An optoelectrochemical detection device and application based on radiation modulation

Abstract

The present invention discloses a photoelectrochemical detection device and application based on radiation modulation, belonging to the technical field of photoelectrochemical sensing and biological detection; a detection device applicable to the detection of alpha-fetoprotein, comprising a photoelectrochemical biosensor and a photoelectrochemical test device; the photoelectrochemical biosensor includes an ATA thin film electrode; the ATA thin film electrode includes an FTO substrate; a Au film layer is provided on the upper surface of the FTO substrate, and a TiO2 film layer is provided on the upper surface of the Au film layer; a AuNPs layer is deposited on the TiO2 film layer; the photoelectrochemical test device includes an optical chopper connected to a lock-in amplifier; the lock-in amplifier is connected to a low-pass filter; the optical chopper is arranged between the light source and the photoelectrochemical biosensor; the photoelectrochemical biosensor is connected to the low-pass filter through an electrochemical workstation; the present invention uses lock-in amplification combined with the ATA thin film electrode to enable the detection of the analyte to have a lower detection limit and a larger detection range.

Description

Technical Field

[0001] The present invention belongs to the technical field of photoelectrochemical sensing and biological detection, and relates to a photoelectrochemical detection device based on radiation modulation and its application. Background Technique

[0002] Currently, in biomedical research, alpha-fetoprotein (AFP) is an important serum biomarker for the early diagnosis and prognosis of primary liver cancer in clinical analysis. Therefore, for the early diagnosis and prognosis of liver cancer in clinical analysis, the detection method of AFP concentration is required to have high sensitivity and a wide linear range. As one of the best candidate methods for the early diagnosis and prognosis of liver cancer in clinical analysis, the low background noise, fast speed, and low cost of photoelectrochemical (PEC) sensors make them have broad application prospects in clinical early diagnosis.

[0003] Photoelectrochemistry (PEC) refers to a method of applying light irradiation to a photoactive material, generating electron-hole pairs and charge transfer on the electrode under the action of photoexcitation, and detecting the generated current using an electrochemical workstation. In PEC sensing detection, light irradiation serves as the excitation signal and the electrical signal serves as the detection signal. Since the excitation source and the detection signal are completely different energy forms, the background noise is low and it has high sensitivity.

[0004] In PEC sensors, the photoactive material can perform photoelectric conversion, converting the absorbed photons into chemical energy, and thus has a large number of applications in fields such as photocatalysis, solar cells, and photoelectrochemical sensors. It is relatively common to use TiO2 in the photoactive materials for PEC sensing. Since TiO2 has advantages such as high stability, non-toxicity, and excellent photoactivity, it is currently the most promising photoactive material. However, the fatal weakness of TiO2 - its wide bandgap - limits its absorption of solar energy (only absorbing about 4%). [1] . Thus, the noble metal doping technology came into being. Research has confirmed that doping noble metal nanoparticles (Au, Pd, Ag, Cu) into semiconductor materials can, under light radiation, excite the collective oscillation of free electrons to achieve the localized surface plasmon resonance (LSPR) effect. The LSPR effect enhances the excitation of active carriers through [2] light scattering [3] local field enhancement [4-5] hot electron injection [6-7] resonance energy transfer and four mechanisms to enhance the separation probability of electron-hole pairs, thereby broadening the spectral response range of the photoactive material and enhancing the absorption efficiency of visible light. However, due to the small characteristic size and strong scattering effect of metal nanoparticles, its absorption coefficient is usually low, seriously limiting its photoelectric conversion efficiency.

[0005] For traditional PEC sensors, the photocurrent response is detected and recorded through the software of an electrochemical workstation, which requires manual on / off (shading / opening) of the light source to achieve. To ensure signal stability, the period is generally selected as 20 s. A square wave signal is generated with 10 s in the dark and 10 s under illumination. However, during the signal sampling process, switching current spikes will occur in the square wave signal due to charge recombination. In addition, since the 0 fundamental frequency and its harmonics are too close, the low-pass filter cannot separate them. Therefore, in this case, it is very difficult to suppress the harmonics. At the same time, the stability of the current response depends on human subjective judgment and is prone to errors. When the concentration of the target protein analyte continuously increases, the resistance of the sensing electrode will continuously increase. At this time, the photocurrent response will be very small and will be submerged in strong noise and cannot be recognized, thus limiting the measurement range of the sensor.

