1. BackgroundSmall molecules, such as neurotransmitters, are essential for biochemical functions in living systems, including cell signaling and gut-brain communication. However, real-time detection and analysis of neurotransmitter release remains challenging. Conventional detection methods, such as UV-Vis spectroscopy and mass spectrometry, require sample collection and offline analysis, and cannot be measured in real time. Rapid scanning cyclic voltammetry, combined with microdialysis, provides time-resolved measurements of neurotransmitter release with nanomolar sensitivity, but sensitivity decreases over time due to electrode contamination. Although recent advances in chemically modified electrodes and engineered waveforms have improved selectivity, these methods still struggle to solve the problem of oxidation of multiple species at overlapping potentials. Fiber optoelectronics, utilizing genetically encoded fluorescent indicators, are hampered by photobleaching and sensitivity limitations, despite their high specificity and resolution. Field-effect transistor (FET) biosensors using aptamers have emerged as a promising solution for detecting small, negligible charged or neutral molecules. Aptamers, i.e., DNA stem loops that undergo conformational changes when combined with small molecules, enhance the sensitivity and selectivity of FET sensors. However, the readout in these devices requires an ensemble signal of the dense probe molecules that connect the gate of the device. This signal lacks kinetic information about molecular interactions. In addition, it is easily disrupted by the transconductance drift inherent in salt solutions, making it difficult to measure temporal changes in analyte concentrations. 2. Research resultsRecently,Columbia Universitykenneth l. shepardCarbon nanotube-based single-molecule field-effect transistors (FETs) are reportedsmfets).It is able toDetect the charge on a single moleculeYesAs a new platform for the identification and analysis of small molecules. SMFETs are formed by the covalent attachment of probe molecules (DNA aptamers) to carbon nanotubes. The conformational change upon binding is manifested as a discrete change in the conductance of the nanotubes. By monitoring the dynamics of conformational changes in the conjugated aptamer, it has been shown that SMFETS can detect and quantify serotonin at the single-molecule level, providing unique insights into the dynamics of the aptamer-ligand system In particular, the researchers demonstrated the formation of G-quadruplexes and the disruption of natural hairpin structures in conformational changes in the serotonin-aptamer complex. SMFETS is a label-free method for analyzing molecular interactions at the single-molecule level with high temporal resolution, providing additional insights into complex biological processes. Related research work to:Carbon-nanotube field-effect transistors for resolving single-molecule aptamer ligand binding kinetics" was published in the top international journal Nature Nanotechnology. 
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Figure 1Device schematics and concentration-dependent current traces.
Figure 2Single-molecule dynamics analysis.
Figure 3Serotonin binding and kinetics for HMM analysis.
Figure 4HMM analysis estimated dose-response curves occupied by bound states.
Figure 5Conformational schematic diagram of the serotonin-aptamer complex on CNT FETs.
4. Conclusions and prospectsThe compact nucleic acid receptor enables dynamic studies of single-molecule interactions within the Debye layer, demonstrating the potential of SMFETS technology for fine molecular analysis. Relying on the G-quadruplex structure for detection alone can unnecessarily complicate the kinetic pathway. Non-G-quadruplex sequences in aptamers can be designed in the future to produce more direct detection with enhanced signal transduction. The electrostatically force-influenced dynamics of the SMFETS system provide the opportunity to modulate the assay affinity, which is critical for the optimization of single-molecule detection over a wide range of dynamic concentrations of various target molecules. High-density sensor arrays with integrated CMOS electronics are expected to be used for low-concentration and multiplexed detection, as well as for spatially distributed analytes in brain regions. The latter is particularly relevant for in vivo neurotransmitter assays, providing insights into the correlation between neural activity and neurotransmitter release, deepening the understanding of local neurodynamics. Literature Links: