Staphylococcus aureus sensing wound dressing system

Abstract

This application relates to the field of medical dressings and wound monitoring technology, and discloses a Staphylococcus aureus sensing wound dressing system, including a sensing hydrogel layer, a fluorescence excitation component, a macro fluorescence imaging component, and an image processing unit. The sensing hydrogel layer includes a hydrogel matrix, an aptamer fluorescent probe, and a quencher chain. The sensing hydrogel layer is used to adhere to the external wound surface. The aptamer fluorescent probe is fixed in the hydrogel network of the hydrogel matrix. The aptamer fluorescent probe contains a fluorescent group, and the quencher chain is in a partially complementary hybridized connection state with the aptamer fluorescent probe. This invention utilizes the Acrydite group at the 5′ end of the aptamer fluorescent probe to participate in the hydrogel cross-linking reaction, anchoring it covalently within a hydrogel network composed of polyethylene glycol diacrylate and acrylamide. This structure can effectively prevent the probe from diffusing and being lost with exudate in a moist wound environment, ensuring the long-term stability of the probe concentration and detection signal in the sensing layer.

Description

Technical Field

[0001] This invention relates to the field of medical dressings and wound monitoring technology, specifically a Staphylococcus aureus sensing wound dressing system. Background Technology

[0002] Burns and chronic wounds disrupt the barrier function, often resulting in significant fluid exudation and making the wounds highly susceptible to infection by pathogens such as Staphylococcus aureus. To manage and monitor wound conditions, hydrogel materials, with their excellent water-containing physical properties and biocompatibility, are widely used in the development of wound dressings. Introducing various biosensor probes into the hydrogel matrix to construct intelligent dressings capable of real-time monitoring of changes in microenvironment biochemical indicators has become a current development direction in the field of wound diagnosis and treatment technology.

[0003] Existing monitoring hydrogel dressings mainly construct a sensing network by embedding specific sequences of nucleic acid aptamers or fluorescent molecules inside the hydrogel. Healthcare workers apply the probe-loaded hydrogel dressing to the patient's wound surface. The hydrogel continuously absorbs wound exudate. When specific target molecules or bacteria in the exudate enter the hydrogel and come into contact with the internally loaded sensing probes, the sensing probes bind and generate changes in optical signals. External photoelectric detection equipment then irradiates the hydrogel area and acquires images, extracting the corresponding optical signals for medical personnel to assess the current state of the wound.

[0004] Existing monitoring hydrogel dressings often use physical doping or physical adsorption to distribute probe molecules within the hydrogel network during the preparation process. However, when faced with moist wound environments such as burns that produce a large amount of exudate, the probe molecules that have not formed stable chemical bonds inside the hydrogel are prone to diffusion and loss to the outside with the exchange and flow of the liquid. The passive loss of probe molecules will lead to a continuous decrease in the effective probe concentration inside the dressing, causing distortion of the optical detection signal in the later stage. At the same time, the free probes entering the fragile wound tissue at the bottom layer may also trigger unknown reactions. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides a Staphylococcus aureus sensing wound dressing system, which solves the problems of probe molecules in monitoring hydrogel dressings easily diffusing and being lost in a moist wound environment, leading to distortion of detection signals and posing biosafety risks.

[0006] To achieve the above objectives, the present invention is implemented through the following technical solution: a Staphylococcus aureus sensing wound dressing system, comprising a sensing hydrogel layer, a fluorescence excitation component, a macro fluorescence imaging component, and an image processing unit; The sensing hydrogel layer includes a hydrogel matrix, an aptamer fluorescent probe, and a quenching chain. The sensing hydrogel layer is used to adhere to the surface of an external wound. The aptamer fluorescent probe is fixed in the hydrogel network of the hydrogel matrix. The aptamer fluorescent probe contains a fluorescent group. The quenching chain and the aptamer fluorescent probe are in a partially complementary hybridization connection state. The binding of Staphylococcus aureus to the aptamer fluorescent probe changes the hybridization state or spatial distance between the aptamer fluorescent probe and the quenching chain, thereby restoring the fluorescence signal of the aptamer fluorescent probe. The luminescent surface of the fluorescence excitation component faces the sensing hydrogel layer, and the incident light side of the macro fluorescence imaging component faces the sensing hydrogel layer. The fluorescence excitation component and the macro fluorescence imaging component form a fluorescence excitation light path and a collection light path for the sensing hydrogel layer in space. The image processing unit is communicatively connected to the macro fluorescence imaging component. The image processing unit receives the two-dimensional fluorescence image with fluorescence signal intensity and spatial distribution information output by the macro fluorescence imaging component, performs calculations and processing, and generates and outputs Staphylococcus aureus-related fluorescence response results.

[0007] Preferably, the aptamer fluorescent probe includes a polymeric linker group, a spacer sequence, a Staphylococcus aureus aptamer sequence, and a Cy3 fluorescent group. The polymeric linker group is disposed at the 5′ end of the aptamer fluorescent probe, the spacer sequence is connected between the polymeric linker group and the Staphylococcus aureus aptamer sequence, and the Cy3 fluorescent group is connected to the 3′ end of the Staphylococcus aureus aptamer sequence. The quenching chain includes a quenching group and short DNA chains Q1-7. The quenching group is attached to the 5′ end of the short DNA chains Q1-7. The quenching group is BHQ2. The short DNA chains Q1-7 are short chains that are complementary to the continuous region at the 3′ end of the Staphylococcus aureus aptamer sequence in the aptamer fluorescent probe.

