Differential low-noise integrated biosensor array for cell culture detection and preparation method of differential low-noise integrated biosensor array

Abstract

The invention discloses a differential low-noise integrated biosensor array for cell culture detection and a preparation method thereof, and the differential low-noise integrated biosensor array integrates an ISHFET device, an REHFET device and an EXHFET device, so that extracellular action potential signals and local field potential signals can be detected during in-vitro cell culture; therefore, interference of factors such as tissue trauma and immune response on cell activity signal detection is avoided, and good reliability is realized; the ISHFET device and the REHFET device can form a differential sensing unit, so that the effect of reducing the detection error is realized; the EXHFET device not only can detect extracellular action potential signals, but also can independently detect local field potential signals, so that more diversified cell activity signals can be detected, and more data support is provided for in-vitro cell culture research. The method is widely applied to the technical field of semiconductors.

Description

Technical Field

[0001] The embodiments of the present invention relate to the field of semiconductor technology, and in particular to a differential low-noise integrated biosensor array for cell culture detection and its fabrication method. Background Technology

[0002] Neuroelectrophysiological techniques, as a key means of detecting electrical signals in brain nerves, play an irreplaceable role in revealing the functional mechanisms of the nervous system under normal and pathological states, providing a crucial foundation for the early diagnosis and precise intervention of various neurological diseases. However, existing neuroelectrophysiological techniques still have significant limitations in terms of signal-to-noise ratio, acquisition rate, and system stability, restricting their further application.

[0003] Currently used neural electrodes can be broadly classified into two categories: non-invasive external electrodes (such as electroencephalography, EEG) and invasive internal electrodes (such as electrocorticography, ECoG, and multichannel electrode membranes, MEAs). Based on their detection mechanisms, they can also be divided into passive and active electrodes. Passive electrodes typically acquire signals based on microelectrode electrochemical impedance spectroscopy. While structurally simple, they have limited acquisition rates and are susceptible to interference from external electric fields and electromagnetic radiation, resulting in poor signal stability and a low signal-to-noise ratio. Furthermore, the biocompatibility of metallic electrode materials also limits their long-term reliability. Active electrodes, such as silicon-based ion-sensitive field-effect transistors (ISFETs), improve acquisition stability by modulating channel conductance with electrical signals, but they still fail to meet the requirements for high-quality neural signal monitoring in terms of signal-to-noise ratio and response speed. Summary of the Invention

[0004] To address the technical problems of low signal-to-noise ratio, low acquisition rate, and poor stability in current neural electrical signal detection technologies, the present invention aims to provide a differential low-noise integrated biosensor array for cell culture detection and its preparation method.

[0005] On one hand, embodiments of the present invention include a differential low-noise integrated biosensor array for cell culture detection, the differential low-noise integrated biosensor array comprising:

[0006] Substrate;

[0007] Multiple ISHFET devices are located on the substrate; the ISHFET devices are used to detect extracellular action potential signals of nerve cells in cell culture solution;

[0008] Multiple REHFET devices are located on the substrate; the REHFET devices are used to detect extracellular action potential signals of nerve cells in cell culture solution;

[0009] Multiple EXHFET devices are located on the substrate; the EXHFET devices are used to detect extracellular action potential signals and / or local field potential signals of nerve cells in cell culture solution.

[0010] Furthermore, the ISHFET device includes:

[0011] AlGaN / GaN heterojunction; the AlGaN / GaN heterojunction includes a GaN buffer layer, an AlN insertion layer and an AlGaN barrier layer stacked sequentially from bottom to top; a two-dimensional electron gas is formed at the interface between the GaN buffer layer and the AlN insertion layer; the GaN buffer layer is etched with an ISHFET recessed gate;

[0012] ISHFET source lead; one end of the ISHFET source lead forms an ohmic contact with the AlGaN / GaN heterojunction;

[0013] ISHFET drain lead; one end of the ISHFET drain lead forms an ohmic contact with the AlGaN / GaN heterojunction;

[0014] A first insulating passivation layer; the first insulating passivation layer covers the AlGaN / GaN heterojunction, the ISHFET source lead and the ISHFET drain lead, and an ISHFET window is formed on the first insulating passivation layer to expose the ISHFET recessed gate.

[0015] Further, the REHFET device includes:

[0016] AlGaN / GaN heterojunction; the AlGaN / GaN heterojunction includes a GaN buffer layer, an AlN insertion layer and an AlGaN barrier layer stacked sequentially from bottom to top; a two-dimensional electron gas is formed at the interface between the GaN buffer layer and the AlN insertion layer;

[0017] REHFET source lead; one end of the REHFET source lead forms an ohmic contact with the AlGaN / GaN heterojunction;

[0018] REHFET drain lead; one end of the REHFET drain lead forms an ohmic contact with the AlGaN / GaN heterojunction;

[0019] A second insulating passivation layer; the second insulating passivation layer covers the AlGaN / GaN heterojunction, the REHFET source lead, and the REHFET drain lead.

[0020] Further, the REHFET device includes:

[0021] AlGaN / GaN heterojunction; the AlGaN / GaN heterojunction includes a GaN buffer layer, an AlN insertion layer and an AlGaN barrier layer stacked sequentially from bottom to top; a two-dimensional electron gas is formed at the interface between the GaN buffer layer and the AlN insertion layer;

[0022] REHFET source lead; one end of the REHFET source lead forms an ohmic contact with the AlGaN / GaN heterojunction;

[0023] REHFET drain lead; one end of the REHFET drain lead forms an ohmic contact with the AlGaN / GaN heterojunction;

[0024] A second insulating passivation layer covers the AlGaN / GaN heterojunction, the REHFET source lead, and the REHFET drain lead. A REHFET window exposing the AlGaN barrier layer is formed on the second insulating passivation layer. A functional material connected to the AlGaN barrier layer is provided in the REHFET window. The functional material has low sensitivity to neural signals and high sensitivity to environmental interference.

[0025] Furthermore, the EXHFET device includes:

[0026] AlGaN / GaN heterojunction; the AlGaN / GaN heterojunction includes a GaN buffer layer, an AlN insertion layer and an AlGaN barrier layer stacked sequentially from bottom to top; a two-dimensional electron gas is formed at the interface between the GaN buffer layer and the AlN insertion layer;

[0027] EXHFET source lead; the EXHFET source lead is located on one side in the first direction of the EXHFET, and one end of the EXHFET source lead forms an ohmic contact with the AlGaN / GaN heterojunction;

[0028] EXHFET drain lead; the EXHFET drain lead is located on the other side of the first direction in the EXHFET, and one end of the EXHFET drain lead forms an ohmic contact with the AlGaN / GaN heterojunction;

[0029] At least one metal extension gate; each of the metal extension gates is distributed along a second direction, and the metal extension gates form an ohmic contact with the AlGaN / GaN heterojunction; the second direction is perpendicular to the first direction;

[0030] A third insulating passivation layer; the third insulating passivation layer covers the AlGaN / GaN heterojunction, and an EXHFET window is opened on the third insulating passivation layer to expose the metal extended gate.

[0031] Furthermore, the metal extension gate is made of an inert metal material.

[0032] Furthermore, the differential low-noise integrated biosensor array also includes:

[0033] Multiple ISHFET source pads; the ISHFET source pads are connected to the other end of the ISHFET source leads;

[0034] Multiple ISHFET drain pads; the ISHFET drain pads are connected to the other end of the ISHFET drain lead;

[0035] Multiple REHFET source pads; the REHFET source pads are connected to the other end of the REHFET source leads;

[0036] Multiple REHFET drain pads; the REHFET drain pads are connected to the other end of the REHFET drain lead;

[0037] Multiple EXHFET source pads; the EXHFET source pads are connected to the other end of the EXHFET source leads;

[0038] Multiple EXHFET drain pads; the EXHFET drain pads are connected to the other end of the EXHFET drain lead.

