Integrated biological photoelectrode of multicolor light source and preparation method of integrated biological photoelectrode

Abstract

The invention relates to the technical field of optogenetics, and particularly discloses an integrated biological photoelectrode of a multicolor light source and a preparation method. The integrated biological photoelectrode of the multicolor light source comprises a first biological photoelectrode and a second biological photoelectrode, the first biological photoelectrode comprises a first transparent substrate, a first photodiode, a microelectrode layer and a first shielding layer from bottom to top; the second biological photoelectrode comprises a second transparent substrate and a second photodiode from bottom to top; a plurality of first light-transmitting structures are arranged on the first shielding layer; the second biological photoelectrode is arranged below the first biological photoelectrode; the first photodiode and the first light-transmitting structure are aligned in the vertical direction; the second photodiode and the first light-transmitting structure are aligned in the vertical direction, and the second photodiode and the first photodiode are staggered in the vertical direction. According to the scheme, the first biological photoelectrode and the second biological photoelectrode can be mutually overlapped and cooperatively work, and efficient integration of a multicolor light source is achieved.

Description

Technical Field

[0001] This application relates to the technical field of optogenetics, and particularly to an integrated bio-optrode with multi-color light sources and a preparation method thereof. Background Art

[0002] Optogenetics is a bioengineering technology that integrates optical technology and genetic engineering. It can introduce light-sensitive protein genes (such as ChR2, NpHR, etc.) into the cell membrane of specific cells (such as nerve cells) through transgenic technology. The specific cells after transgenic can be activated or inhibited by light. Therefore, it can precisely activate or inhibit the ion channel activities of target cells by using specific wavelength light (such as blue light, yellow light, red light, green light), thereby achieving high-precision regulation of cell functions. Through the stimulation and signal collection of cells, optogenetics can conduct biomedical explorations such as studying the signal transmission between nerve cells, providing a manipulation tool with both cell specificity and millisecond-level resolution for biomedical research.

[0003] As an optogenetic tool, the integrated bio-optrode integrates a photodiode for specific targeted light regulation and a metal electrode for nerve signal recording on the top of the probe. Therefore, it can have both the specific targeted light regulation function and the nerve signal recording function required by optogenetics, and achieve precise regulation and monitoring of nerve activities. The integrated bio-optrode generally includes structures such as a substrate, a photodiode, and microelectrodes. When the probe works, the photodiode emits light under the action of a pulsed voltage, stimulating nerve cells to generate nerve impulses; the microelectrodes are used to record the signals generated by nerve cell impulses.

[0004] With the development of optogenetic technology, current technologies can already achieve the expression of two or more photosensitive proteins on the same cell. For example: the co-expression of the photosensitive protein GtACR2 sensitive to blue light (about 450 nm) and the photosensitive protein Chrimson sensitive to orange light (590 - 620 nm) in the same nerve cell. Nerve cells expressing multiple photosensitive proteins can respond to light stimuli of different wavelengths and produce different responses respectively. For example: for nerve cells co-expressing the photosensitive proteins GtACR2 and Chrimson, it is possible to inhibit the nerve cells with blue light and activate them with red light.

[0005] In practical applications, due to process and material limitations, an integrated bio-optical electrode usually has only one type of photodiode with a single color. Specifically, when fabricating an integrated bio-optical electrode, its substrate is used to fabricate photodiodes through epitaxial growth technology. Due to differences in epitaxial temperature, thermal expansion coefficient, lattice constant (such as between GaN and SiC, Si and InP), and the risk of heteroepitaxial cross-interference for different materials, if heteroepitaxial growth of multiple materials is carried out on the substrate, hetero-buffer layer design (to reduce the risk of heteroepitaxial cross-interference) and precise process control are required. This makes the process complex and the success rate low, resulting in a significant increase in cost. At the same time, since the emission color of a photodiode is determined by the properties of its material itself. For example, the bandgap of the epitaxial layer in the gallium arsenide (GaAs) system corresponds to the red light band (600 - 750 nm). To achieve other colors (such as blue-green light), materials such as gallium nitride (GaN) or indium phosphide (InP) need to be introduced. Therefore, a conventional integrated bio-optical electrode only has photodiodes made of one material, which enables the integrated bio-optical electrode to apply only one type of stimulus to nerve tissue.

[0006] In addition to the above-mentioned multi-material epitaxial growth method, in order to enable an integrated bio-optical electrode to have multi-color light sources, in the prior art, an epitaxial layer transfer method can also be used: First, define a photodiode of the first color on the first epitaxial wafer, then coat a transparent photoresist on the photodiode of the first color to protect the first-layer photodiode and bond the second epitaxial wafer. After that, bond the epitaxial side of the second epitaxial wafer downward on the photoresist, and remove the substrate of the second epitaxial wafer through a lift-off process and fabricate a photodiode of the second color. In this method, an etchant is required to etch the substrate of the second epitaxial wafer to achieve wet substrate lift-off. In the current photodiode epitaxial system, a gallium arsenide substrate is suitable for the epitaxial layer transfer method, which is achieved by the high etching selectivity of aqueous ammonia to the gallium arsenide substrate; a high etching selectivity means that the etching rate of aqueous ammonia to gallium arsenide is faster than that of the epitaxial layer, so that the substrate can be peeled off without damaging the epitaxial layer as much as possible. Since the epitaxial layer structure of the gallium arsenide system is only used for the production of red-light photodiodes, this solution can only achieve combinations of red light and other colors, and cannot achieve free combinations of arbitrary color light sources; moreover, since the thickness of the epitaxial wafer is in the micron range, the risk of damage and failure during the substrate etching process is high. Therefore, there are also problems of complex process and low success rate. Summary of the Invention

[0007] In view of this, the purpose of this application is to provide an integrated bio-optical electrode with multi-color light sources and a preparation method, which are used to solve the problems of complex preparation process and low success rate of existing integrated bio-optical electrodes with multi-color light sources.

