All chemical reagents in this experiment were of analytical grade. Na₂S·9H₂O, CdCl₂·2.5H₂O, I₂, KI, and HAuCl₄·4H₂O were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China). NaOH and trisodium citrate dihydrate (Na₃C₆H₅O₇·2H₂O) were obtained from Shanghai Aladdin biochemical technology Co., Ltd. (Shanghai, China). H₄[Si(W₃O₁₀)₄]·xH₂O, CS was obtained from Shanghai Maclin Biochemical Technology Co., Ltd. Synthetic oligodeoxy-nucleotides corresponding to partial sequences of the gene of HPV type 16 and TE buffer were received from Sangon Biotech (Shanghai) Co., Ltd. T7 Exo and NE buffer were received from New England Biotechnology (Beijing) Co., Ltd. HPV samples of vaginal swab scraping with different infection subtypes (HPV 16, 18, 33 DNA) were supplied by Suzhou KunTao Intelligent Manufacturing Technology Co., Ltd. All reagent solutions were prepared using ultrapure water (resistivity as 18 MΩ⋅cm at 25°C). The nucleotide sequences of the oligonucleotides are listed in Table 1.

First, CdS QDs were synthesized according to our previously published literature (Wang F. et al., 2022). Afterward, a post-modification procedure of CdS QDs with SiW₁₂ proceeded. H₄[Si(W₃O₁₀)₄]·xH₂O (0.1435 g) were dissolved in a CdS QD solution (15 mL), and the obtained mixed solution was vigorously stirred at 40°C for 12 h to obtain the final products, a homogeneous yellow solution. The product solution was stored at 4°C for the next step.

Au NPs were synthesized according to the previously published literature (Wang F. et al., 2022). First, 1-mg CS and a 20-mL gold solution were added to 30-mL ultrapure water. After that, the obtained solution was stirred at room temperature (25°C) for 24 h to achieve a CS hydrogel. The obtained Au NP-based CS hydrogel was stored at 4°C (Suginta et al., 2013; Liu et al., 2018; Feyziazar et al., 2020; Vesel, 2023).

5 mL of a 9-μM armDNA solution and 5 mL of a 9-μM cDNA solution were mixed and heated at 95°C for 5 min. After cooling down to room temperature, arm-cDNA was received. For the fabrication of the PEC biosensor, 40 μL of the Au NP-based CS hydrogel (Au NP/CS gel) was embellished on the ITO slide surface, and then, 2 μL of the arm-cDNA solution and 3 μL of 3-μM pDNA were sequentially modified on the electrode. When they were successfully connected to the electrode, 3 μL of the SiW₁₂@CdS QD/NP DNA solution was successively modified and incubated for 2 h, and unstable residues on the electrode surface were washed with buffer. Finally, the prepared electrode was incubated with a series of concentrations of HPV 16 DNA for 2 h. Then, the electrode was dropped in 50-U mL⁻¹ T7 exonuclease and incubated for 2 h. The electrode was rinsed with buffer and dried under nitrogen atmosphere for PEC measurements in a 5-mM I 3 − / I − working solution. Here, an HPV 16 DNA-detecting biosensor was successfully fabricated.

The detection performance and reliability of the as-prepared PEC biosensor were investigated with the samples from vaginal swab scraping. Different specimens of the as-prepared biosensor were incubated with different patients’ samples of HPV 16 DNA for 2 h. Then, the electrode was washed and dropped in 50-U mL⁻¹ T7 exonuclease. Finally, the electrode was rinsed with buffer and dried under nitrogen atmosphere for PEC measurements in 5-mmol L⁻¹ of I 3 − and 0.5-mol L⁻¹ of an I⁻ working solution, and each sample was detected three times. Similarly, to detect the samples of HPV 18 DNA and HPV 33 DNA to evaluate the selectivity and stability of the as-prepared biosensor, the same PEC measurement procedure was performed with different HPV DNA subtypes.

Transmission electron microscopy (TEM) images of the NPs were obtained using an HT7800 transmission electron microscope at 200-kV acceleration voltages. The Zeta potentials and particle size distribution of the SiW₁₂@CdS QDs were obtained by Malvern nano-ZS NP size and Zeta potential analyses, and UV–vis absorption spectra were recorded with a UV-26001 UV–vis spectrophotometer. Electrochemical data were obtained with a three-electrode-system CHI 660E electrochemical workstation. PEC signals were obtained from a PL-X500D Simulated solar xenon lamp source and electrochemical word station (CHI 660e). A single-sided glass electrode (1 cm × 3 cm) was coated with ITO as the working electrode. A platinum wire and a saturated calomel electrode were used as the counter and reference electrodes, respectively; PEC measurements were performed in a 5-mM I 3 − / I − working solution.