[0006] References

[0007] [1] Choi H., Shin D., Yeo B.C., Song T., Han S.S., Park N., & Kim S. Simultaneously controllable doping sites and the activity of a W–N codoped TiO2 photocatalyst. 2016, ACS Catalysis, 6(5):2745 - 2753.

[0008] [2] Zhu H., Xie H., Yang Y., Wang K., Zhao F., Ye W., & Ni W. Mapping hot electron response of individual gold nanocrystals on a TiO2 photoanode. Nano Letters, 2020, 20(4):2423 - 2431.

[0009] [3] Ko W.Y, Tien T.J, Hsu C.Y, et al. Ultrasensitive label - and amplification - free photoelectric protocols based on sandwiched layer - by - layer plasmonic nanocomposite films for the detection of alpha - fetoprotein[J]. Biosensors and Bioelectronics, 2019, 126:455 - 462.

[0010] [4] Iida K., Noda M., Ishimura K., & Nobusada K. First-principles computational visualization of localized surface plasmon resonance in gold nanoclusters. The Journal of Physical Chemistry A, 2014, 118(47): 11317-11322.

[0011] [5] Li X.D., Chen T.P., Liu Y., & Leong K.C. Influence of localized surface plasmon resonance and free electrons on the optical properties of ultrathin Au films: a study of the aggregation effect. Optics Express, 2014, 22(5): 5124-5132.

[0012] [6] Jia H., Li Z., Wang B., Xing G., Wong Y.L., Ren H. & Zhang X. Plasmonic nanohole arrays with enhanced visible light photoelectrocatalytic activity. Acs Photonics, 2022, 9(2): 652-663.

[0013] [7] Jia H., Wong Y.L., Wang B., Xing G., Tsoi C.C., Wang M., & Zhang, X. Enhanced solar water splitting using plasmon-induced resonance energy transfer and unidirectional charge carrier transport. Optics express, 2021, 29(21): 34810-34825. Summary of the Invention

[0014] The present invention overcomes the deficiencies of the prior art and provides a photoelectrochemical detection device and application based on radiation modulation. The technical problem to be solved is how to utilize the advantages of lock-in amplification, and at the same time propose a biotechnological detection device with a simple structure and easy implementation, so that the detection of alpha-fetoprotein has a lower detection limit and a larger detection range.

[0015] To achieve the above object, the present invention is realized through the following technical solutions.

[0016] A photoelectrochemical detection device based on radiation modulation includes a photoelectrochemical biosensor and a photoelectrochemical testing device; the photoelectrochemical biosensor includes an ATA thin film electrode; the ATA thin film electrode includes an FTO substrate; a Au film layer is provided on the upper surface of the FTO substrate, and a TiO2 film layer is provided on the upper surface of the Au film layer; an AuNPs layer is deposited on the TiO2 film layer; the photoelectrochemical testing device includes a lock-in amplifier, an optical chopper, and a low-pass filter; the optical chopper is connected to the lock-in amplifier; the lock-in amplifier is connected to the low-pass filter; the optical chopper is provided between the light source and the photoelectrochemical biosensor; the photoelectrochemical biosensor is connected to the low-pass filter through an electrochemical workstation.

[0017] Preferably, the thickness of the TiO2 film layer is 60-80 nm.

[0018] Preferably, the thickness of the AuNPs layer is 5-10 nm.

[0019] More preferably, the AuNPs layer is multiple AuNPs nanoparticles.

[0020] Preferably, the thickness of the Au film layer is 100-120 nm.

[0021] Preferably, the FTO substrate is conductive glass.

[0022] More preferably, the AuNPs layer is modified with alpha-fetoprotein antibody.

[0023] Preferably, the light source is a xenon lamp light source, 300 nm < λ < 1100 nm, 300 mW / cm 2 .

[0024] Preferably, the photoelectrochemical biosensor is connected as a working electrode to a counter electrode and a reference electrode.

[0025] A photoelectrochemical detection device based on radiation modulation that can be applied to the detection of alpha-fetoprotein.