[0008] Preferably, the polymeric linker is Acrydite, the spacer sequence is a sequence composed of consecutive thymine deoxyribonucleotides, and the base arrangement of the Staphylococcus aureus aptamer sequence is 5′-TCGGCACGTTCTCAGTAGCGCTCGCTGGTCATCCCACAGCTACGTC-3′. The base sequence of the short DNA chain Q1-7 is 5′-GACGTAG-3′; The aptamer fluorescent probe participates in the cross-linking process of the hydrogel matrix through the 5′ end of Acrydite, and is fixed in the hydrogel network of the hydrogel matrix by covalent bonding.

[0009] Preferably, the hydrogel matrix is ​​formulated from polyethylene glycol diacrylate, acrylamide, and carboxymethyl chitosan. The polyethylene glycol diacrylate and acrylamide undergo a polymerization reaction under the action of a photoinitiator to form a hydrogel network, and the carboxymethyl chitosan is interspersed within the hydrogel network.

[0010] Preferably, both the fluorescence excitation component and the macro fluorescence imaging component are integrated inside the encapsulation substrate; The fluorescent excitation component includes a green LED and an excitation filter. The excitation filter is disposed on the light-emitting side of the green LED. The light emitted by the green LED passes through the excitation filter to form excitation light. An excitation optical window is disposed inside the encapsulation substrate at the position corresponding to the optical path of the excitation light. The macro fluorescence imaging component includes an emission filter, a macro lens, and an image sensor. The macro lens is disposed on the light-incident side of the image sensor, the emission filter is disposed on the light-incident side of the macro lens, and an imaging optical window is disposed inside the encapsulation substrate corresponding to the optical path position of the macro fluorescence imaging component.

[0011] Preferably, the excitation filter is a 530nm narrowband excitation filter, and the emission filter is a 585nm emission filter; The packaging substrate has a light-shielding structure located outside the excitation optical window and the imaging optical window. The light-shielding structure includes a light shield or a dark cavity structure. The Staphylococcus aureus sensing wound dressing system also includes a microcontroller unit and a wireless communication module, both of which are integrated inside the encapsulation substrate.

[0012] Preferably, after acquiring the two-dimensional fluorescence image, the image processing unit determines the region of interest corresponding to the sensing hydrogel layer in the two-dimensional fluorescence image, extracts the red channel data corresponding to the Cy3 fluorescent group in the region of interest as the original gray-scale intensity matrix, and retrieves the baseline dark-field image matrix under the condition of no Staphylococcus aureus. The image processing unit performs pixel-by-pixel subtraction between the original grayscale intensity matrix and the baseline dark field image matrix to perform background subtraction and brightness normalization processing, thereby obtaining a normalized effective fluorescence intensity matrix.

[0013] Preferably, the image processing unit calculates the average fluorescence intensity and the maximum fluorescence intensity within the region of interest based on the normalized effective fluorescence intensity matrix; The image processing unit retrieves a pre-stored reference high signal determination threshold, compares the values ​​in the normalized effective fluorescence intensity matrix with the reference high signal determination threshold to construct an indicator function, and calculates the area ratio of the high signal region and extracts the spatial distribution parameters of the high signal region based on the indicator function. The image processing unit obtains the average fluorescence intensity, maximum fluorescence intensity, high signal area ratio, and spatial distribution parameters, and then combines these with the time point of the two-dimensional fluorescence image to generate data on the changes of the average fluorescence intensity, maximum fluorescence intensity, high signal area ratio, and spatial distribution parameters over time.

[0014] Preferably, the Staphylococcus aureus sensing wound dressing system further includes a therapeutic hydrogel region, an electrode assembly, and an electrical stimulation driving module. The therapeutic hydrogel region is disposed on the outer surface of the sensing hydrogel layer or arranged adjacent to the sensing hydrogel layer, and the therapeutic hydrogel region is loaded with lysostaphin. The electrode assembly is disposed on the surface of the therapeutic hydrogel area or extends into the interior of the therapeutic hydrogel area. The electrode assembly is electrically connected to the electrical stimulation driving module, and the electrical stimulation driving module is communicatively connected to the image processing unit.

[0015] Preferably, the image processing unit compares the generated Staphylococcus aureus-related fluorescence response results with preset conditions, and outputs a control signal to the electrical stimulation driving module when the Staphylococcus aureus-related fluorescence response results meet the preset conditions; After receiving the control signal, the electrical stimulation drive module starts the preset electrical stimulation program and applies an electrical signal with a preset duration, preset voltage, or preset pulse parameters to the treatment hydrogel area through the electrode assembly, so that the lysostaphin enzyme loaded in the treatment hydrogel area is released under the action of the electric field.

[0016] This invention provides a Staphylococcus aureus-sensing wound dressing system. It has the following beneficial effects: 1. This invention utilizes an aptamer fluorescent probe that participates in the cross-linking reaction of the hydrogel matrix via the 5' end of its Acrydite, covalently fixing it within the hydrogel network formed by polyethylene glycol diacrylate and acrylamide. This covalent connection structure anchors the probe molecules within the hydrogel matrix, preventing the probe from being lost and diffused outward with exudate in the moist, fluid-rich wound environment, thus ensuring the structural stability and probe concentration of the sensing hydrogel layer during continuous operation.

