A chip for rapidly detecting sterilization effect

Abstract

The application discloses a chip for detecting the effect of rapid sterilization. The chip is composed of a substrate and a cover plate; the substrate is provided with a bacteria piece cavity and a culture solution cavity, and a safety cavity can be optionally arranged; the bacteria piece cavity is provided with a pinhole type bacteria piece placing groove similar to a pinhole type SIM card groove; the culture solution cavity is provided with a culture solution injection pipeline; the chip provided with the safety cavity is provided with a safety pipeline; a micro-fluidic channel is arranged between the bacteria piece cavity and the culture solution cavity; the culture solution injection pipeline, the micro-fluidic channel and the safety pipeline are all provided with check valves. The chip can be used for not only the rapid detection of the effect of hydrogen peroxide plasma sterilization, but also the rapid detection of the effect of pressure steam sterilization and ethylene oxide sterilization. While realizing the effect of sterilization, i.e. "sterilization is completed, and the result is obtained", the chip can be repeatedly used, and the problem of "disadvantages" of ampoules in the prior art is avoided, and the chip provided with the safety cavity is conducive to environmental protection and cost reduction.

Description

Technical Field

[0001] This invention belongs to the field of detection chip technology, specifically, it relates to a chip for detecting the effect of rapid sterilization. Background Technology

[0002] With the rapid development of modern science and technology, low-temperature plasma sterilizers have begun to be widely used in the cleaning, disinfection, and sterilization of medical devices. The People's Republic of China has also successively introduced national standards such as GB 27955-2020 (Hygienic Requirements for Low-Temperature Hydrogen Peroxide Gas Plasma Sterilizers) and GB / T 15981-2021 (Evaluation Methods for Sterilization Effect of Disinfection Instruments) to guide and standardize the evaluation of low-temperature plasma sterilization effects. However, due to the limitations of existing evaluation techniques, existing sterilization effect evaluation methods cannot definitively determine whether each sterilization effect is qualified or meets national requirements. Furthermore, current verification devices used for hydrogen peroxide plasma sterilization not only suffer from problems such as ampoule compression causing probe light interception and affecting normal verification, but also have long result processing times and are prone to false negatives and false positives.

[0003] Therefore, it is essential to develop a rapid sterilization effect detection chip that can achieve "results immediately after sterilization," enabling rapid determination of the sterilization effect of each low-temperature plasma sterilizer, improving the integration and automation of low-temperature plasma sterilizer sterilization effect detection, and simultaneously solving the problems of "false negatives" and "false positives" in the sterilization effect evaluation process. Summary of the Invention

[0004] The technical problem this invention aims to solve is to overcome the shortcomings of existing technologies for evaluating the sterilization effect of low-temperature plasma sterilizers. It provides a reusable, compatible, and rapid detection chip for low-temperature plasma sterilization effects that integrates sterilization, culture, and fluorescence detection, overcoming or partially solving the aforementioned problems. This chip is compatible with existing commercially available biometric readers, providing strong support and reducing costs for laboratories using plasma sterilizers to efficiently evaluate sterilization effects. Furthermore, this invention further provides a multi-chip design that facilitates rapid detection of sterilization effects.

[0005] To achieve the above objectives, the technical solution of the present invention is as follows: A rapid sterilization effect detection chip is disclosed. The chip comprises a substrate and a cover plate. The substrate includes a bacterial culture chamber, a culture medium chamber, and a safety chamber. This chip enables rapid and accurate evaluation of sterilization effectiveness, improving the integration and automation of rapid sterilization effect detection while eliminating false negatives and false positives in the evaluation process. The chip provided by this invention is compatible with various bio-readers currently on the market. Furthermore, this invention provides a chip with multiple bacterial culture chambers that facilitates rapid sterilization effect detection.

[0006] According to embodiments of the present invention, the present invention can be further optimized, and the optimized technical solution is as follows: In one preferred embodiment, a rapid sterilization effect detection chip includes a substrate and a cover plate that are bonded together.

[0007] In one preferred embodiment, a rapid sterilization effect detection chip is provided, wherein the substrate and cover plate of the chip are manufactured by thermo-press bonding.

[0008] In one preferred embodiment, a rapid sterilization effect detection chip is provided, wherein the substrate and cover plate of the chip are formed by plasma bonding encapsulation.

[0009] In one preferred embodiment, a rapid sterilization effect detection chip is provided, wherein the cover plate of the chip is made of a mixture of silicon carbide and PDMS, wherein the mass content of silicon carbide in the mixture is 1%-45%.

[0010] In one preferred embodiment, a rapid sterilization effect detection chip is provided, wherein the cover plate of the chip is made of a mixture of PDMS and graphene, wherein the graphene content in the mixture is 0.5%-15% by mass.

[0011] In one preferred embodiment, a rapid sterilization effect detection chip is provided, wherein the substrate of the chip is made of a mixture of silicon dioxide and PDMS, wherein the mass content of PDMS in the mixture is 1%-20%.

[0012] In one preferred embodiment, a rapid sterilization effect detection chip is provided, wherein the substrate of the chip is made of a mixed material of boron carbide and PDMS, wherein the mass content of PDMS in the mixed material is 15%-50%.

[0013] In one preferred embodiment, a rapid sterilization effect detection chip is provided, wherein the substrate of the chip is provided with a bacterial strip cavity and a culture medium cavity, the bacterial strip cavity is provided with a pinhole bacterial strip placement slot, and the pinhole bacterial strip placement slot is shaped like a pinhole SIM card slot.

[0014] In one preferred embodiment, a rapid sterilization effect detection chip is provided. The substrate of the chip is provided with a bacterial strip cavity, a culture medium cavity and a safety cavity. The bacterial strip cavity is provided with a pinhole bacterial strip placement slot, which is shaped like a pinhole SIM card slot.

[0015] In one preferred embodiment, a rapid sterilization effect detection chip has 1-20 bacterial cells. In one embodiment, the number of bacterial cells is 2. In some embodiments, the number of bacterial cells is 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19.

[0016] In one preferred embodiment, a rapid sterilization effect detection chip has 1-20 culture chambers. In one embodiment, the number of culture chambers is 2. In some embodiments, the number of culture chambers is 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19.

[0017] In one preferred embodiment, a rapid sterilization effect detection chip has 0-20 safety cavities. In one embodiment, the number of safety cavities is 0, i.e., no safety cavities are provided. In one embodiment, the number of safety cavities is 1. In one embodiment, the number of safety cavities is 2. In some embodiments, the number of safety cavities is 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19.

[0018] In some embodiments of the present invention, a chip for rapidly detecting sterilization effect, without a safety cavity, has the words "bacterial plate cavity" and "culture medium cavity" marked on one side of a cover plate that is not attached to the substrate of the chip.

[0019] In some embodiments of the present invention, a chip for rapidly detecting sterilization effect, when a safety cavity is provided, has the words "bacterial cavity", "culture medium cavity" and "safety cavity" marked on one side of the cover plate that is not attached to the substrate of the chip.

[0020] In one preferred embodiment, a rapid sterilization effect detection chip is provided, wherein the substrate of the chip is provided with an injection channel for a culture medium chamber.

[0021] In one preferred embodiment, a rapid sterilization effect detection chip, when a safety cavity is provided, has a safety conduit on its substrate.

[0022] In one preferred embodiment, a microfluidic channel is provided between the bacterial cell cavity of the chip substrate and the culture medium cavity of the substrate; the microfluidic channel is provided with a check valve.

[0023] In one preferred embodiment, when a safety cavity is provided, a microfluidic channel is provided between the microbial cavity of the chip substrate and the safety cavity of the substrate; the microfluidic channel is provided with a check valve.

[0024] In one preferred embodiment, a rapid sterilization effect detection chip, without a safety cavity, has injection lines and microfluidic channels connecting different cavities with the same or different diameters.

[0025] In one preferred embodiment, a rapid sterilization effect detection chip, when a safety cavity is provided, has the same or different diameters for the injection tubing, microfluidic channel and safety tubing connecting different cavities.

[0026] In one preferred embodiment, a rapid sterilization effect detection chip is provided, wherein the diameter of the injection conduit connecting the culture medium chamber is 0.5 mm to 5 mm. In one embodiment, the diameter of the injection conduit connecting the culture medium chamber is 0.51 mm. In some embodiments, the diameter of the injection conduit connecting the culture medium chamber is 0.52 mm, 0.53 mm, 0.54 mm, 0.55 mm, 0.56 mm, 0.57 mm, 0.58 mm, 0.59 mm, or 0.6 mm. In some embodiments, the diameter of the injection conduit connecting the culture medium chamber is 0.61 mm to 0.7 mm. In some embodiments, the diameter of the injection conduit connecting the culture medium chamber is 0.71 mm to 0.8 mm. In some embodiments, the diameter of the injection conduit connecting the culture medium chamber is 0.81 mm to 0.9 mm. In some embodiments, the diameter of the injection conduit connecting the culture medium chamber is 0.91 mm to 1 mm. In some embodiments, the diameter of the injection tubing connecting the culture medium chamber is 1.1 mm to 2 mm. In some embodiments, the diameter of the injection tubing connecting the culture medium chamber is 2.1 mm to 3 mm. In some embodiments, the diameter of the injection tubing connecting the culture medium chamber is 3.1 mm to 4 mm. In some embodiments, the diameter of the injection tubing connecting the culture medium chamber is 4.1 mm to 4.9 mm.

[0027] In one preferred embodiment, a rapid sterilization effect detection chip is provided, wherein the cross-sectional area of ​​the microfluidic channel connecting the bacterial culture chamber and the culture medium chamber on the chip substrate is 1 / 4 to 3 / 4 of the cross-sectional area of ​​the culture medium injection channel in the culture medium chamber. In one embodiment, the cross-sectional area of ​​the microfluidic channel is 1 / 3 of the cross-sectional area of ​​the culture medium injection channel in the culture medium chamber. In some embodiments, the cross-sectional area of ​​the microfluidic channel is 1 / 2 to 2 / 3 of the cross-sectional area of ​​the culture medium injection channel in the culture medium chamber.