[0026] Based on the AuNPs / TiO2 / Au (abbreviated as ATA) structure, the generated dipole mirror effect photoelectrochemical sensor can be used for the specific detection of alpha-fetoprotein. The principle utilized is the separation of the excitation source and detection signal of the PEC sensor, which improves the performance of the PEC sensor in the spatial domain. In the space, a sandwich structure is used to generate magnetic resonance and enhance light absorption; the dipole image effect generated by the ATA electrode significantly enhances light absorption and photoelectric conversion efficiency between AuNPs and the Au film.

[0027] The principle utilized is the separation of the excitation source and detection signal of the PEC sensor, which improves the performance of the PEC sensor in the time domain. In the time domain, the method of intensity modulation of modulation (IMM) is used to replace the photocurrent of the traditional manually switched light source. This modulated signal can be coherently amplified and extracted from the strong noise background, improving the signal-to-noise ratio of the photocurrent signal, thereby increasing the measurement range of the substance to be detected and reducing the detection limit.

[0028] When the fabricated photoelectrochemical sensor reaches the optimal index, the photocurrent signal is measured based on the IMM system. This method uses a chopper to modulate the light source. On the one hand, the on / off of the light source can generate a transistor-transistor logic (TLL) signal, which is then sent to a lock-in amplifier and converted into a sine signal as a reference. On the other hand, when the irradiated object flashes at the electrode, the generated photocurrent signal is collected by an electrochemical workstation. Then, phase-sensitive detection (PD) technology is used to detect and measure very small AC signals obscured by noise sources. The photocurrent signal after IMM processing is clear and spike-free, and the quasi-sine current response can be easily measured using the peak-to-peak detection method.

[0029] The beneficial effects of the present invention compared with the prior art are as follows:

[0030] According to the physical mechanism of the PEC sensor, its photocurrent response (determined by the interaction between chemical and biological analytes) can be enhanced in the spatial domain. To enhance the light-matter interaction in the spatial domain, the AuNPs / TiO2 / Au sandwich nanocomposite film is used as the electrode for AFP analysis. In this structure, the top metal layer includes a large number of noble metal nanostructures that can generate strong LSPRs, the middle dielectric layer can confine the electromagnetic field in a narrow gap, inducing electromagnetic coupling between the metal layers, and the bottom thicker metal layer serves as an opaque mirror.

[0031] The absorption efficiency of visible light has been greatly improved, mainly due to the plasmonic coupling between the metal layer and the nanoparticles in this structure, namely the dipole mirror interaction. Due to the LSPR effect of the noble metal nanoparticles in the ATA structure, the electrons in the AuNPs absorb the photon energy to excite hot electron-hole pairs, and the hot electrons cross the Schottky barrier between the AuNPs and TiO2 and enter TiO2. Then the electrons are transferred to the Au film. Since the conduction band of the FTO conductive layer is very low, the electrons will enter the FTO conductive layer along the way, thus effectively hindering the recombination of electron-hole pairs and greatly improving the absorption efficiency. Due to the presence of AuNPs, the ATA structure will generate plasmon resonance, and due to the presence of the Au film, it will produce a dipole mirror effect, thus significantly improving the absorption efficiency of the photoactive material for solar energy and the photoelectric conversion efficiency.

[0032] Therefore, the present invention adopts the AuNPs / TiO2 / Au sandwich-structured nanocomposite film. The plasmonic coupling and dipole mirror interaction between the Au thin film and the nanoparticles make up for the self-defects of TiO2 and greatly improve the optoelectronic performance of TiO2, making the prepared optoelectrochemical biosensor have good selectivity, stability, simple fabrication, low cost and high sensitivity. The sensitivity for alpha-fetoprotein detection has been increased by more than 20 times. The present invention uses the AuNPs / TiO2 / Au structure and the measurement strategy for generating magnetic resonance and enhancing light absorption to improve the sensor performance to achieve the specific detection of alpha-fetoprotein.

