to the connecting tube 13. The second chamber 3 has two retaining tubes 9, also for inlet and outlet of liquid (right side for inlet, left side for outlet).

[0050] like Figure 3 The first chamber 2 shown mainly includes a flow channel seat 21 and a cover plate 22. A flow channel 211 is provided inside the flow channel seat 21. The flow channel 211 extends downward from the upper surface of the flow channel seat 21 to a certain depth. The left and right ends of the flow channel 211 are respectively connected to the retaining tubes 9 on the outer sides of both ends of the flow channel seat 21 via inlet channels 26 and outlet channels 25 formed on the side walls of both ends of the flow channel seat 21 (see...). Figure 7 They are connected together. Each ferrule tube 9 has a through channel between its two ends.

[0051] like Figure 7 As shown, the flow channel 211 includes three placement slots 2111 and an elongated channel connecting adjacent placement slots 2111. The placement slots 2111 accommodate the lower end of the sensor, allowing the probe at the lower end of the sensor to contact the cell culture medium within the flow channel 211. As the cell culture medium flows from the left end to the right end within the flow channel 211, it contacts the sensor probe, thereby allowing relevant parameters in the culture medium to be measured by multiple sensors. Figure 3 From left to right are pH sensor 5, dissolved oxygen sensor 6, and dissolved carbon dioxide sensor 7, which allow the pH, dissolved oxygen, and dissolved carbon dioxide in the culture medium to be measured.

[0052] like Figure 3A cover plate 22 is provided on the flow channel seat 21. The cover plate 22 seals the upper end of the flow channel 211. The cover plate 22 has three holes: two threaded mounting holes 222 and one smooth mounting hole 223 from left to right. The pH sensor 5 and the dissolved oxygen sensor 6 are threaded into the two threaded mounting holes 222, and the lower ends of the sensors extend downward through the cover plate 22 and into the placement groove 2111. The dissolved carbon dioxide sensor 7 is tightly inserted into the smooth mounting hole 223, and the lower end extends downward through the cover plate 22 and into the placement groove 2111 to contact the culture medium.

[0053] The inner surface of the smooth mounting hole 223 is smooth. The outer peripheral surface of the dissolved carbon dioxide sensor 7 is also smooth, and the dissolved carbon dioxide sensor 7 is fixed in the smooth mounting hole 223 by the gasket 24. Figure 20 and Figure 21 The upper section of the smooth mounting hole 223 is flared, and its diameter gradually decreases from top to bottom. The lower section of the smooth mounting hole 223 is cylindrical. The peripheral surface of the dissolved carbon dioxide sensor 7 is a fastening surface 71, and a washer 24 is wrapped around the fastening surface 71 and inserted into the smooth mounting hole 223 from top to bottom. The washer 24 can be a soft, waterproof material. The washer 24 has a sufficient thickness so that when the lower end of the washer 24 is inserted into the lower section of the smooth mounting hole 223, the washer 24 is squeezed and presses the dissolved carbon dioxide sensor 7 tightly into the smooth mounting hole 223, thereby fixing the dissolved carbon dioxide sensor 7. A pressure plate 23 is provided at the upper end of the smooth mounting hole 223. The height of the dissolved carbon dioxide sensor 7 can be adjusted by adjusting the height of the fastening surface 71. Furthermore, because the washer 24 has a certain amount of elasticity, even if the outer diameter of the dissolved carbon dioxide sensor 7 has different sizes of deviation, it can be adapted to the smooth mounting hole 223 by the deformation of the washer 24.

[0054] like Figure 3 The pressure plate 23 at the upper end of the smooth mounting hole 223 is a square plate. A round hole, i.e., a through hole 232, is opened in the middle of the pressure plate 23, through hole 232 for the dissolved carbon dioxide sensor 7 to pass through. Small through holes 231 are also opened on both sides of the through hole 232. Small screw holes 224 are opened on the cover plate 22 corresponding to the positions of the small through holes 231. The inner wall of the small through hole 231 is smooth, while the inner wall of the small screw hole 224 is threaded. The pressure plate 23 is fixed by passing a bolt (not shown) from top to bottom through the small through hole 231 and threading it into the small screw hole 224.