[0028] In one preferred embodiment, a rapid sterilization effect detection chip is provided, wherein the volume of the culture medium chamber is 0.5 ml to 20 ml. In one embodiment, the volume of a single culture medium chamber is 1 ml. In some embodiments, the volume of a single culture medium chamber is 2 ml. In some embodiments, the volume of a single culture medium chamber is 3 ml, 4 ml, 5 ml, 6 ml, 7 ml, 8 ml, 9 ml, 10 ml, 11 ml, 12 ml, 13 ml, 14 ml, 15 ml, 16 ml, 17 ml, 18 ml, or 19 ml.

[0029] In one preferred embodiment, a rapid sterilization effect detection chip has a cover plate with a thickness of 0.1-0.5 mm. In one embodiment, the cover plate has a thickness of 0.11 mm. In some embodiments, the cover plate has thicknesses of 0.12 mm, 0.13 mm, 0.14 mm, 0.15 mm, 0.16 mm, 0.17 mm, 0.18 mm, 0.19 mm, and 0.2 mm. In some embodiments, the cover plate has a thickness of 0.21 mm-0.3 mm. In some embodiments, the cover plate has a thickness of 0.31 mm-0.4 mm. In some embodiments, the cover plate has a thickness of 0.41 mm-0.5 mm.

[0030] A method for fabricating a rapid sterilization effect detection chip, without providing a safety cavity, includes the following steps: The first step involves using photolithography to fabricate a silicon wafer mold, etching out the edge lines of the bacterial cell cavity and culture medium cavity, as well as the injection channels, microfluidic channels, and pinhole bacterial cell placement slots that sequentially connect the cavities. The second step involves casting the wafer onto the mold to form a chip substrate. Channels are created in the bacterial cell cavity and culture medium cavity, and these channels are functionalized. Pinhole bacterial cell placement slots and check valves are then installed. The third step uses bonding and encapsulation technology to bond one side of the functionalized chip substrate with the bacterial cell cavity and culture medium cavity to a cover plate, completing chip fabrication. The fourth step involves installing an antibacterial protective plug for the culture medium injection port and an antibacterial sealing ring for the pinhole bacterial cell placement slot.

[0031] A method for fabricating a rapid sterilization effect detection chip, with a safety cavity provided, includes the following steps: The first step involves using photolithography to fabricate a silicon wafer mold, etching out the edge lines of the bacterial cell cavity, culture medium cavity, and safety cavity, as well as the injection lines, microfluidic channels, safety lines, and pinhole bacterial cell placement slots that sequentially connect to each cavity. The second step involves casting onto the silicon wafer mold to form a chip substrate. Channels are created in the bacterial cell cavity, culture medium cavity, and safety cavity, and these channels are functionalized. A pinhole bacterial cell placement slot, a check valve, and a safety valve are then installed. The third step uses bonding and encapsulation technology to bond one side of the functionalized chip substrate with the bacterial cell cavity, culture medium cavity, and safety cavity to a cover plate, completing chip fabrication. The fourth step involves installing a safety outlet antibacterial plug, an antibacterial protective plug for the culture medium injection port, and an antibacterial sealing ring for the pinhole bacterial cell placement slot.

[0032] A method for using a rapid sterilization effect detection chip, without a safety cavity, includes the following steps: Step 1: Sequentially open the antibacterial protective plug and antibacterial check valve of the inlet of the chip culture medium chamber, and inject sterile culture medium until the level of sterile culture medium in the culture medium chamber is two-thirds of the chamber height. Then, sequentially reset the antibacterial check valve and the antibacterial protective plug of the inlet. Step 2: Open the antibacterial sealing ring, use a chuck to eject the pinhole-type bacterial substrate placement slot, and place the sterilized bacterial substrate or control bacterial substrate using sterilized tweezers. Then, sequentially reset the pinhole-type bacterial substrate placement slot and the antibacterial sealing ring. Step 3: Open the check valve of the microfluidic channel between the culture medium chamber and the bacterial substrate chamber, allowing the sterile culture medium in the culture medium chamber to completely immerse the pinhole-type bacterial substrate placement slot. Step 4: Place the chip in a constant temperature incubator, culture instrument, or bio-automatic reader for a period of time to incubate. Step 5: Use a bio-automatic reader to automatically detect fluorescence, or transfer it to a fluorescence microscope for analysis using software. Compare it with the control chip and report the positive (fluorescence, i.e., "+") and negative (no fluorescence, i.e., "-") test results. Result judgment: If the control chip shows "+" and the experimental chip shows "+", it is judged as sterilization unqualified; if the control chip shows "+" and the experimental chip shows "-", it is judged as sterilization qualified. Step 6: Place the tested chip into a medical waste recycling container with safety measures in place.

[0033] A method for using a rapid sterilization effect detection chip, in the presence of a safety cavity, includes the following steps: Step 1: Sequentially open the antibacterial protective plug and antibacterial check valve of the chip culture medium chamber, inject sterile culture medium until the sterile culture medium level in the chamber is two-thirds of the chamber height, then sequentially reset the antibacterial check valve and the antibacterial protective plug of the injection port; Step 2: Open the antibacterial sealing ring, use a chuck to eject the pinhole-type bacterial substrate placement slot, and place the sterilized bacterial substrate or control bacterial substrate into the slot using sterilized tweezers, then sequentially reset the pinhole-type bacterial substrate placement slot and the antibacterial sealing ring; Step 3: Open the connection between the culture medium chamber and the bacterial substrate chamber. Step 4: The microfluidic channel's check valve is used to completely immerse the bacterial chip placed in the pinhole-type bacterial chip placement slot through the sterile culture medium in the culture medium chamber, and then the check valve is reset; Step 5: The chip is placed in an incubator, culture instrument, or bio-automatic reader for a period of time for cultivation; Step 6: The fluorescence is automatically detected using a bio-automatic reader, or transferred to a fluorescence microscope for analysis using software, comparing it with the control chip, and reporting positive (fluorescence present, i.e., displaying "+") and negative (no fluorescence present, i.e., displaying "-") detection results. Result interpretation: The control chip displays "+". If the experimental chip displays "+", sterilization is deemed unqualified; if the control chip displays "+", and the experimental chip displays "-", sterilization is deemed qualified. Step Six: If a safety malfunction occurs with the tested chip, the safety outlet antibacterial plug and safety valve can be opened to troubleshoot the safety malfunction through the safety outlet and safety pipeline, ensuring operational safety. Step Seven: For the tested chip, open the check valve of the microfluidic channel between the bacterial chamber and the safety chamber to allow the liquid to completely enter the safety chamber, and then sequentially open the safety outlet antibacterial plug and safety valve. Step 8: Open the antibacterial protective plug and antibacterial check valve at the inlet of the chip culture medium chamber, inject medical disinfectant, and disinfect the culture medium chamber, the chip chamber, the safety chamber, and the corresponding pipes and valves one by one. After disinfection and cleaning, the liquid flowing out of the safety outlet is judged to be qualified after disinfection and sterilization. Dry it, and reset each valve and the qualified disinfection and sterilization protective plug for later use.

[0034] Compared with the prior art, the beneficial effects of the present invention are: (1) The rapid sterilization effect detection chip integrates the functions of mixing bacterial strips and culture medium and detecting sterilization effect, realizing the entire process of "sterilization completed and results obtained", avoiding the problem in the existing method where the ampoule squeezes the bacterial strips, causing the detection light to be blocked and affecting the detection of sterilization effect; (2) The rapid sterilization effect detection chip realizes the different functions of multiple series-connected chambers by means of microfluidic channel valves with anti-reverse function between each chamber, and the liquid between each chamber can be fully isolated, without cross-contamination, avoiding the "destructive" operation that requires breaking the ampoule in the existing method. (3) The rapid sterilization effect detection chip is an assembled and detachable design. After proper treatment and sterilization, it can be reused repeatedly. The chip with a safety cavity can avoid the generation of too much medical waste such as biological monitoring reagent tubes in the existing methods, which is beneficial to environmental protection and reduces detection costs. (4) The design of the multi-chambered chip for rapid sterilization effect detection can greatly shorten the preoperative waiting time for some special medical devices, improve the accuracy of the start time of surgery, shorten the duration of surgery, increase the turnover rate of operating rooms, accelerate the turnover of beds, and improve the bed turnover rate. At the same time, the sterilization effect can be determined on a large scale in the shortest time, shorten the sterilization effect inspection cycle, and recall unqualified sterilized items in the shortest time. It can achieve the final sterilization effect before using medical devices, reduce the cost and adverse effects of item recall due to sterilization failure, and thus reduce and control the occurrence of infection. (5) The rapid sterilization effect detection chip can be used not only for rapid detection of the sterilization effect of hydrogen peroxide plasma, but also for rapid detection of the sterilization effect of pressure steam sterilization and ethylene oxide sterilization. Attached Figure Description

[0035] Figure 1 This is a schematic diagram of the rapid sterilization effect detection chip structure of Embodiment 1 of the present invention.

[0036] Figure 2 yes Figure 1 A schematic diagram of the height adjustment frame structure of the rapid sterilization effect detection chip compatible with existing biological readers on the market.

[0037] Figure 3 yes Figure 1 A three-dimensional structural diagram of the first pinhole-type bacterial substrate placement slot 8 of the rapid detection chip for sterilization effect.

[0038] Figure 4 This is a schematic diagram of the rapid sterilization effect detection chip structure of Embodiment 7 of the present invention.

[0039] Figure 5 This is a schematic diagram of the substrate structure of the rapid sterilization effect detection chip in Embodiment 13 of the present invention.

