Introduction

Photoelectrochemical (PEC) biosensor has attracted considerable interest among other analytical methods due to their promising and burgeoning working principles [1,2,3,4]. Its benefit stems from the combination of optical and electrochemical properties. PEC was based on the light-harvesting electrical response, making it a crucial component to amplify the corresponding signals in biosensing design [5]. This light-harvesting property is closely associated with photoreceptors, charge separation, and the interfacial redox reaction. Recently, most techniques rely significantly on the light-harvesting capability and charge-transfer kinetics of the PEC system. They overlooked the redox reaction at the surface of the photoelectrode/electrolyte interface.

Building a PEC biosensor requires a decent photoactive material with several ideal properties, such as fast exciton generation, migration, increased signal intensity, and stability. Consequently, as the photogenerated carriers accumulate on the interfacial surface, the semiconductor undergoes metal ion leaching due to photocorrosion [6, 7]. This leads to a significant photocurrent decay and hampers the quantitative analysis. Boosting the catalytic kinetics of the reaction at the interface will increase the charge separation of photogenerated electron–hole pairs [8].

Quantum dot (QD)–metal oxide junctions have been explored as a key component in developing next-generation PEC devices, light-emitting diodes, and nanostructured electronic arrays [9]. Semiconducting QDs have received much attention for their size-dependent electronic structure [10]. It offers a broad-spectrum design on various devices and systems with tailored electrical characteristics by altering the size and complexity of component elements [10, 11]. QDs were integrated and functionalized into biomolecules for bioimaging and monitoring devices [12]. Although the electronic interactions between QDs and organic molecules have been well established, a recent trend showcased an interesting annotation on the coupling QDs to other inorganic species. Because inorganic materials have a continuum of electronic states rather than the discrete conditions seen in organic molecule receptors, the pairing is fundamentally different from QD-organic coupling. When QDs are coupled to an inorganic species, it acts either as an electron donor or both electron receptors and donor [13]. In the case of practical application, electron transfer processes are closely engaged in the QDs’ adaptive function.

Titanium dioxide (TiO2) is widely used in PEC biosensors due to its chemical stability, photocatalytic activity, and environmental friendliness [14, 15]. However, it suffers from the rapid interacting photogenerated charges, impeding the progress of TiO2-based PEC devices. Various strategies have been reported to suppress photogenerated electron–hole recombination: nanostructure engineering, doping enrichment, surface treatment, and heterostructure modification [16, 17]. Among the various strategies reported to suppress photogenerated electron–hole recombination, combining other semiconductors has been attempted to improve the overall photocatalytic performance. Designing a TiO2-based catalyst with a heterojunction structure on the surface would enable natural self-sustaining charge management, encouraging the separation of photogenerated carriers, and supplying electrons continuously for the catalysis reaction [18].

We have now elucidated interactions between unique nanostructured semiconducting metal oxides, TiO2 and Co3O4, focusing on a QD-implemented PEC device (Scheme 1). CdSe QD semiconductors produce photointroduced electron–hole pairs (e − /h +) under light irradiation [19, 20]. When combined with metal oxides like TiO2, the high efficiency of this heterostructure is compromised by its limited stability because of the trapping of oxidizing holes in chalcogenide nanocrystal subject to rapid photocorrosion of the material. More importantly, these catalysts cannot produce oxygen at their anodic site. Thus, coupling with a robust oxygen-evolving catalyst (Co3O4) at the hole-trapping site (CdSe) will enhance the overall system’s stability and benefit oxygen evolution [21, 22]. The excellent chemical stability and sensitive response of light irradiation make Co3O4 much attractive to researchers. The combination of CdSe, Co3O4, and TiO2 prevents electron–hole recombination and boosts photoinduced carriers’ transfer. In addition, Co3O4 allows the loading of CdSe QDs and antibodies and acts as a hole collector in catalytic redox, preventing the rapid photocorrosion of the chalcogenide’s nanocrystal by the oxidizing holes. The steric impedance of Co3O4 blocks the transfer of charge. As a result, CdSe-Co3O4 impacts steric impedance and competition, improving signal reduction from the photoactive matrix and enhancing the PEC sensor’s sensitivity.

