Ultrathin Covalent Organic Framework Nanosheets/Ti3C2Tx-Based Photoelectrochemical Biosensor for Efficient Detection of Prostate-Specific Antigen

Designable and ultrathin covalent organic framework nanosheets (CONs) with good photoelectric activity are promising candidates for the construction of photoelectrochemical (PEC) biosensors for the detection of low-abundance biological substrates. However, achieving highly sensitive PEC properties by using emerging covalent organic framework nanosheets (CONs) remains a great challenge due to the polymeric nature and poor photoelectric activity of CONs. Herein, we report for the first time the preparation of novel composites and their PEC sensing properties by electrostatic self-assembly of ultrathin CONs (called TTPA-CONs) with Ti3C2Tx. The prepared TTPA-CONs/Ti3C2Tx composites can be used as photocathodes for PEC detection of prostate-specific antigen (PSA) with high sensitivity, low detection limit, and good stability. This work not only expands the application of CONs but also opens new avenues for the development of efficient PEC sensing platforms.


Introduction
The detection of tumor markers is important for early clinical diagnosis of cancer [1][2][3]. Prostate cancer is one of the most common and fatal diseases in humans. Prostate-specific antigen (PSA) is one of the most reliable cancer markers for early diagnosis of prostate cancer. PSA levels in normal human body serum are below 4 ng mL −1 . The presence of prostate cancer leads to elevated PSA levels [4]. Therefore, there is an urgent need to develop effective methods to achieve sensitive detection of PSA [5][6][7][8]. To date, many techniques have been used for the quantitative detection of PSA, including enzyme-linked immunosorbent assay (ELISA), electrochemical technique, colorimetric technique, and fluorescence technique [9][10][11][12], but the complicated operation and unsatisfactory sensitivity of these methods have greatly hindered their wide application. Therefore, it is essential to explore a reliable, simple, rapid, and sensitive method for prostate-specific antigen detection.
Photoelectrochemical (PEC) sensing is a new sensing technology that uses a singlewavelength light source assembled with an electrochemical detection device [13,14]. Due to its ease of miniaturization and integration, PEC sensing offers higher sensitivity than conventional electrochemical sensing [15][16][17][18]. When a beam of light is irradiated on a semiconductor material and its energy is equal to or greater than the semiconductor band gap, electrons can jump from the valence band to the conduction band under the excitation of light and generate holes in the valence band, achieving the effective separation of electrons and holes [19,20]. After the separation of electrons and holes, a photovoltage is generated and a photocurrent is formed in the external circuit [21][22][23]. Using the interaction between the photoelectrode material and the substances, the quantitative analysis of PEC