[0039] Furthermore, the differential low-noise integrated biosensor array also includes:

[0040] A signal processing module; the signal processing module is used to process the extracellular action potential signal and the local field potential signal.

[0041] Furthermore, each of the ISHFET devices and each of the REHFET devices constitutes a plurality of differential sensing units;

[0042] Each of the differential sensing units includes one of the ISHFET devices and one of the REHFET devices;

[0043] Each of the ISHFET devices belongs to a corresponding differential sensing unit;

[0044] Each of the REHFET devices belongs to at least one of the differential sensing units;

[0045] The signal processing module is used to perform differential processing on the extracellular action potential signal detected by the ISHFET device and the extracellular action potential signal detected by the REHFET device in the same differential sensing unit.

[0046] On the other hand, embodiments of the present invention also include a method for fabricating a differential low-noise integrated biosensor array, the fabrication method comprising:

[0047] An epitaxial wafer is obtained by sequentially depositing a GaN buffer layer, an AlN insertion layer, and an AlGaN barrier layer on a sapphire substrate.

[0048] Under the conditions of Cl2 and CF4 gas environment, the epitaxial wafer is etched down to the GaN buffer layer by photolithography and dry etching processes to form a plurality of spaced ISHFET mesa, REHFET mesa and EXHFET mesa;

[0049] The AlGaN barrier layer is subjected to photolithography and electron beam evaporation to deposit a metal thin film in a high vacuum environment. The metal thin film is then patterned using a lift-off process to obtain source and drain electrodes disposed on the AlGaN barrier layer. The metal thin film is made of Ti, Al, Ni, or Au material. The source and drain electrodes include ISHFET source, ISHFET drain, REHFET source, REHFET drain, EXHFET source, and EXHFET drain.

[0050] The epitaxial wafer and the source and drain electrodes are subjected to rapid thermal annealing to form ohmic contacts;

[0051] A seed layer is deposited in a high-vacuum environment using photolithography and electron beam evaporation processes. A thick gold film is then electroplated using secondary photolithography and electroplating processes. The thick gold film is then patterned using a lift-off process to obtain source / drain leads, pads, and the extended gate. The seed layer is made of Cr, Al, Ti, Pt, Ti, Pt, Au, or Ti material. The source / drain leads include the ISHFET source lead, the ISHFET drain lead, the REHFET source lead, the REHFET drain lead, the EXHFET source lead, and the EXHFET drain lead.

[0052] A silicon dioxide thin film is grown on the surface of the epitaxial wafer by atomic layer deposition. The silicon dioxide isolation layer is prepared by placing it in a gas environment of SiH4 and N2O, a vacuum environment and a high temperature environment to obtain an insulating passivation layer.

[0053] Using a dry etching process, CF4 gas is used to etch the outline of the patterned insulating layer on the insulating passivation layer to ensure its accuracy.

[0054] The insulating passivation layer is patterned using a wet etching process with a buffered oxide etching solution to prepare the ISHFET window, the EXHFET window, and the window corresponding to each of the pads.

[0055] The surface of the AlGaN barrier layer under the ISHFET window is shallowly etched using a photoelectrochemical oxidation process in an environment of ultraviolet lamp irradiation and deionized water solution to obtain the ISHFET recessed gate.

[0056] The beneficial effects of the embodiments of the present invention are as follows: The differential low-noise integrated biosensor array in the embodiments integrates ISHFET, REHFET, and EXHFET devices, thereby enabling the detection of extracellular action potential signals and local field potential signals during in vitro cell culture. This helps to avoid interference from factors such as tissue trauma, immune response, biological contamination, and interface instability on the detection of cell activity signals, achieving good reliability. The ISHFET device can form a differential sensing unit with the REHFET device, thereby reducing detection errors. In addition to assisting the ISHFET and REHFET devices in detecting extracellular action potential signals, the EXHFET device can also independently detect local field potential signals, thereby detecting more diverse cell activity signals and providing more data support for in vitro cell culture research. Attached Figure Description

[0057] Figure 1 This is a schematic diagram illustrating the steps of the fabrication method of the differential low-noise integrated biosensor array in the embodiment;

[0058] Figure 2 This is a schematic diagram of the epitaxial wafer obtained by performing step S1 in the embodiment;

[0059] Figure 3 This is a schematic diagram of the ISHFET mesa, REHFET mesa, and EXHFET mesa obtained by performing step S2 in the embodiment;

[0060] Figure 4 This is a schematic diagram of the process effect obtained by performing step S4 in the embodiment;

[0061] Figure 5 This is a schematic diagram of the process effect obtained by performing step S6 in the embodiment;

[0062] Figure 6 This is a schematic diagram illustrating the process effect obtained by performing step S7 in the embodiment;

[0063] Figure 7 This is a schematic diagram of the process effect obtained by performing step S8 in the embodiment;

[0064] Figure 8 This is a schematic diagram of the ISHFET device in the embodiment;

[0065] Figure 9 This is a schematic diagram of the REHFET device in the embodiment;

[0066] Figure 10 This is a front view of the structure of the EXHFET device in the embodiment;

[0067] Figure 11 This is a top view of the structure of the EXHFET device in the embodiment;

[0068] Figure 12 This is a side view of the structure of the EXHFET device in the embodiment;

[0069] Figure 13 This is a schematic diagram showing the connection between the signal processing module and the ISHFET, REHFET, and EXHFET devices in the embodiment. Detailed Implementation

[0070] Terminology Explanation:

[0071] FET: an abbreviation for Field Effect Transistor, which is a voltage-controlled semiconductor device that utilizes the field effect. It can be further subdivided into several types.

[0072] ISHFET: an abbreviation for Ion-Sensitive Heterojunction FET, which stands for Ion-Sensitive High Electron Mobility Heterojunction Field Effect Transistor. It is a FET that combines heterojunction and ion-sensitive characteristics and can be used for biochemical sensing.

[0073] REHFET: Abbreviation for Recessed-Gate Heterojunction FET, which means a heterojunction field-effect transistor with a recessed gate structure. It is equivalent to an ISHFET with a recessed gate structure. Like ISHFET, it can be used for biochemical sensing. Due to the recessed gate structure, REHFET can optimize radio frequency performance or reduce gate capacitance.

[0074] EXHFET: an abbreviation for Extended Gate Heterojunction FET, which refers to a heterojunction field-effect transistor with an extended gate structure.

[0075] I. Technical Problems and Solutions

[0076] Currently, signal acquisition elements used in neuroelectrophysiology mainly include traditional microelectrodes based on metal / glass materials and various field-effect transistor (FET) structures. Traditional microelectrode arrays (MEAs) rely on passive detection principles, depending on electrochemical impedance spectroscopy to acquire neural electrical signals. The signal quality of this method is highly dependent on the electrode-electrolyte interface impedance: too low an impedance makes signal acquisition difficult, while too high an impedance easily introduces significant noise. Furthermore, this type of technology generally suffers from low signal acquisition rates and insufficient long-term stability.