[0055] from Figure 21As can be seen, the pressure plate 23 limits the upper end of the washer 24, thereby preventing the washer 24 from coming off upwards. Given that the upper part of the smooth mounting hole 223 is flared, and the diameter of the upper part gradually decreases from top to bottom, the washer 24 may come off upwards. This is because the pressure on the upper part of the washer 24 is less than that on the lower part, thus the washer 24 tends to move upwards. After removing the pressure plate 23, the dissolved carbon dioxide sensor 7 can be removed. Setting the upper part of the smooth mounting hole 223 to be flared is a preferred measure. However, it is also possible to omit this flare. In this case, although the washer 24 is not easily inserted into the smooth mounting hole 223, a larger external force can still force the washer 24 into the smooth mounting hole 223. The upper part of the smooth mounting hole 223 is flared, which allows the gasket 24 to be quickly inserted into the smooth mounting hole 223. When the gasket 24 is first inserted, it is easy to insert into the smooth mounting hole 223 because the lower end of the gasket 24 is hardly squeezed. As the downward diameter of the smooth mounting hole 223 gradually decreases, the squeezing force on the lower end of the gasket 24 gradually increases, thereby increasing the clamping force between the fastening surface 71 of the dissolved carbon dioxide sensor 7 and the inner wall of the smooth mounting hole 223, thus ensuring that the dissolved carbon dioxide sensor 7 can be securely installed.

[0056] And such Figure 4 As shown, the lower end of the pH sensor 5 and the dissolved oxygen sensor 6 have a first threaded portion 51 and a second threaded portion 61 on their outer peripheral surfaces. The first threaded portion 51 and the second threaded portion 61 are threadedly engaged with the threaded mounting hole 222 on the cover plate 22 and thus firmly fixed.

[0057] like Figure 9 Taking pH sensor 5 as an example, the lower part of pH sensor 5 is inserted into flow channel 211 and comes into contact with the culture medium (not shown in the figure) in flow channel 211. Specifically, the first probe 52 at the lower end of pH sensor 5 comes into contact with the culture medium.

[0058] A first surrounding plate 53 is provided around the first probe 52. A first notch 531 is provided on the first surrounding plate 53, extending upward from the lower end of the first surrounding plate 53. The first surrounding plate 53 helps to protect the first probe 52 from damage. The first notch 531 on the first surrounding plate 53 allows the culture medium to flow from the outside of the first surrounding plate 53 through the first notch 531 into the first surrounding plate 53 and into contact with the first probe 52.

[0059] Similarly, such as Figure 3The lower end of the dissolved oxygen sensor 6 has a second threaded portion 61 on its outer peripheral surface, which is threadedly engaged with the threaded mounting hole 22 in the middle of the cover plate 22. The lower end of the dissolved oxygen sensor 6 is provided with a second probe 62, and a second surrounding plate 63 is provided around the second probe 62. The second surrounding plate 63 has a plurality of second notches 631 extending upward from the lower end.

[0060] like Figure 7 and Figure 8 The top of the flow channel seat 21 is provided with a recessed platform 212, which is formed by a downward indentation in the middle region of the flow channel seat 21. A retaining ring surrounds the recessed platform 212. For example... Figure 10 and Figure 11 The lower surface of the cover plate 22 is provided with a boss 221. The boss 221 is formed by protruding downward from the middle area of ​​the cover plate 22. The shape of the boss 221 matches the shape of the recess 212. The boss 221 can be inserted into the recess 212 and they are almost in a clearance fit. This allows the cover plate 22 and the flow channel seat 21 to be quickly positioned and limited. After the cover plate 22 is placed on the flow channel seat 21, it is not easy to move laterally.