[0008] To achieve the above technical objectives, a first aspect of the present application provides an integrated biophotoelectrode with a multi-color light source, including: a first biophotoelectrode and a second biophotoelectrode;

[0009] The first biophotoelectrode includes, from bottom to top: a first transparent substrate, a first photodiode, a microelectrode layer, and a first shielding layer;

[0010] The second biophotoelectrode includes, from bottom to top: a second transparent substrate and a second photodiode;

[0011] A plurality of first light-transmitting structures are provided on the first shielding layer;

[0012] The second biophotoelectrode is disposed below the first biophotoelectrode;

[0013] The first photodiode is aligned with the first light-transmitting structure in the vertical direction;

[0014] The second photodiode is aligned with the first light-transmitting structure in the vertical direction, and the second photodiode is staggered from the first photodiode in the vertical direction.

[0015] Further, the first biophotoelectrode includes: a first electrode metal layer and a plurality of first transparent insulating layers;

[0016] The first electrode metal layer is disposed above the first photodiode and is electrically connected to the first photodiode;

[0017] The microelectrode layer is disposed above the first electrode metal layer;

[0018] The first shielding layer is disposed above the microelectrode layer;

[0019] The first transparent insulating layers are provided above and below the first electrode metal layer, the microelectrode layer, and the first shielding layer.

[0020] Further, the second biophotoelectrode includes: a second electrode metal layer and a plurality of second transparent insulating layers;

[0021] The second electrode metal layer is disposed above the second transparent substrate;

[0022] The second transparent insulating layers are provided above and below the second electrode metal layer.

[0023] Further, windows are provided on the first transparent insulating layer and the first shielding layer;

[0024] The windows penetrate through the first electrode metal layer to the top surface of the first biophotoelectrode, and penetrate through the microelectrode layer to the top surface of the first biophotoelectrode;

[0025] A second window is provided on the second transparent insulating layer;

[0026] The second window penetrates through the top surface of the second electrode metal layer and the second bio-optical electrode;

[0027] The bottom surface of the second bio-optical electrode abuts against the bottom surface of the first bio-optical electrode, or, the top surface of the second bio-optical electrode abuts against the bottom surface of the first bio-optical electrode and the second window is located outside the window in the horizontal direction.

[0028] Further, a PCB circuit board is further included;

[0029] A slotted opening is provided on the PCB circuit board;

[0030] The first bio-optical electrode and the second bio-optical electrode are electrically connected to the PCB circuit board through bonding wires;

[0031] The slotted opening is used for the bonding wires to pass through.

[0032] Further, a third bio-optical electrode is further included;

[0033] The third bio-optical electrode includes, from bottom to top: a third transparent substrate and a third photodiode;

[0034] The third bio-optical electrode is disposed below the second bio-optical electrode;

[0035] The third photodiode is staggered from both the second photodiode and the first photodiode in the vertical direction.

[0036] Further, the second bio-optical electrode includes: a second shielding layer;

[0037] The second shielding layer is disposed above the second photodiode;

[0038] A plurality of second light-transmitting structures are provided on the second shielding layer;

[0039] The second light-transmitting structure is aligned with the first light-transmitting structure in the vertical direction;

[0040] The second photodiode is aligned with the second light-transmitting structure in the vertical direction;

[0041] The third photodiode is aligned with the second light-transmitting structure in the vertical direction.

[0042] Further, the third bio-optical electrode includes: a third shielding layer;

[0043] The third shielding layer is disposed above the third photodiode;

[0044] A third light-transmitting structure is provided on the third shielding layer;

[0045] The third photodiode is aligned with the third light-transmitting structure in the vertical direction.

[0046] Further, a plurality of the first light-transmitting structures are provided on the first shielding layer;

[0047] The plurality of first light-transmitting structures are distributed at intervals in a straight line or in an array.

[0048] A second aspect of the present application provides a method for preparing an integrated bio-photoelectrode of a multi-color light source, which is used to prepare the first bio-photoelectrode in the integrated bio-photoelectrode of the multi-color light source described in any one of the above, and includes:

[0049] S1. Growing an n-type gallium nitride, an active layer, and a p-type gallium nitride on a first transparent substrate in sequence, so as to form an epitaxial wafer on the first transparent substrate;

[0050] S2. Etching the epitaxial wafer to the n-type gallium nitride;

[0051] S3. Preparing a transparent conductive layer on the p-type gallium nitride;

[0052] S4. Patterning the transparent conductive layer to be adapted to the shape of the first photodiode;

[0053] S5. Preparing a first layer of the first transparent insulating layer on the transparent conductive layer, and patterning the first layer of the first transparent insulating layer to obtain a cathode contact window and an anode contact window;

[0054] S6. Preparing a first metal thin film on the first transparent insulating layer, and patterning the first metal thin film to obtain a first electrode metal layer;

[0055] S7. Preparing a second layer of the first transparent insulating layer on the first electrode metal layer, and patterning the second layer of the first transparent insulating layer to obtain an anode pad window and a cathode pad window;

[0056] S8. Preparing a second metal thin film on the second layer of the first transparent insulating layer, and patterning the metal thin film to prepare a microelectrode layer;

[0057] S9. Preparing a third layer of the first transparent insulating layer on the microelectrode layer, and patterning the third layer of the first transparent insulating layer to obtain a microelectrode window, a microelectrode pad window, a second anode pad window, and a second cathode pad window;

[0058] S10. Prepare a third metal thin film on the first transparent insulating layer of the third layer, and pattern the third metal thin film to obtain a first shielding layer with a first light-transmitting structure;

[0059] S11. Prepare a fourth layer of the first transparent insulating layer on the first shielding layer, and pattern the fourth layer of the first transparent insulating layer to obtain a shielding layer pad window, a second microelectrode window, a second microelectrode pad window, a third anode pad window, and a third cathode pad window.

[0060] Furthermore, the transparent conductive layer is made of indium tin oxide.