[0017] 2. This invention utilizes the initial hybridization quenching state formed by the Staphylococcus aureus aptamer sequence and a short DNA strand containing BHQ2. The competitive binding force generated upon the appearance of the target bacteria drives the dissociation of the short DNA strand, increasing the physical distance between the fluorescent group and the quenching group to restore the fluorescence signal. This chain displacement process directly translates the specific binding of pathogens at the molecular level into macroscopic optical changes, enabling targeted monitoring of specific bacterial types rather than broad environmental indicators using dressings.

[0018] 3. This invention constructs a near-field area array optical acquisition path using a fluorescence excitation component and a micro-fluorescence imaging component. The image processing unit performs region of interest extraction, background subtraction, and calculates the area ratio of high-signal regions on the received two-dimensional fluorescence image. The area array image-based acquisition and calculation process breaks through the sampling range limitations of single-point probe detection, enabling the extraction and output of spatial distribution parameters of fluorescence signals covering the entire dressing area, providing an analytical basis for assessing the adhesion and colonization status of bacteria at different wound locations. Attached Figure Description

[0019] Figure 1 This is a schematic diagram illustrating the aptamer chain displacement sensing and fluorescence emission principle of the present invention; Figure 2 This is a schematic diagram of the overall system structure and module connections of the present invention; Figure 3 This is a flowchart of the image processing and response algorithm of the present invention.

[0020] Among them, 100 is the sensing hydrogel layer; 110 is the hydrogel matrix; 120 is the aptamer fluorescent probe; 130 is the quenching chain; 200 is the fluorescence excitation component; 210 is the green LED; 220 is the excitation filter; 300 is the macro fluorescence imaging component; 310 is the emission filter; 320 is the macro lens; 330 is the image sensor; 340 is the encapsulation substrate; 350 is the microcontroller unit; 360 is the wireless communication module; 370 is the excitation optical window; 380 is the imaging optical window; 390 is the light-shielding structure; 400 is the image processing unit; 500 is the therapeutic hydrogel area; 600 is the electrode assembly; and 700 is the electrical stimulation driving module. Detailed Implementation

[0021] The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. 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.

[0022] Please see the appendix Figure 1 -Appendix Figure 3 The invention provides a Staphylococcus aureus sensing wound dressing system, including a sensing hydrogel layer 100, a fluorescence excitation component 200, a macro fluorescence imaging component 300, and an image processing unit 400.

[0023] The sensing hydrogel layer 100 is used to adhere to the surface of the external wound. The sensing hydrogel layer 100 includes a hydrogel matrix 110, an aptamer fluorescent probe 120, and a quenching chain 130. The aptamer fluorescent probe 120 is fixed in the three-dimensional network of the hydrogel matrix 110 and contains a Cy3 fluorescent group. The quenching chain 130 and the aptamer fluorescent probe 120 are in a partially complementary hybrid connection state. The fluorescence excitation component 200 is disposed on the side of the sensing hydrogel layer 100 away from the external wound, with the emitting surface of the fluorescence excitation component 200 facing the sensing hydrogel layer 100. The micro-fluorescence imaging component 300 is arranged adjacent to the fluorescence excitation component 200, with the incident light side of the micro-fluorescence imaging component 300 facing the sensing hydrogel layer 100. The fluorescence excitation component 200 and the micro-fluorescence imaging component 300 spatially constitute the fluorescence excitation light path and the acquisition light path for the sensing hydrogel layer 100.

[0024] The image processing unit 400 is communicatively connected to the macro fluorescence imaging component 300. The image processing unit 400 receives the image signal output by the macro fluorescence imaging component 300 and performs calculations. In embodiments including responsive treatment, the Staphylococcus aureus sensing wound dressing system further includes a therapeutic hydrogel region 500, an electrode assembly 600, and an electrical stimulation driving module 700. The therapeutic hydrogel region 500 is disposed on the outer surface of the sensing hydrogel layer 100 or arranged adjacent to the sensing hydrogel layer 100. The therapeutic hydrogel region 500 is loaded with lysostaphin. The electrode assembly 600 is disposed on the surface of the therapeutic hydrogel region 500 or extends into the therapeutic hydrogel region 500. The electrode assembly 600 is electrically connected to the electrical stimulation driving module 700, and the electrical stimulation driving module 700 is communicatively connected to the image processing unit 400.

[0025] After the sensing hydrogel layer 100 is attached to the external wound surface, in the absence of Staphylococcus aureus, the quenching chain 130 and the aptamer fluorescent probe 120 maintain a hybridization state. The quenching chain 130 quenches the Cy3 fluorescent group of the aptamer fluorescent probe 120, resulting in a low fluorescence signal of the aptamer fluorescent probe 120. When Staphylococcus aureus is present in the external wound surface, the Staphylococcus aureus enters the sensing hydrogel layer 100 and binds to the aptamer fluorescent probe 120. The binding of 20 alters the hybridization state or spatial distance between the aptamer fluorescent probe 120 and the quencher chain 130, restoring the fluorescence signal of the aptamer fluorescent probe 120. The fluorescence excitation component 200 is activated and emits excitation light to the sensing hydrogel layer 100. The macro fluorescence imaging component 300 performs two-dimensional imaging of the sensing hydrogel layer 100 at a predetermined working distance, acquiring a two-dimensional fluorescence image with Cy3 fluorescence signal intensity and spatial distribution information. The macro fluorescence imaging component 300 sends the two-dimensional fluorescence image to the image processing unit 400.