[0061] like Figure 2 When the cover plate 22 is placed on the flow channel seat 21, the two pressure strips 10 press down on the cover plate 22 to brake it.

[0062] like Figure 18 The front and rear sides of the first chamber 2 ( Figure 18 The left side of the first chamber 2 is the rear side, and the right side is the front side of the first chamber 2. Both sides are equipped with fixing plates 11. The two ends of the pressure strip 10 are fixed to the fixing plates 11 by bolts 12, thereby pressing the cover plate 22 tightly onto the flow channel seat 21.

[0063] like Figure 19 The fixing plate 11 includes a base 112 at the bottom and rods 113 connected to both sides of the top of the base 112, with a notch 111 formed between the rods 113. The rear ends of two pressure strips 10 are respectively connected to the two rods 113, thereby fixing the ends of the two pressure strips 10 with a fixing plate 11, improving stability. The notch 111 also helps reduce the weight of the fixing plate 11. The base 112 is fixedly connected to the base plate 4 with bolts. Figure 5 and Figure 6 The base plate 4 has multiple through holes 42, the inner walls of which are smooth. Figure 19 The base 112 has a threaded hole corresponding to the through hole 42 at its bottom end. A bolt is used to fix the fixing plate 11 by passing through the through hole 42 from bottom to top and engaging with the threaded hole at the bottom end of the base 112. Figure 5 and Figure 6 The outer diameter of the lower end of the through hole 42 is larger to accommodate the bolt head, thus preventing the bolt from protruding from the through hole 42. For example... Figure 19The device is equipped with two pressure bars 10, which can effectively brake the first chamber 2, thus improving the braking effect compared to having only one pressure bar 10.

[0064] like Figure 17 The pressure strip 10 has a first end 101 and a second end 102 at its two ends. Both the first end 101 and the second end 102 are provided with through holes 103 for bolts 12 to pass through. A sliding groove 104 is also provided on the side of the through hole 103 on the second end 102. The sliding groove 104 extends through the through hole 103 and the edge of the pressure strip 10, and extends through the entire thickness of the pressure strip 10. The width of the sliding groove 104 is slightly larger than the outer diameter of the bolt 12's threaded portion, allowing the bolt 12 to slide out of the sliding groove 104. In actual use, the first end 101 is fixed to a fixing plate 11 by bolts 12, and the second end 102 can swing around the first end 101. This allows the pressure strip 10 to swing without removing the bolt 12 from the corresponding fixing plate 11, thus removing the restriction on the lower components and enabling quick disassembly or assembly of the first chamber 2 and the second chamber 3.

[0065] like Figure 20 The culture medium flows into the flow channel 211 from the left end of the ferrule 9 and comes into contact with the probes of the pH sensor 5, dissolved oxygen sensor 6, and dissolved carbon dioxide sensor 7 in the flow channel 211, and flows out from the right end of the ferrule 9.

[0066] like Figure 20 The flow channel 211 has an inlet channel 26 at its left end and an outlet channel 25 at its right end. The inlet channel 26 and the outlet channel 25 are at the same height, but are lower than the upper edge of the flow channel seat 21. The culture medium level in the flow channel 211 will not be higher than the height of the inlet channel 26 and the outlet channel 25, that is, it will not reach the upper edge of the flow channel seat 21, and therefore will not overflow from the upper edge of the flow channel seat 21. Therefore, a rubber ring seal is not required between the cover plate 22 and the flow channel seat 21.

[0067] like Figure 2 As shown, the second chamber 3 is located in front of the first chamber 2 to make the entire device structure compact.

[0068] like Figure 12 The second chamber 3 mainly includes an upper guide plate 31, a lower guide plate 32, and an electrode plate socket 33. The three components are combined to form an inner chamber, in which four electrochemical biosensors 8 are inserted. The electrochemical biosensors 8 are in contact with the culture medium in the inner chamber, thereby measuring indicators such as glucose, lactic acid, glutamic acid, glutamine, or hydrogen peroxide (selected from four) in the culture medium.