Morphology and Structure Characterization
To realize the above design principle, it is necessary to obtain PEC sensors with hierarchical structures by electrostatic self-assembly. In our work, in order to obtain COF nanosheets directly by a bottom-up approach, we chose N,N,N ,N -tetrakis(4-aminopheneyl)-1,4-phenylenediamine (TAPPDA) and N,N,N ,N -tetrakis(4-formylpheneyl)-1,4-phenylenediamine (TFPPDA) with a distorted diarylamino structure to reduce the interlayer stacking of COF. After sonication, the TTPA-CONs were obtained. The as-synthesized TTPA-CONs were then electrostatically self-assembled with the newly synthesized Ti 3 C 2 T x nanosheets (Scheme 1a). Typically, high yields of TTPA-COF were achieved by [4+4] imine condensation of TAPPDA and TFPPDA at a molar ratio of 1:1 in a mixture of o-dichlorobenzene (o-DCB) and N, N-dimethylacetamide (DMA), using 6 M acetic acid as the catalyst. To improve the yield of CONs, the obtained TTPA-COF was subjected to ultrasonication. Subsequently, the obtained TTPA-CONs were mixed with the freshly obtained Ti 3 C 2 T x nanosheets. After sonication, TTPA-CONs/Ti 3 C 2 T x composites were obtained (see the Materials and Methods section below for details). A typical C=N stretching vibration peak at~1617 cm −1 can be observed in Fourier transform infrared spectroscopy (FT-IR). In addition, the disappearance of N-H stretching bonds in TAPA (3337 cm −1 and 3432 cm −1 ) and the reduction in C=O stretching bonds in TFPA (~1693 cm −1 ) indicate the formation of a Schiff base. Furthermore, TTPA-CONs exhibit FTIR spectra similar to those of TTPA-COF, indicating the retention of chemical structure after exfoliation ( Figure S1).
obtained (see the Materials and Methods section below for details). A typical C=N stretch ing vibration peak at ~1617 cm −1 can be observed in Fourier transform infrared spectros copy (FT-IR). In addition, the disappearance of N-H stretching bonds in TAPA (3337 cm and 3432 cm −1 ) and the reduction in C=O stretching bonds in TFPA (~1693 cm −1 ) indicat the formation of a Schiff base. Furthermore, TTPA-CONs exhibit FTIR spectra similar t those of TTPA-COF, indicating the retention of chemical structure after exfoliation (Figur S1). The crystal structures of the TTPA-CONs/Ti3C2Tx composites were further deter mined by power X-ray diffraction (PXRD) measurements and structural simulations ( Fig  ures 1a and S2a,b). The experimental PXRD patterns of the obtained TTPA-COF matc well with the simulated patterns of the crystal model. As shown in Figure S2a, a promi nent diffraction peak arising from the (110) plane of TTPA-COF at 5.7° can be observed as well as some weak peaks at 8.0° (200), 11.7° (220), and 13.0° (310), which is the same a our previous work [31]. After exfoliation, the main peak was extremely weakened an shifted to 4.0°. In addition, a weak and broad peak around 20.0° appears, indicating πstacking interaction between the vertically stacked 2D layers and the broadening of th layer distance of TTPA-COF, confirming the successful stripping of TTPA-COF (Figur 1a). Moreover, the disappearance of the typical diffraction peak of Al at 38.9° proves th successful etching of multilayer Ti3C2Tx. In addition, the shift of the diffraction peak as signed to the (002) crystal plane from 9.5° to 6.2° originates from the removal of the alu minum atomic layer and the intercalation of lithium ions and H2O molecules after etchin and exfoliation, which also represents the successful acquisition of Ti3C2Tx (Figure S2b The crystal structures of the TTPA-CONs/Ti 3 C 2 T x composites were further determined by power X-ray diffraction (PXRD) measurements and structural simulations ( Figure 1a and Figure S2a,b). The experimental PXRD patterns of the obtained TTPA-COF match well with the simulated patterns of the crystal model. As shown in Figure S2a, a prominent diffraction peak arising from the (110) plane of TTPA-COF at 5.7 • can be observed, as well as some weak peaks at 8.0 • (200), 11.7 • (220), and 13.0 • (310), which is the same as our previous work [31]. After exfoliation, the main peak was extremely weakened and shifted to 4.0 • . In addition, a weak and broad peak around 20.0 • appears, indicating π-π stacking interaction between the vertically stacked 2D layers and the broadening of the layer distance of TTPA-COF, confirming the successful stripping of TTPA-COF ( Figure 1a). Moreover, the disappearance of the typical diffraction peak of Al at 38.9 • proves the successful etching of multilayer Ti 3 C 2 T x . In addition, the shift of the diffraction peak assigned to the (002) crystal plane from 9.5 • to 6.2 • originates from the removal of the aluminum atomic layer and the intercalation of lithium ions and H 2 O molecules after etching and exfoliation, which also represents the successful acquisition of Ti 3 C 2 T x ( Figure S2b). After electrostatic self-assembly, the composites show similar PXRD patterns to Ti 3 C 2 T x without the characteristic peaks of TTPA-CONs, which could be attributed to the weaker peaks of TTPA-CONs compared with Ti 3 C 2 T x .
After electrostatic self-assembly, the composites show similar PXRD patterns to Ti3C without the characteristic peaks of TTPA-CONs, which could be attributed to the weak peaks of TTPA-CONs compared with Ti3C2Tx.  (Figures 1c and S4 a,b). As shown in Figu 1d-g, O, Ti, C, and N are uniformly distributed throughout the nanostructure, showi the homogeneity of TTPA-CONs and Ti3C2Tx. To investigate the composite mode betwe TTPA-CONs and Ti3C2Tx, we measured the zeta potential of TTPA-COF, TTPA-CON and Ti3C2Tx. As shown in Figure S5, Ti3C2Tx exhibits a potential of −29.33 mV, which c be attributed to the presence of a large number of surface end groups (e.g., -O). Althou TTPA-COF also exhibits a negative potential of -18.52 mV, the TTPA-CONs exhibit a p itive potential of 65.13 mV after the functionalization of their exfoliated nanosheets w PEI. This suggests that TTPA-CONs and Ti3C2Tx can be bound by electrostatic adsorpti X-ray photoelectron spectroscopy (XPS) measurements were used to further inve gate the surface characteristics of the composites. The XPS survey spectra of TTP CONs/Ti3C2Tx in Figure S6 show the presence of N, O, Ti, and C elements. As shown Figure 2a, the high-resolution C1s spectra of TTPA-CONs/Ti3C2Tx exhibit a series of ty cal peaks at 281.7 eV, 284.5 eV, 285.8 eV, 286.9 eV, and 288.14 eV, which can be assign to C-Ti bonds, amorphous carbon, C-O bonds, C=N bonds, and C-N bonds, respective indicating the successful formation of TTPA-CONs. Figure 2b shows the high-resoluti Ti 2p spectrum, providing five characteristic peaks located at 455.08 eV, 456.5 eV, 45 eV, 461.2 eV, and 462.5 eV, which can be ascribed to the corresponding bonds of Ti-C,  SEM images of the TTPA-CONs/Ti 3 C 2 T x composites reveal the sheet-like morphology ( Figure 1a and Figure S3a,b). The low contrast of TEM images corresponds to the ultrathin nature of the TTPA-CONs/Ti 3 C 2 T x composites ( Figure 1c and Figure S4a,b). As shown in Figure 1d-g, O, Ti, C, and N are uniformly distributed throughout the nanostructure, showing the homogeneity of TTPA-CONs and Ti 3 C 2 T x . To investigate the composite mode between TTPA-CONs and Ti 3 C 2 T x , we measured the zeta potential of TTPA-COF, TTPA-CONs, and Ti 3 C 2 T x . As shown in Figure S5, Ti 3 C 2 T x exhibits a potential of −29.33 mV, which can be attributed to the presence of a large number of surface end groups (e.g., -O). Although TTPA-COF also exhibits a negative potential of -18.52 mV, the TTPA-CONs exhibit a positive potential of 65.13 mV after the functionalization of their exfoliated nanosheets with PEI. This suggests that TTPA-CONs and Ti 3 C 2 T x can be bound by electrostatic adsorption.
X-ray photoelectron spectroscopy (XPS) measurements were used to further investigate the surface characteristics of the composites. The XPS survey spectra of TTPA-CONs/Ti 3 C 2 T x in Figure S6 show the presence of N, O, Ti, and C elements. As shown in Figure 2a, the high-resolution C1s spectra of TTPA-CONs/Ti 3 C 2 T x exhibit a series of typical peaks at 281.7 eV, 284.5 eV, 285.8 eV, 286.9 eV, and 288.14 eV, which can be assigned to C-Ti bonds, amorphous carbon, C-O bonds, C=N bonds, and C-N bonds, respectively, indicating the successful formation of TTPA-CONs. Figure 2b shows the high-resolution Ti 2p spectrum, providing five characteristic peaks located at 455.08 eV, 456.5 eV, 459.7 eV, 461.2 eV, and 462.5 eV, which can be ascribed to the corresponding bonds of Ti-C, Ti (iii), Ti-O, and Ti (ii) and Ti (iii), respectively. Among them, the presence of Ti-O indicates the implantation of functional groups (-O) on the surface of Ti 3 C 2 T x . In addition to peaks at 399.7 eV (C-N bond) and 398.1 eV (C=N bond), the N 1s spectrum obtained from CONs shows a peak at 403.2 eV, which can be attributed to the N-O bond (Figure 2c). The presence of this bond suggests that TTPA-CONs and Ti 3 C 2 T x are bound by implanting functional groups O implanted on the surface of Ti 3 C 2 T x . The typical peak at 531.6 eV on the O 1s spectrum (N-O bond) also indicates a strong interfacial interaction between TTPA-CONs and Ti 3 C 2 T x (Figure 2d).    Figure 3a shows the UV-vis diffuse reflectance spectra of TTPA-COF and TTPA-CONs/Ti 3 C 2 Tx composites. The TTPA-COF sample reveals a well-established UV absorption spectrum with a shoulder absorption peak at 475 nm, while the TTPA-CONs/Ti 3 C 2 T x composite material shows a distinct absorption at 490 nm. The broad absorption bands of TTPA-CONs/Ti 3 C 2 T x and TTPA-COF in the visible region can be attributed to the distorted diarylamino units of TAPPDA and TFPPDA. Compared with pure TTPA-COF, the absorption capacity of TTPA-CONs/Ti 3 C 2 T x composites (red line) is significantly enhanced. As depicted in Figure 3b, the band gap energies (E g ) of TTPA-COF and TTPA-CONs/Ti 3 C 2 T x composites are 2.24 eV and 2.02 eV, respectively. In addition, the fabrication steps of the various photoelectrodes were characterized. Electrochemical impedance spectrum (EIS) shows the interfacial behavior of the prepared electrodes in 5 mM K 3 [Fe(CN) 6 ] − solution containing 0.1 M KCl (Figure 3c and Figure S7).