Scheme 1
scheme 1

Schematic representation illustrating the (a) fabrication process of CdSe-Co3O4@TiO2 NF heterostructure, (b) application of virus detection, and (c) change in photocurrent response based on virus conjugation

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Further, observing the desired need for virus sensing in recent times, a viable embodiment of sensitive and accurate strategies, especially for detection in the early stage of infection rapidly and giving a reliable signal, is in demand. In addition, based on our previous experience with virus-like particles (VLP) and viruses [23,24,25], we have developed a PEC device based on CdSe-Co3O4@TiO2 heterojunction structure and applied it for HEV-LP detection. As shown in Scheme 1b, the target virus could be captured by antibody-functionalized Ab/CdSe-Co3O4@TiO2||FTO electrode. After washing with PBS, virus-conjugated virus/Ab/CdSe-Co3O4@TiO2||FTO electrode is introduced into the PEC cell and utilized for measuring photocurrent response (Scheme 1c) based on a PEC principle. Optimizing conditions for sensor formulation and its sensing parameters are investigated thoroughly. The designed PEC device based on CdSe-Co3O4@TiO2 was used to quantitatively detect HEV-LPs, HEV, and spike (S) protein of SARS-CoV-2. This PEC device enabled the development of a realistic embodiment of highly sensitive analytical equipment and reliable detection for the suppression of emerging infectious viral diseases.

Materials and methods

Synthesis of cobalt oxide

To synthesize spherical cobalt oxide nanoparticles (Co3O4 NPs), 80 mg cobalt acetate and 3.5 mL benzylamine were mixed under constant stirring for 2 h at room temperature. Further, 3.5 mL ammonium hydroxide solution was added to the above solution under vigorous stirring, and the temperature was increased to 165 °C and maintained for 2 h under continuous stirring. Diethyl ether was added to the above solution, and thus, formed black precipitate was collected by centrifugation and washed several times with ethanol.

Synthesis of cobalt oxide-CdSe complex

To synthesize cobalt oxide–CdSe complex, initially in a separate flask, 85 mg of Co3O4 NPs, 5 mg Se, and 5 mL of dichlorobenzene were mixed in a flask and heated to 180 °C in an Ar atmosphere. Further, cadmium precursor solution, prepared by dissolving 25.6 mg CdO in 0.9 mL oleic acid and 4.5 mL dichlorobenzene at 160 °C under an Ar atmosphere, was injected into the above solution and continued to heat for 5 min. This changed the color of the solution from black to reddish-black. The temperature was reduced to 150 °C, and a solution of 12.5 mg propylphosphonic acid in 0.5 mL dichlorobenzene was added to the above solution. The resulting nanoparticles were allowed to grow for 5 min, and the solution was allowed to cool down to room temperature. After reaching room temperature, the final product as precipitate was isolated and washed with ethanol.

Surface modification of CdSe-Co3O4 NPs by ligand exchange

The obtained nanoparticles were dissolved in a mixture of methanol, mercaptoundecanoic acid, and tetramethylammonium hydroxide. Each concentration was adjusted to ensure sufficient ligand exchange. The nanoparticles were precipitated using toluene as a non-solvent, followed by centrifugation, then dissolved in water.

Synthesis of TiO2 NFs

TiO2 NFs were prepared using the previously reported method [26]. A detailed description of the synthesis method is described in detail in the Electronic Supplementary Material (ESM).

Preparation of CdSe-Co3O4@TiO2 nanocomposite

For CdSe-Co3O4@TiO2 nanocomposite preparation, 10 mg of TiO2 powder and 30 mg of CdSe- Co3O4 powder were dispersed in DI water and mixed overnight, followed by lyophilization for 24 h to obtain CdSe-Co3O4@TiO2 nanohybridization.

Biological sample: These materials are described in detail in the ESM and also in our earlier research works [27, 28].

Fabrication and application of the PEC-based device for virus detection

The anode substrate was fluorine-doped tin oxide (FTO) conductive glass, treated with acetone and 95% ethanol, followed by sonication for 30 min, and dried with pure N2 gas before use. The processed FTO was spin-coated with 20 μL of CdSe-Co3O4@TiO2 (20 mg mL–1) and dried. Further, the CdSe-Co3O4@TiO2-coated electrode was incubated with 10 μL of antibody (16 μg μL–1) and activated by N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDC)/1-hydroxy-2,5-pyrrolidinedione (NHS) for 6 h at 4 °C [29]. The electrode was further cleaned with DI water and used for detection. 0.2 mol L–1 of AA in free-oxygen phosphate-buffered saline (PBS, pH 7.0) was used as the electrolyte solution for detection.

