Accurate and efficient detection of pathogenic bacteria plays a crucial role in the diagnosis of infectious diseases. However, rapid and highly sensitive detection of specific bacteria in clinical samples remains challenging. In this study, we developed a ratiometric dual‐signal electrochemical biosensor for ultrasensitive detection of pathogenic bacteria, based on an aptamer recognition‐induced rolling circle amplification (RCA)/G‐quadruplex strategy. On the surface of a gold electrode chip, we used an aptamer sequence (P1) conjugated to ferrocene (Fc) to capture the target bacterium. This capture released a different, previously bound sequence (P2) sequence that initiated the RCA/G‐quadruplex cascade, which, in the presence of potassium ions, was able to bind the electrochemical indicator methylene blue (MB). We integrated the signals from the loss of Fc and the appearance of MB (IMB/IFC ratio) to quantify the target bacteria concentration. This biosensor showed excellent detection performance and specificity, with a detection limit of 10 CFU/mL. Notably, it showed great diagnostic potential for clinical infectious diseases caused by pathogenic bacteria in precise/personalized medicine applications.

Bioactive small molecules serve as invaluable biomarkers for recognizing modulated organismal metabolism in correlation with numerous diseases. Therefore, sensitive and specific molecular biosensing and imaging in vitro and in vivo are particularly critical for the diagnosis and treatment of a large group of diseases. Herein, a modular DNA tetrahedron-based nanomachine was engineered for the ultrasensitive detection of intracellular small molecules. The nanomachine was composed of three self-assembled modules: an aptamer for target recognition, an entropy-driven unit for signal reporting, and a tetrahedral oligonucleotide for the transportation of the cargo (e.g., the nanomachine and fluorescent markers). Adenosine triphosphate (ATP) was used as the molecular model. Once the target ATP bonded with the aptamer module, an initiator was released from the aptamer module to activate the entropy-driven module, ultimately activating the ATP-responsive signal output and subsequent signal amplification. The performance of the nanomachine was validated by delivering it to living cells with the aid of the tetrahedral module to demonstrate the possibility of executing intracellular ATP imaging. This innovative nanomachine displays a linear response to ATP in the 1 pM to 10 nM concentration range and demonstrates high sensitivity with a low detection limit of 0.40 pM. Remarkably, our nanomachine successfully executes endogenous ATP imaging and is able to distinguish tumor cells from normal ones based on the ATP level. Overall, the proposed strategy opens up a promising avenue for bioactive small molecule-based detection/diagnostic assays.

Separation of micro/nanoparticles, such as cellular, subcellular and biomolecular, has attracted increasing attention because of their remarkable potential applications in various fields, including chemistry, physics, medicine, etc. Among different micro/nanoparticle separation methods, acoustofluidics, which combines acoustics and microfluidics, has drawn the interest of researchers due to its biocompatibility, high efficiency and free labeling. In this review, the basic constitutions, mechanisms, and materials of acoustofluidics are described. Subsequently, sorts of delicately designed acoustofluidic devices, including diverse bulk acoustic wave (BAW) microfluidics and surface acoustic wave (SAW) microfluidics, are discussed, covering principles, advantages, limitations and applications in separation. Besides the introduction of advances of micro/nanoparticle separation in the BAW microfluidics, the SAW microfluidics are elaborated in detail with a focus on various configurations of interdigital transducers (IDTs), comprising straight IDT, slanted‐finger IDT, chirped IDT and focused IDT. Microfluidic systems of the acoustofluidics involve the forms of straight channels, serpentine channels, and droplets. Additionally, besides simply structured acoustofluidics, acoustofluidics integrated with other structures are also mentioned. Finally, the prospects and limitations of acoustofluidics in micro/nanoparticle separation are also discussed. The acoustofluidics reviewed here is envisioned as a versatile tool for micro/nanoparticle separation at the cellular, sub‐cellular, and biomolecular levels.