Design of PEC Sensing Platform
The diameter of the semicircle in the high-frequency region represents the charge transfer resistance (R ct ). Accordingly, the semicircular diameter of TTPA-CONs/Ti 3 C 2 T x composites is significantly smaller than that of the pristine TTPA-CONs, suggesting that the charge carriers in the TTPA-CONs/Ti 3 C 2 T x composites have a smaller charge transfer resistance and faster transfer rate. In addition, Figure 3d depicts the transient cathodic PEC response of different photoelectrodes under visible light irradiation. The photocurrent intensities of TTPA-CONs and Ti 3 C 2 T x show a small value (curve I, II), while the photocurrent intensity of TTPA-CONs/Ti 3 C 2 T x is significantly higher, recorded as 6.9 µA (curve III). As shown in Scheme 1b, light excites photogenerated electrons to the conduction band (CB) of TTPA-CONs, and these electrons can be transferred from the conduction band (CB) of TTPA-CONs to Ti 3 C 2 T x , which can effectively improve the efficiency of photoelectron-hole pair separation due to its excellent electron transport ability at the TTPA-CONs/Ti 3 C 2 T x composites interface, and further suppress the photoelectron-hole pair complexation. The The diameter of the semicircle in the high-frequency region represents the cha transfer resistance (Rct). Accordingly, the semicircular diameter of TTPA-CONs/Ti3C composites is significantly smaller than that of the pristine TTPA-CONs, suggesting t the charge carriers in the TTPA-CONs/Ti3C2Tx composites have a smaller charge tran resistance and faster transfer rate. In addition, Figure 3d depicts the transient catho PEC response of different photoelectrodes under visible light irradiation. The photoc rent intensities of TTPA-CONs and Ti3C2Tx show a small value (curve I, II), while the p tocurrent intensity of TTPA-CONs/Ti3C2Tx is significantly higher, recorded as 6.9 (curve Ⅲ). As shown in Scheme 1b, light excites photogenerated electrons to the cond tion band (CB) of TTPA-CONs, and these electrons can be transferred from the conduct band (CB) of TTPA-CONs to Ti3C2Tx, which can effectively improve the efficiency of p toelectron-hole pair separation due to its excellent electron transport ability at the TTP CONs/Ti3C2Tx composites interface, and further suppress the photoelectron-hole p Afterward, the NH 2 -terminal aptamer was immobilized on the acetic acid-catalyzed TTPA-CONs/Ti 3 C 2 T x surface due to the abundance of aldehyde groups on the surface of TTPA-CONs/Ti 3 C 2 T x , and the photocurrent intensity was measured to be 5.4 µA (curve IV). The decrease in the photocurrent density was due to the restart of electron transfer at the PEC sensing interface by the loading of the poorly conducting aptamer. After treatment with BSA, the photocurrent density was further reduced to 3.6 µA (curve V). Finally, when the target PSA was trapped on the photoelectrode, the photocurrent density further decreased to 1.8 µA (curve VI). The reason for this decrease in photocurrent density may be related to the following reasons. PSA can be trapped at the sensing interface due to the biorecognition reaction between the aptamer and its target, while the formation of PSAaptamer complexes with large steric hindrance increases the electron transfer resistance and adversely affects the photocurrent response. Therefore, significant changes in photocurrent density can be used for the quantitative detection of PSA.