[0069] like Figure 12 and Figure 13The upper guide vane 31 has two through holes, one vertically and one vertically, which are respectively the inlet channel 311 and the outlet channel 312 located at approximately both ends. The inlet channel 311 and the outlet channel 312 are respectively connected to a retaining tube 9. The retaining tubes 9 are vertically connected, and each retaining tube 9 has a through internal channel between its upper and lower ends. The retaining tube 9 on the right side is connected to the retaining tube 9 at the right end of the first chamber 2 via a connecting pipe 13 (see...). Figure 1 This is to receive the culture medium flowing out from the first chamber 2. The left-side tubing 9 connects to the cell culture flask to return the culture medium to the cell culture flask.

[0070] like Figure 12 A liquid collecting groove 321 is provided on the upper surface of the lower guide vane 32. The liquid collecting groove 321 extends downward to a certain depth from the upper surface of the lower guide vane 32. The liquid collecting groove 321 extends almost horizontally along the length of the lower guide vane 32 to simultaneously connect the liquid inlet channel 311 and the liquid outlet channel 312. A second slot 323 is provided around the liquid collecting groove 321, and... Figure 13 The lower surface of the upper guide vane 31 is provided with a first slot 313 corresponding to the second slot 323. When the upper guide vane 31 and the lower guide vane 32 are spliced ​​together, the first slot 313 and the second slot 323 are connected, and a device is placed in the space formed by the first slot 313 and the second slot 323. Figure 12 The first sealing ring 34, which is elastic and deformable by compression, is located in the first slot 313 and the second slot 323 to seal between the upper guide plate 31 and the lower guide plate 32, so that the culture medium in the collection tank 321 will not flow out from the gap between the upper guide plate 31 and the lower guide plate 32.

[0071] like Figure 12 and Figure 14 The bottom of the collection tank 321 is provided with four connecting grooves 322, which extend downwards from the bottom of the collection tank 321 through the lower guide plate 32. The connecting grooves 322 are used to guide the cell culture medium in the collection tank 321 downwards to contact the electrochemical biosensor 8.

[0072] like Figure 12 The upper surface of the electrode plate socket 33 has four slots 331. The slots 331 are used to insert the electrochemical biosensor 8. The electrochemical biosensor 8 is plate-shaped. Figure 23 The upper surface of the inner end of the electrochemical biosensor 8 is the contact area 81, used for contact with the culture medium. For example... Figures 22 to 24The entire lower edge of the second sealing ring 35 is pressed tightly against the surface of the electrochemical biosensor 8, forming a liquid-containing space 351 within the second sealing ring 35. During detection, the liquid-containing space 351 contains culture medium. Because the lower edge of the second sealing ring 35 is pressed tightly against the surface of the electrochemical biosensor 8, there will be no leakage at the bottom of the liquid-containing space 351, and the culture medium will not flow out from between the second sealing ring 35 and the electrochemical biosensor 8.

[0073] like Figure 14 The lower surface of the lower guide plate 32 has a lower end opening of the connecting groove 322. The lower end opening of the connecting groove 322 corresponds to the four recesses 331 on the electrode plate socket 33. A third retaining groove 324 is formed around the lower end opening of the connecting groove 322, and the second sealing ring 35 is embedded in the third retaining groove 324.

[0074] like Figure 15 and Figure 16 When the lower guide plate 32 and the electrode plate socket 33 are put together, the second sealing ring 35 seals the gap between the lower guide plate 32 and the electrode plate socket 33, so that the culture medium in the connecting groove 322 will not flow out from the gap between the lower guide plate 32 and the electrode plate socket 33.

[0075] like Figure 12 and Figure 22 The groove 331 extends outward on both sides to form extensions 3311, which are used to accommodate the two sides of the second sealing ring 35. The shape of the extensions 3311 matches the shape of the two sides of the second sealing ring 35, so that the extensions 3311 can limit the movement of the second sealing ring 35, thereby ensuring the sealing effect of the second sealing ring 35. Furthermore, because the second sealing ring 35 is not easily displaced, the culture medium in the liquid-filled space 351 in the middle of the second sealing ring 35 has a stable environment, which does not interfere with the reaction between the culture medium and the substance coated on the contact area 81 of the electrochemical biosensor 8, thus ensuring the accuracy of the detection.