A series of diluted virus concentrations were incubated for 20 min at 4 °C with the functionalized electrode for virus detection. The electrode was washed three times using PBS solution to eliminate the uncaptured targets. Finally, the electrode was replaced in the chamber, and the signal was checked based on its photocurrent density from the PEC. In running the PEC for the detection of viruses, the irradiation source wavelength for PEC experiments was 430 nm, with an exposure period of 20 s and a bias voltage of 0.1 V. The scan frequency range was 100 mHz–100 kHz for the EIS tests, and an AC sine wave with 5 mV amplitude was utilized. The anode’s functional surface area was 1 cm2.

For detection in the complex media, the sample should be 10X-diluted, and 0.5 mL of the diluted sample needs to be dropped onto the prepared Ab/CdSe-Co3O4@TiO2||FTO electrode and incubated for 20 min (in room temperature around 25 °C) for the conjugation of target virus to the antibodies on the modified electrode. Further, the electrode should be washed three times using PBS solution to remove the uncaptured targets. Finally, the electrode must be placed in the PEC cell, and the photocurrent should be measured based on the same PEC parameter mentioned above. After measuring the photocurrent density, the relationship between the PEC response and the amount of HEV in the sample is plotted and analyzed to obtain the calibration curve. The calculated linear equation would be used further as the standard calibration of the HEV concentration in the sample analysis.

Results and discussion

Characterization of photoelectrode materials

The sequential formation of nanoheterostructure with increasing complexity comprises three stages, as illustrated in Scheme 1a. Figure 1a–d shows characteristic TEM images of heteronanostructure corresponding to consecutive stages of the growth protocol. The synthesis steps begin with forming cobalt oxide nanoparticles (Co3O4 NPs) by decomposition of cobalt nitrate with benzylamine in the presence of ammonium hydroxide. Figure 1a shows the TEM image of Co3O4 NPs. The synthesized Co3O4 NPs are circular, with particle sizes ranging from 5 to 13 nm and an average particle diameter of 9 nm (Fig. 1e). Then, the CdSe QD was grown on the surface of Co3O4 NPs. The TEM image in Fig. 1b clearly demonstrates the CdSe-decorated Co3O4 NPs. The contrast in the TEM image confirmed the subsequent growth of CdSe QD onto Co3O4 NP. The growth of CdSe QD on Co3O4 NP was accompanied by the onset of absorption and emission peak at λ = 576 nm, which is shifted from the 2.4-nm-size bare CdSe peak as shown in the UV–Vis spectra for different stages of growth in Fig. 1f. Figure 1c clearly shows the SEM image of prepared TiO2 NF. The arrangement of cross-linked nanosheets in TiO2 NF increases the number of active sites for photocatalytic reactions [30]. The TiO2 NFs are used as a photoconductive nanomaterial to capture excitation energy and generate photoresponse [31]. The TEM image shown in Fig. S1 reveals that TiO2 inherits the “flower-like” architecture. In addition, the specific surface area of TiO2 NFs will significantly affect the contact area with the light source and the CdSe-Co3O4 NPs. CdSe@Co3O4 NPs are adequately spread over the surfaces of TiO2 NFs, as shown in Fig. 1d, and their flower-like apparent morphologies of TiO2 NFs remain unaltered. The surface of TiO2 NF is negatively charged, while the Co3O4 is positively charged; thus, the two were readily adsorbed together. As shown in Fig. S2a, the lattice fringes observed on TiO2 are 0.352 nm, which agrees with the in-plane lattice spacing pattern of TiO2 (101 facet). In addition, two types of the fringes for TiO2 and Co3O4 are also deciphered, and the distances of 0.352 nm and 0.243 nm clearly match the characteristic lattice plane of TiO2 (101) and Co3O4 (311) (Fig. S2b) respectively, indicating the presence of CdSe-Co3O4 on the surface of TiO2 forming CdSe-Co3O4@TiO2. CdSe-Co3O4 nanoparticles are precipitated uniformly on the surface of TiO2 NF. This can be attributed to the positively charged Co3O4 being adsorbed on the negatively charged TiO2 NF by electrostatic interaction.