[0061] As can be seen from the above technical solutions, the present application provides an integrated bio-optical electrode for a multi-color light source and a preparation method thereof; wherein, the integrated bio-optical electrode for a multi-color light source includes: a first bio-optical electrode and a second bio-optical electrode; the first bio-optical electrode includes, from bottom to top: a first transparent substrate, a first photodiode, a microelectrode layer, and a first shielding layer; the second bio-optical electrode includes, from bottom to top: a second transparent substrate and a second photodiode; a plurality of first light-transmitting structures are provided on the first shielding layer; the second bio-optical electrode is disposed below the first bio-optical electrode; the first photodiode is aligned with the first light-transmitting structure in the vertical direction; the second photodiode is aligned with the first light-transmitting structure in the vertical direction, and the second photodiode is staggered from the first photodiode in the vertical direction.

[0062] In this solution, the first light-transmitting structure can allow the light of the first photodiode to pass through and at the same time allow the light of the second photodiode to pass through, so that the first bio-optical electrode and the second bio-optical electrode can be stacked and work together to realize the integration of any color light source. During preparation, the method of setting the first light-transmitting structure has a simpler operation process than the method of peeling the substrate, which can effectively reduce the process complexity and improve the success rate. Description of the Drawings

[0063] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the following will briefly introduce the drawings required for use in the description of the embodiments or the prior art. Obviously, the following drawings are only some embodiments of the present application. For those of ordinary skill in the art, other drawings can be obtained based on these drawings without creative efforts.

[0064] Figure 1 It is a split view of the first bio-optical electrode in an integrated bio-optical electrode for a multi-color light source provided by an embodiment of the present application;

[0065] Figure 2 It is a combination process diagram of an integrated bio-optical electrode for a multi-color light source provided by an embodiment of the present application;

[0066] Figure 3 Another combination process diagram of an integrated bio - photoelectrode with a multi - color light source provided by an embodiment of the present application;

[0067] Figure 4 Combination process diagram of an integrated bio - photoelectrode with a multi - color light source provided by another embodiment of the present application;

[0068] Figure 5 Connection diagram between a bio - photoelectrode and a PCB circuit board in an integrated bio - photoelectrode with a multi - color light source provided by an embodiment of the present application;

[0069] Figure 6 Another connection diagram between a bio - photoelectrode and a PCB circuit board in an integrated bio - photoelectrode with a multi - color light source provided by an embodiment of the present application

[0070] Figure 7 Connection diagram of an integrated bio - photoelectrode with a multi - color light source provided by an embodiment of the present application when the PCB circuit board is provided with a slot;

[0071] Figure 8 Another connection diagram of an integrated bio - photoelectrode with a multi - color light source provided by an embodiment of the present application when the PCB circuit board is provided with a slot;

[0072] Figure 9 Top view of the first bio - photoelectrode of an integrated bio - photoelectrode with a multi - color light source provided by an embodiment of the present application;

[0073] Figure 10 Another top view of the first bio - photoelectrode of an integrated bio - photoelectrode with a multi - color light source provided by an embodiment of the present application;

[0074] In the figure:

[0075] 10. First bio - photoelectrode; 11. First transparent substrate; 12. First photodiode; 121. n - type gallium nitride; 122. Active layer; 123. p - type gallium nitride; 13. Electrode layer; 14. First shielding layer; 15. First light - transmitting structure; 16. First electrode metal layer; 17. First transparent insulating layer; 171. Cathode contact window; 172. Anode contact window; 18. Transparent conductive layer;

[0076] 20. Second bio - photoelectrode; 21. Second transparent substrate; 22. Second photodiode; 23. Second light - transmitting structure; 24. Second electrode metal layer; 25. Second transparent insulating layer;

[0077] 30. Third bio - photoelectrode; 31. Third transparent substrate; 32. Third photodiode; 33. Third light - transmitting structure;

[0078] 40. Circuit board; 41. Circuit board pad; 42. Bonding wire; 43. Groove. Detailed implementation manner

[0079] The technical solutions of the embodiments of the present application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are part of the embodiments of the present application, rather than all of the embodiments. Based on the embodiments in this specification of the present application, all other embodiments obtained by those of ordinary skill in the art without creative efforts belong to the scope protected by the present application.

[0080] In the description of the embodiments of the present application, it should be noted that the orientation or positional relationship indicated by the terms "center", "upper", "lower", "left", "right", "vertical", "horizontal", "inner", "outer", etc. is based on the orientation or positional relationship shown in the accompanying drawings. It is only for the convenience of describing the embodiments of the present application and simplifying the description, rather than indicating or implying that the device or element referred to must have a specific orientation, be constructed and operated in a specific orientation, and therefore should not be construed as a limitation to the embodiments of the present application. In addition, the terms "first", "second", and "third" are only used for descriptive purposes and cannot be understood as indicating or implying relative importance.

[0081] In the description of the embodiments of the present application, it should be noted that unless otherwise clearly specified and limited, the terms "installation", "connection", and "connection" should be understood in a broad sense. For example, it can be a fixed connection, a replaceable connection, or an integral connection. It can be a mechanical connection or an electrical connection. It can be directly connected or indirectly connected through an intermediate medium. It can be the communication inside two elements. For those of ordinary skill in the art, the specific meanings of the above terms in the embodiments of the present application can be understood according to specific situations.

[0082] Please refer to Figure 1 and Figure 2 , in the first aspect of the embodiments of the present application, an integrated biophotoelectrode of a multi-color light source is provided, including: a first biophotoelectrode 10 and a second biophotoelectrode 20. Among them, the first biophotoelectrode 10 and the second biophotoelectrode 20 have both specific targeting light regulation functions and neural signal recording functions.

[0083] Please refer to Figure 1 , the first biophotoelectrode 10 includes, from bottom to top: a first transparent substrate 11, a first photodiode 12, a microelectrode layer 13, and a first shielding layer 14. The second biophotoelectrode 20 includes, from bottom to top: a second transparent substrate 21 and a second photodiode 22.