[0040] Figure 6 This is a schematic diagram of the cover plate of the rapid sterilization effect detection chip in Embodiment 13 of the present invention. Implementation

[0041] The present invention will be described in detail below with reference to the accompanying drawings and embodiments. It should be noted that, unless otherwise specified, the embodiments and features described in the embodiments of the present invention can be combined with each other. For ease of description, the terms "upper," "lower," "left," and "right" appearing below only indicate that they correspond to the upper, lower, left, and right directions of the drawings themselves, and do not limit the structure. The embodiments of the present invention are merely for explaining the present invention and are not intended to limit the present invention, and the embodiments of the present invention are not limited to the embodiments given in the specification. Unless specific experimental or operating conditions are specified in the embodiments, they are prepared under conventional conditions or according to the conditions recommended by the material supplier. Furthermore, it should be understood that one or more method steps mentioned in the present invention do not preclude the existence of other method steps before or after the combined steps, or the insertion of other method steps between these explicitly mentioned steps, unless otherwise stated; it should also be understood that the combined connection relationships mentioned in the present invention do not preclude the existence of other connections before or after the combination, or the insertion of other connections between these explicitly mentioned connections, unless otherwise stated. Furthermore, unless otherwise stated, the numbering of each method step is merely a convenient tool for identifying each method step, and not intended to limit the order of the method steps or limit the scope of the invention. Changes or adjustments to their relative relationships, without substantially altering the technical content, should also be considered within the scope of the invention. In the following embodiments, the reagents, materials, and instruments used are commercially available unless otherwise specified.

[0042] The substrate, cover plate, culture medium injection port, culture medium injection pipeline and check valve, culture medium chamber, bacterial strip chamber, pinhole bacterial strip placement groove, safety chamber, microfluidic channel and check valve between culture medium chamber and bacterial strip chamber, microfluidic channel and check valve between bacterial strip chamber and safety chamber, safety pipeline, safety valve, safety outlet, antibacterial plug of safety outlet, antibacterial protective plug of injection port and antibacterial sealing ring component of pinhole bacterial strip placement groove of the present invention are all general standard parts or components known to those skilled in the art. Their structure and principle can be known by those skilled in the art through technical manuals or conventional experimental methods.

[0043] Example 1: A rapid sterilization effect detection chip comprising a culture medium chamber, a bacterial culture chamber, and a safety chamber. A rapid sterilization effect detection chip comprising a culture medium chamber, a bacterial culture chamber, and a safety chamber, as shown in the attached diagram. Figure 1As shown, the structure mainly consists of two layers: a substrate 1 and a cover plate 2. The substrate 1 has a culture medium injection port 3, a culture medium injection pipeline 4, a first check valve 5, a culture medium chamber 6, a first bacterial substrate chamber 7, a first pinhole bacterial substrate placement slot 8, and a first safety chamber 9. A first microfluidic channel 10 and a second check valve 11 are provided between the culture medium chamber 6 and the first bacterial substrate chamber 7. A second microfluidic channel 12 and a third check valve 13 are provided between the first bacterial substrate chamber 7 and the first safety chamber 9. Below the first safety chamber 9 are a first safety pipeline 14, a first safety valve 15, a first safety outlet 16, and a first safety outlet antibacterial plug 17. The culture medium injection port 3 is equipped with an antibacterial protective plug 18, and the first pinhole bacterial substrate placement slot 8 is equipped with a first antibacterial sealing ring 19. The diameter of each culture medium injection line 4 is 10 mm, the volume of each culture medium chamber is 2 ml, the cross-sectional area of ​​the microfluidic channel is 3 / 4 of the cross-sectional area of ​​the injection line channel, and the volume of the first bacterial plate chamber 7 is 2 ml. The substrate 1 is made of a mixture of silicon carbide and PDMS, with PDMS comprising 65% by mass. The cover plate 2 is made of a mixture of PDMS and graphene, with graphene comprising 4.1% by mass. The dimensions of the cover plate are 0.2*10*75 mm. To ensure compatibility with existing bio-readers, the rapid sterilization effect detection chip is equipped with a height adjustment frame structure 20 and a telescopic plate 21 on the back of the chip, as shown in the attached diagram. Figure 2 As shown. (Attached) Figure 3 It is attached Figure 1 A three-dimensional structural diagram of the first pinhole-type bacterial strip placement slot 8 of the rapid detection chip for sterilization effect.

[0044] Example 2: Preparation of a rapid sterilization effect detection chip comprising a culture medium chamber, a bacterial substrate chamber, and a safety chamber. The fabrication of a rapid sterilization effect detection chip comprising a culture medium chamber, a bacterial substrate chamber, and a safety chamber includes the following steps: First, the design of the rapid sterilization effect detection chip is drawn using AutoCAD software and a photomask is made for photolithography; using a four-inch single-crystal silicon wafer as a substrate, a silicon wafer mold is fabricated using photolithography technology, and the basic components of the chip, namely the culture medium chamber 6, the first bacterial substrate chamber 7, and the first safety chamber 9, are etched on the photoresist; the edge lines of the culture medium chamber 6, the first bacterial substrate chamber 7, and the first safety chamber 9, and the culture medium injection pipeline 4 connecting each chamber, the first microfluidic channel 10 between the culture medium chamber 6 and the first bacterial substrate chamber 7, the second microfluidic channel 12 between the first bacterial substrate chamber 7 and the first safety chamber 9, the first safety pipeline 14, and the slots and channels for placing the first pinhole bacterial substrate placement slot 8 in the first bacterial substrate chamber 7 are also described. The second step involves casting the culture medium into a mold. After casting, the culture medium cavity 6, the first mycelium plate cavity 7, and the first safety cavity 9 are functionalized. The first check valve 5 in the culture medium injection pipeline 4, the second check valve 11 in the first microfluidic channel 10 between the culture medium cavity 6 and the first mycelium plate cavity 7, the third check valve 13 in the second microfluidic channel 12 between the first mycelium plate cavity 7 and the first safety cavity 9, the first safety valve 15 in the first safety pipeline 14, and the first pinhole mycelium plate placement slot 8 are installed. The third step is to prepare a cover plate 2 corresponding to the substrate 1. The cover plate 2, which does not face the substrate 1, has markings corresponding to each cavity and channel of the substrate 1. The fourth step involves using thermo-press packaging technology to bond the functionalized substrate and cover plate together to form a chip. Fifth step, install the first safety outlet antibacterial plug 17 of the first safety outlet 16, the antibacterial protective plug 18 of the culture medium injection port 3, and the first antibacterial sealing ring 19 of the first pinhole bacterial plate placement groove 8.

[0045] Example 3: A usage example of a rapid sterilization effect detection chip comprising a culture medium chamber, a bacterial culture chamber, and a safety chamber. The use of a rapid sterilization effect detection chip comprising a culture medium chamber, a bacterial substrate chamber, and a safety chamber includes the following steps: Step 1, sequentially open the antibacterial protective plug 18 and the first antibacterial check valve 5 configured on the injection port 3 of the culture medium chamber 6, and inject sterile culture medium. The sterile culture medium formula includes: tryptone, brain and heart extract broth medium, fructose, glucose, potassium chloride, dipotassium hydrogen phosphate, Tween 80, L-alanine, L-lysine, L-valine, 2,6-pyridinedicarboxylic acid, calcium chloride, manganese chloride, 4-methylumbelliferyl-α-D-glucopyranoside, bromocresol purple, and sterile water to 100 ml, adjusting the pH to 7.4-7.8. The sterile culture medium level in the culture medium chamber 6 is two-thirds of the chamber height. Then, sequentially reset the first antibacterial check valve 5 and the antibacterial protective plug 18. Step 2: Open the first antibacterial sealing ring 19, use a chuck to pop out the first pinhole-type bacterial strip placement slot 8, and place a 6X1320 thermophilic Bacillus stearothermophilus bacterial strip or control bacterial strip that has completed its sterilization cycle in the hydrogen peroxide plasma lumen biological sterilization device into the slot using sterilized tweezers. Then, sequentially reset the first pinhole-type bacterial strip placement slot 8 and the first antibacterial sealing ring 19 to form the experimental chip or control chip. Step 3: Open the second check valve 11 of the first microfluidic channel 10 between the culture medium chamber 6 and the first bacterial strip chamber 7, and completely immerse the bacterial strip placed in the first pinhole-type bacterial strip placement slot 8 with the sterile culture medium in the culture medium chamber 6. Then, reset the second check valve 11. Step 4: Place the experimental chip and control chip together in the culture well of the bio-automatic reader at 56 degrees Celsius for 10 minutes. During the culture process, the height of the chip can be adjusted to align the bacterial strip chamber with the fluorescence detection port of the bio-automatic reader. Step 5: Automated fluorescence detection is performed using a bio-reader, or the sample is transferred to a fluorescence microscope for software analysis. The sample is compared to a control chip, and the results are reported as positive (fluorescence, indicated by "+") and negative (no fluorescence, indicated by "-"). Result determination: If both the control chip and the experimental chip show "+", sterilization is deemed unqualified; if both show "+", sterilization is deemed qualified. Step 6: After testing the chip, the third check valve 13 of the second microfluidic channel 12 between the first bacterial chamber 7 and the first safety chamber 9 is opened to allow the liquid to fully enter the first safety chamber 9. Then, the first safety outlet antibacterial plug 17 and the first safety valve 15 are opened sequentially. The liquid is discharged into a medical waste liquid recycling container with safety measures in place through the first safety pipeline 14 and the first safety outlet 16. Finally, the first safety outlet antibacterial plug 17 and the first safety valve 15 are reset.Step 7: Open the antibacterial protective plug 18 and the antibacterial first check valve 5 at the injection port 3 of the chip culture medium chamber 6. Inject a 3% hydrogen peroxide aqueous solution or 0.2% peracetic acid or a disinfectant. Disinfect the culture medium chamber 6, the first bacterial substrate chamber 7, the first safety chamber 9, and the corresponding pipes and valves one by one. Place the remaining bacterial substrate carrier into a medical waste liquid recycling container with safety measures. After disinfection and cleaning, the liquid flowing out from the first safety outlet 16 is dried after the disinfection and sterilization effect is determined to be qualified. Then, reset all valves and the disinfection and sterilization qualified protective plugs for later use.