Fig. 1
figure 1

Synthesis and characterizations of CdSe-Co3O4@TiO2 NFs. The TEM images of Co3O4 NPs (a), CdSe-Co3O4 (b), and CdSe-Co3O4@TiO2 NF (d). SEM image of TiO2 NFs (c); corresponding particle size distributions of Co3O4 NPs (e); absorption spectra of Co3O4, CdSe-Co3O4, and CdSe-Co3O4@TiO2 (f), XRD pattern (g); deconvoluted XPS spectra of Co 2p of Co3O4 (h); O 1 s of TiO2 (i); and CdSe-Co3O4@TiO2 NFs (j)

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The formation of CdSe-Co3O4@TiO2 NF was evidenced by sequential X-ray diffraction (XRD) pattern. The diffraction pattern of the TiO2 NFs (black in Fig. 1g) shows multiple prominent diffraction peaks at 2θ = 25.3°, 37.7°, 38.5°, 48.0°, 53.8°, 55.0°, 62.7°, 68.7°, 70.2°, and 75.0° attributing to (101), (004), (112), (200), (105), (211), (204), (116), (220), and (215) planes, respectively. The diffraction peaks are correlated with the JCPDS card no. 84–1285, indicating the successful formation of the anatase phase of TiO2 NF [32]. Figure 1g (green) shows the XRD patterns of Co3O4 NPs with diffraction peaks at 31.27°, 36.85°, 44.81°, 59.35°, and 64.61° attributing to the (220), (311), (400), (511), and (440) crystal planes of cubic spinel Co3O4 NPs respectively which are correlated to the JCPDS no. 43–1003 [33]. In addition, peaks appearing at (2 theta) of 25.36°, 42.02°, and 49.72° in the XRD patterns of CdSe-Co3O4 (red in Fig. 1g) can be attributed to (111), (220), and (311) crystal planes of CdSe (JCPDS no. 88–2346) [34]. In the XRD pattern of CdSe-Co3O4@TiO2, all peaks of TiO2 and CdSe@Co3O4 can be noticed. The peak positions remain unaltered after the formation of the CdSe-Co3O4@TiO2 NF heterostructure. This indicates the preservation of the nature of the CdSe-Co3O4 and the successful construction of the heterostructure.

The detailed bonding state of the material was further characterized by XPS measurement. Figure 1h shows the Co 2p spectra of Co3O4. The Co 2p spectra exhibit a doublet with a low energy band (Co 2p3/2) at 780.1 and a high energy band (Co 2p1/2) at 795.5 eV. The energy difference between the Co 2p3/2 − 2p1/2 peaks is approximately 15.4 eV which is in corroboration with the reported values of Co (II, III) [35]. The absence of the prominent shake-up satellite peak in the Co 2p spectra suggests a dominant Co3O4 phase. The peak deconvolution of O 1 s spectra of TiO2 and CdSe-Co3O4@TiO2 is further performed (Fig. 1i and j). The deconvolution spectra of O 1 s of synthesized TiO2 NF (Fig. 1i) show three peaks at 531.9, 533.1, and 529.3 eV, which belong to H–O–H, Ti–OH, and Ti–O–Ti, respectively. Furthermore, the weak shoulder peak at 531.2 eV of O 1 s in the CdSe-Co3O4@TiO2 is attributed to the Co–OH bonds.

Performance of PEC-based biosensing device

Furthermore, the binding ability and the detection performance of the anti-HEV antibodies conjugated to CdSe-Co3O4@TiO2 toward HEV-LPs are investigated (Fig. 2a and b). A time-based i − t approach was used to understand the performance of the PEC device following the assembly of Co3O4, CdSe-Co3O4, and CdSe-Co3O4@TiO2 NFs, respectively. When CdSe-Co3O4 or TiO2 NFs were used alone, the photocurrent density response remained at 2.21 or 4.9 μA cm−2, respectively. However, after the CdSe-Co3O4@TiO2 NF nanocomposite formation, the current density improved drastically to 15.5 μA cm−2, owing to the synergistic effect among CdSe-Co3O4 and TiO2 NFs. The heterostructure formation boosted the interfacial electron-transfer efficiency while suppressing e−/h+ recombination in converting light to electrical energy. The photocurrent was reduced to 14.1 μA cm−2 after the conjugating of the anti-HEV antibody, probably due to the antibody acting as an insulator. After incubation with HEV-LPs (100 pg mL–1), the current density further decreased to 7.1 μA cm−2, indicating that the antibody successfully captured HEV-LPs on the CdSe-Co3O4@TiO2 NFs electrode. Modifying the nanocomposite with an anti-HEV antibody helped in the specific binding of the HEV-LPs when incubated with samples containing HEV-LPs. These HEV-LPs bound on the surface of the nanocomposite acted as insulating agents and hindered the light source, preventing the excitation of the CdSe-Co3O4@TiO2. This resulted in a reduced photocurrent response of the CdSe-Co3O4@TiO2 electrode hybridized with HEV-LPs compared with the CdSe-Co3O4@TiO2 electrode without HEV-LPs.