The morphology and size distribution of the as-prepared CdS QDs and SiW₁₂@CdS NPs were characterized via TEM and Malvern nano-ZS NP size analysis. As shown in Figures 1A, C black curve, uniform CdS QDs were obtained using the solvothermal method and observed as a yellow solution (insert photograph on Figure 1A), and the average particle size of the obtained CdS QDs was ∼10 nm; note that the size of the as-prepared SiW₁₂@CdS NPs has increased to ∼100 nm, making the color brighter yellow, as shown in Figures 1B, C red curve. The larger size resulted from the aggregation of the SiW₁₂-modified CdS QDs, as shown in Figure 1B (Liu et al., 2018; Feyziazar et al., 2020; Zhang J. et al., 2022; Vesel, 2023). Here, the SiW₁₂@CdS NPs were successfully synthesized. The photographs of the as-prepared CdS QDs and SiW₁₂@CdS NPs under UV light irradiation are shown in Supplementary Figure S1.

After the modification of SiW₁₂ to CdS QDs, a new absorption wave appeared at ∼250 nm on the UV–vis absorption spectrum of the SiW₁₂@CdS NPs (Figure 1D, red curve) compared with that of the CdS QDs (Figure 1D, black curve), and both underwent absorption at ∼400 nm. Thus, improved light absorption was achieved according to this phenomenon. Otherwise, from Figure 1E of the fluorescence emission spectrograms of the CdS QDs and SiW₁₂@CdS QDs, an enhanced emission spectrum and a 90-nm blue shift were observed from ∼590 nm of the CdS QDs to ∼500 nm of SiW₁₂@CdS QDs under the same excitation light at 340 nm, indicating a strong interaction between SiW₁₂ and CdS QDs with a broader band gap (Guo et al., 2016; Ji et al., 2017; Zang et al., 2022; Nie et al., 2020). To summarize the UV–vis absorption and fluorescence emission spectroscopic studies, the improved light response ability was successfully achieved by the strong association between SiW₁₂ and CdS QDs, indicating remarkable photoelectric properties.

As an important role of the first layer in the construction of the as-prepared PEC biosensor, Au NPs were synthesized using the solvothermal method, as shown in Figure 2A, with a size of ∼24 nm in Figure 2C. Notably, during the preparation of the Au NP/CS gel (Au NP/CS gel), the Zeta potential of the Au NPs decreased by 20 mV from −45 to −25 mV (Figure 2B), indicating a strong accumulating capacity, which would endow it with good adhesive ability as the first layer on the ITO slide to fabricate the PEC biosensor, while the visible light absorption ability of the Au NPs was well maintained without any changes in the UV–vis absorption spectrum at 524 nm (as shown in Figure 2D), high conductivity and visible light absorption for the PEC biosensor.

Electrochemical impedance spectroscopy (EIS) changes associated with the modification of the ITO slide and the PEC response of the as-prepared PEC biosensor were measured for each modification layer. As shown in Figure 3A, the first layer of the Au NP/CS gel on the ITO slide showed the largest R_et value (Figure 3A; a: red curve) because the poor conductivity of the CS gel obstructed electron transfer to the ITO electrode. Au NP addition not only increases conductivity but also PEC response via the LSPR effect, emphasizing its importance (Aiken and Finke, 1999; Lee et al., 2013; Chou et al., 2017; Wen et al., 2017; Domingues et al., 2018; Shi et al., 2019; Figueiredo et al., 2021). After the successive modification with arm-c DNA and pDNA (Figure 3A, b: green curve), the obtained R_et value decreased because of the association between Au NPs and the primers, offering an electron transfer pathway with reasonable steric hindrance to the ITO electrode. The smallest R_et value appeared after the NP DNA–SiW₁₂@CdS NP modification (Figure 3A, c: dark blue curve), for the reason that the remarkable conductivity of the SiW₁₂@CdS NPs significantly improved electron transfer. The R_et value was increased when the HPV 16 DNA was added, as shown in Figure 3A, d: light blue curve, owing to the high steric hindrance of HPV DNA. The photocurrent response (PEC) was consistent with the EIS investigation, as shown in Figure 3. The ITO/Au NP/CS sample exhibited the smallest photocurrent (Figure 3B, a: red curve), and the photocurrent increased when the primers were continuously anchored onto the modified ITO slide (Figure 3B, b: green curve). The largest photocurrent was achieved by the NP DNA–SiW₁₂@CdS NP modification (Figure 3B, c: dark blue curve) because of the strong synergistic effect on the photoelectronic phenomenon that occurred within the association of SiW₁₂@CdS NPs. Here, the photocurrent response of the as-prepared PEC biosensor with the outmost layer of NP DNA–SiW₁₂@CdS NPs established the maximum photocurrent monitoring range.