[0046] Example 4: Detection results of a rapid sterilization effect detection chip comprising a culture medium chamber, a bacterial strip chamber, and a safety chamber on a sample bacterial strip of a hydrogen peroxide plasma tubular biological sterilization device. In Example 1 of this invention, a rapid sterilization effect detection chip comprising a culture medium chamber, a bacterial strip chamber, and a safety chamber yielded detection results for a sample bacterial strip from a hydrogen peroxide plasma tubular biological sterilization device. These results were consistent with those obtained using a commercially available ultra-fast biological reader and its accompanying biological monitoring indicator reagent. The sterile culture medium formula is as follows: 1 g tryptone, 1 g brain heart extract broth medium, 0.3 g fructose, 0.2 g glucose, 0.1 g potassium chloride, 0.1 g dipotassium hydrogen phosphate, 0.1 g Tween 80, 5 g L-alanine, 1 g L-lysine, 1 g L-valine, 2.1 g 2,6-pyridinedicarboxylic acid, 1.4 g calcium chloride, 0.001 g manganese chloride, 0.2 g 4-methylumbelliferyl-α-D-glucopyranoside, 0.002 g bromocresol purple, and sterile water to a final volume of 100 ml. The pH was adjusted to 7.4-7.8. The detection method for the commercial ultra-fast biological reader's accompanying biological monitoring indicator reagent is as follows: Place the bacterial substrate that has completed one sterilization cycle into the test tube on the automatic reader's culture rack. Press the cap vertically to crush the culture medium ampoule, allowing the culture medium to flow out and submerge the substrate. Incubate together with the control tube at a constant temperature of 56 degrees Celsius. Results for the control tube are available within 30 minutes, and results for the test tube are available within 45 minutes (the reader automatically displays "+" or "-"). Result interpretation: If both the control and test tubes show "+", sterilization is considered unqualified; if both show "+", sterilization is considered qualified. Furthermore, this invention reduces the operation time required for sample detection by 13 minutes compared to the commercial ultra-fast biological reader's accompanying biological monitoring indicator reagent, significantly improving the efficiency of the detection process. In addition, this invention achieves a 100% accuracy rate in detecting negative or positive samples from 100 bacterial substrate samples.

[0047] Example 5: An example of using a rapid sterilization effect detection chip comprising a culture medium chamber, a bacterial strip chamber, and a safety chamber to detect pressure steam sterilized bacterial strip samples. The sterilization effect rapid detection chip prepared in Example 1 of this invention includes a culture medium chamber, a bacterial strip chamber and a safety chamber. It can rapidly detect the sterilization effect of pressure steam sterilized bacterial strip samples. The experimental chip and the control chip are placed together in the culture well of the bio-automatic reader at 57 degrees Celsius and incubated for 25 minutes. The culture medium formula is as follows: 0.5 g tryptone, 0.5 g brain and heart extract broth, 0.2 g fructose, 0.2 g glucose, 0.05 g potassium chloride, 0.05 g dipotassium hydrogen phosphate, 0.05 g Tween 80, 0.3 g L-alanine, 0.5 g L-lysine, 0.1 g L-valine, 0.1 g 2,6-pyridinedicarboxylic acid, 0.1 g CaCl2, 0.001 g manganese chloride, 0.01 g 4-methylumbelliferyl-α-D-glucopyranoside, 0.001 g bromocresol purple, and sterile water to make up to 100 ml. Adjust the pH to 7.4-7.8. During the cultivation process, the height of the chip can be adjusted to align the bacterial cell cavity with the fluorescence detection port of the bio-automatic reader for automatic fluorescence detection. Alternatively, the sample can be transferred to a fluorescence microscope for software analysis, comparing it with a control chip and reporting positive (fluorescence, indicated by "+") and negative (no fluorescence, indicated by "-") results. Result interpretation: If both the control chip and the experimental chip show "+", sterilization is deemed unqualified; if both show "+", sterilization is deemed qualified. The results indicate that the rapid sterilization effect detection chip constructed in Example 1 accurately reflects the true condition of the pressure steam sterilized samples. Compared with commercially available pressure steam sterilization biomonitoring indicator reagents, the rapid sterilization effect detection chip constructed in Example 1 achieves a 100% detection accuracy for 100 pressure steam sterilized bacterial cell samples, with a short detection time. Based on cost price, the average detection cost per bacterial cell sample is reduced by 7.5 yuan, which not only benefits hospital medical insurance procurement and reuse but also saves testing fees for the testers.

[0048] Example 6: A rapid sterilization effect detection chip comprising a culture medium chamber, a bacterial strip chamber, and a safety chamber, used to detect ethylene oxide sterilized bacterial strip samples. The rapid sterilization effect detection chip prepared in Example 1 of this invention was used to rapidly detect the sterilization effect of ethylene oxide sterilized bacterial tablets according to the method of this invention. The experimental chip and the control chip were placed together in the culture well of a bio-automatic reader at 58 degrees Celsius for 30 minutes. The culture medium formula was as follows: 1.5 g tryptone, 1.5 g brain heart extract broth, 0.4 g fructose, 0.3 g glucose, 0.15 g potassium chloride, 0.1 g dipotassium hydrogen phosphate, 0.2 g Tween 80, 0.7 g L-alanine, 0.15 g L-lysine, 0.15 g L-valine, 0.25 g 2,6-pyridinedicarboxylic acid, 0.18 g calcium chloride, 0.0015 g manganese chloride, 0.03 g 4-methylumbelliferyl-α-D-glucopyranoside, 0.003 g bromocresol purple, and sterile water was added to 100 ml. The pH was adjusted to 7.4-7.8. During the cultivation process, the height of the chip can be adjusted to align the bacterial cell cavity with the fluorescence detection port of a bio-automatic reader for automatic fluorescence detection. Alternatively, the sample can be transferred to a fluorescence microscope for analysis using software, comparing it with a control chip and reporting positive (fluorescence, indicated by "+") and negative (no fluorescence, indicated by "-") results. Result interpretation: If both the control chip and the experimental chip show "+", sterilization is deemed unqualified; if both show "+", sterilization is deemed qualified. The results indicate that the rapid sterilization effect detection chip constructed in Example 1 accurately reflects the true condition of the bacterial cell samples. Compared with commercially available ethylene oxide sterilization biomonitoring indicator reagents, the rapid sterilization effect detection chip constructed in Example 1 achieves a 100% detection accuracy for 100 ethylene oxide-sterilized bacterial cell samples, with a shorter detection time.

[0049] Example 7: A rapid sterilization effect detection chip comprising one culture medium chamber, two bacterial substrate chambers, and one safety chamber. A rapid sterilization effect detection chip comprising one culture medium chamber, two bacterial substrate chambers, and one safety chamber, as shown in the attached diagram. Figure 4As shown, the structure mainly consists of two layers: a substrate 1 and a cover plate 2. The substrate 1 has a culture medium injection port 3, a culture medium injection pipe 4, a first check valve 5, a culture medium chamber 6, a first bacterial substrate chamber 7, a first pinhole bacterial substrate placement slot 8, a second bacterial substrate chamber 101, a second pinhole bacterial substrate placement slot 102, and a first safety chamber 9. A first microfluidic channel 10 and a second check valve 11 are provided between the culture medium chamber 6 and the first bacterial substrate chamber 7. A third microfluidic channel 201 and a fourth check valve 202 are provided between the culture medium chamber 6 and the second bacterial substrate chamber 101. A second microfluidic channel 12 and a third check valve 13 are provided between the first bacterial substrate chamber 7 and the first safety chamber 9. A fourth microfluidic channel 203 and a fifth check valve 204 are provided between the second bacterial substrate chamber 101 and the first safety chamber 9. Below the first safety chamber 9, a first safety pipe 14, a first safety valve 15, a first safety outlet 16, and a first safety outlet antibacterial plug 17 are provided. The culture medium injection port 3 is equipped with an antibacterial protective plug 18, the first pinhole bacterial strip placement slot 8 is equipped with a first antibacterial sealing ring 19, and the second pinhole bacterial strip placement slot 102 is equipped with a second antibacterial sealing ring 111.