[0084] Among them, the first transparent substrate 11 and / or the second transparent substrate 21 can be a sapphire substrate, which has strong corrosion resistance and oxidation resistance, and has a high optical transmittance. The first photodiode 12 and / or the second photodiode 22 can be working devices based on the PN junction photoelectric effect, capable of converting electrical signals and optical signals, so as to emit light under the action of a pulsed voltage and generate a stimulus to two pairs of nerve cells. The microelectrode layer 13 is used to record the signals generated by nerve cell impulses.

[0085] The first shielding layer 14 is located above the microelectrode layer 13, and is used to reduce the interference of external signals to the microelectrode layer 13. A plurality of first light-transmitting structures 15 are provided on the first shielding layer 14; the first photodiode 12 is aligned with the first light-transmitting structure 15 in the vertical direction, the second photodiode 22 is aligned with the first light-transmitting structure 15 in the vertical direction, and the second photodiode 22 is staggered from the first photodiode 12 in the vertical direction, so that the first light-transmitting structure 15 not only allows the light emitted by the photodiode on this probe to pass through, but also allows the light from the corresponding position below to pass through.

[0086] In this embodiment, please refer to Figure 2 and Figure 3 , the second biophotoelectrode 20 is disposed below the first biophotoelectrode 10. Specifically, the first biophotoelectrode 10 can be stacked on the second biophotoelectrode 20 to form an integrated biophotoelectrode. In the first biophotoelectrode 10, the light emitted by the first photodiode 12 can pass through the first light-transmitting structure 15 to achieve optical stimulation of external cells; the light of the second photodiode 22 can also pass through the first light-transmitting structure 15 to achieve optical stimulation of external cells.

[0087] The first biophotoelectrode 10 and the second biophotoelectrode 20 can be bonded using transparent epoxy resin to achieve stacking.

[0088] As an implementation manner, as shown in Figure 2 , the bonding surface between the second biophotoelectrode 20 and the first biophotoelectrode 10 is the top surface of the second biophotoelectrode 20. Generally, the top surface of the second biophotoelectrode 20 is the second transparent insulating layer 25; therefore, in this implementation manner, the top surface of the uppermost second transparent insulating layer 25 abuts against the bottom surface of the first transparent substrate 11.

[0089] As another implementation manner, as shown in Figure 3 , the bonding surface between the second biophotoelectrode 20 and the first biophotoelectrode 10 is the bottom surface of the second biophotoelectrode 20, that is, the bottom surface of the first transparent substrate 11 abuts against the bottom surface of the second transparent substrate 21.

[0090] In practical applications, the first photodiode 12 and the second photodiode 22 can be fabricated using different materials to achieve light source outputs of different wavelengths. The way they are stacked generally does not impose restrictions on material selection like the substrate peeling method, so that any combination of light sources of different colors can be freely realized on the integrated bio-optical electrode. Through this stacking design, the already fabricated first photodiode 12 and second photodiode 22 can be directly used without going through steps such as etching and peeling the substrate, simplifying the production process, reducing the manufacturing cost, and at the same time increasing the preparation success rate.

[0091] Similarly, in the case of needing more color light sources, taking the need for three colors as an example; please refer to Figure 4 , the integrated bio-optical electrode with multi-color light sources further includes: a third bio-optical electrode 30; the third bio-optical electrode 30 includes, from bottom to top: a third transparent substrate 31 and a third photodiode 32; the third bio-optical electrode 30 is disposed below the second bio-optical electrode 20; the third photodiode 32 is staggered from both the second photodiode 22 and the first photodiode 12 in the vertical direction.

[0092] Similarly, during fabrication, only the third bio-optical electrode 30 needs to be stacked below the first photodiode 12 and the second photodiode 22, and it is ensured that each photodiode is aligned with the first light-transmitting structure 15. Through this flexible stacking method, the integrated bio-optical electrode can not only achieve precise control of multi-color light sources but also be quickly adjusted according to different experimental requirements, greatly improving the efficiency and accuracy of bio-optical experiments.

[0093] In addition, when a single bio-optical electrode is used as a stimulation probe, a light-transmitting structure can also be opened on the probe to facilitate combination with bio-optical electrodes of different colors in other cases.

[0094] As an implementation manner, the first light-transmitting structure 15 can be an optical window opened on the first shielding layer 14, which can be obtained by patterning the first shielding layer 14 through a peeling process. Moreover, the size and shape of the first light-transmitting structure 15 are configured to be the same as the shape of the photodiode.

[0095] In one embodiment, please refer to Figure 1 , the first bio-optical electrode 10 includes: a first electrode metal layer 16 and a plurality of first transparent insulating layers 17; the first electrode metal layer 16 is disposed above the first photodiode 12 and is electrically connected to the first photodiode 12; the microelectrode layer 13 is disposed above the first electrode metal layer 16; the first shielding layer 14 is disposed above the microelectrode layer 13; first transparent insulating layers 17 are disposed above and below the first electrode metal layer 16, the microelectrode layer 13, and the first shielding layer 14.

[0096] In a more specific embodiment, please refer to Figure 2 , the second biological optoelectrode 20 includes: a second electrode metal layer 24 and a plurality of second transparent insulating layers 25; the second electrode metal layer 24 is disposed above the second transparent substrate 21; second transparent insulating layers 25 are disposed both above and below the second electrode metal layer 24.

[0097] In practical applications, the first transparent insulating layer 17 and / or the second transparent insulating layer 25 is a silicon dioxide layer.

[0098] Pad can be disposed on the second electrode metal layer 24, the first electrode metal layer 16 and the electrode layer 13, and can be connected to an external circuit through the pad.

[0099] In one embodiment, windows are provided on the first transparent insulating layer 17 and the first shielding layer 14 (the cathode contact window 171 and the anode contact window 172 are shown in Figure 1 , and other windows are not shown in the figure); a second window is provided on the second transparent insulating layer 25.