[0077] In contrast, field-effect transistors (FETs) employ an active detection mechanism, modulating the potential at the interface between the gate-sensitive membrane and the electrolyte by varying neural potentials, thereby controlling the channel conductance and converting it into a measurable current signal output. This approach offers significant advantages in signal stability, signal-to-noise ratio, and acquisition speed; its overall performance primarily depends on the transconductance, noise level, and interface characteristics of the FET device. Therefore, current technological development focuses on utilizing novel semiconductor materials to construct FET structures with superior electrical properties, aiming to achieve more accurate, rapid, and stable neural signal acquisition.

[0078] Traditional microelectrodes require direct insertion into biological tissues for measurement, which not only easily causes tissue trauma and immune responses during implantation, but also often leads to signal degradation and decreased detection reliability in the in vivo environment due to factors such as biological contamination and interface instability.

[0079] AlGaN / GaN heterojunction field-effect transistors (ISHFETs) have shown great potential in the field of biochemical sensing due to their material advantages. The two-dimensional electron gas formed at their interface possesses high carrier concentration and high mobility, enabling label-free, rapid, and stable electrical responses to charged biomolecules. Given that neural signals are essentially the flow of charged ions across the cell membrane, using ISHFET devices for neural electrical signal acquisition is both theoretically sound and technically feasible.

[0080] In summary, current neuroelectrophysiological techniques still face significant challenges in detecting the electrical activity of single cells and populations of neurons, particularly in terms of signal-to-noise ratio, acquisition speed, and stability. In vitro cell culture biosensor arrays based on ISHFET, REHFET, and EXHFET devices can be used. These arrays are designed to achieve high signal-to-noise ratio, rapid, and stable neural signal recording while maintaining cell viability. Furthermore, they integrate simultaneous detection of multiple parameters, including pH, specific biomolecule concentrations, and neural electrical signals in the culture environment, without increasing process complexity. This provides an advanced integrated solution for neuroscience research and high-throughput drug screening.

[0081] Therefore, it is possible to consider manufacturing a differential low-noise integrated biosensor array for cell culture detection. Nerve cells are cultured in biological buffer (PBS), which simulates the in vivo environment. The differential low-noise integrated biosensor array is then placed in the PBS, allowing nerve cells to be cultured on the array. The electrical potential signals generated by nerve cell activity can then be detected by the differential low-noise integrated biosensor array. Because the differential low-noise integrated biosensor array detects electrical potential signals from in vitro cultured nerve cells, it avoids interference from factors such as tissue trauma, immune responses, biological contamination, and interfacial instability, achieving good reliability.

[0082] II. Fabrication Method of Differential Low-Noise Integrated Biosensor Array

[0083] To fabricate a differential low-noise integrated biosensor array suitable for cell culture detection, the following steps can be performed: Figure 1 The method for fabricating a differential low-noise integrated biosensor array is shown.

[0084] Reference Figure 1 The fabrication method of the differential low-noise integrated biosensor array includes the following steps:

[0085] S1. A GaN buffer layer, an AlN insertion layer, and an AlGaN barrier layer are sequentially deposited on a sapphire substrate to obtain an epitaxial wafer;

[0086] S2. Under the conditions of Cl2 and CF4 gas environment, the epitaxial wafer is etched down to the GaN buffer layer through photolithography and dry etching processes to form multiple spaced ISHFET mesa, REHFET mesa and EXHFET mesa;

[0087] S3. Photolithography and electron beam evaporation are performed on the AlGaN barrier layer to deposit a metal thin film in a high vacuum environment, and the metal thin film is patterned using a lift-off process to obtain the source and drain electrodes on the AlGaN barrier layer.

[0088] S4. Perform rapid thermal annealing on the epitaxial wafer and source / drain electrodes to form ohmic contacts;

[0089] S5. Seed crystal layer is deposited in a high vacuum environment by photolithography and electron beam evaporation process, thick gold film is electroplated by secondary photolithography and electroplating process, and the thick gold film is patterned by lift-off process to obtain source and drain leads, pads and extended gate.

[0090] S6. A silicon dioxide thin film is grown on the surface of the epitaxial wafer using atomic layer deposition. The silicon dioxide isolation layer is prepared by placing it in a gas environment of SiH4 and N2O, a vacuum environment, and a high temperature environment to obtain an insulating passivation layer.

[0091] S7. Using a dry etching process, CF4 gas is used to etch the outline of the patterned insulating layer on the insulating passivation layer to ensure its accuracy.

[0092] S8. Using a wet etching process, the insulating passivation layer is patterned with a buffer oxide etching solution to prepare the ISHFET window, EXHFET window and the window corresponding to each pad.

[0093] S9. By using photoelectrochemical oxidation, the surface of the AlGaN barrier layer under the ISHFET window is lightly etched in an environment of ultraviolet lamp irradiation and deionized water solution to obtain the ISHFET recessed gate.

[0094] The process effects of steps S1-S9 are as follows: Figures 2-7 As shown.

[0095] In step S1, refer to Figure 2 An epitaxial wafer is obtained by sequentially depositing a GaN buffer layer, an AlN insertion layer, and an AlGaN barrier layer on a sapphire substrate.

[0096] In step S2, the etching depth can be set to 320nm to etch the epitaxial wafer down to the GaN buffer layer, thereby achieving the desired result. Figure 3 As shown, multiple ISHFET mesa, REHFET mesa, and EXHFET mesa are obtained. Among them, the ISHFET mesa is the basis for the final fabricated ISHFET device, the REHFET mesa is the basis for the final fabricated REHFET device, and the EXHFET mesa is the basis for the final fabricated EXHFET device.

[0097] In step S3, Figure 3 Based on this, a metal thin film is deposited on the AlGaN barrier layer. Specifically, the metal thin film is made of Ti (150 nm thick), Al (800 nm thick), Ni (200 nm thick), or Au (600 nm thick). The metal thin film is patterned using a lift-off process to obtain the source and drain electrodes located on the AlGaN barrier layer.

[0098] Next, step S4 is performed, where rapid thermal annealing is applied to the epitaxial wafer and the source / drain electrodes to form ohmic contacts between the source / drain electrodes and the heterojunction of the epitaxial wafer (composed of a GaN buffer layer, an AlN insertion layer, and an AlGaN barrier layer). The resulting structure is as follows: Figure 4 As shown.

[0099] Specifically, the source and drain on the ISHFET mesa eventually become the ISHFET source and ISHFET drain (i.e., the source and drain on the ISHFET device), the source and drain on the REHFET mesa eventually become the REHFET source and REHFET drain (i.e., the source and drain on the REHFET device), and the source and drain on the EXHFET mesa eventually become the EXHFET source and EXHFET drain (i.e., the source and drain on the EXHFET device).

[0100] In step S5, Figure 4 The image shows a seed layer deposited on one side. Specifically, the seed layer can be made of Cr (3 nm thick), Al (100 nm thick), Ti (50 nm thick), Pt (30 nm thick), Ti (50 nm thick), Pt (100 nm thick), Au (650 nm thick), or Ti (30 nm thick). Next, a thick gold film is electroplated onto the seed layer. This thick gold film is then patterned to form the source / drain leads, pads, and extended gate.

[0101] Specifically, the source and drain leads formed by the thick gold film include an ISHFET source lead (one end leading out from the ISHFET source), an ISHFET drain lead (one end leading out from the ISHFET drain), a REHFET source lead (one end leading out from the REHFET source), a REHFET drain lead (one end leading out from the REHFET drain), an EXHFET source lead (one end leading out from the EXHFET source), and an EXHFET drain lead (one end leading out from the EXHFET drain); the source and drain leads formed by the thick gold film include... The pads specifically include the ISHFET source pad (connected to the other end of the ISHFET source lead), the ISHFET drain pad (connected to the other end of the ISHFET drain lead), the REHFET source pad (connected to the other end of the REHFET source), the REHFET drain pad (connected to the other end of the REHFET drain lead), the EXHFET source pad (connected to the other end of the EXHFET source lead), and the EXHFET drain pad (connected to the other end of the EXHFET drain lead).