From the spectrum analysis in Figures 1D, E and photocurrent response analysis in Figure 3B, the intense PEC response occurrence mechanism for the largest photocurrent with SiW₁₂@CdS QDs can be illustrated, as shown in Figure 3C. The SiW₁₂@CdS QDs were stimulated under Xe light irradiation, in detail. The highest occupied molecular orbital (HOMO) of both the SiW₁₂ and CdS QDs were stimulated to generate photo-electrons (e⁻) and photo-holes (h⁺) simultaneously; I 3 − / I − electrolytes donating electrons via I⁻ to I⁰ occurred on the excited HOMO of SiW₁₂ through the generated photo-holes (h⁺). The photo-electrons naturally transfer to the lowest unoccupied molecular orbital (LUMO) of SiW₁₂ and then to the LUMO of CdS QDs due to the close contact between them, with a matched energy level (0.4 eV difference between 5.87 and 5.47 eV). Successfully, the photo-electrons finally moved to the ITO external circuit to generate photocurrent, and the generated photo-holes (h⁺) from the HOMO of CdS QDs are transferred to the HOMO of SiW₁₂ to complete the PEC procedure (Liang et al., 2015; Kokal et al., 2016; Li et al., 2016; Shi et al., 2018; Su S. et al., 2022; Kar et al., 2023). The enhanced PEC response of SiW₁₂@CdS QDs to bare CdS QDs is attributed to the increased photogenerated electron energy of 0.4 eV from the strong synergistic effect resulting from the association of SiW₁₂@CdS NPs realizing the outstretched band gap from a HOMO of 3.26 eV to LUMO of 5.87 eV.

For the detection of HPV 16 DNA, a series of target DNA samples of different concentrations were incubated in the PEC biosensor, and PEC detection was performed accordingly. EIS spectra after this target DNA incubation were also investigated, as depicted in Figure 4 A, and the observed R_et value increased gradually with increasing HPV 16 DNA concentrations (15–130 nm), which was predictable because of the increasing steric hindrance. Conversely, the measured PEC response of the biosensor gradually decreased (Figure 4B), exhibiting a PEC quenching phenomenon caused by the addition of T7 exonuclease in the DNA walker cycle process (Scheme 1B) to release the corresponding associated SiW₁₂@CdS NPs from the PEC biosensor. Finally, the detection of HPV 16 DNA achieved the expected performance of linear quantitative determination, as shown in Figure 4C. The PEC response had a linear relationship with the concentration of the target DNA, ranging from 15 to 130 nM, with an LOD of 0.8 nM, according to Eq. 1: C L = K ∙ S b m ; (1) C L : LOD; S b : Blank standard deviation; m : The slope of the calibration curve in the low concentration range that was analyzed; K : Confidence coefficient; the value is 3.

All required photocurrent values are listed in Supplementary Table S1.

Specificity is critical to verify the accuracy and sensitivity of PEC biosensors, indicating their anti-jamming capability. Different HPV subtypes of HPV 18 DNA and HPV 33 DNA were selected as potential disruptors for specific studies of the as-prepared PEC biosensor. As shown in Figure 5 A, the presence of the target HPV 16 DNA (Figure 5A, green column) or target DNA-containing mixture (Figure 5A, yellow-green column) showed an obvious decrease in the PEC response, whereas the highly homologous interference of HPV 18 DNA and HPV 33 DNA would not promote the biological DNA walker cycle process to decrease the photocurrent, indicating the remarkable specificity of the proposed PEC biosensor. For the stability test of the as-prepared PEC biosensor, the fabricated ITO electrodes were preserved at 4°C for 1–4 weeks, and three parallel experiments were conducted every week. As shown in Figure 5B, the PEC response was 90.77% of the initial value after 4 weeks of storage, indicating the significant stability of the as-prepared PEC biosensor. Moreover, within one PEC measurement process, 7 times consecutive light “on/off” cycles were performed to evaluate its repeatability. As shown in Supplementary Figure S2, the photocurrent response exhibited a steady signal with an incredibly small variation, and the relative standard deviation was 6.37%, indicating the distinguished reproducibility of the as-prepared PEC biosensor.