Nanopipettes, as a class of solid-state nanopores, have evolved into universal tools in biomedicine for the detection of biomarkers and different biological analytes. Nanopipette-based methods combine high sensitivity, selectivity, single-molecule resolution, and multifunctionality. The features have significantly expanded interest in their applications for the biomolecular detection, imaging, and molecular diagnostics of real samples. Moreover, the ease of manufacturing nanopipettes, coupled with their compatibility with fluorescence and electrochemical methods, makes them ideal for portable point-of-care diagnostic devices. This review summarized the latest progress in nanopipette-based nanopore technology for the detection of biomarkers, DNA, RNA, proteins, and peptides, in particular β-amyloid or α-synuclein, emphasizing the impact of technology on molecular diagnostics. By addressing key challenges in single-molecule detection and expanding applications in diverse biological areas, nanopipettes are poised to play a transformative role in the future of personalized medicine.

Background/Objectives: One of the hallmarks of Alzheimer’s disease (AD) is the accumulation of aggregated beta-amyloid (Aβ) protein in the form of senile plaques within brain tissue. Senile plaques contain various post-translational modifications of Aβ, including prevalent isomerization of Asp7 residue. The Asp7 isomer has been shown to exhibit increased neurotoxicity and induce amyloidogenesis in brain tissue of transgenic mice. The toxicity of Aβ peptides may be partly mediated by their structure and morphology. In this respect, in this study we analyzed the structural and aggregation characteristics of the Asp7 isoform of Aβ₄₂ and compared them to those of synthetic Aβ₄₂. We also investigated the effects of intracerebroventricular (i.c.v.) administration of these peptides, a method often used to induce AD-like symptoms in rodent models. Methods: Atomic force microscopy (AFM) was conducted to compare the morphological and aggregation properties of Aβ₄₂ and Asp7 iso-Aβ₄₂. The effects of i.c.v. stereotaxic administration of the proteins were assessed via behavioral analysis and reactive oxygen species (ROS) estimation in vivo using a scanning ion-conductance microscope with a confocal module. Results: AFM measurements revealed structural differences between the two peptides, most notably in their soluble toxic oligomeric forms. The i.c.v. administration of Asp7 iso-Aβ₄₂ induced spatial memory deficits in rats and elevated oxidative stress levels in vivo, suggesting a potential of ROS in the pathogenic mechanism of the peptide. Conclusions: The findings support the further investigation of Asp7 iso-Aβ₄₂ in translational research on AD and suggest its involvement in neurodegenerative processes.

Alzheimer’s disease (AD) is a worldwide problem due to the lack of effective therapy and accurate methods for timely diagnosis. The complexity of AD’s pathophysiology complicates the development of effective therapeutic agents, as most drugs act on only one therapeutic target, bypassing others. The design and development of multifunctional agents capable of altering metal ion-induced abnormalities, oxidative stress, and toxic beta amyloid (Aβ) aggregates is of interest. Herein, we report the first boron dipyrromethene (BODIPY) based bifunctional copper chelator with clioquinol, BDP-CLQ, capable of both optical detection of Aβ fibrils and copper chelation, with multiple anti-AD properties. Foremost, BDP-CLQ demonstrated a 3-fold and 5-fold fluorescence increase at 650 nm and 565 nm in the presence of Aβ and effective copper chelation (pK_d = 16.6 ± 0.3). In addition, BDP-CLQ demonstrated a potent inhibition of Aβ aggregation, reduction in Aβ-induced stiffness of neuronal cells, and antioxidant activity. BDP-CLQ is the first BODIPY-based fluorescent probe with multiple anti-AD activities, as well as the first clioquinol-based probe capable of Aβ optical visualization. This study demonstrates the prospects of the development of clioquinol-based theranostic probes since this allows combining several promising anti-AD actions in a single molecule and developing multi-targeted drugs.