[0026] Image processing unit 400 acquires a two-dimensional fluorescence image, determines the region of interest (ROI) corresponding to the sensing hydrogel layer 100, performs background subtraction and brightness normalization on the ROI, calculates the average fluorescence intensity, maximum fluorescence intensity, high-signal area ratio, and spatial distribution parameters within the ROI, and generates and outputs Staphylococcus aureus-related fluorescence response results based on the time-varying data of the average fluorescence intensity, maximum fluorescence intensity, high-signal area ratio, and spatial distribution parameters. In embodiments including the therapeutic hydrogel region 500, image processing unit 400 will generate... The Staphylococcus aureus-related fluorescence response results are compared with preset conditions. When the Staphylococcus aureus-related fluorescence response results meet the preset conditions, the image processing unit 400 outputs a control signal to the electrical stimulation driving module 700. After receiving the control signal, the electrical stimulation driving module 700 starts the preset electrical stimulation program. The electrical stimulation driving module 700 applies an electrical signal with a preset voltage and preset duration to the treatment hydrogel region 500 through the electrode assembly 600. The lysostaphin enzyme loaded in the treatment hydrogel region 500 is released under the action of the electric field. After the electrical stimulation program ends, the image processing unit 400 controls the micro-fluorescence imaging component 300 to re-acquire the two-dimensional fluorescence image of the sensing hydrogel layer 100.

[0027] The present invention provides a Staphylococcus aureus sensing wound dressing system whose workflow may include: a sensing hydrogel layer 100 is attached to the surface of an external wound; when Staphylococcus aureus is present in the external wound, the Staphylococcus aureus enters the hydrogel matrix 110 of the sensing hydrogel layer 100; the Staphylococcus aureus binds to the aptamer fluorescent probe 120 immobilized in the hydrogel matrix 110; the binding of Staphylococcus aureus to the aptamer fluorescent probe 120 changes the hybridization state or spatial distance between the aptamer fluorescent probe 120 and the quencher chain 130; and the fluorescence signal of the aptamer fluorescent probe 120 is restored.

[0028] The fluorescence excitation component 200 is activated and emits excitation light to the sensing hydrogel layer 100. The macro fluorescence imaging component 300 performs two-dimensional imaging of the sensing hydrogel layer 100 at a predetermined working distance. The macro fluorescence imaging component 300 acquires a two-dimensional fluorescence image with Cy3 fluorescence signal intensity and spatial distribution information. The macro fluorescence imaging component 300 transmits the two-dimensional fluorescence image to the image processing unit 400.

[0029] The image processing unit 400 receives a two-dimensional fluorescence image, determines the region of interest corresponding to the sensing hydrogel layer 100, performs background subtraction and brightness normalization on the region of interest, calculates the average fluorescence intensity, maximum fluorescence intensity, high signal area ratio, and spatial distribution parameters within the region of interest, and generates and outputs Staphylococcus aureus-related fluorescence response results based on the changes in average fluorescence intensity, maximum fluorescence intensity, high signal area ratio, and spatial distribution parameters over time.

[0030] In an embodiment including the therapeutic hydrogel region 500, the image processing unit 400 compares the Staphylococcus aureus-related fluorescence response results with preset conditions. When the Staphylococcus aureus-related fluorescence response results meet the preset conditions, the image processing unit 400 outputs a control signal to the electrical stimulation driving module 700. After receiving the control signal, the electrical stimulation driving module 700 starts an electrical stimulation program with a preset value. The electrical stimulation driving module 700 applies an electrical signal with a preset voltage and a preset duration to the therapeutic hydrogel region 500 through the electrode assembly 600. The lysostaphin loaded in the therapeutic hydrogel region 500 is released under the action of the electric field. After the electrical stimulation program ends, the image processing unit 400 controls the macro fluorescence imaging component 300 to re-acquire the two-dimensional fluorescence image of the sensing hydrogel layer 100.

[0031] The sensing hydrogel layer 100 may include a hydrogel matrix 110, an aptamer fluorescent probe 120, and a quenching chain 130. The hydrogel matrix 110 constitutes a hydrogel network of the sensing hydrogel layer 100. The aptamer fluorescent probe 120 is fixed in the hydrogel network of the hydrogel matrix 110. The quenching chain 130 hybridizes with the corresponding sequence portion of the aptamer fluorescent probe 120.

[0032] The hydrogel matrix 110 is formulated from polyethylene glycol diacrylate, acrylamide and carboxymethyl chitosan. The polyethylene glycol diacrylate and acrylamide undergo a polymerization reaction under the action of a photoinitiator to form a hydrogel network. Carboxymethyl chitosan is interspersed in the hydrogel network. The hydrogel matrix 110 provides an adhesion basis for the aptamer fluorescent probe 120 and maintains structural stability and optical transparency in a water-containing environment.

[0033] The aptamer fluorescent probe 120 includes a polymeric linker group, a spacer sequence, a Staphylococcus aureus aptamer sequence, and a Cy3 fluorescent group. The polymeric linker group is located at the 5′ end of the aptamer fluorescent probe 120. The spacer sequence is connected between the polymeric linker group and the Staphylococcus aureus aptamer sequence. The Cy3 fluorescent group is connected to the 3′ end of the Staphylococcus aureus aptamer sequence. The polymeric linker group is Acrydite. The spacer sequence is a sequence composed of consecutive thymine deoxyribonucleotides. The base arrangement of the Staphylococcus aureus aptamer sequence is 5′-TCGGCACGTTCTCAGTAGCGCTCGCTGGTCATCCCACAGCTACGTC-3′. The sequence of the aptamer fluorescent probe 120 is 5′-Acrydite-TTTTTTT-TCGGCACGTTCTCAGTAGCGCTCGCTGGTCATCCCACAGCTACGTC-Cy3-3′.