Optimization of Experimental Conditions
In order to obtain better PEC performance, we investigated the effect of each experimental parameter on the photocurrent intensity. As shown in Figure S8, the photocurrent intensity increased with increasing concentration of TTPA-CONs/Ti 3 C 2 T x in the range of 1 to 3.0 mg L −1 . When the concentration of TTPA-CONs/Ti 3 C 2 T x was further increased, the photocurrent intensity decreased rapidly, which may be due to the thicker nanocoating hindering electron transfer. Therefore, a TTPA-CONs/Ti 3 C 2 T x concentration of 3.0 mg mL −1 was chosen for subsequent experiments. Next, we optimized the pH of the Tris-HCl solution. As shown in Figure S9, the photocurrent intensity gradually increased and then gradually decreased as the pH increased from 6.0 to 7.4, which indicated that the Tris-HCl solution with pH 7.4 was the most favorable for electron transfer, so we chose the solution with pH 7.4 to detect PSA. In addition, the incubation time of the PSA aptamer, BSA, and PSA had important effects on the performance of the PEC sensor. The photocurrent intensity decreased rapidly with increasing incubation time and reached the lowest values at 2.5 h, 50 min, and 120 min, respectively (Figures S10-S12). Subsequently, the photocurrent did not change greatly with the extension of time, so the incubation times for the PSA aptamer, BSA, and PSA were finally determined to be 2.5 h, 50 min, and 120 min, respectively. Since the PSA aptamer can bind to P4, which can directly affect the intensity of the photocurrent, the concentration of the P4 aptamer was optimized in Figure S13. The photocurrent intensity reached the highest value when the concentration of PSA aptamer was 5 µM. Finally, we chose the concentration of P4 aptamer to be 5 µM for subsequent experiments.