[0102] In step S6, Figure 5 Based on this, an atomic layer deposition method is used to grow a silicon dioxide thin film on the surface of the epitaxial wafer. The silicon dioxide isolation layer is prepared by placing it in a gas environment of SiH4 and N2O, a vacuum environment, and a high temperature environment, thus obtaining an insulating passivation layer of silicon dioxide (SiO2).

[0103] The insulating passivation layer obtained in step S6 covers the heterojunction, source and drain leads, and various pads. To expose these pads for connection to the signal processing module and to create openings such as ISHFET and EXHFET windows, steps S7 and S8 are performed. First, a dry etching process is used with CF4 gas to etch a patterned outline of the insulating layer onto the passivation layer to ensure its precision. Figure 6 The structure shown is then used in a wet etching process to pattern the insulating passivation layer with a buffered oxide etchant, thereby fabricating the ISHFET window, EXHFET window, and the windows corresponding to each pad, thus obtaining... Figure 7 The structure shown.

[0104] Finally, step S9 is performed, in which the surface of the AlGaN barrier layer under the ISHFET window is shallowly etched in an environment of ultraviolet lamp irradiation and deionized water solution through photoelectrochemical oxidation process to obtain the ISHFET recessed gate.

[0105] By executing steps S1-S9, multiple ISHFET devices, multiple REHFET devices, and multiple EXHFET devices can be fabricated on the same substrate. These devices can be used to detect potential signals generated by nerve cell activity. III. Differential Low-Noise Integrated Biosensor Array

[0106] In this embodiment, the structure of each ISHFET device, each REHFET device, and each EXHFET device in the differential low-noise integrated biosensor array fabricated through steps S1-S9 will be described. Since the structures of different ISHFET devices, different REHFET devices, and different EXHFET devices are identical when errors are ignored, the description will focus on the structure of one ISHFET device, one REHFET device, and one EXHFET device. For example, unless explicitly stated otherwise, when referring to the ISHFET source lead and the ISHFET drain lead, it refers to the ISHFET source lead and the ISHFET drain lead within the same ISHFET device.

[0107] In this embodiment, since each ISHFET device, each REHFET device, and each EXHFET device are fabricated simultaneously on the same substrate during steps S1-S9, the substrate in different ISHFET, REHFET, and EXHFET devices is actually equivalent to the portion of the same substrate located in these devices. The same applies to structures such as the GaN buffer layer, AlN insertion layer, and AlGaN barrier layer. In this embodiment, the portion of the insulating passivation layer located in the ISHFET device is called the first insulating passivation layer, the portion located in the REHFET device is called the second insulating passivation layer, and the portion located in the EXHFET device is called the third insulating passivation layer.

[0108] 1. Structure of ISHFET device

[0109] In this embodiment, the structure of the ISHFET device is as follows: Figure 8 As shown. (Refer to...) Figure 8 An ISHFET device comprises an AlGaN / GaN heterojunction, an ISHFET source lead, an ISHFET drain lead, and a first insulating passivation layer.

[0110] Reference Figure 8 The AlGaN / GaN heterojunction comprises a GaN buffer layer, an AlN insertion layer, and an AlGaN barrier layer stacked sequentially from bottom to top. A two-dimensional electron gas is formed at the interface between the GaN buffer layer and the AlN insertion layer.

[0111] Reference Figure 8 As a result of step S9, the GaN buffer layer in the ISHFET device is etched with an ISHFET recessed gate.

[0112] Reference Figure 8 One end of the ISHFET source lead forms an ohmic contact with the AlGaN / GaN heterojunction, and the connection point that forms the ohmic contact is the ISHFET source.

[0113] Reference Figure 8 One end of the drain lead of the ISHFET forms an ohmic contact with the AlGaN / GaN heterojunction, and the connection point that forms the ohmic contact is the drain of the ISHFET.

[0114] Reference Figure 8 The first insulating passivation layer covers the AlGaN / GaN heterojunction, the ISHFET source lead, and the ISHFET drain lead, and an ISHFET window is opened on the first insulating passivation layer to expose the ISHFET recessed gate.

[0115] Reference Figure 8 The ISHFET source lead is ultimately led out to the ISHFET source pad (e.g., Figure 7 One of the pads in the ISHFET (e.g., one of the pads in the ISHFET) forms the source interface of the ISHFET device; the ISHFET drain lead is finally led out to the ISHFET drain pad (e.g., one of the pads in the ISHFET). Figure 7 One of the pads in the circuit forms the drain interface of the ISHFET device. During in vitro cell culture, the ISHFET gate and the pad located in the circuit... Figure 8 In the ISHFET device, the electrolyte solution (e.g., biological buffer) above it forms the gate, meaning the electrolyte solution is part of the gate.

[0116] 2. Structure of REHFET device

[0117] In this embodiment, the structure of the REHFET device is as follows: Figure 9 As shown. (Refer to...) Figure 8 and Figure 9 The structure of a REHFET device is basically the same as that of an ISHFET, the difference being that in a REHFET device, the second insulating passivation layer does not have a window. Figure 9 The shaded area, like the surrounding second insulating passivation layer, is SiO2. Alternatively, similar to ISHFET devices, a window is created in the second insulating passivation layer to form a REHFET window. However, this REHFET window is modified with a functional material that is low in sensitivity to neural signals and high in sensitivity to environmental interference, connected to the AlGaN barrier layer. Figure 9 The shaded areas in the image represent functional materials.

[0118] Reference Figure 9 The REHFET source lead is ultimately led out to the REHFET source pad (e.g., Figure 7 One of the pads in the middle), forming the source interface of the REHFET device; the REHFET drain lead is finally led out to the REHFET drain pad (e.g., Figure 7 One of the pads in the circuit forms the drain interface of the REHFET device. During in vitro cell culture, the pad located in... Figure 9 In a REHFET device, the electrolyte solution (e.g., a biological buffer) above the device forms the gate, meaning the electrolyte solution is part of the gate.

[0119] 3. Structure of EXHFET devices

[0120] In this embodiment, the structure of the EXHFET device is as follows: Figure 10 , Figure 11 and Figure 12 As shown. Among them. Figure 10 This is a front view of the EXHFET device (viewed from the Y-axis of the XYZ coordinate system). Figure 11 This is a top view of the EXHFET device (viewed from the Z-axis of the XYZ coordinate system). Figure 12 This is a side view of the EXHFET device (viewed from the X-axis of the XYZ coordinate system). In this embodiment, the direction of the X-axis is referred to as the first direction, and the direction of the Y-axis is referred to as the second direction.

[0121] Reference Figure 10 , Figure 11 and Figure 12 An EXHFET device comprises an AlGaN / GaN heterojunction, an EXHFET source lead, an EXHFET drain lead, a third insulating passivation layer, and a metal extended gate.

[0122] Reference Figure 10 , Figure 11 and Figure 12 The AlGaN / GaN heterojunction comprises a GaN buffer layer, an AlN insertion layer, and an AlGaN barrier layer stacked sequentially from bottom to top. A two-dimensional electron gas is formed at the interface between the GaN buffer layer and the AlN insertion layer.

[0123] Reference Figure 10 , Figure 11 and Figure 12 One end of the EXHFET source lead forms an ohmic contact with the AlGaN / GaN heterojunction, and the connection point that forms the ohmic contact is the EXHFET source.