[0050] Example 8: Preparation of a rapid sterilization effect detection chip comprising one culture medium chamber, two bacterial plate chambers, and one safety chamber. The fabrication of a rapid sterilization effect detection chip comprising one culture medium chamber, two bacterial substrate chambers, and one safety chamber includes the following steps: First, the designed rapid sterilization effect detection chip is drawn using AutoCAD software and fabricated into a photomask for photolithography; using a four-inch single-crystal silicon wafer as a substrate, a silicon wafer mold is fabricated using photolithography, and the basic components of the chip—the culture medium chamber 6, the first bacterial substrate chamber 7, the second bacterial substrate chamber 101, and the first safety chamber 9—are etched onto the photoresist, including the edge lines of the first bacterial substrate chamber 7, the second bacterial substrate chamber 101, and the first safety chamber 9, and the connecting lines between each chamber. Culture medium injection line 4, first microfluidic channel 10 between culture medium chamber 6 and first mycelium plate chamber 7, third microfluidic channel 201 between culture medium chamber 6 and second mycelium plate chamber 101, second microfluidic channel 12 between first mycelium plate chamber 7 and first safety chamber 9, fourth microfluidic channel 203 between second mycelium plate chamber 101 and first safety chamber 9, first safety line 14, slot and channel of first pinhole mycelium plate placement groove 8 placed in first mycelium plate chamber 7, slot and channel of second pinhole mycelium plate placement groove 102 placed in second mycelium plate chamber 101; The second step involves casting the culture medium into a mold. After casting, the culture medium cavity 6, the first mycelium plate cavity 7, the second mycelium plate cavity 101, and the first safety cavity 9 are functionalized. The first check valve 5 in the culture medium injection pipeline 4, the second check valve 11 in the first microfluidic channel 10 between the culture medium cavity 6 and the first mycelium plate cavity 7, the fourth check valve 202 in the third microfluidic channel 201 between the culture medium cavity 6 and the second mycelium plate cavity 101, the third check valve 13 in the second microfluidic channel 12 between the first mycelium plate cavity 7 and the first safety cavity 9, the fifth check valve 204 in the fourth microfluidic channel 203 between the second mycelium plate cavity 101 and the first safety cavity 9, the first safety valve 15 in the first safety pipeline 14, the first pinhole mycelium plate placement slot 8, and the second pinhole mycelium plate placement slot 102 are installed. The third step is to prepare a cover plate 2 corresponding to the substrate 1. The cover plate 2, which does not face the substrate 1, has markings corresponding to each cavity and channel of the substrate 1. The fourth step involves using thermo-press packaging technology to bond the functionalized substrate and cover plate together to form a chip. Fifth step, install the first safety outlet antibacterial plug 17 of the first safety outlet 16, the antibacterial protective plug 18 of the culture medium injection port 3, and the first antibacterial sealing ring 19 of the first pinhole bacterial tablet placement groove 8; and the second antibacterial sealing ring 111 of the second pinhole bacterial tablet placement groove 102.

[0051] Example 9: A usage example of a rapid sterilization effect detection chip comprising one culture medium chamber, two bacterial substrate chambers, and one safety chamber. The use of a multi-chip for rapid sterilization effect detection, comprising one culture medium chamber, two bacterial substrate chambers, and one safety chamber, includes the following steps: Step 1: Sequentially open the antibacterial protective plug 18 and the first antibacterial check valve 5 configured on the injection port 3 of the culture medium chamber 6, and inject sterile culture medium. The sterile culture medium contains appropriate amounts of tryptone, brain and heart extract broth medium, fructose, glucose, potassium chloride, dipotassium hydrogen phosphate, Tween 80, L-alanine, L-lysine, L-valine, 2,6-pyridinedicarboxylic acid, calcium chloride, manganese chloride, 4-methylumbelliferyl-α-D-glucopyranoside, bromocresol purple, and sterile water. Step 1: The sterile culture medium in the culture medium chamber 6 is two-thirds the height of the chamber. Then, the first antibacterial check valve 5 and the antibacterial protective plug 18 are reset sequentially. Step 2: Open the first antibacterial sealing ring 19, use a chuck to pop out the first pinhole-type bacterial strip placement slot 8, and place the thermophilic Bacillus stearothermophilus 6X1320 bacterial strip, which has completed the sterilization cycle in the hydrogen peroxide plasma lumen biological sterilization device, into the chamber using sterilized tweezers. Then, reset the first pinhole-type bacterial strip placement slot 8 and the first antibacterial sealing ring 19 sequentially to form the experimental bacterial strip chamber. Step 3: Open the second antibacterial sealing ring 111, use a chuck to pop out the second pinhole-type bacterial strip placement slot 102, and place the thermophilic Bacillus stearothermophilus 6X1320 control bacterial strip into the chamber using sterilized tweezers. Then, reset the second pinhole-type bacterial strip placement slot 102 and the second antibacterial sealing ring 111 sequentially to form the control bacterial strip chamber. Step 4: Sequentially open the first antibacterial sealing ring 19 between the culture medium chamber 6 and the first bacterial strip chamber 7. The second check valve 11 of the microfluidic channel 10 and the fourth check valve 202 of the third microfluidic channel 201 between the culture medium chamber 6 and the second bacterial plate chamber 101 are used to completely immerse the experimental bacterial plates placed in the first pinhole bacterial plate placement slot 8 and the control bacterial plates placed in the second pinhole bacterial plate placement slot 102 through the sterile culture medium in the culture medium chamber 6. Then, the second check valve 11 and the fourth check valve 202 are reset in sequence. Step 5: Place the experimental chip and the control chip together in a constant temperature incubator or incubator at 56 degrees Celsius for 20 minutes. Use a fluorescence detector to automatically detect fluorescence, or transfer it to a fluorescence microscope and analyze the positive (fluorescent) and negative (no fluorescence) detection results through software. Use a bio-automatic reader to automatically detect fluorescence, or transfer it to a fluorescence microscope and analyze it through software. Compare with the control chip and report the positive (fluorescent, i.e., display "+") and negative (no fluorescence, i.e., display "-") detection results.Result determination: If both the control chip and the experimental chip show "+", the sterilization is deemed unqualified; if both show "+", the sterilization is deemed qualified. Step six: For the tested chips, open the third check valve 13 of the second microfluidic channel 12 between the first bacterial chamber 7 and the first safety chamber 9, and open the fifth check valve 204 of the fourth microfluidic channel 203 between the second bacterial chamber 101 and the first safety chamber 9, allowing the liquid to completely enter the first safety chamber 9. Then, sequentially open the first safety outlet antibacterial plug 17 and the first safety valve 15, and discharge the liquid through the first safety pipeline 14 and the first safety outlet 16 into the safe area. The medical waste liquid recycling container is secured, and then the first safety outlet antibacterial plug 17 and the first safety valve 15 are reset; Step 7: Open the antibacterial protective plug 18 and the antibacterial first check valve 5 configured in the injection port 3 of the chip culture medium chamber 6, inject medical disinfectant, and disinfect the culture medium chamber 6, the first bacterial sheet chamber 7, the second bacterial sheet chamber 101, the first safety chamber 9 and the corresponding pipes and valves one by one. Place the remaining bacterial sheet carrier into the medical waste liquid recycling container with safety measures. After disinfection and cleaning, the liquid flowing out from the first safety outlet 16 is dried after the disinfection and sterilization effect is judged to be qualified, and each valve and the disinfection and sterilization qualified protective plug are reset for later use.

[0052] Example 10: Detection results of sample bacterial sheets from a hydrogen peroxide plasma tubular biological sterilization device comprising one culture medium chamber, two bacterial sheet chambers, and one safety chamber, using a multi-chip method for rapid sterilization effect detection. In Example 7 of this invention, a rapid sterilization effect detection chip comprising one culture medium chamber, two bacterial plate chambers, and one safety chamber yielded detection results for bacterial plates from a hydrogen peroxide plasma lumen biological sterilization device. These results were consistent with those from a commercially available ultra-fast biological reader and its accompanying biological monitoring indicator reagent. The sterile culture medium formula was as follows: 1 g tryptone, 1 g brain heart extract broth medium, 0.3 g fructose, 0.2 g glucose, 0.1 g potassium chloride, 0.1 g dipotassium hydrogen phosphate, 0.1 g Tween 80, 5 g L-alanine, 1 g L-lysine, 1 g L-valine, 2.1 g 2,6-pyridinedicarboxylic acid, 1.4 g calcium chloride, 0.001 g manganese chloride, 0.2 g 4-methylumbelliferyl-α-D-glucopyranoside, 0.002 g bromocresol purple, and sterile water to a final volume of 100 ml. The pH was adjusted to 7.4-7.8. The detection method for the commercial ultra-fast biological reader with its accompanying biological monitoring indicator reagent is as follows: Place the bacterial strips, having completed one sterilization cycle, into the test tube on the automatic reader's culture rack. Press the cap vertically to crush the culture medium ampoule, allowing the culture medium to flow out and submerge the bacterial strips. Incubate together with the control tube at a constant temperature of 56 degrees Celsius. Results for the control tube are available within 30 minutes, and results for the test tube are available within 45 minutes (the reader automatically displays "+" or "-"). Result interpretation: If both the control and test tubes show "+", sterilization is considered unqualified; if both show "+", sterilization is considered qualified. However, compared to commercial ultra-fast biological readers with their accompanying biological monitoring indicator reagents, this invention reduces the required operation time by 20 minutes because the control and test bacterial strips are detected as a single unit, significantly improving the efficiency of the detection process. Furthermore, this invention achieves 100% accuracy in detecting negative or positive samples from 100 bacterial strips.

[0053] Example 11: A rapid sterilization effect detection chip comprising one culture medium chamber, two bacterial strip chambers, and one safety chamber is used to detect pressure steam sterilized bacterial strip samples. The rapid sterilization effect detection chip prepared in Example 7 of this invention includes one culture medium chamber, two bacterial plate chambers, and one safety chamber. It rapidly detects the sterilization effect of pressure steam sterilized bacterial plate samples. The experimental chip and control chip were placed together in the culture wells of a bio-automatic reader at 57 degrees Celsius for 25 minutes. The culture medium formula was: 0.5 g tryptone, 0.5 g brain and heart extract broth, 0.2 g fructose, 0.2 g glucose, 0.05 g potassium chloride, 0.05 g dipotassium hydrogen phosphate, 0.05 g Tween 80, 0.3 g L-alanine, 0.5 g L-lysine, 0.1 g L-valine, 0.1 g 2,6-pyridinedicarboxylic acid, and 0.1 g CaCl2. The sample contained 0.001 g of manganese chloride, 0.01 g of 4-methylumbelliferyl-α-D-glucopyranoside, 0.001 g of bromocresol purple, and 100 ml of sterile water, adjusting the pH to 7.4-7.8. During cultivation, the bacterial cell cavity could be aligned with the fluorescence detection port of a bio-automatic reader by adjusting the chip height, or the sample could be transferred to a fluorescence microscope for analysis using software. The chip was compared with a control chip, and positive (fluorescence, i.e., "+") and negative (no fluorescence, i.e., "-") results were reported. Result interpretation: If both the control chip and the experimental chip showed "+", the sterilization was deemed unqualified; if both showed "+", the sterilization was deemed qualified. The results indicate that the rapid sterilization effect detection chip constructed in Example 7 accurately reflects the true condition of the pressure steam sterilized samples. Compared with commercially available biomonitoring indicator reagents for pressure steam sterilization, the rapid detection chip for sterilization effect constructed in Example 7 has a detection accuracy of up to 100% for 100 pressure steam sterilized bacterial tablet samples, and the detection time is short.