Analytical Behavior of the Proposed PEC Sensing Platform
The photocurrent intensities of the fabricated PEC sensors with the addition of different concentrations of PSA were recorded under optimal conditions. As shown in Figure 4a, the photocurrent intensities gradually decreased with increasing concentrations of PSA in the range of 0.001-10,000 ng/mL. This can be explained by the fact that a large amount of poorly conducting PSA-aptamer complexes is generated by being trapped on the sensing surface, hindering electron transfer. As shown in Figure 4b, the logarithm of photocurrent intensity and PSA concentration showed a good linear relationship in the range of 0.001 to 10,000 ng/mL. The corresponding linear regression equation was I (µA) = 0.298 log C PSA (ng/mL)-2.119 (correlation coefficient R 2 = 0.995). In addition, the limit of detection (LOD) was estimated to be 0.0003 ng/mL based on the analytical function LOD = Kσ/S, where K is 3, σ is the standard deviation of the blank solution (n = 10), and S is the slope of the regression line. Compared with many previously reported sensors for the determination of PSA (Table S1), the PEC biosensor exhibited an acceptable linear range and low detection limit. The formation of the composite excited excellent photoelectrochemical activity, including intense visible light collection, effective suppression of the recombination rate of light-generated electron-hole pairs, and accelerated charge separation/transfer, leading to the high-performance of the proposed PEC sensor. On the other hand, the coupling of TTPA-CONs and Ti 3 C 2 T x generates composites that minimize the mismatch between the interface and the lattice, contributing to the easy separation and transfer of photogenerated carriers. This feature is also an advantage for establishing an efficient PEC bioassay platform.
transfer of photogenerated carriers. This feature is also an advantage for establishing an efficient PEC bioassay platform.