[0033] Based on the physical mechanism of the PEC sensor, its photocurrent response (determined by the interaction between chemical and biological analytes) can be enhanced in the time domain. In the case of the time domain, based on the AuNPs / TiO2 / Au (ATA) sandwich nanocomposite film electrode structure, an IMM-based method is proposed as a measurement strategy to further improve the sensor performance. The proposed IMM can easily remove the switching current spikes generated by charge recombination, and a chopper is used to modulate the light source, reducing the labor cost and making the detection more accurate. In the IMM, the illumination source of the light is modulated by a reference signal in the time domain. A lock-in amplifier is used to select and amplify the components with the same frequency and phase as the reference signal. Therefore, through differentiation and integration in lock-in detection, the signal-to-noise ratio (SNR) of the output signal can be significantly improved. In addition, through the detection by the system built in the electrochemical workstation, it can be seen that the shoulder profile of the quasi-square signal is blurred and almost unrecognizable, and is interfered by strong spikes. While the photocurrent signal processed by the IMM proposed in the present invention is smoother and spike-free, and the quasi-sinusoidal current response can be easily measured by the peak-to-peak detection method. It shows that the automatic operation of the proposed IMM provides a good foundation for achieving more stable and accurate results. The comparison of the photoelectrochemical sensor described in the present invention with other AFP sensors based on various sensing principles shows that the photocurrent response of the electrode can greatly improve the signal-to-noise ratio through IMM detection, and this sensing strategy is one of the best methods for early diagnosis and prognosis of liver cancer in clinical analysis. This amplification strategy has the advantages of simple preparation process, automatic operation and significantly improved performance, and can be applied to various PEC sensors for detecting other proteins.

[0034] For the IMM method used, it can replace the photocurrent of the traditional manually switched light source. This modulation signal can be coherently amplified and extracted from the strong noise background, improving the signal-to-noise ratio of the photocurrent signal and increasing the measurement range. Brief Description of the Drawings

[0035] Figure 1 It is a schematic diagram of the AuNPs / TiO2 / Au (ATA) sandwich structure.

[0036] Figure 2 It is a schematic diagram of the charge transfer mechanism of the AuNPs / TiO2 / Au (ATA) film.

[0037] Figure 3 It is the manufacturing process of the AFP-modified ATA film PEC sensor.

[0038] Figure 4 It is a schematic diagram of the IMM.

[0039] Figures 1-4Among them, 1 is AuNPs, 2 is the TiO2 thin film, 3 is the Au film, 4 is the FTO white glass, 6 is the lock-in amplifier, 7 is the xenon lamp, 8 is the optical chopper, 9 is the three-electrode, 10 is the oscilloscope, 11 is the electrochemical workstation, and 12 is the low-pass filter.

[0040] Figure 5 When it is the total output in the time domain of the lock-in amplifier, the signal is highly periodic.

[0041] Figure 6 It is the EDS data display. Ti, O, and Au signals are detected from the ATA film ( Figure 5 (inserted part in (a)).

[0042] Figure 7 The original signal in (a) comes from the analog port of the electrochemical workstation. (b) is the signal after being processed by the lock-in amplifier. (c) is the signal after harmonic suppression by a 3Hz low-pass digital filter. (d - f) are Figure 7 the corresponding power spectral density PSDs of (a - c) in sequence.

[0043] Figure 8 They are the sampled signals recorded by the typical method and their corresponding PSDs.

[0044] Figure 9 It is the comparison of the signal responses of the intensity modulation of radiation (IMM), the conventional method (CM), and the original number.

[0045] Figure 10 (a) is the photocurrent response after the designed ATA film electrode is incubated with target proteins of different concentrations. (b) is the measurement comparison diagram of the AuNPs / TiO2 electrode under the same experimental conditions. (c) is the ATA sandwich structure, and different concentrations of AFP are detected by the IMM method. (d) is the linear fitting of the logarithm of the AFP concentration c and the photocurrent.

[0046] Figure 11 (a) is the photocurrent response of the ATA structure sensor to alpha-fetoprotein and other proteins. (b) is the stability evaluation of the ATA structure sensor for detecting 0.01 ng / mL AFP, and the error bars show the standard deviation of 5 repeated tests.

[0047] Figure 12 It is the performance comparison between the sensor designed by the present invention and other sensors.

