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Abnormal expression of hydrogen peroxide (H2O2) elucidates cell dysfunctions and might induce the occurrence and deterioration of various diseases. However, limited by its ultralow level under pathophysiological conditions, intracellular and extracellular H2O2 was difficult to be detected accurately. Herein, a colorimetric and homogeneous electrochemical dual-mode biosensing platform was constructed for intracellular/extracellular H2O2 detection based on FeSx/SiO2 nanoparticles (FeSx/SiO2 NPs) with high peroxidase-like activity. In this design, FeSx/SiO2 NPs were synthesized with excellent catalytic activity and stability compared to natural enzymes, which improved the sensitivity and stability of sensing strategy. 3,3',5,5'-Tetramethylbenzidine (TMB), as a multifunctional indicator, was oxidized in the presence of H2O2, generated color changes and realized visual analysis. In this process, the characteristic peak current of TMB decreased, which could realize the ultrasensitive detection of H2O2 by homogeneous electrochemistry. Accordingly, by integrating visual analysis ability of colorimetry and the high sensitivity of homogeneous electrochemistry, the dual-mode biosensing platform exhibited high accuracy, sensitivity and reliability. The detection limits of H2O2 were 0.2 µM (S/N = 3) for the colorimetric method and 2.5 nM (S/N = 3) for the homogeneous electrochemistry assay. Therefore, the dual-mode biosensing platform provided a new opportunity for highly accurate and sensitive detection of intracellular/extracellular H2O2.

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Human epidermal growth factor receptor 2 (HER2) has been regarded as the considerable biomarker of breast and gastric cancer. Thus, precise detection of HER2 is of significance for the early diagnosis and treatment. Here, a photofuel cell-based self-powered biosensor (PFC-SPB) was constructed for the ultrasensitive HER2 detection, which was composed of a plasmonic gold nanoparticles (Au NPs)/organic semiconductor hybrid photoanode and a cathode with biosensing strategy of electrochemical sandwich structure. The localized surface plasmon resonance effect of Au NPs can obviously enhance the separation efficiency of photo-generated electron/hole pair, which was beneficial to the sensitivity and stability of PFC-SPB. Meanwhile, the cathodic sandwich structure not only was used for the target recognition, but also can guarantee the enrichment of electroactive molecules (molybdophosphate). Consequently, with the open circuit voltage (EOCV) as the output signal, the PFC-SPB can achieve the HER2 detection in the range of 0.1-500 pg mL-1 with a low detection limit of 0.02 pg mL-1. Moreover, the as-proposed bioassay can be applied in cell lysate sample without any pretreatment, providing a promising and powerful tool early clinical diagnosis of cancer.

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Biofuel cells (BFCs)-based self-powered biosensors suffer from the limited stability of bioenzymes. Meanwhile, the poor performance of self-powered biosensors affects the sensitivity of biosensing, thus, it is significant and challenging to improve their stability and sensitivity. In our work, a BFC-based self-powered biosensor, with simultaneously enhanced stability and sensitivity, was constructed utilizing dual metal-organic frameworks (MOFs) as the carriers of the bioenzyme and the electroactive probe, respectively. Anodic enzyme, glucose dehydrogenase (GDH), was encapsulated in zeolitic imidazolate framework-8 (ZIF-8) to form GDH@ZIF-8 composites, enhancing the catalytic activity and stability of GDH. Meanwhile, another zirconium metal-organic frameworks (UiO-66-NH2) loaded with electroactive molecules (K3[Fe(CN)6]) served as nano-enrichment carriers and improved the capability of the cathode to accept electrons from the anode, further improving the sensitivity of the as-proposed biosensor. Herein, the "signal-on" BFC-based self-powered biosensing of exosomes, the model analyte, with excellent stability and outstanding sensitivity was realized with the assistance of dual MOFs, and the detection limit was down to 300 particles mL-1 (based on 3s/k), which was superior to those previously reported in literatures. Furthermore, the developed protocol was capable of detecting exosomes derived from cancer cells in complex biological samples. Overall, in this work the enhancement of both stability and sensitivity has been achieved by utilizing two types of MOFs, which laid the foundation for expanding the applications of BFC-based self-powered biosensors.

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Enzymatic biofuel cells (EBFCs) with or without a membrane to separate the anodic and cathodic compartments generally suffered from high internal resistance or interactive interference, both of which restricted the improvement of their performance. Herein, a smart membrane-less EBFC was engineered based on anode-driven controlled release of cathodic acceptor via pH-responsive metal-organic framework ([Fe(CN)6]3-@ZIF-8) nanocarriers. The glucose anodic oxidation would produce gluconic acid accompanied by the change in pH value from neutral to the acidic case, which could drive the degradation of [Fe(CN)6]3-@ZIF-8 nanocarriers and further realize the controlled release of cathodic acceptor [Fe(CN)6]3-. More importantly, compared with controlled EBFC with or without membrane, the power output of the as-proposed EBFC enhanced at least 700 times due to the seamless electronic communication. Therefore, the ingenious strategy not only realized the successful engineering of the membrane-less EBFC but also provided an appealing idea for constructing smart devices.