[0124] Reference Figure 10 , Figure 11 and Figure 12 One end of the EXHFET drain lead forms an ohmic contact with the AlGaN / GaN heterojunction, and the connection point that forms the ohmic contact is the EXHFET drain.

[0125] Reference Figure 11 Two metal extension grilles are provided in the second direction (Y-axis). Alternatively, only one metal extension grille can be provided, or more metal extension grilles can be arranged in the second direction (Y-axis).

[0126] Reference Figure 10 , Figure 11 and Figure 12 The third insulating passivation layer covers the AlGaN / GaN heterojunction, the EXHFET source lead, and the EXHFET drain lead, and an EXHFET window with an exposed metal extended gate is opened on the third insulating passivation layer.

[0127] Reference Figure 10 , Figure 11 and Figure 12 The EXHFET source lead is ultimately led out to the EXHFET source pad (e.g., Figure 7One of the pads in the EXHFET (e.g., one of the pads in the EXHFET) forms the source interface of the EXHFET device; the EXHFET drain lead is finally led out to the EXHFET drain pad (e.g., one of the pads in the EXHFET). Figure 7 One of the pads in the circuit forms the drain interface of the EXHFET device. During in vitro cell culture, the metal extended gate and the pad located in the circuit... Figure 8 In the case of an EXHFET device, an electrolyte solution (e.g., a biological buffer solution) is placed above the device to form the gate, meaning the electrolyte solution is part of the gate.

[0128] In an EXHFET device, the extended metal gate, together with the electrolyte solution, forms the gate of the EXHFET device. Similarly, in an ISHFET device, the recessed ISHFET gate, also together with the electrolyte solution, forms the gate of the ISHFET device. Compared to the recessed ISHFET gate, the extended metal gate has a larger gate sensing area, enabling the EXHFET device to respond to changes in the local field potential signal of cells in the solution, ensuring stable and reliable operation of the EXHFET device under the control of the extended metal gate.

[0129] In this embodiment, inert metal materials such as gold (Au) and platinum (Pt) or inert materials such as indium oxide (In2O3) nanowires can be used to manufacture metal extended gates, thereby reducing the toxic effects on nerve cells.

[0130] 4. Component connection method

[0131] In this embodiment, the differential low-noise integrated biosensor array may include the same number of ISHFET devices and REHFET devices (e.g., 40 ISHFET devices and 40 REHFET devices), and may include fewer EXHFET devices (e.g., 8 EXHFET devices) as auxiliary to the ISHFET devices and REHFET devices.

[0132] In this embodiment, a processor chip with multiple input pins is used as the signal processing module, and each ISHFET device, REHFET device, and EXHFET device is connected to the signal processing module. For example, refer to... Figure 13 The source pads of each ISHFET, REHFET, and EXHFET device can share one input pin of the signal processing module, while the drain pads of different devices are each connected to one input pin of the signal processing module, thus saving pins.

[0133] In this embodiment, when using a differential low-noise integrated biosensor array, the gates of the ISHFET, REHFET, and EXHFET devices are all electrolyte solutions used in in vitro cell culture. The activity of the cells will affect the gates of each device, thereby affecting parameters such as the source-drain voltage of each device, which are read and processed by the signal processing module.

[0134] In this embodiment, refer to Figure 13 When the number of ISHFET devices and REHFET devices is the same, they can be combined one by one to form a differential sensing unit. That is, a differential sensing unit contains one ISHFET device and one REHFET device, and any ISHFET device or REHFET device belongs to only one corresponding differential sensing unit.

[0135] In this embodiment, different numbers of ISHFET devices and REHFET devices can be used, for example, fewer REHFET devices and more ISHFET devices. In this way, each ISHFET device and each REHFET device can be combined into a differential sensing unit in a one-to-many configuration. That is, a differential sensing unit contains one ISHFET device and one REHFET device. Each ISHFET device belongs to only one differential sensing unit, while each REHFET device can belong to multiple different differential sensing units.

[0136] In this embodiment, even with a differential sensing unit, the signal processing module can still independently read the source-drain voltage and other signals detected by each ISHFET device, REHFET device, and EXHFET device. For the source-drain voltage and other signals detected by the ISHFET and REHFET devices within the same differential sensing unit, the signal processing module can perform differential processing, thereby helping to reduce errors.

[0137] IV. Working Principle of Differential Low-Noise Integrated Biosensor Array

[0138] Cellular neural electrical signals mainly include two categories: one is the extracellular action potential, which characterizes neuronal firing, and the other is the local field potential (LFP), which reflects the synchronous activity of the neural network.

[0139] ISHFET device structure and sensing mechanism:

[0140] AlGaN / GaN heterojunction and formation of two-dimensional electron gas (2DEG):

[0141] Both AlGaN and GaN possess a wurtzite crystal structure, and their low symmetry endows the materials with significant spontaneous polarization characteristics. When an AlGaN layer is epitaxially grown on GaN to form a heterojunction, the stress caused by lattice mismatch induces additional piezoelectric polarization. The combined effect of spontaneous polarization and piezoelectric polarization induces a high-concentration, high-mobility two-dimensional electron gas (2DEG) near the GaN side of the heterojunction interface. This 2DEG layer is extremely sensitive to changes in gate voltage. The charges (positive or negative ions) adsorbed on the gate surface directly modulate the electron concentration in the channel 2DEG. Since the electron mobility in the 2DEG is much higher than the charge mobility on the gate surface (usually by orders of magnitude), even small charge fluctuations on the gate surface can cause significant changes in the 2DEG conductivity, thereby effectively "amplifying" the signal and ultimately reflecting it as source-drain current (Idrain). DS ) changes.

[0142] Active detection mechanism of neural signals:

[0143] In neural sensing applications, the gate region (solution gate) of an ISHFET device is in close contact with the neuron. When the neuron is stimulated and generates an action potential, it is mainly manifested as Na+. + Inflow (depolarization) and K + Outflow (repolarization) is usually accompanied by Ca 2+ Inflow (such as during synaptic transmission). This transmembrane flow of ions instantly alters the local potential of the electrolyte solution in the gap between the neuron and the ISHFET gate region. This change in solution potential is equivalent to the effective gate voltage (V) applied to the ISHFET solution gate. G The gate voltage change highly sensitively modulates the conductivity of the channel 2DEG, which in turn affects the source-drain current (Id) of the ISHFET. DS ) undergoes corresponding changes. Detection I DS Changes in these parameters can enable highly sensitive and rapid detection of neuronal action potentials.

[0144] When the differential low-noise integrated biosensor array is in operation, nerve cells are cultured on the array, and their sensing sites are all placed in biological buffered saline (PBS) to simulate the in vivo environment. The top of the ISHFET gate in the ISHFET device is closest to the nerve cell. After power is turned on, biological neural signals are acquired through the source and drain electrodes of the ISHFET, REHFET, and EXHFET devices.