Fig. 2
figure 2

Time-based photocurrent (a), histogram (b), of (1) CdSe-Co3O4||FTO, (2) TiO2/FTO, (3) CdSe-Co3O4@TiO2||FTO, (4) Anti-HEV-Ab/CdSe-Co3O4@TiO2||FTO, and (5) HEV-LPs/Anti-HEV-Ab/CdSe-Co3O4@TiO2||FTO (irradiation source wavelength of 430 nm and a bias voltage of 0.1 V). Nyquist plots (c) of CdSe-Co3O4@TiO2||FTO (1), anti-HEV-Ab/CdSe-Co3O4@TiO2||FTO (2), and HEV-LPs/Anti-HEV-Ab/CdSe-Co3O4@TiO2||FTO (3). Inset in (c) is Randles equivalent circuit. Schematic illustration of the energy band structure and the proposed charge-transfer mechanism (d). Error bars in (b) denote data as average ± SD (n = 3)

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As shown in Fig. 2c, electrochemical impedance spectroscopy (EIS) analyzes the electrochemical activity, the antibody-modified electrode’s capturing property, and the antibody’s detection ability toward HEV-LPs. Randle’s equivalent circuit model is used for fitting the EIS curves as shown in the inset in Fig. 2c. It consists of the circuit components like charge-transfer resistance (Rct), solution resistance (Rs), double-layer capacitance (CPE), and the Warburg impedance (W). Rct was majorly affected during the construction of the biosensor and HEV-LP detection. Therefore, changes in Rct can be monitored to analyze the interfacial electron transfer during the fabrication of biosensing devices with individual components and the detection process. The Rct of the electrode upon deposition of CdSe-Co3O4@TiO2 NFs is 447 Ω, and further conjugation of the anti-HEV antibody increased the Rct to 772 Ω. The Rct value further increased to 1125 Ω upon incubation of anti-HEV antibody–conjugated electrode which was HEV-LPs (10 pg mL–1). The changes in Rct values evidenced the successful formation of the biosensing device with the apparent detection ability of HEV-LPs. Further, modified ELISA was performed to confirm the conjugation of anti-HEV Abs onto anti-HEV-Ab/CdSe-Co3O4@TiO2||FTO electrode. As shown in Fig. S4, only the anti-HEV-Ab-conjugated anti-HEV-Ab/CdSe-Co3O4@TiO2||FTO electrode generated high absorbance compared with bare FTO and CdSe-Co3O4@TiO2||FTO electrodes. Because anti-rabbit IgG-conjugated HRP could only bind with the anti-HEV-Abs present on anti-HEV-Ab/CdSe-Co3O4@TiO2||FTO electrode, this indicated the conjugation of anti-HEV-Ab on the anti-HEV-Ab/CdSe-Co3O4@TiO2||FTO electrode.

Photovoltaic working mechanism of the device

The possible mechanism of photocurrent response for the developed PEC device is shown in Fig. 2d. The CdSe-Co3O4@TiO2-modified FTO showed good photocurrent intensity, attributed to the excellent matching cascade band-edge levels of CdSe-Co3O4@TiO2 and its increased photocurrent conversion efficiency by minimizing electron–hole recombination. TiO2 NFs have a conduction band (CB) edge potential of about − 0.44 V and a wide bandgap of 3.2 eV [36]. Using the equation Eg = VVB – VCB, the valance band (VB) edge potential of TiO2 NFs is calculated to be about 2.76 V. In addition, previous reports claim that CdSe QDs possess a bandgap of 1.8 − 1.9 eV [13, 37]. Figure 2d depicts the energy level diagram with well-matched energy levels of TiO2 NFs and CdSe QDs forming a heterojunction alignment structure.

The developed PEC device generated an electrical signal under visible light excitation in this report. Because of the external energy provided by visible light, the carriers in the VB of CdSe QDs were excited and injected into the CB of CdSe QDs. In most semiconductors, the occupied VB and the vacant CB existed simultaneously [38]. Electrons were injected from the VB to the CB upon excitation by the external energy. This result generates holes in the VB because of the bandgap. However, the excited electrons and the holes could quickly recombine because this course was very fast [39]. The recombination occurred quickly when TiO2 NFs were alone. However, by forming the CdSe-Co3O4@TiO2, the recombination of excited electrons and holes was inhibited.