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Aberrant DNA methylation catalyzed by DNA methyltransferases (MTase) has proved to be associated with human diseases such as cancers. Thus, the development of an efficient strategy to accurately detect DNA MTase is highly desirable in medical diagnostics. Herein, we proposed a robust "signal-on" enzymatic biofuel cell (EBFC)-based self-powered biosensing platform with excellent anti-interference ability for DNA MTase activity analysis and inhibitor screening. In the presence of target MTase, the MTase-catalyzed DNA methylation occurred and hindered the HpaII endonuclease-catalyzed dsDNA dissociation, which enabled more bilirubin oxidase (BOD) to immobilize at the cathode surface via amidation. Then, BOD-catalyzed oxygen reduction took place by accepting electrons generated at the anode via glucose oxidation, thus leading to an elevated open-circuit voltage value, the amplitude of which was directly related to MTase concentration. The direct detection limit of the M.SssI assay was down to 0.005 U/mL, which was lower than that of those reported results. Notably, the as-proposed protocol was competent to detect DNA MTase activity directly in human serum samples without enrichment and separation, and applicable to the screening of M.SssI inhibitors. Considering the virtues of the excellent anti-interference ability, no requirement of external power, simplicity, and high accuracy, the biosensing platform would hold great potential in DNA MTase bioassay and clinical diagnosis of cancers.

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Enzymatic biofuel cell (EBFC)-based self-powered biosensors could offer significant advantages: no requirement for an external power source, simple instruments, and easy miniaturization. However, they also suffered from the limitations of lower sensitivity or specific targets. In this study, a self-powered biosensor for the ultrasensitive and selective detection of single nucleotide polymorphisms (SNPs) produced by combining the toehold-mediated strand displacement reaction (SDR) and DNA hybridization chain reaction (HCR) was proposed. Herein, the capture probe (CP) with an external toehold was designed to switch on the sensing system. In the presence of target sequence, both SDR and DNA HCR reaction would happen to produce a long double-helix chain. Because of the electrostatic interaction between [Ru(NH3)6]3+ and the double-helix chain described above, the open circuit voltage ( EOCV) of the as-proposed biosensor was significantly elevated, thus realizing the detection of SNPs. Overall, in this work, an ingeniously constructed self-powered biosensor for the detection of SNPs was created by integrating EBFCs with a DNA amplification strategy. Furthermore, the as-proposed self-powered biosensor not only showed prominent specificity to distinguish the p53 gene fragment from random sequences (e.g., single-base mutant sequences) but exhibited excellent sensitivity with the detection limit of 20 aM. More importantly, the results obtained from the real cell lysate sample have laid a strong foundation for disease diagnostics and, potentially, as a powerful tool for even more fields.

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Herein, a light-driven, membrane-less and mediator-less self-powered cytosensing platform via integration of biofuel cells (BFCs) and a photoelectrochemical strategy was developed for ultrasensitive detection of circulating tumor cells (CTCs). To construct cytosensors, an elaborately designed SH-Sgc8c aptamer/AuNP/g-C3N4 photoelectrode was used as an alternative anode for glucose oxidation, avoiding the introduction of anodic enzymes. Initially, glucose could favorably reach the photoanode surface and be easily oxidized by the photogenerated holes, while the photogenerated electrons would transfer to the biocathode and achieve biocatalytic reduction of O2, leading to a high EOCV. However, in the presence of CTCs, they could preferentially interact with the Sgc8c aptamer via specific recognition, and then complexes with large steric hindrance were immobilized on the photoanode surface, which could greatly affect the electron transfer between glucose and the photoanode surface. In this case, the EOCV decreased sharply. Encouragingly, this self-powered cytosensor exhibited an ultrasensitive response to the target CTCs in a wide concentration range from 20 to 2 × 105 cells mL-1 with a low detection limit of 10 cells mL-1 (S/N = 3), being superior to those of the reported methods. Moreover, this as-proposed self-powered cytosensor integrated with a photoelectrochemical strategy possessed unique advantages of not requiring an external power source, being anodic enzyme-free, having a simple construction process, facile miniaturization, and high selectivity and sensitivity, providing a promising and powerful tool for fundamental biochemical research and clinical diagnosis.

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We developed a facile and ultrasensitive enzymatic biofuel cell (EBFC)-based self-powered biosensor of protein kinase A (PKA) activity and inhibition via thiophosphorylation-mediated interface engineering. The detection limit was down to 0.00022 U mL-1 (S/N = 3). In addition, the PKA activities from MCF-7 and A549 cell lysates were analyzed and achieved reliable results.

