2.6. Selectivity, stability, and reproducibility
For practical biosensors, it is vital
to have outstanding selectivity, stability, and reproducibility. To
investigate the selectivity of PEC biosensors, a range of potential
interferents including Omicron BA.2, Wild-type, MERS,
H1N1,
H3N2, Influenza B, and HRSV were
utilized.
The selectivity of the PEC biosensor was determined by holding the cDNA
concentration constant at 1.0 nM and comparing the ∆I value produced by
the Omicron BA.5 cDNA template with the ∆I values produced by these
potential interferents. According to Figure 7A, the photocurrent change
generated by the Omicron BA.5 cDNA template was significantly greater
compared to other potential interferents, demonstrating the notable
selectivity of the PEC biosensor.
The investigation of the PEC biosensor’s stability was also conducted.
The photocurrent of the CdTe/ZnS
QDs/dsDNA/Au NPs/rGO biosensor was evaluated in the detection buffer
through multiple consecutive off–on–off irradiation cycles (10 cycles,
200 s; Figure 7B). The photocurrent response remained largely unchanged
throughout the cycle test, with a minimal relative standard deviation
(RSD) of only 0.58%, indicating the high stability and reliability of
the PEC signal. To further study the biosensor’s stability, a long-term
storage stability study was performed (Figure 7C). Upon storage in a
dark and humid environment at 4°C for 1, 2, 3, and 4 weeks, the PEC
biosensor exhibited photocurrent retention rates of 96.20%, 90.66%,
85.63%, and 73.99% of its initial response, highlighting its
exceptional stability. The reproducibility of the PEC biosensor was
evaluated by measuring the photocurrents of six biosensors assembled
from the same batch and comparing their results. The photocurrents
obtained from the six individual PEC biosensors exhibited negligible
variation (RSD = 1.96%), demonstrating the remarkable reproducibility
of the biosensor (Figure 7D).
2.7. Real sample
analysis
To assess the sensitivity and specificity of the photoelectrochemical
sensing platform for detecting the BA.5 variant in
sewage, practical sample analysis
was conducted. Initially, 24 sewage samples collected by automated
robots were identified using RT-PCR and Sanger sequencing. The
sequencing results revealed that among the collected
sewage samples, only the BA.2 and
BA.5 variants were detected. Specifically, there were 6 negative
samples, 6 samples positive for the BA.2 variant, and 12 samples
positive for the BA.5 variant. The RT-PCR analysis for the 18 positive
sewage samples (Figure 8A) all
showed positive results, with Ct values ranging from 15 to 31.
Conversely, the results for the 6 negative sewage samples were all
negative. Additionally, using the same categorized samples, the
photoelectrochemical sensing platform was employed to detect the L452R
mutation site (unique to the BA.5 variant and absent in the BA.2
variant). The positive threshold was defined as a ΔI value of 6.37 nA,
representing the average of the blank signals plus triple its standard
deviation. Positive results (ΔI > 6.37 nA) were observed
exclusively in the samples containing the BA.5 variant (Figure 8B).
These findings demonstrate that the photoelectrochemical sensing
platform can specifically monitor the crucial L452R variant of
SARS-CoV-2. In summary, the photoelectrochemical sensing platform
demonstrates remarkable sensitivity and specificity in detecting the
BA.5 variant in sewage.
2.8.
PEC reaction mechanism
In order to elucidate the PEC reaction mechanism of the constructed PEC
sensing platform, a thorough investigation into the photogenerated
carrier behavior of CdTe/ZnS QDs
was conducted using first-principle calculations based on density
functional theory. The crystal and band structures of CdTe/ZnS QDs were
depicted in the Figure 9A and 9B, respectively. The conduction band (CB)
and valence band (VB) energy levels of CdTe were found to be
approximately 0.80 and -0.34 eV,
respectively. Besides, for ZnS, the CB and VB energy levels were
approximately 2.53 and -0.19 eV. The energy gaps (Eg) of CdTe and ZnS
were estimated to be 1.14 and 2.72 eV, respectively. Consequently,
CdTe/ZnS QDs exhibit a type-II heterostructure, resulting in effective
spatial separation of photoexcited electrons and holes. This spatial
separation leads to prolonged carrier lifetimes due to reduced
recombination rates. Based on these research findings, CdTe/ZnS QDs
exhibit exceptional photoelectric response characteristics, which
significantly enhance the sensitivity of the constructed biosensor.
The accompanying Figure 9C illustrates the specific mechanism of charge
carrier transfer. Upon illumination, electrons from the VB of the
semiconductors transition to the CB,
resulting in an accumulation of holes in the VB. Subsequently, there is
an electron transfer from ZnS to CdTe, while concurrently, holes are
transferred from CdTe to ZnS due to the existing potential difference,
thus realizing efficient transfer of photo generated carriers.
Meanwhile, photoexcited electrons undergo a transfer process from the
conduction bands of CdTe to Au NPs. Ultimately, the electrons are
conveyed to vertically aligned graphene, resulting in a remarkable
photocurrent response.