[0034] The aptamer fluorescent probe 120 participates in the cross-linking process of the hydrogel matrix 110 through the 5′ end of Acrydite. Acrydite undergoes a copolymerization reaction with polyethylene glycol diacrylate and acrylamide. The aptamer fluorescent probe 120 is fixed in the hydrogel network of the hydrogel matrix 110 by covalent bonding. The covalent bonding structure anchors the aptamer fluorescent probe 120 inside the hydrogel matrix 110 to reduce the risk of the aptamer fluorescent probe 120 leaking outward.

[0035] Quenching strand 130 includes a quenching group and a short DNA strand Q1-7. The quenching group is attached to the 5′ end of the short DNA strand Q1-7. The quenching group is BHQ2. The short DNA strand Q1-7 is a short strand complementary to the continuous region at the 3′ end of the Staphylococcus aureus aptamer sequence in the aptamer fluorescent probe 120. The base arrangement of the short DNA strand Q1-7 is 5′-GACGTAG-3′, and the sequence of quenching strand 130 is 5′-BHQ2-GACGTAG-3′. In the absence of Staphylococcus aureus, the short DNA strand Q1-7 of quenching strand 130 hybridizes with the Staphylococcus aureus aptamer sequence, thereby bringing BHQ2 closer to the Cy3 fluorophore and reducing the fluorescence signal of the Cy3 fluorophore.

[0036] The aptamer chain displacement sensing mechanism includes the initial hybridization quenching process between quenched chain 130 and aptamer fluorescent probe 120, as well as the chain displacement fluorescence recovery process triggered by Staphylococcus aureus.

[0037] In the absence of Staphylococcus aureus, the short DNA strand Q1-7 of quenched strand 130 maintains hybridization with the continuous 3′ region of the aptamer fluorescent probe 120. The complementary base pairing between the short DNA strand Q1-7 and the aptamer fluorescent probe 120 causes BHQ2 of quenched strand 130 to be close to the Cy3 fluorescent group of the aptamer fluorescent probe 120. The spatial distance between BHQ2 and the Cy3 fluorescent group is within the effective quenching range. BHQ2 absorbs the energy emitted by the Cy3 fluorescent group, thereby reducing the initial fluorescence signal of the sensing hydrogel layer 100.

[0038] When Staphylococcus aureus from the external wound enters the sensing hydrogel layer 100, it binds to the aptamer fluorescent probe 120. The binding force between Staphylococcus aureus and the aptamer fluorescent probe 120 is greater than the hybridization force between the short DNA strand Q1-7 and the aptamer fluorescent probe 120. The binding of Staphylococcus aureus drives a conformational change in the aptamer fluorescent probe 120. This conformational change alters the hybridization state between the aptamer fluorescent probe 120 and the quencher strand 130. The conformational change causes the short DNA strand Q1-7 to dissociate from the continuous 3′ end region of the aptamer fluorescent probe 120. The dissociation of the quencher strand 130 increases the spatial distance between the BHQ2 and Cy3 fluorescent groups. The Cy3 fluorescent group leaves the quenching range of BHQ2 and resumes its fluorescence emission state, thereby generating a fluorescence signal.

[0039] The preparation method of the sensing hydrogel layer 100 may include preparing a prepolymer solution for the hydrogel matrix 110, mixing and stirring 3 wt% polyethylene glycol diacrylate, 15 wt% acrylamide and 1 wt% carboxymethyl chitosan at room temperature and in the dark until homogeneous to obtain the prepolymer solution, and adding 0.1 wt% phenyl (2,4,6-trimethylbenzoyl) lithium phosphate as a photoinitiator to the prepolymer solution.

[0040] The aptamer fluorescent probe 120 and the quencher chain 130 were added to a phosphate buffer containing 0.9% sodium chloride at a molar ratio of 1:1.5 to 1:2. The phosphate buffer containing the aptamer fluorescent probe 120 and the quencher chain 130 was kept at 70°C for 5 minutes and then allowed to cool naturally. The cooled phosphate buffer was placed at 4°C to allow the aptamer fluorescent probe 120 and the quencher chain 130 to complete the hybridization annealing process through base complementary pairing to form a composite probe.

[0041] The hybridized annealed composite probe was added to the prepolymer solution and mixed evenly. The prepolymer solution containing the composite probe was injected into the mold and irradiated under a 365nm ultraviolet light source for 1 minute to carry out photocrosslinking polymerization. During the photocrosslinking polymerization process, the aptamer fluorescent probe 120's Acrydite participated in the polymerization reaction of polyethylene glycol diacrylate and acrylamide, thereby fixing the aptamer fluorescent probe 120 in the hydrogel network of the hydrogel matrix 110. After the photocrosslinking polymerization was completed, the hydrogel matrix 110 was eluted and equilibrated using phosphate buffer to remove unfixed free probes and unreacted polyethylene glycol diacrylate and acrylamide to finally obtain the sensing hydrogel layer 100.