[0145] The working principle of an ISHFET device is as follows: The source of an ISHFET device shares a common lead and pad, while the drain has its own separate lead. Current flows into the ISHFET device through the source lead and the ohmic contact electrode, passes through the two-dimensional electron gas structure between the AlN insertion layer and the GaN buffer layer, and then flows out through the drain ohmic contact and the drain lead. When a living organism receives stimulation, the neuron at the top of the concave gate 9 of the ISHFET device generates an action potential (nerve signal). The extracellular potential (nerve signal) of the neuron rapidly and sensitively modulates the two-dimensional electron gas structure between the AlN insertion layer and the GaN buffer layer, causing a change in the conductivity of the two-dimensional electron gas, which in turn causes a change in the source-drain current of the ISHFET device. In other words, the change in the source-drain current of the ISHFET device reflects the change in the neural signal of the living neuron. The extracellular potential V can be calculated using a formula. GS Size:

[0146]

[0147] Among them, I ds For source and drain current, μ n C represents the electron mobility of a two-dimensional electron gas. ox V is the capacitance of the two-dimensional electron gas to the surface of the AlGaN barrier layer, W is the width of the ISHFET device, L is the channel length of the ISHFET device, and V is the capacitance of the two-dimensional electron gas to the surface of the AlGaN barrier layer. GS Extracellular potential (nerve signal), V TH V is the threshold voltage of the ISHFET device. DS The source-drain voltage is the extracellular action potential signal detected by the ISHFET device.

[0148] Since the ISHFET device is essentially a gallium nitride low-noise sensor, its noise analysis reveals a larger V. G Voltage increases the 1 / f noise of the sensor. A fixed bias voltage V is applied to the source and drain of the gallium nitride sensor device. DS Sampling and detection I DS The current flows for a period of time. I is calculated. DS The standard deviation is used to characterize the sensor noise σ. N Sensor signal-to-noise ratio:

[0149]

[0150] In the signal-to-noise ratio (SNR), Ψ0 is determined by the interfacial properties between the sensor and the solution, and therefore can be considered a constant. The SNR of the sensor is related to g. m / σ N Proportional. By g m Size dominates, while noise σ NThe impact is minimal; therefore, the sensor's maximum signal-to-noise ratio operating point occurs at V, which is almost the same as the sensor's maximum transconductance. G Voltage bias is applied, and the transconductance of this device can be controlled through a recessed gate, thus reducing the impact of noise. The recessed gate structure in the ISHFET device reduces the heterojunction polarization effect by thinning the AlGaN barrier layer, thereby decreasing the two-dimensional electron gas concentration and causing a positive shift in the threshold voltage. This structure allows the ISHFET device to operate normally within the sensitive range without additional metal gate or differential electrode bias.

[0151] The REHFET device operates as follows: After depositing an insulating passivation layer, the gate region of the reference field-effect transistor is not windowed, and the gate surface remains covered by the insulating passivation layer. Therefore, its sensitivity to neural signals is low, but it can still collect ambient common-mode noise and background noise from the ISHFET device. In a preferred embodiment, the REHFET gate region is windowed during the fabrication of the insulating passivation layer, and a functional material that is insensitive to neural signals but still sensitive to environmental interference is modified on the AlGaN barrier layer surface. By differentially processing the signals collected by the ISHFET device and the REHFET device, common-mode noise can be effectively suppressed, and a purer neural signal can be extracted, thereby improving the system signal-to-noise ratio.

[0152] The working principle of the EXHFET device is as follows: The EXHFET device has a metal gate structure, and its sensing region surface is covered with an Au layer after BOE etching. This structure not only has excellent solution stability but also has no significant toxicity to cells. When subjected to biological stimulation, neurons near the EXHFET extended gate generate local field potentials. This potential change can rapidly modulate the two-dimensional electron gas between the AlN insertion layer and the GaN buffer layer, causing a change in conductivity, which in turn modulates the source-drain current of the EXHFET. Therefore, the source-drain current response of the EXHFET directly reflects the neuronal electrical activity signal. This device can not only detect extracellular action potentials in conjunction with ISHFETs but also independently acquire local field potential (LFP) signals. The extended gate structure increases the gate sensing area, enabling it to respond to overall local field potential changes in cells in solution, ensuring the stable and reliable operation of the EXHFET device under metal gate modulation.

[0153] Based on the above principles, when nerve cells are cultured in vitro in a cell culture solution and a differential low-noise integrated biosensor array is placed in the cell culture solution, the activity of the nerve cells will affect the gates of the ISHFET, REHFET, and EXHFET devices. This allows the source and drain of the ISHFET device to detect the extracellular action potential signal of the nerve cell; similarly, the source and drain of the REHFET device and the EXHFET device can also detect the extracellular action potential signal of the nerve cell. Furthermore, because the EXHFET device has a larger metal extended gate, its source and drain can also detect the local field potential signal of the nerve cell. Therefore, ISHFET and REHFET devices can be combined to form a differential sensing unit. The signal processing module can perform differential processing on the extracellular action potential signals detected by the ISHFET and REHFET devices in the differential sensing unit to obtain extracellular action potential signals with small errors. The signal processing module can also detect the local field potential signal through the EXHFET device.

[0154] For extracellular action potential signals and local field potential signals, the signal processing module can further perform processing such as storage, graphics, filtering, and waveform analysis. For example, for an EXHFET device, the signal processing module can acquire the V detected by the device. DS and I DS By calculating V DS / I DS The channel resistance of the extended gate in an EXHFET device is calculated. Since the extended gate surface is topped with inert metal Au, different organic substances can adhere better after external modification, thus altering the channel resistance. Because the channel resistance of the extended gate in an EXHFET device is typically related to the concentration of the electrolyte solution used in in vitro cell culture, the signal processing module can use the detected channel resistance to look up tables or calculate using formulas to determine the concentration of the electrolyte solution.

[0155] This invention provides a low-noise, integrated bio-neural sensor based on an AlGaN / GaN heterojunction ion-sensitive field-effect transistor (ISHFET) and its fabrication method. This sensor utilizes a high-mobility two-dimensional electron gas at the AlGaN / GaN heterojunction interface to actively detect neural signals, exhibiting high signal-to-noise ratio, fast response, and high stability. To effectively suppress common-mode environmental noise, this invention also integrates an AlGaN / GaN reference field-effect transistor (REHFET) with a structure similar to the ISHFET but different only in the gate region, and an AlGaN / GaN extended-gate field-effect transistor (EXHFET) with a metal layer deposited above the gate to form an extended gate structure. When a potential change occurs outside the nerve cell membrane, the ISHFET and EXHFET can jointly sense the potential change and effectively cancel external interference through a differential detection mechanism, significantly improving signal quality. Furthermore, thanks to the excellent biocompatibility and chemical stability of AlGaN and GaN materials, this sensing system supports long-term in vitro culture of nerve cells and can continuously and stably acquire weak neural electrical signals. The ISHFET, REHFET, and EXHFET devices used have the following effects:

[0156] 1. AlGaN / GaN ISHFET device design employs photoelectrochemical etching with recessed gates to adjust the device operating point and reduce device size; reduces signal crosstalk between devices by adjusting the spacing between AlGaN / GaN ISHFETs; and improves the signal-to-noise ratio by adjusting the shape of the recessed gate region.

[0157] 2. AlGaN / GaN REHFET device design: By adjusting the distance between AlGaN / GaN ISHFET, AlGaN / GaN REHFET, and AlGaN / GaN EXHFET, crosstalk between devices is reduced, further improving the acquisition signal-to-noise ratio of AlGaN / GaN ISHFET and AlGaN / GaN EXHFET;

[0158] 3. Share the source leads of AlGaN / GaN ISHFET devices, AlGaN / GaN REHFET devices, and AlGaN / GaN EXHFET devices in the same group to reduce the number of leads and further reduce the device size;

[0159] 4. AlGaN / GaN EXHFET device design: An extended gate structure is adopted. Due to the larger area of ​​the sensing region, it can better sense the local field potential signal, which can cooperate with the extracellular action potential sensed by the AlGaN / GaN ISHFET to better acquire the corresponding signal.