Selectivity, Stability, and Reproducibility of the PEC Sensor
The stability of the cathodic PEC conformal sensor was evaluated. Figure 4c shows that the photocurrent response of the PEC sensor remains stable under continuous illumination and 1000 s of repeated on/off illumination, showing significant stability. To investigate the interference of other hormones and metal ions on the assay, we performed interference measurements including Progesterone (P4), Kanamycin (KANA), glucose oxidase (GXO), Bovine Serum Albumin (BSA), Vitamin B6 (B6), and Alkaline phosphatase (ALP). As shown in Figure 4d, there is little change in the PEC response signal before and after the presence of the above-mentioned interferents. The photocurrent intensity was significantly reduced only in the presence of the target or in the coexistence of the target and the potential interferer. These results indicate that our constructed PEC platform has good selectivity for PSA detection.
To further evaluate the stability of this PEC sensor, we checked the reproducibility of the sensor by comparing five individual modified electrodes. As shown in Figure S14, the photocurrent response of the PEC sensor was tested in a PSA containing 10 ng/mL. All electrodes exhibited similar photocurrent intensities and the calculated relative

Selectivity, Stability, and Reproducibility of the PEC Sensor
The stability of the cathodic PEC conformal sensor was evaluated. Figure 4c shows that the photocurrent response of the PEC sensor remains stable under continuous illumination and 1000 s of repeated on/off illumination, showing significant stability. To investigate the interference of other hormones and metal ions on the assay, we performed interference measurements including Progesterone (P4), Kanamycin (KANA), glucose oxidase (GXO), Bovine Serum Albumin (BSA), Vitamin B6 (B6), and Alkaline phosphatase (ALP). As shown in Figure 4d, there is little change in the PEC response signal before and after the presence of the above-mentioned interferents. The photocurrent intensity was significantly reduced only in the presence of the target or in the coexistence of the target and the potential interferer. These results indicate that our constructed PEC platform has good selectivity for PSA detection.
To further evaluate the stability of this PEC sensor, we checked the reproducibility of the sensor by comparing five individual modified electrodes. As shown in Figure S14, the photocurrent response of the PEC sensor was tested in a PSA containing 10 ng/mL. All electrodes exhibited similar photocurrent intensities and the calculated relative standard deviation (RSD) was 2.79%. This indicates that the sensor has good repeatability and stability. In addition, the modified electrode was stored in a refrigerator at 4 • C and its photocurrent response was observed and measured every three days ( Figure S15). The results showed that after 15 days of storage, the PEC sensor still maintained 95% of its initial response. This indicates that the PEC sensor has satisfactory storage stability.
To further evaluate the applicability of the prepared PEC sensor in real complex samples, we performed spiked recovery experiments on this PEC sensor. A cathodic PEC biosensor was used to determine the amount of PSA in bovine serum to simulate the amount of PSA in the prepared human serum. A certain concentration of standard PSA solution was added to the diluted serum sample, and the photocurrent response of the prepared electrode was recorded. As shown in Table S2, the recoveries ranged from 94.3% to 103.2% with RSD values less than 1.76%, demonstrating the feasibility and acceptable accuracy of the designed method for the determination of PSA in real samples. Fourier transform infrared spectroscopy (FTIR) spectra were obtained with a Nicolet 6700 Thermo FT-IR spectrometer. Powder X-ray diffraction (PXRD) patterns were obtained on an X-ray powder diffractometer (D/max-Ultima IV) equipped with a Cu sealed tube (λ = 1.54178 Å) at a scan rate of 0.02 deg s −1 . Scanning electron microscopy (SEM) was conducted on a FE-SEM (Nova NanoSEM 450) equipped with an energy dispersive spectrometer. Samples were treated via Pt sputtering before observation. Transmission electron microscopy (TEM) images were obtained using a transmission electron microscope (Talos S-FEG (FEI, Hillsboro, OR, USA)). X-ray photoelectron spectroscopy (XPS) analysis was measured on Thermo ESCALAB 250Xi spectrometer equipped with a light source of Al Ka X-ray (1486.6 eV) (Thermoelectricity Instruments, USA). UV-Vis diffuse reflectance spectroscopy (UV-Vis DRS) spectra were obtained by a UV-Vis spectrophotometer (UV-Vis-NIR Cary 5000) and the data were converted to Kubelka-Munk functions for the band gap extraction. PEC measurements were carried out on a CHI760 E electrochemical workstation (Shanghai Chenhua Instrument Co., Shanghai, China) equipped with PLS-FX300HU xenon lamp parallel light source system (Beijing Perfect light Technology Co., Ltd., Beijing, China). The mixture was sonicated for 10 min to get a homogenous dispersed russet solution. Subsequently, acetic acid (6 M, 0.12 mL) was added and the vial was then flash frozen at 77 K using a liquid N 2 bath and degassed by three freeze-pump-thaw cycles. Subsequently, the tube was sealed under vacuum, and then heated at 120 • C for 3 days. The yielded orange precipitate was collected by centrifugation and immersed with N, N-Dimethylformamide (DMF) for 6 h under 80 • C by 2 cycles, separately. The collected powder was then activated by solvent exchange with anhydrous tetrahydrofuran (THF) and anhydrous acetone in Soxhlet extractor for 2 days, and dried at 80 • C under vacuum for 12 h to give an orange powder with 85 % isolated yield.