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Biofuel cell (BFC)-based self-powered biosensors have attracted substantial attentions because of their unique merits such as having no need for power sources (only two electrodes are needed). More importantly, in case it can also work in a homogeneous system, more efficient and easy-to-use bioassays could come true. Thus, herein, we proposed a novel homogeneous self-powered biosensing strategy via the integration of BFCs and a homogeneous electrochemical method, which was further utilized for ultrasensitive microRNA (miRNA) detection. To construct such an assay protocol, the cathodic electron acceptor [Fe(CN)6]3- was entrapped in the pores of positively charged mesoporous silica nanoparticles and capped by the biogate DNAs. Once the target miRNA existed, it would trigger the controlled release of [Fe(CN)6]3-, leading to the dramatic increase of the open circuit voltage. Consequently, the "signal-on" homogeneous self-powered biosensor for the ultrasensitive miRNA assay was realized. Encouragingly, the limit of detection for the miRNA-21 assay was down to 2.7 aM (S/N = 3), obviously superior to those of other analogous reported approaches. This work not only provides an ingenious idea to construct the ultrasensitive and easy-to-use bioassays of miRNA but also exhibits a successful prototype of a portable and on-site biomedical sensor.

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A new label-free and enzyme-free ratiometric homogeneous electrochemical microRNA biosensing platform was constructed via target-triggered Ru(III) release and redox recycling. To design the effective ratiometric dual-signal strategy, [Ru(NH3)6]3+ (Ru(III)), as one of the electroactive probes, was ingeniously entrapped in the pores of the positively charged mesoporous silica nanoparticle (PMSN), and another electroactive probe, [Fe(CN)6]3- (Fe(III)), was selected to facilitate Ru(III) redox recycling due to its distinctly separated reduction potential and different redox properties. Owing to the liberation of the formed RNA-ssDNA complex from PMSN, the target miRNA triggered the Ru(III) release and was quickly electroreduced to Ru(II), and then, the in-site-generated Ru(II) could be chemically oxidized back to Ru(III) by Fe(III). Thus, with the release of Ru(III) and the consumption of Fe(III), a significant enhancement for the ratio of electroreduction current [Ru(NH3)6]3+ over [Fe(CN)6]3- (IRu(III)/IFe(III)) value was observed, which was dependent on the concentration of the target miRNA. Consequently, a simple, accurate, and ultrasensitive method for the miRNA assay was readily realized. Furthermore, the limit of detection (LOD) of our method was down to 33 aM (S/N = 3), comparable or even superior to other approaches reported in literature. More importantly, it also exhibited excellent analytical performance in the complex biological matrix cell lysates. Therefore, this homogeneous biosensing strategy not only provides an ingenious idea for realizing simple, rapid, reliable, and ultrasensitive bioassays but also has a great potential to be adopted as a powerful tool for precision medicine.

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Enzymatic fuel cell (EFC)-based self-powered biosensors have attracted considerable attention because of their unique feature of no need for extra power sources during the entire detection process, which endows them with the merits of simplicity, rapidness, low cost, anti-interference, and ease of use. Herein, we proposed, for the first time, an EFC-based self-powered homogeneous immunosensing platform by integrating the target-induced biofuel release and bioconjugate immunoassay for ultrasensitive melamine (ME) detection. In this design, the biofuel, i.e., glucose molecules, was entrapped in the pores of positively charged mesoporous silica nanoparticles and capped by the biogate AuNPs-labeled anti-ME antibody (AuNPs-Ab). The presence of the target ME triggered the entrapped glucose release due to the removal of the biogate via immunoreaction, which resulted in the transfer of electrons produced by glucose oxidation at the bioanode to the biocathode, and thus, the open-circuit voltage of the EFC-based self-powered immunosensor dramatically increased, realizing the ultrasensitive turn-on assay for ME. The limit of detection for ME assay was down to 2.1 pM (S/N = 3), superior to those previously reported in the literature. Notably, real milk samples need no special sample pretreatment for the detection of ME because of the good anti-interference ability of EFC-based self-powered biosensors and the excellent selectivity of the homogeneous immunoassay. Therefore, this appealing self-powered homogeneous immunosensing platform holds great promise as a successful prototype of portable and on-site bioassay in the field of food safety.

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Herein, we reported a novel ultrasensitive one-compartment enzyme biofuel cells (EBFCs)-based self-powered aptasensing platform for antibiotic residue detection. By taking full advantage of the unique features of both EBFCs-based self-powered sensors and aptamers, the as-proposed aptasensing platform has the merits of simple instrumentation, anti-interference ability, high selectivity, and low cost. In this study, DNA bioconjugate, i.e., SiO2@gold nanoparticles-complementary strand of aptamer (SiO2@AuNPs-csDNA), was elaborately designed and played a key role in blocking the mass transport of glucose to the bioanode. While in the presence of the target antibiotic, SiO2@AuNPs-csDNA bioconjugate broke away from the bioanode due to the aptamer recognition of the target. Without the blocking of glucose by the DNA bioconjugate, a significantly elevated open circuit voltage of the EBFCs-based aptasensor was obtained, whose amplitude was dependent on the antibiotic concentration. In addition, this proposed aptasensor was the first reported self-powered aptasensing platform for antibiotic determination and featured high sensitivity owing to the elaborate design of the DNA bioconjugate modified bioanode of EBFC, which was superior to those previously reported in the literature. Furthermore, due to the anti-interference ability and the excellent selectivity of the aptasensor, no special sample pretreatment was needed for the detection of antibiotics in milk samples. Therefore, the proposed EBFCs-based self-powered aptasensor has a great promise to be applied as a powerful tool for on-site assay in the field of food safety.

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