[0076] like Figure 15 and Figure 16 The culture medium flows into the second chamber 3 through the tubing 9, then into the collection tank 321, and finally into the four connecting grooves 322 at the bottom of the collection tank 321. It then contacts the electrochemical biosensor 8 at the bottom of the connecting grooves 322, where it measures indicators such as glucose, lactic acid, glutamic acid, glutamine, or hydrogen peroxide. The electrochemical biosensor 8 has an exposed portion outside the second chamber 3, which serves as a signal collection terminal. This exposed portion can be connected to an adapter (not shown) to collect the detection signal.

[0077] like Figure 12 and Figure 15A pad 36 is connected below the electrode socket 33. The pad 36 can raise the position of the electrode socket 33, thereby raising the position of the electrochemical biosensor 8 so that the electrochemical biosensor 8 can be connected to the adapter.

[0078] The pad 36 includes a bottom support column 362 and a connecting plate 361 disposed on top of the support column 362. The area of ​​the connecting plate 361 is approximately the same as the area of ​​the electrode plate socket 33, thereby allowing for maximum fit with the electrode plate socket 33 to provide support. The support column 362 has a smaller outer diameter, and its upper end is connected to the middle of the lower surface of the connecting plate 361. The support column 362 and the connecting plate 361 are integral. The lower end of the support column 362 is detachably connected to the base plate 4. Figure 7 The two through holes 42 within the frame of the base plate 4, indicated by H, are used to fix the support column 362. The bolts pass through the through holes 42 from the bottom of the base plate 4 and are threaded into the threaded holes on the lower end face of the support column 362.

[0079] like Figure 18 and Figure 19 As shown, the upper guide vane 31, lower guide vane 32, electrode plate socket 33, and other components of the second chamber 3 are pressed together by two pressure strips 10. The two ends of the pressure strips 10 are connected to the fixing plates 11 on the front and rear sides of the second chamber 3 by bolts 12. The front fixing plate 11 has a base 112 connected to rods 113 on both sides of the top surface of the base 112, and the rods 113 and the base 112 are integral. A notch 111 is formed between the two rods 113. The notch 111 reduces the weight of the fixing plate 11, and since the position of the notch 111 corresponds to the position of the electrochemical biosensor 8, the fixing plate 11 avoids the electrochemical biosensor 8, facilitating the connection between the electrochemical biosensor 8 and the adapter. The fixing plate 11 on the rear side of the second chamber 3 is shared with the fixing plate 11 on the front side of the first chamber 2, thus making the entire device compact. Furthermore, the fixing plate 11 on the front side of the second chamber 3 is thinner than the fixing plate 11 on the rear side. This is because the overall mass of the second chamber 3 is smaller than that of the first chamber 2. The function can be achieved by using a thinner fixing plate 11 on the front side, which helps to reduce the mass of the entire device.

[0080] In this example, apart from the sensor, sealing ring, and gasket, the main components are made of 316 medical-grade steel, which has good biocompatibility. To maintain the temperature of the culture medium, the tests are conducted in a constant temperature chamber, and 316 medical-grade steel has good thermal conductivity, allowing it to adapt to the temperature inside the chamber more quickly.

[0081] In this example, pump mechanism 1 can be any type of pump that can drive liquid flow, including peristaltic pumps.

[0082] The above embodiments are only for illustrating the technical concept and features of the present invention. Their purpose is to enable those skilled in the art to understand the content of the present invention and implement it accordingly. They should not be used to limit the scope of protection of the present invention. All equivalent changes or modifications made in accordance with the spirit and essence of the present invention should be covered within the scope of protection of the present invention.