Further, these excited carriers were injected instantly into the CB of TiO2 NFs. CdSe QDs served as electron donors. The absorbed light energy was transformed into an electric current and quickly delivered to the external circuit. The carriers in the VB of TiO2 NFs and CdSe QDs were reduced by Co3O4 and ascorbic acid (AA), which improved the energy conversion efficiency and reduced carrier residence time across the energy band. The UV–Vis spectra (Fig. 1f) indicated that the hybridization between CdSe-Co3O4 and TiO2 facilitated carrier transmission efficiency and energy conversion efficiency enhancement. In addition, photocurrent response was enhanced several folds in the CdSe-Co3O4@TiO2 nanocomposite, thereby providing evidence for the working mechanism mentioned above. Prolonged irradiation of the CdSe-based system caused photocorrosion in the presence of oxygen on the surface of the photocatalyst by the photogenerated hole [40]. In our system, the photogenerated hole transfers to the Co3O4; thus, an oxidation reaction occurs at the Co3O4 surface, increasing the system’s stability. Thus, the CdSe-Co3O4@TiO2 carrier-transfer nanohybridization boosts external energy and photocatalytic capability. As explained below, the photocurrent response from the CdSe-Co3O4@TiO2-based device was successfully exported for virus detection.

Detection of HEV-LPs

The application of the developed PEC device for HEV-LP detection is based on the specific interactions of anti-HEV antibodies and HEV-LPs. Different concentrations of HEV-LPs are initially added to the anti-HEV antibody–conjugated CdSe-Co3O4@TiO2 NF–modified FTO electrode. The photocurrent response at various concentration ranges of HEV-LP continuously decreased with the increase in HEV-LP concentration (Fig. 3a). The linear relationship between the photocurrent density and the HEV-LP concentration ranging from 10 fg mL–1 to 10 ng mL–1 shows a correlation coefficient of 0.998 (Fig. 3b). The limit of detection (LOD) was 3.5 fg mL−1, as calculated by the 3σ/S where S is the slope of the linear calibration plot and σ is the standard deviation of the lowest signal [41,42,43]. The sensitivity of the developed PEC-based sensor for HEV-LPs detection is competitive with recently developed analytical sensing methods comprising optical methods, as shown in Table S1 of the ESM. These results indicate that the developed PEC sensor for virus detection exhibits benefits, including ultra-sensitivity of the electrochemical method and wide linear range detection, which can offer accurate and reliable quantitative results.

Fig. 3
figure 3

The photocurrent response of the electrode after incubation with various concentrations of HEV-LPs ranging from 10 fg mL–1 to 10 ng mL–1 (irradiation source wavelength of 430 nm and a bias voltage of 0.1 V) (a), calibration curve obtained using change in photocurrent density vs. concentration of HEV-LPs (b), calibration curve for HEV-LPs ranging from 10 fg mL–1 to 10 ng mL–1 in 10% human serum as a sensing medium (c), and the change in photocurrent density of anti-HEV-Ab/CdSe-Co3O4@TiO2||FTO electrode when incubated with 1 pg mL–1 HEV-LPs and other common interferences at a concentration of 10 ng mL–1 using 10% human serum as a sensing medium (d). Error bars in (b), (c), and (d) denote data as average ± SD (n = 3)

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The applicability of the proposed sensor was evaluated in a complex biological matrix using 10% human serum as a sensing medium. The calibration curve obtained for HEV-LP detection in 10% human serum (Fig. 3c) shows a comparable detection pattern, indicating the suitability of the proposed sensor for the clinical sample analysis. Compared with the calibration curve obtained in DI water, the slope of the calibration curve in 10% human serum is slightly flattened. The complex human serum matrix caused a modest decrease in the slope of the calibration curve, lowering the LOD value to 8.5 fg mL–1. However, the sensitivity of the proposed sensor in the serum matrix is considerable compared to previous reports for its actual application.