[0100] The windows penetrate the top surface of the first electrode metal layer 16 and the first biological optoelectrode 10, and penetrate the top surface of the microelectrode layer 13 and the first biological optoelectrode 10. The second window penetrates the top surface of the second electrode metal layer 24 and the second biological optoelectrode 20. Herein, penetration means that an electrical connector can penetrate into the window or the second window. Therefore, through the window and the second window, an external electrical connector can be electrically connected to the second electrode metal layer 24, the first electrode metal layer 16 and the electrode layer 13.

[0101] In this embodiment, the bottom surface of the second biological optoelectrode 20 abuts against the bottom surface of the first biological optoelectrode 10, or, the top surface of the second biological optoelectrode 20 abuts against the bottom surface of the first biological optoelectrode 10 and the second window is located outside the window in the horizontal direction, so that both the second biological optoelectrode 20 and the first biological optoelectrode 10 can be electrically connected to the outside. Taking their electrical connection to the PCB circuit board 40 as an example:

[0102] In, please refer to Figure 2 And Figure 5 , the first biological optoelectrode 10 covers the top surface of the second biological optoelectrode 20. Since the second window is located outside the first window, the bonding wire 42 for electrical connection can be electrically connected to the second electrode metal layer 24, the first electrode metal layer 16 and the electrode layer 13 simultaneously. Among them, the bonding wire 42 can be an ultrasonic bonding wire. After welding, epoxy resin can be coated at the position of the bonding wire 42 and the epoxy resin can be air-dried and cured to prevent the bonding lead from failing during the use of the device.

[0103] It should be noted that when the single first biological optoelectrode 10 works, similarly, it can be electrically connected to the circuit board pad 41 on the PCB circuit board 40 through the bonding wire 42.

[0104] Taking the former as an example above, please refer to Figure 3 and Figure 6 , when the first transparent substrate 11 and the second transparent substrate 21 are in contact, the integrated biological optoelectrode can be welded to the PCB circuit board 40 by means of flip-chip bonding. In practical applications, the integrated biological optoelectrode can be first placed with the second biological optoelectrode 20 facing downwards, so that the pads of the second electrode metal layer 24 on the second biological optoelectrode are aligned with some circuit board pads 41 on the PCB circuit board 40 to achieve electrical connection. Then, the pads on the first biological optoelectrode 10 are electrically connected to some other circuit board pads 41 on the PCB circuit board 40 through the bonding wire 42. Finally, epoxy resin is applied to the position of the bonding wire 42 and allowed to dry and cure to prevent the bonding wire from failing during the use of the device.

[0105] As a way, please refer to Figure 7 and Figure 8 , a slot 43 is provided on the PCB circuit board 40; the first biological optoelectrode 10 and the second biological optoelectrode 20 are electrically connected to the PCB circuit board 40 through the bonding wire 42; the slot 43 is used for the bonding wire 42 to pass through.

[0106] In this embodiment, since the slot 43 is provided on the PCB circuit board 40, regardless of whether the top surface or the bottom surface of the second biological optoelectrode 20 is in contact with the first biological optoelectrode 10, the integrated biological optoelectrode can be conveniently electrically connected to the PCB circuit board 40.

[0107] As an implementation manner, as Figure 7 shown, the integrated biological optoelectrode can be disposed on the front surface of the PCB circuit board 40, and the pads are exposed through the slot 43.

[0108] As another implementation manner, as Figure 8 shown, the integrated biological optoelectrode can be disposed on the back surface of the PCB circuit board 40, and the pads on the integrated biological optoelectrode are aligned with the slot 43.

[0109] In the above embodiments, the integrated biological optoelectrode and the PCB circuit board 40 can be bonded with epoxy resin, and after bonding, the epoxy resin is allowed to dry and cure.

[0110] In another embodiment provided in the present application, the second biological photoelectrode 20 includes: a second shielding layer; the second shielding layer is arranged above the second photodiode 22; a plurality of second light-transmitting structures 23 are arranged on the second shielding layer; the second light-transmitting structure 23 is aligned with the first light-transmitting structure 15 along the vertical direction; the second photodiode 22 is aligned with the second light-transmitting structure 23 along the vertical direction; the third photodiode 32 is aligned with the second light-transmitting structure 23 along the vertical direction.

[0111] In application, the second light-transmitting structure 23 may be a window structure. By providing a second shielding layer on the second biophotoelectrode 20, the anti-electromagnetic interference capability of the second photodiode 22 can be enhanced. In this embodiment, the second light-transmitting structure 23 is aligned with the first light-transmitting structure 15, so that the light emitted by the second photodiode 22 can pass through the second light-transmitting structure 23 and the first light-transmitting structure 15. Among them, the first light-transmitting structure 15 is configured to be more in number than the second light-transmitting structure 23, so that the first light-transmitting structure 15 can take into account both the first photodiode 12 and the second photodiode 22.

[0112] It should be noted that, in the present application, the up-down orientation relationship between the layers refers to the up-down orientation relationship when the bio-photoelectrode is placed horizontally in the front. For example, the second shielding layer is arranged above the second photodiode 22, which refers to the orientation relationship when the second bio-photoelectrode 20 is placed forward, that is, the second transparent substrate 21 is located at the bottom layer. When the second bio-photoelectrode 20 is inverted at the bottom of the first bio-photoelectrode 10, the second shielding layer is located below the second photodiode 22.

[0113] In one embodiment, see Figure 4 The third bio-photoelectrode 30 includes: a third shielding layer; the third shielding layer is arranged above the third photodiode 32; a third light-transmitting structure 33 is arranged on the third shielding layer; the third photodiode 32 and the third light-transmitting structure 33 are aligned in the vertical direction.

[0114] Similarly, the third light-transmitting structure 33 may be a window structure, which can enhance the anti-electromagnetic interference capability of the third photodiode 32 .