Claims

1. A real-time cell metabolism measurement device for measuring metabolic cells in cell culture flasks, characterized in that, It includes a pump mechanism (1) and a first chamber (2) and a second chamber (3) connected in sequence to the outlet of the pump mechanism (1). The inlet of the pump mechanism (1) is used to connect to a cell culture flask. The second chamber (3) has an outlet for connecting to the cell culture flask. It also includes necessary connecting pipes so that the pump mechanism (1), the first chamber (2), the second chamber (3) and the cell culture flask form a circulation path. The first chamber (2) is provided with probes of at least three of the following sensors: pH sensor (5), dissolved oxygen sensor (6), dissolved carbon dioxide sensor (7) and ion sensor. The second chamber (3) is provided with multiple electrochemical biosensors (8). The first chamber (2) includes a flow channel seat (21) and a cover plate (22) detachably connected to the flow channel seat (21). A flow channel (211) is provided in the flow channel seat (21). The pH sensor (5), dissolved oxygen sensor (6), dissolved carbon dioxide sensor (7) or the ion sensor is combined on the cover plate (22) and the probe extends into the flow channel (211). The first chamber (2) has a placement slot (2111) corresponding to the sensor. The flow channel between adjacent placement slots (2111) is long and narrow. The two ends of the flow channel in the flow channel seat (21) are respectively provided with an inlet channel (26) and an outlet channel (25). The height of the inlet channel (26) and the outlet channel (25) is lower than the top edge of the flow channel seat (21). The second chamber (3) includes an upper guide plate (31), a lower guide plate (32), and an electrode plate socket (33) stacked sequentially from top to bottom. The upper surface of the lower guide plate (32) is provided with a liquid collection groove (321) extending from top to bottom. The bottom of the liquid collection groove (321) is provided with a plurality of connecting grooves (322) that communicate with the liquid collection groove (321) and extend downward. A plurality of electrochemical biosensors (8) facing the connecting grooves (322) are placed in the electrode plate socket (33). The upper guide plate (31) The liquid collection tank (321) is provided with an inlet channel (311) and an outlet channel (312) that are respectively connected to both ends of the liquid collection tank (321). A first sealing ring (34) is provided around the liquid collection tank (321) and abuts between the upper guide plate (31) and the lower guide plate (32). A second sealing ring (35) is provided around the sensing area of ​​the electrochemical biosensor (8) and abuts between the lower guide plate (32) and the electrode plate socket (33). The second sealing ring (35) abuts against the outside of the connecting groove (322).

2. The real-time cell metabolism measurement device according to claim 1, characterized in that: The first chamber (2) is provided with a fixing plate (11) on both sides, and a pressure strip (10) is connected between the two fixing plates (11). The pressure strip (10) is connected to the fixing plate (11) by bolts (12), and the pressure strip (10) is pressed tightly against the upper surface of the cover plate (22).

3. The real-time cell metabolism measurement device according to claim 1, characterized in that: The second chamber (3) is provided with fixing plates (11) on both sides, and pressure strips (10) are connected between the two fixing plates (11). The pressure strips (10) are fixed to the fixing plates (11) by bolts (12).

4. The real-time cell metabolism measurement device according to claim 3, characterized in that: The pressure strip (10) has a first end (101) and a second end (102). Both the first end (101) and the second end (102) are provided with through holes (103) for bolts to pass through. The second end (102) is also provided with a slip groove (104) extending to the edge of the pressure strip (10) on the side of the through hole (103). The width of the slip groove (104) is greater than the outer diameter of the bolt (12) screw portion, and the slip groove (104) penetrates the pressure strip (10) in the thickness direction.

5. The real-time cell metabolism measurement device according to claim 3, characterized in that: The signal collection end of the electrochemical biosensor (8) extends forward from between the lower guide plate (32) and the electrode plate socket (33), and a notch (111) is provided in the fixing plate (11) on the front side of the second chamber (3), the position of the notch (111) corresponding to the position of the signal collection end of the electrochemical biosensor (8).

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