[0054] Example 12: A rapid sterilization effect detection chip comprising one culture medium chamber, two bacterial strip chambers, and one safety chamber is used to detect ethylene oxide sterilized bacterial strip samples. The rapid sterilization effect detection chip prepared in Example 7 of this invention was used to rapidly detect the sterilization effect of ethylene oxide sterilized bacterial tablets according to the method of this invention. The experimental chip and the control chip were placed together in the culture well of a bio-automatic reader at 58 degrees Celsius for 30 minutes. The culture medium formula was as follows: 1.5 g tryptone, 1.5 g brain heart extract broth, 0.4 g fructose, 0.3 g glucose, 0.15 g potassium chloride, 0.1 g dipotassium hydrogen phosphate, 0.2 g Tween 80, 0.7 g L-alanine, 0.15 g L-lysine, 0.15 g L-valine, 0.25 g 2,6-pyridinedicarboxylic acid, 0.18 g calcium chloride, 0.0015 g manganese chloride, 0.03 g 4-methylumbelliferyl-α-D-glucopyranoside, 0.003 g bromocresol purple, and sterile water was added to 100 ml. The pH was adjusted to 7.4-7.8. During the cultivation process, the height of the chip can be adjusted to align the bacterial cell cavity with the fluorescence detection port of a bio-automatic reader for automatic fluorescence detection. Alternatively, the sample can be transferred to a fluorescence microscope for analysis using software, comparing it with a control chip and reporting positive (fluorescence, indicated by "+") and negative (no fluorescence, indicated by "-") results. Result interpretation: If both the control chip and the experimental chip show "+", sterilization is deemed unqualified; if both show "+", sterilization is deemed qualified. The results indicate that the rapid sterilization effect detection chip constructed in Example 7 accurately reflects the true condition of the bacterial cell samples. Compared with commercially available ethylene oxide sterilization biomonitoring indicator reagents, the rapid sterilization effect detection chip constructed in Example 7 achieves a 100% detection accuracy for 100 ethylene oxide-sterilized bacterial cell samples, with a shorter detection time.

[0055] Example 13: A rapid sterilization effect detection chip comprising one culture medium chamber, five bacterial substrate chambers, and three safety chambers. A rapid sterilization effect detection chip comprising one culture medium chamber, five bacterial substrate chambers, and three safety chambers, as shown in the attached diagram. Figure 5 and attached Figure 6As shown, it mainly consists of a two-layer structure of substrate 1 and cover plate 2. The substrate 1 has a culture medium injection port 3, a culture medium injection pipeline 4 and a first check valve 5, a culture medium chamber 6, a first bacterial strip chamber 7, a first pinhole bacterial strip placement slot 8, a second bacterial strip chamber 101, a second pinhole bacterial strip placement slot 102, a third bacterial strip chamber 103, a third pinhole bacterial strip placement slot 104, a fourth bacterial strip chamber 105, a fourth pinhole bacterial strip placement slot 106, a fifth bacterial strip chamber 107, a fifth pinhole bacterial strip placement slot 108, a first safety chamber 9, a second safety chamber 109, and a third safety chamber 110. A first microfluidic channel 10 and a second check valve 11 are provided between the culture medium chamber 6 and the first mycelium plate chamber 7. A third microfluidic channel 201 and a fourth check valve 202 are provided between the culture medium chamber 6 and the second mycelium plate chamber 101. A fifth microfluidic channel 205 and a sixth check valve 206 are provided between the culture medium chamber 6 and the third mycelium plate chamber 103. A seventh microfluidic channel 209 and an eighth check valve 210 are provided between the culture medium chamber 6 and the fourth mycelium plate chamber 105. A ninth microfluidic channel 213 and a tenth check valve 214 are provided between the culture medium chamber 6 and the fifth mycelium plate chamber 107.

[0056] A second microfluidic channel 12 and a third check valve 13 are provided between the first mycelium chamber 7 and the first safety chamber 9. A fourth microfluidic channel 203 and a fifth check valve 204 are provided between the second mycelium chamber 101 and the first safety chamber 9. A sixth microfluidic channel 207 and a seventh check valve 208 are provided between the third mycelium chamber 103 and the second safety chamber 109. An eighth microfluidic channel 211 and a ninth check valve 212 are provided between the fourth mycelium chamber 105 and the third safety chamber 110. A tenth microfluidic channel 215 and an eleventh check valve 216 are provided between the fifth mycelium chamber 107 and the third safety chamber 110. A first safety conduit 14, a first safety valve 15, a first safety outlet 16, and a first safety outlet plug 17 are provided below the first safety chamber 9. A second safety conduit 217, a second safety valve 218, a second safety outlet 219, and a second safety outlet plug 220 are provided below the second safety chamber 109. Below the third safety chamber 110, a third safety pipeline 221, a third safety valve 222, a third safety outlet 223, and a third safety outlet plug 224 are provided. The culture medium injection port 3 is equipped with an antibacterial protective plug 18. The first pinhole-type bacterial strip placement slot 8 is equipped with a first antibacterial sealing ring 19. The second pinhole-type bacterial strip placement slot 102 is equipped with a second antibacterial sealing ring 111. The third pinhole-type bacterial strip placement slot 104 is equipped with a third antibacterial sealing ring 112. The fourth pinhole-type bacterial strip placement slot 106 is equipped with a fourth antibacterial sealing ring 113. The fifth pinhole-type bacterial strip placement slot 108 is equipped with a fifth antibacterial sealing ring 114.

[0057] Example 14: Preparation of a rapid sterilization effect detection chip comprising one culture medium chamber, five bacterial plate chambers, and three safety chambers. The fabrication of a rapid sterilization effect detection chip comprising one culture medium chamber, five bacterial substrate chambers, and three safety chambers includes the following steps: First, the design of the rapid sterilization effect detection chip is drawn using AutoCAD software and a photomask is fabricated for photolithography; using a four-inch single-crystal silicon wafer as a substrate, a silicon wafer mold is fabricated using photolithography, and the basic components of the chip, namely the culture medium chamber 6, the first bacterial substrate chamber 7, the second bacterial substrate chamber 101, the third bacterial substrate chamber 103, the fourth bacterial substrate chamber 105, the fifth bacterial substrate chamber 107, the first safety chamber 9, the second safety chamber 109, and the third safety chamber 110, are etched onto the photoresist. The edge lines of the culture medium chamber 6, the first mycelium plate chamber 7, the second mycelium plate chamber 101, the third mycelium plate chamber 103, the fourth mycelium plate chamber 105, the fifth mycelium plate chamber 107, the first safety chamber 9, the second safety chamber 109, and the third safety chamber 110, and the culture medium injection pipeline 4 connecting each chamber, the first microfluidic channel 10 between the culture medium chamber 6 and the first mycelium plate chamber 7, the third microfluidic channel 201 between the culture medium chamber 6 and the second mycelium plate chamber 101, the fifth microfluidic channel 205 between the culture medium chamber 6 and the third mycelium plate chamber 103, the seventh microfluidic channel 209 between the culture medium chamber 6 and the fourth mycelium plate chamber 105, the ninth microfluidic channel 213 between the culture medium chamber 6 and the fifth mycelium plate chamber 107, the second microfluidic channel 12 between the first mycelium plate chamber 7 and the first safety chamber 9, the second mycelium plate chamber 101 and the first safety chamber 110, the first microfluidic channel 12 between the first mycelium plate chamber 7 and the first safety chamber 9, the second mycelium plate chamber 101 and the first safety chamber 110, the first microfluidic channel 10 ... safety chamber 101 and the first safety chamber 110, the second safety chamber 109 and the third safety chamber 110, the second safety chamber 109 and the third safety chamber 110, the second safety chamber 109 and the third safety chamber 110, the second safety chamber The fourth microfluidic channel 203 between cavities 9, the sixth microfluidic channel 207 between the third bacterial cell cavity 103 and the second safety cavity 109, the eighth microfluidic channel 211 between the fourth bacterial cell cavity 105 and the third safety cavity 110, the tenth microfluidic channel 215 between the fifth bacterial cell cavity 107 and the third safety cavity 110, the first safety conduit 14, the second safety conduit 217, the third safety conduit 221, and the first bacterial cell cavity 9. The first pinhole-type mycelium plate placement groove 8 is placed in the cavity 7; the second mycelium plate placement groove 101 is placed in the cavity 102 of the second pinhole-type mycelium plate placement groove 102; the third mycelium plate placement groove 103 is placed in the cavity 104 of the third pinhole-type mycelium plate placement groove 104; the fourth mycelium plate placement groove 105 is placed in the cavity 106 of the fourth pinhole-type mycelium plate placement groove; and the fifth mycelium plate placement groove 107 is placed in the cavity 108 of the fifth pinhole-type mycelium plate placement groove. The second step involves casting the mixture into a mold. After casting, the first mycelium chamber 7, the second mycelium chamber 101, the third mycelium chamber 103, the fourth mycelium chamber 105, the fifth mycelium chamber 107, the first safety chamber 9, the second safety chamber 109, and the third safety chamber 110 are functionalized. A first check valve 5 is installed in the culture medium injection pipeline 4, and a second check valve 11 is installed in the first microfluidic channel 10 between the culture medium chamber 6 and the first mycelium chamber 7. The fourth check valve 202 of the third microfluidic channel 201 between the culture medium chamber 6 and the second mycelial plate chamber 101; the sixth check valve 206 of the fifth microfluidic channel 205 between the culture medium chamber 6 and the third mycelial plate chamber 103; the eighth check valve 210 of the seventh microfluidic channel 209 between the culture medium chamber 6 and the fourth mycelial plate chamber 105; the tenth check valve 214 of the ninth microfluidic channel 213 between the culture medium chamber 6 and the fifth mycelial plate chamber 107; the first mycelial plate chamber 7 and the first... The third check valve 13 of the second microfluidic channel 12 between the safety chambers 9; the fifth check valve 204 of the fourth microfluidic channel 203 between the second mycelium chamber 101 and the first safety chamber 9; the seventh check valve 208 of the sixth microfluidic channel 207 between the third mycelium chamber 103 and the second safety chamber 109; the ninth check valve 212 of the eighth microfluidic channel 211 between the fourth mycelium chamber 105 and the third safety chamber 110; and the fifth mycelium chamber 107 and the third safety chamber... The eleventh check valve 216 of the tenth microfluidic channel 215 between 110, the first safety valve 15 of the first safety line 14, the second safety valve 218 of the second safety line 217, the third safety valve 222 of the third safety line 221, the first pinhole-type bacterial tablet placement slot 8, the second pinhole-type bacterial tablet placement slot 102, the third pinhole-type bacterial tablet placement slot 104, the fourth pinhole-type bacterial tablet placement slot 106, and the fifth pinhole-type bacterial tablet placement slot 108; The third step is to prepare a cover plate 2 corresponding to the substrate 1. The cover plate 2, which does not face the substrate 1, has markings corresponding to each cavity and channel of the substrate 1. The fourth step involves using thermo-press packaging technology to bond the functionalized substrate and cover plate together to form a chip. Step 5: Install the first safety outlet antibacterial plug 17 of the first safety outlet 16, the second safety outlet antibacterial plug 220 of the second safety outlet 219, the third safety outlet antibacterial plug 224 of the third safety outlet 223, the antibacterial protective plug 18 of the culture medium injection port 3, the first antibacterial sealing ring 19 of the first pinhole bacterial tablet placement groove 8, the second antibacterial sealing ring 111 of the second pinhole bacterial tablet placement groove 102, the third antibacterial sealing ring 112 of the third pinhole bacterial tablet placement groove 104, the fourth antibacterial sealing ring 113 of the fourth pinhole bacterial tablet placement groove 106, and the fifth antibacterial sealing ring 114 of the fifth pinhole bacterial tablet placement groove 108.