Preparation of TTPA-CONs
A 200 mL glass vial was charged with TTPA-COF (10 mg) and 100 mL DI water. Then 2 mL polyethyleneimine (PEI) solution (30 wt %, M.W. 70,000) was added and stirred at room temperature for 0.5 h. Subsequently, the mixture was sonicated for 8 h. After sonication, the dispersion was centrifuged at 3500 rpm for 10 min to obtain the supernatant. Then the obtained supernatant was centrifuged at 10,000 rpm for 10min to obtain TTPA-CONs, which was subsequently re-dispersed in DI water with a concentration of 500 µg /L.

Preparation of Ti 3 C 2 T x
Ti 3 C 2 T x was prepared according to reported methods [40,41]. Specifically, LiF (0.5 g) was dissolved in 10 mL HCl (9 M) under room temperature. Subsequently, 0.5g Ti 3 AlC 2 powder was slowly added within 5min and stirred at 35 • C for 24 h. After reaction, the suspension was washed with DI water by centrifugation repeatedly until the pH of the supernatant was greater than 6.0 was ultrasonic dispersed in DI water for 20 min under N 2 atmosphere for 20 min. Finally, the supernatant containing Ti 3 C 2 T x nanosheets (10 mg/mL) was collected by centrifugation at 7500 rpm for 20 min. The powder with ultra-thin and low-layer Ti 3 C 2 T x nanosheets was obtained by freeze-dried for yield measurement.

Preparation of TTPA-CONs/Ti 3 C 2 T x
The TTPA-CONs/Ti 3 C 2 T x composite material was synthesized by electrostatic selfassembly. Specifically, the as-obtained Ti 3 C 2 T x suspension (50 µL) was diluted with DI water to 1 mL. Then, the Ti 3 C 2 T x suspension was mixed with 1 mL TTPA-CONs suspension. The TTPA-CONs/Ti 3 C 2 T x composite materials was obtained after sonicated for 10 min.

Fabrication of the PEC sensor
Scheme 1 illustrates the fabrication of the PEC biosensor based on the TTPA-CONs/Ti 3 C 2 T x composites for PSA detection. Prior to modification, the glassy carbon electrodes (GCE) were polished with 0.3 µm and 0.05 µm alumina slurry and sonicated in ethanol and ultrapure water. Then, the TTPA-CONs/Ti 3 C 2 T x composites were sonicated in ultrapure water for 1 min to obtain a uniformly distributed suspension with a concentration of

Conclusions
In summary, we herein demonstrated a novel PEC sensor based on composites constructed by electrostatic self-assembly of TTPA-CONs and Ti 3 C 2 T x . The prepared TTPA-CONs/Ti 3 C 2 T x composites show enhanced photoelectric properties compared with TTPA-CONs, making them effective candidates for fabricating cathodic PEC sensors. Due to the effective separation of photogenerated electrons and holes, the well-designed PEC sensor exhibits a linear response to PSA in the range of 0.001 to 10,000 ng/mL, with a low detection limit of 0.0003 ng/mL under optimal conditions. It is believed that this work may pave the way toward the design of high-performance PEC sensing platforms and broaden the application of CONs in biomimetic sensing and analysis.