[0115] It should be noted that when the number of biophotoelectrodes included in the integrated biophotoelectrode of the multicolor light source is greater than two, each biophotoelectrode can be configured in a manner of increasing length from top to bottom. Taking the number of biophotoelectrodes as three as an example, the length of the third biophotoelectrode 30 is greater than the second biophotoelectrode 20, so that its pad is located outside the second biophotoelectrode 20 in the horizontal direction; the length of the second biophotoelectrode 20 is greater than the first biophotoelectrode 10, so that its pad is located outside the first biophotoelectrode 10 in the horizontal direction.

[0116] A plurality of first light-transmitting structures 15 are provided on the first shielding layer 14.

[0117] As an implementation manner, please refer to Figure 9 , the plurality of first light-transmitting structures 15 are arranged at intervals in a straight line. Among them, some of the first light-transmitting structures 15 are aligned with the first photodiodes 12, and some are aligned with the photodiodes of other biological photoelectrodes.

[0118] As an implementation manner, please refer to Figure 10 , the plurality of first light-transmitting structures 15 are arranged in an array. And, as Figure 10 shown, in this embodiment, the photodiodes on the biological photoelectrode can be arranged in a curve, and the photodiodes located in the upper layer and the lower layer are staggered both longitudinally and laterally, so that there are light sources of two colors in both the horizontal and vertical directions for the integrated biological photoelectrode.

[0119] Thus, there are light sources of two colors in both the horizontal and vertical directions

[0120] The second aspect of the present application provides a preparation method for an integrated biological photoelectrode with a multi-color light source, which is used to prepare the first biological photoelectrode 10 in the integrated biological photoelectrode with a multi-color light source in any one of the above embodiments, and includes:

[0121] S1. Grow an n-type gallium nitride 121, an active layer 122, and a p-type gallium nitride 123 on the first transparent substrate 11 in sequence, so that an epitaxial wafer is formed on the first transparent substrate 11.

[0122] Among them, the first transparent substrate 11 can be a sapphire substrate.

[0123] S2. Etch the epitaxial wafer to the n-type gallium nitride 121.

[0124] Among them, step S2 can etch the epitaxial wafer through a photolithography process and a dry etching process in a gas environment of Cl2 and BCl3, so that part of the epitaxial wafer is etched to the n-type gallium nitride 121. As Figure 1 shown, in the unetched part, the p-type gallium nitride 123 is exposed, and the remaining n-type gallium nitride 121, active layer 122, and p-type gallium nitride 123 form the first photodiode 12.

[0125] S3. Prepare a transparent conductive layer 18 on the p-type gallium nitride 123;

[0126] Among them, step S3 can prepare the transparent conductive layer 18 through a photolithography and magnetron sputtering process in a high-vacuum argon gas environment. The transparent conductive layer 18 can be made of indium tin oxide material.

[0127] S4. Pattern the transparent conductive layer 18 to be adapted to the shape of the first photodiode 12.

[0128] Among them, in step S4, the transparent conductive layer 18 can be patterned by a wet etching process to prepare the transparent anode of the first photodiode 12; then, the resistance of the transparent conductive layer 18 is reduced and the light transmittance of the transparent conductive layer 18 is increased through a rapid thermal annealing process, so as to form a good ohmic contact between the transparent conductive layer 18 and the p-type gallium nitride 123.

[0129] S5. Prepare the first layer of the first transparent insulating layer 17 on the transparent conductive layer 18, and pattern the first layer of the first transparent insulating layer 17 to obtain a cathode contact window 171 and an anode contact window 172.

[0130] Among them, in step S5, the first layer of the first transparent insulating layer 17 can be prepared by a photolithography process and a plasma-enhanced chemical vapor deposition process in a gas environment of SiH4 and N2O, a high vacuum and a high temperature environment of 350 °C, and the first layer of the first transparent insulating layer 17 is patterned by an inductively coupled plasma etching process to prepare the cathode contact window 171 and the anode contact window 172. The cathode contact window 171 and the anode contact window 172 can expose the cathode and anode of the first photodiode 12 on the first layer of the first transparent insulating layer 17. The cathode contact window 171 and the anode contact window 172 enable the following first electrode metal layer 16 to contact the first photodiode 12.

[0131] S6. Prepare a first metal thin film on the first transparent insulating layer 17, and pattern the first metal thin film to obtain the first electrode metal layer 16.

[0132] Among them, in step S6, the first metal thin film can be prepared by a photolithography process and an electron beam evaporation process in a high vacuum environment, and the first metal thin film is patterned by a lift-off process to form the first electrode metal layer 16. The first electrode metal layer 16 includes a photodiode anode electrode, a photodiode anode wire, a photodiode anode pad, a photodiode cathode electrode, a photodiode cathode wire, and a photodiode cathode pad.

[0133] S7. Prepare the second layer of the first transparent insulating layer 17 on the first electrode metal layer 16, and pattern the second layer of the first transparent insulating layer 17 to obtain an anode pad window and a cathode pad window.

[0134] Among them, in step S7, the second layer of the first transparent insulating layer 17 can be prepared by a photolithography process and a plasma-enhanced chemical vapor deposition process in a gas environment of SiH4 and N2O, a high vacuum and a high temperature environment of 350 °C, and the second layer of the first transparent insulating layer 17 is patterned by an inductively coupled plasma etching process to prepare the anode pad window and the cathode pad window. The anode pad window and the cathode pad window can expose the pads on the first electrode metal layer 16.

[0135] S8. Prepare a second metal thin film on the first transparent insulating layer 17 of the second layer, and pattern the metal thin film to obtain the microelectrode layer 13.

[0136] Among them, step S8 can prepare the second metal thin film in a high-vacuum environment through a photolithography process and an electron beam evaporation process, and pattern the second metal thin film by using a lift-off process to obtain the microelectrode layer 13. The microelectrode layer 13 includes microelectrode leads and microelectrode pads;

[0137] S9. Prepare a third layer of the first transparent insulating layer 17 on the microelectrode layer 13, and pattern the third layer of the first transparent insulating layer 17 to obtain a microelectrode window, a microelectrode pad window, a second anode pad window, and a second cathode pad window.