[0058] Example 15: A usage example of a rapid sterilization effect detection chip comprising one culture medium chamber, five bacterial substrate chambers, and three safety chambers. The use of a rapid sterilization effect detection chip comprising one culture medium chamber, five bacterial substrate chambers, and three safety chambers includes the following steps: Step 1: Sequentially open the antibacterial protective plug 18 and the first antibacterial check valve 5 configured on the injection port 3 of the culture medium chamber 6 of the chip, and inject sterile culture medium. The sterile culture medium contains appropriate amounts of tryptone, brain and heart extract broth culture medium, fructose, glucose, potassium chloride, dipotassium hydrogen phosphate, Tween 80, L-alanine, L-lysine, L-valine, 2,6-pyridinedicarboxylic acid, calcium chloride, manganese chloride, 4-methylumbelliferyl-α-D-glucopyranoside, bromocresol purple, and sterile water. Step 1: Fill the culture medium chamber 6 with sterile culture medium to two-thirds of its height, then reset the first antibacterial check valve 5 and the antibacterial protective plug 18 in sequence. Step 2: Open the first antibacterial sealing ring 19, the second antibacterial sealing ring 111, the third antibacterial sealing ring 112, and the fourth antibacterial sealing ring 113 in sequence. Use a chuck to allow the first pinhole bacterial substrate placement slot 8, the second pinhole bacterial substrate placement slot 102, the third pinhole bacterial substrate placement slot 104, and the fourth pinhole bacterial substrate placement slot 106 to... Step 1: First, pop out the culture medium chamber 6 and place the four sterilized bacterial slides into the chamber using sterilized tweezers. Then, sequentially reset the pinhole-type bacterial slide placement slot and the corresponding antibacterial sealing ring. Step 2: Open the fifth antibacterial sealing ring 114, use a chuck to pop out the fifth pinhole-type bacterial slide placement slot 108, place the control bacterial slide into the slot using sterilized tweezers, and then sequentially reset the fifth pinhole-type bacterial slide placement slot 108 and the fifth antibacterial sealing ring 114. Step 3: Sequentially open the culture medium chamber 6 and the first bacterial slide chamber. The second check valve 11 of the first microfluidic channel 10 between the culture medium chamber 6 and the second microbial plate chamber 101, the fourth check valve 202 of the third microfluidic channel 201 between the culture medium chamber 6 and the second microbial plate chamber 101, the sixth check valve 206 of the fifth microfluidic channel 205 between the culture medium chamber 6 and the third microbial plate chamber 103, the eighth check valve 210 of the seventh microfluidic channel 209 between the culture medium chamber 6 and the fourth microbial plate chamber 105, and the ninth check valve 210 of the seventh microfluidic channel 209 between the culture medium chamber 6 and the fifth microbial plate chamber 107. The tenth check valve 214 of the microfluidic channel 213 is completely immersed in the experimental bacterial strips and control bacterial strips placed in the pinhole-type bacterial strip placement slot, and then the check valves are reset sequentially. Step 5: The chip is placed in an incubator or culture instrument at 56 degrees Celsius for 20 minutes for incubation. Fluorescence is automatically detected using a bio-automatic reader, or transferred to a fluorescence microscope and compared with the control chip using software. The test results are reported as positive (fluorescence present, i.e., "+") and negative (no fluorescence present, i.e., "-"). Result interpretation: If both the control chip and the experimental chip show "+", the sterilization is deemed unqualified; if both the control chip and the experimental chip show "+", the sterilization is deemed qualified.If a safety malfunction occurs, the corresponding safety outlet antibacterial plug and safety valve can be opened to troubleshoot the safety malfunction through the safety outlet and safety pipeline, ensuring operational safety; Step Six: For the tested chips, open the check valves of the microfluidic channels between all the bacterial chambers and the safety chambers to allow the liquid to completely enter all safety chambers. Then, sequentially open the safety outlet antibacterial plugs and safety valves, and discharge the liquid into a medical waste liquid recycling container with safety measures in place through the safety pipeline and safety outlet. Afterwards, reset all safety outlet antibacterial plugs and safety valves; Step Seven Open the antibacterial protective plug 18 and the first antibacterial check valve 5 at the injection port 3 of the chip culture medium chamber 6. Inject a 3% hydrogen peroxide aqueous solution or 0.2% peracetic acid or a disinfectant. Disinfect all culture medium chambers, bacterial cell chambers, safety chambers, and corresponding pipes and valves one by one. Place the remaining bacterial cell carriers into a medical waste liquid recycling container with safety measures in place. After disinfection and cleaning, the liquids flowing out from different safety outlets are deemed to be qualified after disinfection and sterilization. Dry them and reset all valves and disinfection and sterilization qualified protective plugs for later use.

[0059] Example 16: Detection results of sample bacterial sheets from a hydrogen peroxide plasma lumen biological sterilization device comprising one culture medium chamber, five bacterial sheet chambers, and three safety chambers, using a multi-chip rapid sterilization effect detection system. In Example 13 of this invention, a rapid sterilization effect detection chip comprising one culture medium chamber, five bacterial plate chambers, and three safety chambers showed consistent detection results of bacterial plates from a hydrogen peroxide plasma lumen biological sterilization device with the detection results of a commercially available ultra-fast biological reader's accompanying biological monitoring indicator reagent. The sterile culture medium formula is as follows: 1 g tryptone, 1 g brain heart extract broth medium, 0.3 g fructose, 0.2 g glucose, 0.1 g potassium chloride, 0.1 g dipotassium hydrogen phosphate, 0.1 g Tween 80, 5 g L-alanine, 1 g L-lysine, 1 g L-valine, 2.1 g 2,6-pyridinedicarboxylic acid, 1.4 g calcium chloride, 0.001 g manganese chloride, 0.2 g 4-methylumbelliferyl-α-D-glucopyranoside, 0.002 g bromocresol purple, and sterile water to 100 ml, adjusting the pH to 7.4-7.8. The detection method for the commercial ultra-fast biological reader with its accompanying biological monitoring indicator reagent is as follows: Place the bacterial strips that have completed one sterilization cycle into the test tube on the automatic reader's culture rack. Press the cap vertically to crush the culture medium ampoule, allowing the culture medium to flow out and submerge the bacterial strips. Incubate together with the control tube at a constant temperature of 56 degrees Celsius. Results for the control tube are available within 30 minutes, and results for the test tubes are available within 45 minutes (the reader automatically displays "+" or "-"). Result interpretation: If both the control and test tubes show "+", sterilization is considered unqualified; if both show "+", sterilization is considered qualified. However, compared to the commercial ultra-fast biological reader with its accompanying biological monitoring indicator reagent, this invention reduces the total operation time by 30 minutes because the control and four test bacterial strips are detected as a single unit, significantly improving the efficiency of the detection process. Furthermore, this invention achieves 100% accuracy in detecting negative or positive samples from 100 bacterial strips.