[0042] In another embodiment of the preparation of the sensing hydrogel layer 100, the aptamer fluorescent probe 120 that has not hybridized with the quencher chain 130 can be directly added to the prepolymer solution for photocrosslinking polymerization under ultraviolet light irradiation. After the aptamer fluorescent probe 120 is fixed to the hydrogel network of the hydrogel matrix 110, the hydrogel matrix 110 is immersed in phosphate buffer containing the quencher chain 130. During the immersion process, the quencher chain 130 enters the interior of the hydrogel matrix 110 and hybridizes with the aptamer fluorescent probe 120 to obtain the sensing hydrogel layer 100.

[0043] The structure of the fluorescence excitation component 200 and the macro fluorescence imaging component 300 may include an encapsulation substrate 340, a green LED 210, an excitation filter 220, an emission filter 310, a macro lens 320, an image sensor 330, a microcontroller unit 350, and a wireless communication module 360. The encapsulation substrate 340 is made of polydimethylsiloxane. The green LED 210 and the excitation filter 220 constitute the fluorescence excitation component 200. The emission filter 310, the macro lens 320, and the image sensor 330 constitute the macro fluorescence imaging component 300. The fluorescence excitation component 200, the macro fluorescence imaging component 300, the microcontroller unit 350, and the wireless communication module 360 ​​are all integrated inside the encapsulation substrate 340.

[0044] An excitation filter 220 is disposed on the light-emitting side of the green LED 210. The excitation filter 220 is a 530nm narrowband excitation filter. The light emitted by the green LED 210 passes through the excitation filter 220 to form excitation light. An excitation optical window 370 is disposed inside the encapsulation substrate 340 corresponding to the optical path position of the excitation light. The excitation light passes through the excitation optical window 370 and irradiates the sensing hydrogel layer 100 to excite the Cy3 fluorescent group in the sensing hydrogel layer 100.

[0045] The macro lens 320 is located on the light-incident side of the image sensor 330, and the emission filter 310 is located on the light-incident side of the macro lens 320. The image sensor 330 is an OV2640, and the emission filter 310 is a 585nm emission filter. An imaging optical window 380 is provided inside the encapsulation substrate 340 corresponding to the optical path position of the macro fluorescence imaging component 300. The fluorescence signal emitted by the Cy3 fluorescent group in the sensing hydrogel layer 100 passes through the imaging optical window 380 and the emission filter 310 to enter the macro lens 320. The macro lens 320 focuses the fluorescence signal onto the surface of the image sensor 330.

[0046] The packaging substrate 340 has a light-shielding structure 390 located outside the excitation optical window 370 and the imaging optical window 380. The light-shielding structure 390 includes a light shield or a dark cavity structure. The light-shielding structure 390 blocks ambient light from entering the imaging optical window 380 and blocks the excitation light from leaking directly to the macro fluorescence imaging component 300. The microcontroller unit 350 adopts an ESP32. The microcontroller unit 350 is electrically connected to the green LED 210, the image sensor 330 and the wireless communication module 360. The microcontroller unit 350 controls the opening and closing of the green LED 210 and the acquisition parameters of the image sensor 330. The wireless communication module 360 ​​transmits the two-dimensional fluorescence image acquired by the image sensor 330 to the image processing unit 400.

[0047] The present invention provides an image processing hardware and communication structure that may include a microcontroller unit 350, a wireless communication module 360, and an image processing unit 400. The microcontroller unit 350 is electrically connected to the wireless communication module 360, and the image processing unit 400 is communicatively connected to the microcontroller unit 350 through the wireless communication module 360.

[0048] The microcontroller unit 350 acquires the two-dimensional fluorescence image collected by the macro fluorescence imaging component 300. The microcontroller unit 350 transmits the two-dimensional fluorescence image to the wireless communication module 360. The wireless communication module 360 ​​sends the two-dimensional fluorescence image to the image processing unit 400. The image processing unit 400 receives the two-dimensional fluorescence image sent by the wireless communication module 360. The image processing unit 400 saves the two-dimensional fluorescence image as the basic image data for subsequent calculation of average fluorescence intensity, maximum fluorescence intensity, high signal area ratio, and spatial distribution parameters. The image processing unit 400 provides a physical computing environment for region of interest extraction and brightness normalization processing.

[0049] The fluorescence image processing and response algorithm executed by the image processing unit 400 may include steps such as acquiring a two-dimensional fluorescence image and calculating the fluorescence response results related to Staphylococcus aureus. After acquiring the two-dimensional fluorescence image, the image processing unit 400 determines the region of interest (ROI) corresponding to the sensing hydrogel layer 100 in the two-dimensional fluorescence image. The image processing unit 400 extracts the red channel data corresponding to the Cy3 fluorophore within the ROI as the original grayscale intensity matrix. The image processing unit 400 retrieves the baseline dark-field image matrix under Staphylococcus aureus-free conditions. Let the total number of pixels contained in the ROI be... The spatial coordinates of the pixels are marked as The original gray-level intensity matrix is ​​denoted as The baseline dark field image matrix is ​​denoted as The image processing unit 400 performs pixel-by-pixel subtraction between the original grayscale intensity matrix and the baseline dark field image matrix to perform background subtraction and brightness normalization. The image processing unit 400 obtains the normalized effective fluorescence intensity matrix through background subtraction and brightness normalization. The normalized effective fluorescence intensity matrix is ​​denoted as... .