[0160] 5. The process is less complex, allowing for better mass production.

[0161] 6. In electrophysiology, the potential generated by nerve cells is divided into changes in extracellular action potential and changes in local field potential. Therefore, compared with existing nerve electrodes in electrophysiological techniques, such as passive nerve electrodes and active silicon-based nerve electrodes, the AlGaN / GaN ISHFET of this invention can acquire biological nerve signals (extracellular action potential of nerve cells) more quickly and stably. After differential processing with AlGaN / GaN REHFET, a high signal-to-noise ratio nerve signal can be obtained. Compared with AlGaN / GaN ISHFET, AlGaN / GaN EXHFET, although less sensitive to extracellular action potential, can better sense local field potential (LFP). The biological nerve signals acquired by AlGaN / GaN ISHFET and AlGaN / GaN EXHFET are transconductance amplified signals, rather than the original weak signals, thus reducing the difficulty of back-end circuit design to a certain extent. AlGaN has better biocompatibility with GaN materials, enabling longer signal acquisition. Furthermore, since the top layer of the extended gate is made of inert metal Au, it also has excellent biocompatibility. In addition, changes in the corresponding cells in the solution will make the changes on the extended gate exceptionally obvious.

[0162] 7. Integrating sensor arrays without adding process steps increases the number of signal acquisition channels, allowing multiple signals to be acquired on a single sensor array, such as weak biological electrical signals, solution pH value, solution substance concentration, and other data.

[0163] 8. When preparing source and drain leads and pads in step S5, using a non-annealing process can better suppress interface reactions;

[0164] 9. Because the likelihood of identical devices appearing in the manufacturing process is low, each sensor chip has a corresponding optimal transconductance and optimal signal-to-noise ratio. Therefore, the self-adjustment of the signal-to-noise ratio could be achieved during the detection process by comparing different source-drain voltages (V). DS ) and the corresponding source-drain current (I DS This allows for adjustment of the signal-to-noise ratio to its optimal value;

[0165] 10. In addition to being used to detect weak electrical signals generated by nerve cell activity during in vitro cell culture, the differential low-noise integrated biosensor array in this embodiment can also be used as a pH detection chip or a solution substance concentration detection chip for cell culture medium.

[0166] In a preferred embodiment of the present invention, the differential low-noise integrated biosensor array chip integrates 40 ISHFET devices and 40 REHFET devices. Each pair of ISHFETs and REHFETs constitutes a differential sensing unit for acquiring electrical signals from single or multiple nerve cells. Additionally, eight EXHFET devices are used to detect the local field potential of nerve cells in solution. Furthermore, four (4) devices are arranged in two groups to monitor the pH value and specific biomolecule concentrations of the cell culture environment in real time. This design achieves multi-parameter, high signal-to-noise ratio, and rapid and stable synchronous detection of neural signals on a single chip.

[0167] It should be noted that, unless otherwise specified, when a feature is referred to as "fixed" or "connected" to another feature, it can be directly fixed or connected to the other feature, or indirectly fixed or connected to the other feature. Furthermore, the descriptions of "upper," "lower," "left," and "right" used in this disclosure are only relative to the relative positional relationships of the components of this disclosure in the accompanying drawings. The singular forms "a," "an," and "the" used in this disclosure are also intended to include the plural forms, unless the context clearly indicates otherwise. Moreover, unless otherwise defined, all technical and scientific terms used in this embodiment have the same meaning as commonly understood by one of ordinary skill in the art. The terminology used in this embodiment specification is only for describing specific embodiments and is not intended to limit the embodiments of the invention. The term "and / or" as used in this embodiment includes any combination of one or more of the associated listed items.

[0168] It should be understood that although the terms first, second, third, etc., may be used to describe various elements in this disclosure, these elements should not be limited to these terms. These terms are only used to distinguish elements of the same type from each other. For example, a first element may also be referred to as a second element without departing from the scope of this disclosure, and similarly, a second element may also be referred to as a first element. The use of any and all instances or exemplary language (“e.g.,” “such as,” etc.) provided in this embodiment is intended only to better illustrate embodiments of the invention and, unless otherwise required, does not impose a limitation on the scope of embodiments of the invention.

[0169] It should be recognized that embodiments of the present invention can be implemented or carried out by computer hardware, a combination of hardware and software, or by computer instructions stored in a non-transitory computer-readable storage medium. The method can be implemented using standard programming techniques—including a non-transitory computer-readable storage medium configured with a computer program, wherein such a storage medium causes the computer to operate in a specific and predefined manner—according to the methods and drawings described in the specific embodiments. Each program can be implemented in a high-level procedural or object-oriented programming language to communicate with the computer system. However, if desired, the program can be implemented in assembly or machine language. In any case, the language can be a compiled or interpreted language. Furthermore, for this purpose, the program can run on a programmed application-specific integrated circuit (ASIC).

[0170] Furthermore, the procedures described in this embodiment can be performed in any suitable order, unless otherwise indicated by this embodiment or otherwise obviously contradictory to the context. The procedures (or variations and / or combinations thereof) described in this embodiment can be executed under the control of one or more computer systems configured with executable instructions, and can be implemented by hardware or a combination thereof as code (e.g., executable instructions, one or more computer programs, or one or more applications) that commonly executes on one or more processors. A computer program includes multiple instructions executable by one or more processors.

[0171] Furthermore, the method can be implemented in any suitable type of computing platform, including but not limited to personal computers, minicomputers, mainframes, workstations, networked or distributed computing environments, standalone or integrated computer platforms, or in communication with charged particle tools or other imaging devices, etc. Aspects of embodiments of the invention can be implemented as machine-readable code stored on a non-transitory storage medium or device, whether removable or integrated into a computing platform, such as a hard disk, optical read and / or write storage medium, RAM, ROM, etc., such that it is readable by a programmable computer, and when the storage medium or device is read by the computer, it can be used to configure and operate the computer to perform the processes described herein. Furthermore, the machine-readable code, or portions thereof, can be transmitted via wired or wireless networks. The invention of this embodiment includes these and other different types of non-transitory computer-readable storage media when such media comprises instructions or programs that implement the steps above in conjunction with a microprocessor or other data processor. Embodiments of the invention also include the computer itself when programmed according to the methods and techniques of embodiments of the invention.

[0172] A computer program can be applied to input data to perform the functions of this embodiment, thereby transforming the input data to generate output data stored in non-volatile memory. The output information can also be applied to one or more output devices, such as a display. In a preferred embodiment of the invention, the transformed data represents physical and tangible objects, including a specific visual depiction of physical and tangible objects generated on the display.

[0173] The above are merely preferred embodiments of the present invention. The embodiments of the present invention are not limited to the above-described implementations. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the embodiments of the present invention, as long as they achieve the same technical effects, should be included within the scope of protection of the embodiments of the present invention. Within the scope of protection of the embodiments of the present invention, the technical solutions and / or implementation methods can have various modifications and variations.

Claims

1. A differential low-noise integrated biosensor array for cell culture detection, characterized in that, The differential low-noise integrated biosensing array comprises: a substrate; a plurality of ISHFET devices on the substrate; the ISHFET devices are used to detect extracellular action potential signals of nerve cells in a cell culture solution; a plurality of REHFET devices on the substrate; the REHFET devices are used to detect extracellular action potential signals of nerve cells in a cell culture solution; a plurality of EXHFET devices on the substrate; the EXHFET devices are used to detect extracellular action potential signals and / or local field potential signals of nerve cells in a cell culture solution.

