Sensitive and selective monitoring of sialic acid (SA) in cerebral nervous system is of great importance for studying the role that SA plays in the pathological process of Alzheimer's disease (AD). In this work, we first reported an electrochemical biosensor based on a novel stimuli-responsive copolymer for selective and sensitive detection of SA in mouse brain. Notably, through synergetic hydrogen-bonding interactions, the copolymer could translate the recognition of SA into their conformational transition and wettability switch, which facilitated the access and enrichment of redox labels and targets to the electrode surface, thus significantly improving the detection sensitivity with the detection limit down to 0.4 pM. Besides amplified sensing signals, the proposed method exhibited good selectivity toward SA in comparison to potential interference molecules coexisting in the complex brain system due to the combination of high affinity between phenylboronic acid (PBA) and SA and the directional hydrogen-bonding interactions in the copolymer. The electrochemical biosensor with remarkable analytical performance was successfully applied to evaluate the dynamic change of SA level in live mouse brain with AD combined with in vivo midrodialysis. The accurate concentration of SA in different brain regions of live mouse with AD has been reported for the first time, which is beneficial for progressing our understanding of the role that SA plays in physiological and pathological events in the brain.

Cyanine dyes are widely used in biological labeling and imaging because of their narrow near infrared emission, good brightness and high flexibility in functionalization, which not only enables multiplex analysis and multi-color imaging, but also greatly reduces autofluorescence from biological matter and increases signal-to-noise ratio. Unfortunately, their poor chemical- and photo-stability strongly limits their applications. The incorporation of cyanine dyes in silica nanoparticles provides a solution to the problem. On one hand, the incorporation of cyanine dyes in silica matrix can enhance their chemical- and photo-stability and increase brightness of the nanomaterials. On the other hand, silica matrix provides an optimized condition to host the dye, which helps to maintain their fluorescent properties during application. In addition, the well-established silane technique provides numerous functionalities for diverse applications. However, commercially available cyanine dyes are not very stable at high alkaline conditions, which will gradually lose their fluorescence over time. Our results showed that cyanine dyes are very vulnerable in the reverse micelle system, in which they will lose their fluorescence in less than half an hour. The existence of surfactant could greatly promote degradation of cyanine dyes. Fluorescent silica nanoparticles cannot be obtained at the high alkaline condition with the existence of surfactant. In contrast, the cyanine dyes are relatively stable in Stöber media. Owing to the fast formation of silica particles in Stöber media, the exposure time of cyanine dye in alkaline solution was greatly reduced, and highly fluorescent particles with good morphology and size distribution could be obtained via Stöber approach. However, the increasing water content in the Stöber could reduce the stability of cyanine dyes, which should be avoided. This research here provides a clear guidance on how to successfully synthesize cyanine dye-doped silica nanoparticles with good morphology, size distribution, stability and brightness.

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Luminescence lifetime-based nanosensors for chloride ions were designed by incorporating a luminescent ruthenium dye [Ru­(1,10-phenanthroline)3] inside silica nanoparticles and chemically labelling their outer surface with chloride ion-sensitive fluorescent dyes (N,N′-bis­(carboxypropyl)-9,9′-biacridine). The nanosensor surface was further functionalized with positively charged amino groups to facilitate intracellular uptake via endocytosis and target lysosomes. The nanosensors have an average diameter of 52 nm and are monodispersed in aqueous solutions. Because of the long lifetime of the reference ruthenium dye, the sensor response can be analyzed using the time-domain dual-lifetime referencing (td-DLR) approach. The use of pulsed excitation in td-DLR rather than intense continuous illumination in ratiometric measurements greatly prevents the dye from photobleaching which significantly improves its measurement stability and reproducibility for long-term monitoring. At optimum conditions, the sensor can measure chloride concentration in the range of 0–200 mM with a large ratiometric signal change from 140.9 to 40.2. Combined with our custom-built microscopic td-DLR system, variations of intracellular chloride concentration in lysosomes were imaged quantitatively with a high spatial resolution and accuracy.