Selectivity and stability of the PEC sensor

The selectivity of the biosensor is crucial for its practical application [44, 45]. To evaluate the selectivity of the developed sensor, several VLPs and viruses, including norovirus-like particles (NoV-LPs), NS1 protein lysate, white spot syndrome virus (WSSV), influenza virus, and Zika virus at 10 ng mL–1, were spiked in 10% human serum. The interfering effect was evaluated by comparing the response with 1 pg mL–1 HEV-LPs. The photocurrent intensity of anti-HEV antibody–conjugated CdSe-Co3O4@TiO2 electrode decreases only in the presence of target HEV-LP (Fig. 3d), demonstrating that interfering materials cannot bind to anti-HEV antibody–conjugated CdSe-Co3O4@TiO2 electrode.

To examine the stability of the developed PEC sensor, photocurrent response and long-time stability are evaluated. As shown in Fig. 4a, there is no discernible fluctuation in the photocurrent response in the recurring 15 times on/off irradiation cycles for 475 s with incubating 10 pg mL–1 of HEV-LPs. This indicates exceptional photocurrent stability for HEV-LP detection. In addition, the stability of the developed PEC sensor is also examined by storing the modified electrodes at 4 °C and measuring the photocurrent response of the modified electrode every week. After storage in a refrigerator at 4 °C for 2 weeks, the photocurrent response of the sensor electrode remained steady at 96.1% compared to its original photocurrent, at 91.3% and 89.5% for 3 and 4 week storage, respectively (Fig. 4b), suggesting good long-term stability.

Fig. 4
figure 4

Stability evaluation of the PEC sensor (a), long-time stability of PEC microfluidic sensor (irradiation source wavelength of 430 nm and a bias voltage of 0.1 V)) (b), the calibration curve for detection of G7 HEV ranging from 102 to 107 RNA copies mL–1 in cell culture supernatants using PEC sensor (c), and detection of Spike protein of SARS-CoV-2 ranging from 10 fg mL–1 to 10 ng mL.–1 in cell lysate (d). Error bars in (b), (c), and (d) denote data as average ± SD (n = 3)

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Furthermore, the repeatability of the constructed PEC-based sensor is investigated by fabricating five separate electrodes to detect HEV-LPs at 10 pg mL–1. The detected concentrations of HEV-LPs utilizing five different electrodes vary from 10.2 to 11.3 pg mL–1, with an average of 10.7 pg mL–1, demonstrating that the PEC sensor’s accuracy is acceptable. Furthermore, the relative standard deviation (RSD) of the measurements is 6.7%, indicating that the PEC sensor has excellent repeatability.

PEC-based detection of HEVs and SARS-CoV-2 Spike protein

The photocurrent responses are recorded to evaluate the PEC sensor’s performance for HEV detection from the cell culture supernatant. As shown in Fig. 4c, G7 HEV shows a correlation coefficient (R2) of 0.983 with LOD of 69 RNA copies mL–1, confirming the high accuracy and sensitivity of the developed PEC sensor. The photocurrent response continuously decreased with the increase in HEV concentration from 102–107 RNA copies mL–1.

In addition to the HEV detection, they substitute the corresponding antibodies from anti-HEV Ab into anti-SARS-CoV-2 Ab, targeting S protein of SARS-CoV-2 (Fig. 4d). The developed Ab-SARS-CoV-2-CdSe-Co3O4@TiO2-based PEC sensor demonstrated a good response of SARS-CoV-2 protein biomarker in cell lysate with a detection limit of 7.8 fg mL−1 with an R2 value of 0.991. These results strongly showcased how the developed PEC sensor is reliable for detecting the target viruses and their biomarker protein, exhibiting the potential application in detecting HEVs and other targets.

Conclusions

A photoelectrochemical (PEC) device based on CdSe-Co3O4@TiO2 is developed, and ultrasensitive virus detection is demonstrated. The fabricated device exhibited a stable and continuous photocurrent response up to 15.5 μA cm−2 with a longer exciton lifetime. Based on the specific strong recognition ability between the anti-HEV antibody and HEV-LPs, the device demonstrated a wide detection range, a low detection limit of 3.5 fg mL−1, and ultrahigh selectivity and sensitivity. In addition, it could detect HEVs in the complex cell culture supernatant and sub-femtogram level of SARS-CoV-2 S protein in cell lysate solution in high precision. Therefore, the proposed device should aid in developing quantitative detection strategies for biomolecule detection by replacing the corresponding antibodies and broadening the applications of PEC-based biosensing devices. It is worth mentioning that although this method has achieved good results, however, there are still distances to the clinical application, such as the detection limit being affected by the substrate type, exploitation of corresponding kits, and the development of portable equipment.