2. The differential low-noise integrated biosensor array of claim 1, wherein, The ISHFET device comprises: an AlGaN / GaN heterojunction; the AlGaN / GaN heterojunction comprises a GaN buffer layer, an AlN insertion layer and an AlGaN barrier layer stacked in order from bottom to top; a two-dimensional electron gas is formed at the interface between the GaN buffer layer and the AlN insertion layer; the GaN buffer layer is etched with an ISHFET recessed gate; an ISHFET source lead; one end of the ISHFET source lead forms an ohmic contact with the AlGaN / GaN heterojunction; an ISHFET drain lead; one end of the ISHFET drain lead forms an ohmic contact with the AlGaN / GaN heterojunction; a first insulating passivation layer; the first insulating passivation layer covers the AlGaN / GaN heterojunction, the ISHFET source lead and the ISHFET drain lead, and an ISHFET window exposing the ISHFET recessed gate is opened on the first insulating passivation layer.

3. The differential low-noise integrated biosensor array of claim 2, wherein, The REHFET device comprises: an AlGaN / GaN heterojunction; the AlGaN / GaN heterojunction comprises a GaN buffer layer, an AlN insertion layer and an AlGaN barrier layer stacked in order from bottom to top; a two-dimensional electron gas is formed at the interface between the GaN buffer layer and the AlN insertion layer; a REHFET source lead; one end of the REHFET source lead forms an ohmic contact with the AlGaN / GaN heterojunction; a REHFET drain lead; one end of the REHFET drain lead forms an ohmic contact with the AlGaN / GaN heterojunction; a second insulating passivation layer; the second insulating passivation layer covers the AlGaN / GaN heterojunction, the REHFET source lead and the REHFET drain lead.

4. The differential low-noise integrated biosensor array of claim 2, wherein, The REHFET device comprises: an AlGaN / GaN heterojunction; the AlGaN / GaN heterojunction comprises a GaN buffer layer, an AlN insertion layer and an AlGaN barrier layer stacked in order from bottom to top; a two-dimensional electron gas is formed at the interface between the GaN buffer layer and the AlN insertion layer; a REHFET source lead; one end of the REHFET source lead forms an ohmic contact with the AlGaN / GaN heterojunction; a REHFET drain lead; one end of the REHFET drain lead forms an ohmic contact with the AlGaN / GaN heterojunction; A second insulating passivation layer covers the AlGaN / GaN heterojunction, the REHFET source lead and the REHFET drain lead, and a REHFET window exposing the AlGaN barrier layer is formed in the second insulating passivation layer, and a functional material connected with the AlGaN barrier layer is arranged in the REHFET window, the functional material having low sensitivity to nerve signals and high sensitivity to environmental interference.

5. The differential low-noise integrated biosensor array according to claim 3 or 4, characterized in that, The EXHFET device comprises: An AlGaN / GaN heterojunction comprising a GaN buffer layer, an AlN insertion layer and an AlGaN barrier layer arranged in sequence from bottom to top, and a two-dimensional electron gas formed at the interface between the GaN buffer layer and the AlN insertion layer; An EXHFET source lead arranged on one side of the EXHFET in a first direction, and one end of the EXHFET source lead forming an ohmic contact with the AlGaN / GaN heterojunction; An EXHFET drain lead arranged on the other side of the EXHFET in the first direction, and one end of the EXHFET drain lead forming an ohmic contact with the AlGaN / GaN heterojunction; At least one metal extended gate arranged along a second direction perpendicular to the first direction, and the metal extended gate forming an ohmic contact with the AlGaN / GaN heterojunction; A third insulating passivation layer covering the AlGaN / GaN heterojunction, and an EXHFET window exposing the metal extended gate formed in the third insulating passivation layer.

6. The differential low-noise integrated biosensor array of claim 5, wherein, The metal extended gate is an inert metal material.

7. The differential low-noise integrated biosensor array of claim 5, wherein, The differential low-noise integrated biosensing array further comprises: A plurality of ISHFET source pads connected with the other end of the ISHFET source lead; A plurality of ISHFET drain pads connected with the other end of the ISHFET drain lead; A plurality of REHFET source pads connected with the other end of the REHFET source lead; A plurality of REHFET drain pads connected with the other end of the REHFET drain lead; A plurality of EXHFET source pads connected with the other end of the EXHFET source lead; A plurality of EXHFET drain pads connected with the other end of the EXHFET drain lead.

8. The differential low-noise integrated biosensor array according to any one of claims 1-7, wherein, The differential low-noise integrated biosensing array further comprises: A signal processing module for processing the extracellular action potential signal and the local field potential signal.

9. The differential low-noise integrated biosensing array according to claim 8, wherein: Each ISHFET device and each REHFET device form a plurality of differential sensing units. Any of the differential sensing units comprises one of the ISHFET devices and one of the REHFET devices; Any of the ISHFET devices belongs to a corresponding one of the differential sensing units; Any of the REHFET devices belongs to at least one of the differential sensing units respectively; The signal processing module is configured to perform differential processing on the extracellular action potential signals detected by the ISHFET device and the REHFET device in the same differential sensing unit.

10. The method of claim 7, wherein the differential low-noise integrated biosensor array is prepared by, The preparation method comprises: Depositing a GaN buffer layer, an AlN insertion layer and an AlGaN barrier layer on a sapphire substrate in sequence to obtain an epitaxial wafer; Under the condition that the gas environment is Cl2 and CF4, the epitaxial wafer is etched to the GaN buffer layer by photolithography and dry etching process to form a plurality of ISHFET mesas, REHFET mesas and EXHFET mesas arranged at intervals; The AlGaN barrier layer is subjected to photolithography and electron beam evaporation, a metal thin film is evaporated in a high vacuum environment, and the metal thin film is patterned using a stripping process to obtain a source-drain electrode arranged on the AlGaN barrier layer; the metal thin film is made of Ti material, Al material, Ni material or Au material; the source-drain electrode comprises an ISHFET source electrode, an ISHFET drain electrode, a REHFET source electrode, a REHFET drain electrode, an EXHFET source electrode and an EXHFET drain electrode; The epitaxial wafer and the source-drain electrode are subjected to rapid thermal annealing to form an ohmic contact; A seed layer is evaporated in a high vacuum environment by photolithography and electron beam evaporation process, a thick gold film is plated using secondary lithography and electroplating process, and the thick gold film is patterned using a stripping process to obtain a source-drain electrode lead, a pad and an extended gate; the seed layer is made of Cr material, Al material, Ti material, Pt material, Ti material, Pt material, Au material or Ti material; the source-drain electrode lead comprises the ISHFET source electrode lead, the ISHFET drain electrode lead, the REHFET source electrode lead, the REHFET drain electrode lead, the EXHFET source electrode lead and the EXHFET drain electrode lead; A layer of silicon dioxide thin film is grown on the surface of the epitaxial wafer using atomic layer deposition, and a silicon dioxide isolation layer is prepared in a gas environment of SiH4 and N2O, vacuum and high temperature environment to obtain an insulating passivation layer; A dry etching process is used to etch the insulating passivation layer to obtain a patterned insulating layer profile using CF4 gas to ensure its accuracy; A wet etching process is used to pattern the insulating passivation layer using a buffer oxide etching solution to prepare the ISHFET window, the EXHFET window and the windows corresponding to each pad; A photoelectrochemical oxidation process is used to perform shallow etching on the surface of the AlGaN barrier layer under the ISHFET window in an ultraviolet lamp irradiation and deionized water solution environment to obtain the ISHFET recessed gate.

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