[0048] In the figure, 1 is AuNPs, 2 is the TiO2 thin film, 3 is the Au film, 4 is the FTO white glass, 6 is the lock-in amplifier, 7 is the xenon lamp, 8 is the optical chopper, 9 is the three-electrode, 10 is the oscilloscope, 11 is the electrochemical workstation, and 12 is the low-pass filter. Detailed implementation mode

[0049] In order to make the technical problems, technical solutions and beneficial effects to be solved by the present invention clearer and more understandable, the present invention will be further described in detail in combination with embodiments and drawings. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention. The technical solutions of the present invention will be described in detail below in combination with embodiments and drawings, but the protection scope is not limited by this.

[0050] Example 1

[0051] This example provides a photoelectrochemical biosensor based on an AuNPs-TiO2-Au (ATA) sandwich-structured nanocomposite film. As Figure 1 shown, the photoelectrochemical biosensor includes an FTO substrate at the bottom, and the FTO substrate is conductive glass; a Au film layer with a thickness of 100 nm is provided on the upper surface of the FTO substrate. A TiO2 film layer with a certain thickness is provided on the upper surface of the Au film layer; the thickness of the TiO2 film layer is set to 40, 60, 80, 110 nm (represented by (T40, T60, T80, T110)); a AuNPs layer is deposited on the TiO2 film layer; the AuNPs layer is a nano-Au particle layer uniformly provided on the TiO2 film layer, and the thickness of the AuNPs layer is 5 nm; the AuNPs layer is modified with alpha-fetoprotein AFP antibody (see Figure 3 ), wherein, fluorescein isothiocyanate (FITC) reacts with the amino group of the protein to label the fluorescein-labeled alpha-fetoprotein antibody.

[0052] With the increase of the TiO2 thickness, due to the magnetic resonance effect, strong magnetic fields gradually appear in the TiO2 layer and the gold film. The ATA thin film has the strongest magnetic resonance, which indicates that due to the image dipole mirror effect, the irradiation absorption is extremely strong. When the thickness of the TiO2 interlayer continues to increase to 100 nm, the magnetic field area and intensity decrease significantly, indicating a decrease in energy absorption. A locally enhanced electric field can be observed at the bottom of the AuNPs, and its intensity is consistent with the trend of the magnetic field intensity. The strong electric field and strong magnetic field of the ATA sandwich structure greatly enhance the light absorption and further contribute to the improvement of the photocurrent. The thickness of the TiO2 thin film in the ATA sandwich structure is selected to be 60 nm.

[0053] As Figure 2 shown, under visible light irradiation, the photoelectrochemical biosensor described in this example generates hot electrons by using local surface plasmon resonance (LSPR). The hot electrons skip the Schottky barrier formed at the metal-semiconductor interface, are injected into the TiO2 film layer, and finally transferred to the Au film layer and the FTO substrate. The dipole mirror effect generated by the ATA electrode enhances the light absorption and improves the photoelectric conversion efficiency in the case of the spatial domain.

[0054] Example 2

[0055] Preparation method of optoelectrochemical biosensor based on AuNPs-TiO2-Au, comprising the following steps:

[0056] S1: Growing an Au film layer on an FTO substrate by magnetron sputtering;

[0057] S2: Depositing a TiO2 sol-gel solution on the surface of the Au film layer by spin coating to prepare a TiO2 film layer; carefully adjusting the spin coating speed to control the thickness of the TiO2 film;

[0058] S3: In order to deposit AuNPs on the formed TiO2 thin film, another Au thin film with a thickness of 5 nm was prepared by magnetron sputtering to obtain an ATA sandwich structure;

[0059] S4: After annealing the ATA sandwich structure at 480 °C for 1 h, the Au thin film was prepared into a uniform nanoparticle layer, obtaining 32 nanoparticles with uniform diameters, and an optoelectrochemical biosensor was prepared.

[0060] S5: Incubating the AFP antibody on the surface of the optoelectrochemical biosensor electrode, specifically: as Figure 3 shown, after the sensor electrode was washed with deionized water, it was immersed in a 40 mM β-cysteamine solution, placed in a dark environment at room temperature for 12 h, rinsed 5 times with deionized water, and the unbound β-cysteamine was dried in nitrogen. The antibody solution was mixed with a PBS solution containing 4 mg / mL EDC and 4 mg / mL NHS in a shaker at 37 °C for 30 min. The carboxyl groups in the antibody were activated into NHS esters and effectively bound to the amino groups on the sensing electrode. Then the sensor was immersed in the activated antibody solution and placed in an incubator at 37 °C for 1 h, and the antibody was uniformly fixed on the surface of the sensor electrode. In order to avoid electrostatic adsorption and steric hindrance between antibodies, the non-binding sites and non-specific sites on the antibody surface were blocked with 0.1% BSA for 30 min.