[0138] Among them, step S9 can prepare the third layer of the first transparent insulating layer 17 in a gas environment of SiH4 and N2O, a high-vacuum and 350°C high-temperature environment through a photolithography process and a plasma-enhanced chemical vapor deposition process, and pattern the third layer of the first transparent insulating layer 17 by using an inductively coupled plasma etching process to obtain a microelectrode window, a microelectrode pad window, a second anode pad window, and a second cathode pad window. The microelectrode window exposes the circular microelectrode; the microelectrode pad window exposes the microelectrode pad; the second anode pad window and the second cathode pad window are respectively aligned with the above-mentioned anode pad window and cathode pad window for exposing the pads on the first electrode metal layer 16.

[0139] S10. Prepare a third metal thin film on the third layer of the first transparent insulating layer 17, and pattern the third metal thin film to obtain the first shielding layer 14 with the first light-transmitting structure 15.

[0140] Among them, step S10 can prepare the third metal thin film in a high-vacuum environment through a photolithography process and an electron beam evaporation process, and pattern the third metal thin film by using a lift-off process to obtain the first shielding layer 14 with the first light-transmitting structure 15.

[0141] S11. Prepare a fourth layer of the first transparent insulating layer 17 on the first shielding layer 14, and pattern the fourth layer of the first transparent insulating layer 17 to obtain a shielding layer pad window, a second microelectrode window, a second microelectrode pad window, a third anode pad window, and a third cathode pad window.

[0142] Among them, step S11 can prepare the fourth layer of the first transparent insulating layer 17 through a photolithography process and a plasma-enhanced chemical vapor deposition process in a gas environment of SiH4 and N2O, a high-vacuum and 350 °C high-temperature environment, and pattern the fourth layer of the first transparent insulating layer 17 by using an inductively coupled plasma etching process to prepare a shielding layer pad window, a second microelectrode window, a second microelectrode pad window, a third anode pad window, and a third cathode pad window. The shielding layer pad window exposes the pad of the first shielding layer 14; the second microelectrode window is aligned with the microelectrode window to expose the microelectrode wire; the second microelectrode pad window is aligned with the above-mentioned microelectrode pad window to expose the microelectrode pad; the third anode pad window and the third cathode pad window are aligned with the above-mentioned second anode pad window and the second cathode pad window to expose the pads on the first electrode metal layer 16.

[0143] When it is necessary to prepare the second biological optoelectrode 20 and the third biological optoelectrode 30, the above steps can be repeated, and the processes of removing the microelectrode layer 13 and the third layer of the first transparent insulating layer 17 in steps S8 and S9 can be selectively removed.

[0144] After multiple biological optoelectrodes are prepared, they are combined by stacking multiple biological optoelectrodes. The combination steps include:

[0145] Use transparent epoxy resin to stack and bond multiple biological optoelectrodes together to achieve rough alignment of the biological optoelectrodes;

[0146] Put the roughly aligned multiple biological optoelectrodes into deionized water. Due to the hydrophobicity of the probe surface, the two probes will automatically align in the water.

[0147] In the embodiment of the present application, when the integrated biological optoelectrode of the multi-color light source is working, the device can be first inserted into the biological brain tissue that has undergone optogenetic gene manipulation so that the device is immersed in the biological solution, and then different colored photodiodes are lit to stimulate nerve cells and collect bioelectrical signals through the microelectrode layer 13.

[0148] Among them, the working principle of the photodiode is that a forward voltage is applied to the anode pad of the photodiode, the cathode pad is grounded, and the current will flow into the P-type gallium nitride region of the photodiode through the wire, flow through the active layer into the N-type gallium nitride, and the current flows out from the cathode pad. During this process, a large number of electron-hole pairs will recombine in the active region and emit a large amount of light of a specific wavelength. The photodiode is lit and emits a specific color to activate biological nerve cells. At the same time, the change in the potential of the biological nerve cells will also generate a signal on the microelectrode layer 13.

[0149] The working principle of the microelectrode layer 13 is that when the probe is inserted into the biological nerve tissue, the microelectrode layer 13 is in direct contact with the tissue fluid. The potential generated by the action of nerve cells will be transmitted into the microelectrode layer 13 through the tissue fluid and finally connected to the external circuit through the pads on the microelectrode layer 13. Thus, the biological nerve cell signals are collected.

[0150] In the integrated biophotoelectrode with a multi-color light source provided in this embodiment, the light emitted by the photodiodes of different probes can pass through another probe to illuminate the front target area of another probe, so as to achieve the stimulation of the same target tissue with different effects in one implantation. Moreover, the choice of color is not restricted, the preparation operation is simpler, and optogenetic regulation can be carried out within a larger range.

[0151] The above are the preferred embodiments of the present application and are not used to limit the present invention. Although the present application has been described in detail with reference to the examples, for those skilled in the art, they can still modify the technical solutions described in the foregoing examples, or perform equivalent replacements for some of the technical features. However, any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. An integrated bio - optoelectrode of a multi - color light source, characterized in that, Comprising: A first biological optoelectrode (10) and a second biological optoelectrode (20); The first biological optoelectrode (10) includes, from bottom to top: a first transparent substrate (11), a first photodiode (12), a microelectrode layer (13), and a first shielding layer (14); The second biological optoelectrode (20) includes, from bottom to top: a second transparent substrate (21) and a second photodiode (22); A plurality of first light-transmitting structures (15) are provided on the first shielding layer (14); The second biological optoelectrode (20) is disposed below the first biological optoelectrode (10); The first photodiode (12) is aligned with the first light-transmitting structure (15) in the vertical direction; The second photodiode (22) is aligned with the first light-transmitting structure (15) in the vertical direction, and the second photodiode (22) is staggered from the first photodiode (12) in the vertical direction.