[0050] The formula for calculating the normalized effective fluorescence intensity matrix is:

[0051] Image processing unit 400 calculates the average fluorescence intensity and maximum fluorescence intensity within the region of interest based on the normalized effective fluorescence intensity matrix. The average fluorescence intensity is denoted as... The formula for calculating the average fluorescence intensity is:

[0052] In the formula This represents the summation of the normalized effective fluorescence intensity values ​​corresponding to all pixels within the region of interest, with the maximum fluorescence intensity denoted as . The maximum fluorescence intensity is the normalized effective fluorescence intensity matrix. The maximum value in.

[0053] The image processing unit 400 retrieves a pre-stored reference height signal determination threshold, which is denoted as: The image processing unit 400 compares the values ​​in the normalized effective fluorescence intensity matrix with the reference high signal determination threshold to construct an indicator function, denoted as . The construction logic of the indicator function is as follows:

[0054] Image processing unit 400 calculates the area ratio of high-signal regions and extracts the spatial distribution parameters of high-signal regions based on the indicator function. The area ratio of high-signal regions is denoted as... .

[0055] The formula for calculating the area ratio of the high-signal region is:

[0056] After acquiring the average fluorescence intensity, maximum fluorescence intensity, high signal area ratio, and spatial distribution parameters, the image processing unit 400 combines the time node of the acquired two-dimensional fluorescence image to generate data on the changes of the average fluorescence intensity, maximum fluorescence intensity, high signal area ratio, and spatial distribution parameters over time. The image processing unit 400 outputs Staphylococcus aureus-related fluorescence response results containing the changes as the basis for determining the preset conditions for triggering the electrical stimulation drive module 700.

[0057] The present invention provides a Staphylococcus aureus sensing wound dressing system, which may further include a therapeutic hydrogel region 500, an electrode assembly 600, and an electrical stimulation driving module 700. The therapeutic hydrogel region 500 is disposed on the outer surface of the sensing hydrogel layer 100 or arranged adjacent to the sensing hydrogel layer 100. The therapeutic hydrogel region 500 and the sensing hydrogel layer 100 are different regions of the same continuous gel, or the therapeutic hydrogel region 500 and the sensing hydrogel layer 100 are two hydrogel regions that are spatially separated and ionically connected. The therapeutic hydrogel region 500 is loaded with lysostaphin. The electrode assembly 600 is disposed on the surface of the therapeutic hydrogel region 500 or extends into the interior of the therapeutic hydrogel region 500. The electrode assembly 600 is electrically connected to the electrical stimulation driving module 700, and the electrical stimulation driving module 700 is communicatively connected to the image processing unit 400.

[0058] The image processing unit 400 compares the generated Staphylococcus aureus-related fluorescence response results with preset conditions. When the Staphylococcus aureus-related fluorescence response results meet the preset conditions, the image processing unit 400 outputs a control signal to the electrical stimulation driving module 700. After receiving the control signal, the electrical stimulation driving module 700 starts the preset electrical stimulation program. The electrical stimulation driving module 700 applies an electrical signal with a preset duration, preset voltage, or preset pulse parameters to the treatment hydrogel region 500 through the electrode assembly 600. The lysostaphin enzyme loaded in the treatment hydrogel region 500 is released under the action of the electric field. After the electrical stimulation program ends, the image processing unit 400 controls the micro-fluorescence imaging component 300 to re-acquire the two-dimensional fluorescence image of the sensing hydrogel layer 100.

[0059] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A Staphylococcus aureus-sensing wound dressing system, characterized in that, It includes a sensing hydrogel layer, a fluorescence excitation component, a macro fluorescence imaging component, and an image processing unit; The sensing hydrogel layer includes a hydrogel matrix, an aptamer fluorescent probe, and a quenching chain. The sensing hydrogel layer is used to adhere to the surface of an external wound. The aptamer fluorescent probe is fixed in the hydrogel network of the hydrogel matrix. The aptamer fluorescent probe contains a fluorescent group. The quenching chain and the aptamer fluorescent probe are in a partially complementary hybridization connection state. The binding of Staphylococcus aureus to the aptamer fluorescent probe changes the hybridization state or spatial distance between the aptamer fluorescent probe and the quenching chain, thereby restoring the fluorescence signal of the aptamer fluorescent probe. The luminescent surface of the fluorescence excitation component faces the sensing hydrogel layer, and the incident light side of the macro fluorescence imaging component faces the sensing hydrogel layer. The fluorescence excitation component and the macro fluorescence imaging component form a fluorescence excitation light path and a collection light path for the sensing hydrogel layer in space. The image processing unit is communicatively connected to the macro fluorescence imaging component. The image processing unit receives the two-dimensional fluorescence image with fluorescence signal intensity and spatial distribution information output by the macro fluorescence imaging component, performs calculations and processing, and generates and outputs Staphylococcus aureus-related fluorescence response results.

2. The Staphylococcus aureus sensing wound dressing system according to claim 1, characterized in that, The aptamer fluorescent probe is a single-stranded deoxyribonucleic acid (DNA) containing a polymeric linker group, a spacer sequence, a Staphylococcus aureus aptamer sequence, and a Cy3 fluorescent group. The polymeric linker group is located at the 5′ end of the aptamer fluorescent probe, the spacer sequence is connected between the polymeric linker group and the Staphylococcus aureus aptamer sequence, and the Cy3 fluorescent group is attached to the 3′ end of the Staphylococcus aureus aptamer sequence. The quenching strand is a type of 7-nucleotide DNA short chain Q1-7 containing a quenching group attached to the 5′ end of the DNA short chain Q1-7. The quenching group is BHQ2. The DNA short chain Q1-7 is a short chain that is continuously complementary to the 3′ end region of the Staphylococcus aureus aptamer sequence in the aptamer fluorescent probe.