[0061] The specificity study of the sensors for detecting AFP designed in Example 1 and 2 is as Figure 11 , the anti-AFP / ATA-film / FTO electrode of the sensor was applied to solutions containing different proteins, including the target protein AFP, carcinoembryonic antigen (CEA), human chorionic gonadotropin (HCG) and bovine serum albumin (BSA), all with the same concentration of 10 ng / mL. As Figure 11As shown in (a), no obvious change in photocurrent response was observed when the sensor was exposed to CEA, HCG, and BSA antigens. However, under the same experimental conditions, significant photocurrent was obtained after incubation of the antibody with AFP antigen. At the same time, there was a significant difference between the AFP antigen group and other antigen groups (P < 0.001). The experimental results indicate that the sensor has good selectivity for AFP antigen.

[0062] Based on the above-prepared photoelectrochemical sensor, photoelectrochemical detection of AFP antigen was carried out. Figure 10 (a) shows the photocurrent response after the designed ATA thin-film electrode was incubated with target proteins at different concentrations. As the concentration of AFP increased, the photocurrent gradually decreased. The AuNPs / TiO2 electrode was measured under the same experimental conditions for comparison ( Figure 10 (b)). The dipole mirror effect increased the photocurrent of the ATA sensor by 14 times compared with Au / TiO2 (22 μA and 1.5 μA, respectively). The enhancement of the photocurrent increased the sensitivity of the ATA-film sensor by more than 20 times. As Figure 10 (d) shows, there is an excellent linear relationship between the photocurrent and the logarithm of the AFP concentration. The specific parameters are as Figure 12 shown. Compared with other AFP detection methods, the sensor described in this embodiment has a wider detection range and a lower detection level. The above data fully illustrate the excellent performance of the ATA sensing structure.

[0063] Example 3

[0064] A device for specific detection of alpha-fetoprotein: It includes a photoelectrochemical biosensor and a photocurrent detection device. The photoelectrochemical biosensor is the photoelectrochemical biosensor based on AuNPs-TiO2-Au described in Examples 1 and 2, and the photoelectrochemical biosensor is modified with AFP antibody; the photocurrent detection device is based on the IMM system; the IMM system is as Figure 4 shown: It includes a lock-in amplifier 6, an optical chopper 8, and a low-pass filter 12; the optical chopper 8 is connected to the lock-in amplifier 6; the lock-in amplifier 6 is connected to the low-pass filter 12; the optical chopper 8 is arranged between the light source and the photoelectrochemical biosensor; in this embodiment, the light source is a xenon light source (300 nm < λ < 1100 nm, 300 mW / cm 2 , and a visible light filter with λ = 420 nm is installed to simulate visible light irradiation); the photoelectrochemical biosensor is connected to the low-pass filter 12 through an electrochemical workstation 11. Among them, the photoelectrochemical biosensor is used as the working electrode and is connected to the counter electrode (platinum wire) and the reference electrode (calomel electrode) to form a three-electrode system. The lock-in amplifier 6 is connected to an oscilloscope 10, and the electrochemical workstation 11 is connected to a computer.

[0065] The detection device described in this embodiment uses an optical chopper 8 to modulate the light source. On the one hand, the on / off of the light source can generate a transistor-transistor logic (TLL) signal, which is then sent to a lock-in amplifier and converted into a sine signal as a reference. On the other hand, when the irradiated object flashes at the electrode, a photocurrent signal is generated and collected by an electrochemical workstation.