[0060] Example 17: A rapid sterilization effect detection chip comprising one culture medium chamber, five bacterial strip chambers, and three safety chambers demonstrates its application in detecting pressure steam-sterilized bacterial strip samples. The rapid sterilization effect detection chip prepared in Example 13 of this invention includes one culture medium chamber, five bacterial plate chambers, and three safety chambers. It rapidly detects the sterilization effect of pressure steam sterilized bacterial plate samples. The experimental chip and control chip were placed together in the culture wells of a bio-automatic reader at 57 degrees Celsius for 25 minutes. The culture medium formula was: 0.5 g tryptone, 0.5 g brain and heart extract broth, 0.2 g fructose, 0.2 g glucose, 0.05 g potassium chloride, 0.05 g dipotassium hydrogen phosphate, 0.05 g Tween 80, 0.3 g L-alanine, 0.5 g L-lysine, 0.1 g L-valine, 0.1 g 2,6-pyridinedicarboxylic acid, and CaCl2. 0.1 g of manganese chloride, 0.001 g of 4-methylumbelliferyl-α-D-glucopyranoside, 0.01 g of bromocresol purple, and sterile water were added to 100 ml to adjust the pH to 7.4-7.8. During the culture process, the bacterial cell cavity can be aligned with the fluorescence detection port of the bio-automatic reader by adjusting the chip height, and the fluorescence can be automatically detected by the bio-automatic reader. Alternatively, the sample can be transferred to a fluorescence microscope for analysis by software, comparing it with the control chip, and reporting positive (fluorescence, i.e., displaying "+") and negative (no fluorescence, i.e., displaying "-") detection results. Result interpretation: If both the control chip and the experimental chip show "+", the sterilization is deemed unqualified; if both the control chip and the experimental chip show "+", the sterilization is deemed qualified. The results show that the detection results of the rapid sterilization effect detection chip constructed in Example 13 can accurately reflect the true situation of the pressure steam sterilized samples. Compared with commercially available biomonitoring indicator reagents for pressure steam sterilization, the rapid detection chip for sterilization effect constructed in Example 13 has a detection accuracy of up to 100% for 100 pressure steam sterilized bacterial tablet samples, and the detection time is short.

[0061] Example 18: A rapid detection chip for sterilization effectiveness, comprising one culture medium chamber, five bacterial substrate chambers, and three safety chambers, is used to detect ethylene oxide sterilized bacterial substrate samples. The rapid sterilization effect detection chip prepared in Example 13 of this invention was used to rapidly detect the sterilization effect of ethylene oxide sterilized bacterial tablets according to the method of this invention. The experimental chip and the control chip were placed together in the culture well of a bio-automatic reader at 58 degrees Celsius for 30 minutes. The culture medium formula was as follows: 1.5 g tryptone, 1.5 g brain heart extract broth, 0.4 g fructose, 0.3 g glucose, 0.15 g potassium chloride, 0.1 g dipotassium hydrogen phosphate, 0.2 g Tween 80, 0.7 g L-alanine, 0.15 g L-lysine, 0.15 g L-valine, 0.25 g 2,6-pyridinedicarboxylic acid, 0.18 g calcium chloride, 0.0015 g manganese chloride, 0.03 g 4-methylumbelliferyl-α-D-glucopyranoside, 0.003 g bromocresol purple, and sterile water was added to 100 ml. The pH was adjusted to 7.4-7.8. During the cultivation process, the height of the chip can be adjusted to align the bacterial cell cavity with the fluorescence detection port of a bio-automatic reader for automatic fluorescence detection. Alternatively, the sample can be transferred to a fluorescence microscope for analysis using software, comparing it with a control chip and reporting positive (fluorescence, indicated by "+") and negative (no fluorescence, indicated by "-") results. Result interpretation: If both the control chip and the experimental chip show "+", sterilization is deemed unqualified; if both show "+", sterilization is deemed qualified. The results indicate that the rapid sterilization effect detection chip constructed in Example 13 accurately reflects the true condition of the bacterial cell samples. Compared with commercially available ethylene oxide sterilization biomonitoring indicator reagents, the rapid sterilization effect detection chip constructed in Example 13 achieves a 100% detection accuracy for 100 ethylene oxide-sterilized bacterial cell samples, with a shorter detection time.

[0062] Example 19 Based on cost price, the cost of testing each bacterial sample using the rapid sterilization effect detection chip of this invention can save an average of 13.5, 10.2, and 11.7 yuan respectively compared to commercially available sterilization biological monitoring indicator reagents for hydrogen peroxide plasma, pressure steam sterilization, and ethylene oxide sterilization. This not only benefits hospital medical insurance procurement and reuse but also saves testing fees for those being tested.

[0063] The above embodiments should be understood as being used only to illustrate the present invention more clearly, and not to limit the scope of the present invention. After reading the present invention, any modifications of the present embodiments by those skilled in the art will fall within the scope defined by the appended claims.

Claims

1. A chip for rapid detection of sterilization effect, characterized in that, The chip is composed of a substrate and a cover plate; the substrate is provided with a bacterial cell cavity, a culture medium cavity and a safety cavity, the bacterial cell cavity is provided with a pinhole bacterial cell placement groove, the culture medium cavity is provided with a culture medium injection pipeline, and the safety cavity is provided with a safety pipeline; A microfluidic channel is provided between the bacterial cell chamber and the culture medium chamber, and a microfluidic channel is provided between the bacterial cell chamber and the safety chamber; the culture medium injection line, the microfluidic channel and the safety line are all equipped with check valves.

2. The rapid sterilization effect detection chip according to claim 1, characterized in that, The cover plate of the chip is made of a mixture of PDMS and graphene, wherein the graphene content in the mixture is 4.1% by mass.

3. The rapid sterilization effect detection chip according to claim 1, characterized in that, The substrate of the chip is made of a mixture of silicon carbide and PDMS, wherein the mass content of silicon carbide in the mixture is 35%.

4. The rapid sterilization effect detection chip according to claim 1, characterized in that, The pinhole-type bacterial substrate placement slot is shaped like a pinhole-type SIM card slot.

5. The rapid sterilization effect detection chip according to claim 1, characterized in that, The number of bacterial cell chambers is 1-20, the number of culture medium chambers is 1-20, and the number of safety chambers is 1-20.

6. The rapid sterilization effect detection chip according to claim 1, characterized in that, The cross-sectional area of ​​the microfluidic channel connecting the bacterial cell chamber and the culture medium chamber on the chip substrate is 1 / 4 to 3 / 4 of the cross-sectional area of ​​the culture medium inlet channel in the culture medium chamber.

7. The rapid sterilization effect detection chip according to claim 1, characterized in that, The chip fabrication method includes the following steps: First, a silicon wafer mold is fabricated using photolithography to etch the edge lines of the bacterial cell cavity, culture medium cavity, and safety cavity, as well as the injection pipeline, microfluidic channel, safety pipeline, and pinhole bacterial cell placement slot that sequentially connect each cavity; Second, the chip substrate is formed by casting on the silicon wafer mold and then molding it. Channels are made in the bacterial cell cavity, culture medium cavity, and safety cavity, and functionalized. A pinhole bacterial cell placement slot, a check valve, and a safety valve are installed; Third, the chip substrate with the functionalized bacterial cell cavity, culture medium cavity, and safety cavity is bonded to a cover plate using bonding and packaging technology to complete the chip fabrication; Fourth, an antibacterial plug for the safety outlet, an antibacterial protective plug for the culture medium injection port, and an antibacterial sealing ring for the pinhole bacterial cell placement slot are installed.

8. The rapid sterilization effect detection chip according to claim 1, characterized in that, The method of using the chip includes the following steps: Step 1, sequentially open the antibacterial protective plug and the antibacterial check valve of the chip culture medium chamber, inject sterile culture medium until the sterile culture medium level in the culture medium chamber is two-thirds of the chamber height, and then sequentially reset the antibacterial check valve and the antibacterial protective plug of the injection port; Step 2, open the antibacterial sealing ring, use a jack to pop out the pinhole-type bacterial tablet placement slot, and place the sterilized bacterial tablet or control bacterial tablet into the slot using sterilized tweezers, and then sequentially reset the pinhole-type bacterial tablet placement slot and the antibacterial sealing ring; Step 3, open the... The check valve of the microfluidic channel between the culture medium chamber and the bacterial substrate chamber is used to completely immerse the bacterial substrate placed in the pinhole-type bacterial substrate placement slot through the sterile culture medium in the culture medium chamber, and then the check valve is reset; Step four, the chip is placed in an incubator, culture instrument or bio-automatic reader for a period of time for cultivation; Step five, the fluorescence is automatically detected by the bio-automatic reader, or transferred to a fluorescence microscope for analysis by software, compared with the control chip, and the detection results are reported as positive (fluorescence present, i.e., "+") and negative (no fluorescence present, i.e., "-"), and the result is determined as follows: control If the control chip displays "+" and the experimental chip displays "+", sterilization is deemed unqualified; if the control chip displays "+" and the experimental chip displays "-", sterilization is deemed qualified. Step six: If a safety malfunction occurs with the tested chip, the safety outlet antibacterial plug and safety valve can be opened to troubleshoot the malfunction through the safety outlet and safety pipeline, ensuring operational safety. Step seven: For the tested chip, open the check valve of the microfluidic channel between the bacterial chamber and the safety chamber to allow liquid to completely enter the safety chamber, then sequentially open the safety outlet antibacterial plug. The liquid is discharged into a medical waste liquid recycling container with safety measures in place through safety pipelines and safety outlets. Then, the antibacterial plug and safety valve of the safety outlet are reset. Step eight: Open the antibacterial protective plug and antibacterial check valve of the inlet of the chip culture medium chamber, inject medical disinfectant, and disinfect the culture medium chamber, the chip chamber, the safety chamber and the corresponding pipelines and valves one by one. After disinfection and cleaning, the liquid flowing out of the safety outlet is judged to be qualified after disinfection and sterilization. It is then dried, and each valve and the qualified disinfection and sterilization protective plug are reset for later use.

Images