2. The integrated bio-photoelectrode of the multi-color light source according to claim 1, characterized in that The first biological optoelectrode (10) includes: a first electrode metal layer (16) and a plurality of first transparent insulating layers (17); The first electrode metal layer (16) is disposed above the first photodiode (12) and is electrically connected to the first photodiode (12); The microelectrode layer (13) is disposed above the first electrode metal layer (16); The first shielding layer (14) is disposed above the microelectrode layer (13); The first transparent insulating layers (17) are provided above and below the first electrode metal layer (16), the microelectrode layer (13), and the first shielding layer (14).

3. The integrated bio-optical electrode of the multi-color light source according to claim 2, characterized in that The second biological optoelectrode (20) includes: a second electrode metal layer (24) and a plurality of second transparent insulating layers (25); The second electrode metal layer (24) is disposed above the second transparent substrate (21); The second transparent insulating layers (25) are provided above and below the second electrode metal layer (24).

4. The integrated bio-photoelectrode of the multi-color light source according to claim 3, characterized in that, Windows are provided on the first transparent insulating layer (17) and the first shielding layer (14); The windows penetrate through the first electrode metal layer (16) to the top surface of the first biological optoelectrode (10), and penetrate through the microelectrode layer (13) to the top surface of the first biological optoelectrode (10); A second window is provided on the second transparent insulating layer (25); The second window penetrates through the second electrode metal layer (24) to the top surface of the second biological optoelectrode (20); The bottom surface of the second biological optoelectrode (20) abuts against the bottom surface of the first biological optoelectrode (10), or, the top surface of the second biological optoelectrode (20) abuts against the bottom surface of the first biological optoelectrode (10) and the second window is located outside the window in the horizontal direction.

5. The integrated bio-photoelectrode of the multi-color light source according to claim 4, characterized in that It further includes a PCB circuit board (40); A slot (43) is provided on the PCB circuit board (40); The first biological optoelectrode (10) and the second biological optoelectrode (20) are electrically connected to the PCB circuit board (40) through bonding wires (42); The slot (43) is used for the bonding wires (42) to pass through.

6. The integrated bio-optrode of the multi-color light source according to claim 3, wherein It further includes: The third biological optoelectrode (30); The third biological optoelectrode (30) includes, from bottom to top: a third transparent substrate (31) and a third photodiode (32); The third biological optoelectrode (30) is disposed below the second biological optoelectrode (20); The third photodiode (32) is staggered from both the second photodiode (22) and the first photodiode (12) in the vertical direction.

7. The integrated bio-optical electrode of the multi-color light source according to claim 6, characterized in that, The second biological optoelectrode (20) includes: a second shielding layer; The second shielding layer is disposed above the second photodiode (22); A plurality of second light-transmitting structures (23) are disposed on the second shielding layer; The second light-transmitting structures (23) are aligned with the first light-transmitting structures (15) in the vertical direction; The second photodiode (22) is aligned with the second light-transmitting structures (23) in the vertical direction; The third photodiode (32) is aligned with the second light-transmitting structures (23) in the vertical direction; The third biological optoelectrode (30) includes: a third shielding layer; The third shielding layer is disposed above the third photodiode (32); A third light-transmitting structure (33) is disposed on the third shielding layer; The third photodiode (32) is aligned with the third light-transmitting structure (33) in the vertical direction.

8. The integrated bio-photoelectrode of the multi-color light source according to claim 1, wherein, A plurality of the first light-transmitting structures (15) are disposed on the first shielding layer (14); The plurality of first light-transmitting structures (15) are distributed at intervals in a straight line or in an array.

9. A preparation method of an integrated bio-photoelectrode of a multi-color light source, characterized in that, For preparing the first biological optoelectrode (10) in the integrated biological optoelectrode of the multi-color light source according to any one of claims 1 to 8, and includes: S1. Grow n-type gallium nitride (121), an active layer (122), and p-type gallium nitride (123) in sequence on a first transparent substrate (11) so that an epitaxial wafer is formed on the first transparent substrate (11); S2. Etch a part of the epitaxial wafer to the n-type gallium nitride (121) to form a first photodiode (12) on the first transparent substrate (11); S3. Prepare a transparent conductive layer (18) on the p-type gallium nitride (123); S4. Pattern the transparent conductive layer (18) to be adapted to the shape of the first photodiode (12); S5. Prepare a first layer of the first transparent insulating layer (17) on the transparent conductive layer (18), and pattern the first layer of the first transparent insulating layer (17) to obtain a cathode contact window (171) and an anode contact window (172); S6. Prepare a first metal thin film on the first transparent insulating layer (17), and pattern the first metal thin film to obtain a first electrode metal layer (16); S7. Prepare a second layer of the first transparent insulating layer (17) on the first electrode metal layer (16), and pattern the second layer of the first transparent insulating layer (17) to obtain an anode pad window and a cathode pad window; S8. Prepare a second metal thin film on the second layer of the first transparent insulating layer (17), and pattern the metal thin film to prepare a microelectrode layer (13); S9. Prepare a third layer of the first transparent insulating layer (17) on the microelectrode layer (13), and pattern the third layer of the first transparent insulating layer (17) to obtain a microelectrode window, a microelectrode pad window, a second anode pad window, and a second cathode pad window; S10. Prepare a third metal thin film on the third layer of the first transparent insulating layer (17), and pattern the third metal thin film to obtain a first shielding layer (14) having a first light-transmitting structure (15); S11. Prepare a fourth layer of the first transparent insulating layer (17) on the first shielding layer (14), and pattern the fourth layer of the first transparent insulating layer (17) to obtain a shielding layer pad window, a second microelectrode window, a second microelectrode pad window, a third anode pad window, and a third cathode pad window.

10. The preparation method of the integrated bio-optrode of the multi-color light source according to claim 9, characterized in that, The transparent conductive layer (18) is made of indium tin oxide.

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