[0066] The specific steps of the above-mentioned photocurrent detection technology based on the IMM system are as follows: The schematic diagram of the IMM system is as Figure 4 shown. On the one hand, the on / off of the light source can generate a TLL signal V 1s (t), which is then sent to a lock-in amplifier 6 and converted into a sine signal V ref (t) as a reference. On the other hand, when the irradiated object flashes at the electrode, a photocurrent signal I in (t) is collected by an electrochemical workstation. I in (t) is also converted into a proportional voltage signal V in (t), which can be obtained from the analog port of the electrochemical workstation.

[0067] Then, phase-sensitive detection (PD) technology is used to detect and measure very small AC signals obscured by noise sources. The PD technology realizes the amplification and acquisition of extremely small AC signals obscured by noise, that is, the voltage signal Vin(t) corresponding to the photocurrent signal measured by the sensor in the same proportion. The implementation of phase-sensitive detection (PD) technology can be simply described as the following three steps:

[0068] First, multiply V in (t) and V ref (t) to get

[0069] V psd (t) = V in (t)V ref (t) (1)

[0070] According to Fourier theory, any signal can be decomposed into several sine components, so V in (t) is represented by only one of these sine components, that is, V in (t) = A in sin(ω in t + θ sig ).

[0071] In addition, assume that the reference V ref (t) can be expressed as V ref (t) = A ref sin(w ref t + θ ref ), for V in (t) and V ref(t), A, w, and θ are the amplitude, frequency, and phase of the sine signal respectively. Then Equation (1) can be transformed into Equation (2).

[0072]

[0073] Obviously, V PD has two AC signal components, one is the difference frequency w in -w ref , and the other is the sum frequency w in +w ref . Since the on-off time of the chopper is on the order of seconds and the response time of the electrode is on the order of picoseconds, the difference between these two time scales is about 12 orders of magnitude. Therefore, the response speed of the electrode is fast enough, and w in is considered to be the same as w ref .

[0074] Secondly, the sum frequency component is removed by the low-pass filter 12. Also considering that w in = w ref , so V PD is converted to V′ PD . Therefore, V′ PD can be expressed as:

[0075]

[0076] Thirdly, in order to eliminate θ in -θ ref , according to the internal algorithm of the lock-in amplifier, following the steps similar to Equation (1), another PD signal V PD = 1 / 2A in A ref cos(θ in -θ ref ) can be obtained, but V ref (t) has an additional 90° phase shift. The total output R of the lock-in amplifier 6 is

[0077]

[0078] In the ideal case, R only contains DC components. However, limited by the cut-off frequency accuracy of the internal filter of the lock-in amplifier, the AC components cannot be completely removed. Therefore, R is still a time-varying signal and needs to be further processed by an external digital low-pass filter F(n). The output of the filter F(n) is represented by y(n). The modulation frequency (i.e., f ref ) is set to 1.8 Hz to avoid insufficient current response. Therefore, the cut-off frequency of F(n) is set to 3 Hz to effectively suppress the 2nd harmonic and higher harmonics of 1.8 Hz.

[0079] The advantages of the above-mentioned photocurrent detection using the IMM system are as follows Figure 5 , by repeatedly measuring the peak-to-peak value y(n) in several consecutive time spans, each time span containing at least two cycles (when f ref = 1.8 HZ, two cycles of y(n) are equal to 1.1 s). Although more repeated measurements and averaging operations will give more accurate results, in this embodiment, relatively stable results can be obtained after 2 - 3 cycles. In addition, the current spikes during the light source on / off switching due to charge recombination can be easily eliminated.

[0080] The second advantage of the above-mentioned photocurrent detection using the IMM system is as follows Figure 7 , Figure 7 The original signal V in (t) comes from the analog port of the electrochemical workstation and is processed by a lock-in amplifier ( Figure 7 (b)), and then filtered by a 3 Hz low-pass digital filter for harmonic suppression ( Figure 7 (c)). Intuitively, the detection device described in this embodiment can reduce noise and gradually smooth the signal.

[0081] Figure 7 The power spectral density (PSD) corresponding to the signals in (a)-(c) is as shown in Figure 7 (d)-(f). In Figure 7 (d), the power levels of the 1.8 Hz fundamental frequency and its second harmonic are basically the same, and the 3 - 5th harmonics and 50 Hz noise are also presented. Using the device of this embodiment, all orders of harmonics and 50 Hz noise are significantly suppressed ( Figure 7 (f)).