3. The Staphylococcus aureus sensing wound dressing system according to claim 2, characterized in that, The polymeric linker is Acrydite, the spacer sequence is a sequence composed of consecutive thymine deoxyribonucleotides, and the base arrangement of the Staphylococcus aureus aptamer sequence is 5′-TCGGCACGTTCTCAGTAGCGCTCGCTGGTCATCCCACAGCTACGTC-3′. The base sequence of the short DNA chain Q1-7 is 5′-GACGTAG-3′; The aptamer fluorescent probe participates in the cross-linking process of the hydrogel matrix through the 5′ end of Acrydite, and is fixed in the hydrogel network of the hydrogel matrix by covalent bonding.

4. The Staphylococcus aureus sensing wound dressing system according to claim 1, characterized in that, The hydrogel matrix is ​​formulated from polyethylene glycol diacrylate, acrylamide, and carboxymethyl chitosan. The polyethylene glycol diacrylate and acrylamide undergo a polymerization reaction under the action of a photoinitiator to form a hydrogel network, and the carboxymethyl chitosan is interspersed within the hydrogel network.

5. The Staphylococcus aureus sensing wound dressing system according to claim 1, characterized in that, Both the fluorescence excitation component and the macro fluorescence imaging component are integrated inside the encapsulation substrate. The fluorescent excitation component includes a green LED and an excitation filter. The excitation filter is disposed on the light-emitting side of the green LED. The light emitted by the green LED passes through the excitation filter to form excitation light. An excitation optical window is disposed inside the encapsulation substrate at the position corresponding to the optical path of the excitation light. The macro fluorescence imaging component includes an emission filter, a macro lens, and an image sensor. The macro lens is disposed on the light-incident side of the image sensor, the emission filter is disposed on the light-incident side of the macro lens, and an imaging optical window is disposed inside the encapsulation substrate corresponding to the optical path position of the macro fluorescence imaging component.

6. The Staphylococcus aureus sensing wound dressing system according to claim 5, characterized in that, The excitation filter is a 530nm narrowband excitation filter, and the emission filter is a 585nm emission filter; The packaging substrate has a light-shielding structure located outside the excitation optical window and the imaging optical window. The light-shielding structure includes a light shield or a dark cavity structure. The Staphylococcus aureus sensing wound dressing system also includes a microcontroller unit and a wireless communication module, both of which are integrated inside the encapsulation substrate.

7. The Staphylococcus aureus sensing wound dressing system according to claim 2, characterized in that, After acquiring the two-dimensional fluorescence image, the image processing unit determines the region of interest corresponding to the sensing hydrogel layer in the two-dimensional fluorescence image, extracts the red channel data corresponding to the Cy3 fluorescent group in the region of interest as the original gray-scale intensity matrix, and retrieves the baseline dark field image matrix under the condition of no Staphylococcus aureus. The image processing unit performs pixel-by-pixel subtraction between the original grayscale intensity matrix and the baseline dark field image matrix to perform background subtraction and brightness normalization processing, thereby obtaining a normalized effective fluorescence intensity matrix.

8. The Staphylococcus aureus sensing wound dressing system according to claim 7, characterized in that, The image processing unit calculates the average fluorescence intensity and the maximum fluorescence intensity within the region of interest based on the normalized effective fluorescence intensity matrix. The image processing unit retrieves a pre-stored reference high signal determination threshold, compares the values ​​in the normalized effective fluorescence intensity matrix with the reference high signal determination threshold to construct an indicator function, and calculates the area ratio of the high signal region and extracts the spatial distribution parameters of the high signal region based on the indicator function. The image processing unit obtains the average fluorescence intensity, maximum fluorescence intensity, high signal area ratio, and spatial distribution parameters, and then combines these with the time point of the two-dimensional fluorescence image to generate data on the changes of the average fluorescence intensity, maximum fluorescence intensity, high signal area ratio, and spatial distribution parameters over time.

9. The Staphylococcus aureus sensing wound dressing system according to claim 1, characterized in that, The Staphylococcus aureus sensing wound dressing system also includes a therapeutic hydrogel area, an electrode assembly, and an electrical stimulation driving module. The therapeutic hydrogel area is disposed on the outer surface of the sensing hydrogel layer or arranged adjacent to the sensing hydrogel layer, and the therapeutic hydrogel area is loaded with lysostaphin. The electrode assembly is disposed on the surface of the therapeutic hydrogel area or extends into the interior of the therapeutic hydrogel area. The electrode assembly is electrically connected to the electrical stimulation driving module, and the electrical stimulation driving module is communicatively connected to the image processing unit.

10. A Staphylococcus aureus-sensing wound dressing system according to claim 9, characterized in that, The image processing unit compares the generated Staphylococcus aureus-related fluorescence response results with preset conditions. When the Staphylococcus aureus-related fluorescence response results meet the preset conditions, it outputs a control signal to the electrical stimulation drive module. After receiving the control signal, the electrical stimulation drive module starts the preset electrical stimulation program and applies an electrical signal with a preset duration, preset voltage, or preset pulse parameters to the treatment hydrogel area through the electrode assembly, so that the lysostaphin enzyme loaded in the treatment hydrogel area is released under the action of the electric field.

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