[0082] The comparison of the signal quality between the IMM detection device proposed in this embodiment and the typical method is as shown in Figure 9 . When the analyte concentration increases, due to the continuous increase of the electrode resistance, the original photocurrent signal collected from the analog port of the electrochemical workstation 11 is overwhelmed by strong noise. It is difficult to identify the required signal V in (t). By the signal amplification and processing method built in the lock-in amplifier 6, most of the noise is removed, but the shoulders of the quasi-square signal are severely blurred and almost unidentifiable. Worse still, the signal is interfered by strong spike signals. The photocurrent signal after IMM processing proposed by the device of this embodiment is clear and spike-free, and the quasi-sinusoidal current response can be easily measured using the peak-to-peak detection method (as shown in Figure 5 ).

[0083] Sequential tests were performed on the ATA sandwich sample using IMM. Here, the voltage signal output by the lock-in amplifier 6 was converted into a current value for comparison. As the AFP concentration increased, the peak-to-peak value of the sine signal gradually decreased. After the combination of ATA-film and IMM, the signal-to-noise ratio of the photocurrent signal was significantly improved, the sensitivity was increased by more than 2 times, the detection limit was 4.5 pg / mL, and the detection range was increased by one order of magnitude. As Figure 10 (d) shows, there is an excellent linear relationship between the photocurrent and the logarithm of the AFP concentration. The specific parameters are as Figure 12 shown.

[0084] Compared with other AFP detection methods, the PEC sensor described in this embodiment has a wider detection range and a lower detection level. The above data fully illustrate the excellent performance of the ATA sensing structure.

[0085] The above content is a further detailed description of the present invention in combination with specific preferred embodiments. It cannot be determined that the specific embodiments of the present invention are limited thereto. For those of ordinary skill in the technical field to which the present invention pertains, without departing from the premise of the present invention, several simple deductions or substitutions can still be made, which should all be regarded as belonging to the patent protection scope determined by the claims submitted by the present invention.

Claims

1. A photoelectrochemical detection device based on radiation modulation, characterized in that, It includes a photoelectrochemical biosensor and a photoelectrochemical testing device; the photoelectrochemical biosensor includes an ATA thin film electrode; the ATA thin film electrode includes an FTO substrate; a Au film layer is provided on the upper surface of the FTO substrate, and a TiO2 film layer is provided on the upper surface of the Au film layer; a AuNPs layer is deposited on the TiO2 film layer; the photoelectrochemical testing device includes a lock-in amplifier (6), an optical chopper (8) and a low-pass filter (12); the optical chopper (8) is connected to the lock-in amplifier (6); the lock-in amplifier (6) is connected to the low-pass filter (12); the optical chopper (8) is arranged between the light source and the photoelectrochemical biosensor; the photoelectrochemical biosensor is connected to the low-pass filter (12) through an electrochemical workstation (11).

2. The optoelectrochemical detection device based on radiation modulation according to claim 1, wherein The thickness of the TiO2 film layer is 60-80 nm.

3. The optoelectrochemical detection device based on radiation modulation according to claim 1, characterized in that, The thickness of the AuNPs layer is 5-10 nm.

4. The optoelectrochemical detection device based on radiation modulation according to claim 3, characterized in that, The AuNPs layer is composed of multiple AuNPs.

5. The optoelectrochemical detection device based on radiation modulation according to claim 1, wherein The thickness of the Au film layer is 100-120 nm.

6. The optoelectrochemical detection device based on radiation modulation according to claim 1, characterized in that, The FTO substrate is conductive glass.

7. A photoelectrochemical detection device based on radiation modulation according to any one of claims 1-6, characterized in that, The AuNPs layer is modified with alpha-fetoprotein antibody.

8. The optoelectrochemical detection device based on radiation modulation according to claim 1, wherein The light source described is a xenon lamp light source, where 300 nm < λ < 1100 nm and 300 mW / cm 2 .

9. The optoelectrochemical detection device based on radiation modulation according to claim 1, characterized in that, The photoelectrochemical biosensor is connected as a working electrode to a counter electrode and a reference electrode.

10. Use of a photoelectrochemical detection device based on radiation modulation according to any one of claims 1-6 in the detection of alpha-fetoprotein.

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