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Dual-emissive protein-capped metal nanoclusters are emerging nanoprobes for ratiometric sensing. Here, we report dual-emissive bovine serum albumin (BSA)-capped silver nanoclusters (AgNCs) with a blue emission band at 450 nm and a far-red emission band at 680 nm. The blue emission is attributed to the BSA capping, while the far-red emission originates from the AgNCs. The intensity ratio of the two bands is susceptible to the presence of IO4– and Cu2+, allowing selective detection of these analytes with a limit of detection (LOD) of 18.8 and 21.6 nM, respectively. These two analytes display different effects on the emission bands. The blue emission band increases with simultaneous diminution of the far-red band in the presence of IO4–; however, only the far-red emission band decreases in the presence of Cu2+ without altering the blue emission. Both the analytes manifested distinct visual changes in the emission color of the AgNCs solution under a UV lamp, which can also be evident from the shift of the Commission Internationale d'Eclairage (CIE) coordinates from (0.48, 0.32) to (0.18, 0.17) and (0.24, 0.25) for IO4– and Cu2+, respectively. IO4– induced a conformational change in BSA, intensifying the blue emission band along with a dynamic quenching of the AgNCs' emission at 680 nm. In contrast, Cu2+ triggered only a metal exchange method-based quenching of the 680 nm emission with almost no effect on the BSA capping. A smartphone-based on-site detection of IO4– and Cu2+ was also achieved. Inspired by the contrasting fluorescence (Fl) response signals of AgNCs toward IO4– concentration, a simple logic gate circuit was constructed. This investigation will provide important insight into the one-pot synthesis of dual-emissive silver nanoclusters and the ratiometric detection mechanism of IO4– and Cu2+. [ABSTRACT FROM AUTHOR]

Dual-emissive carbon dots (D-CDs) are in high demand for multipurpose applications. Herein, nitrogen-doped carbon dots possessing inherent dual emissions at green (490 nm) and yellow (570 nm) are synthesized hydrothermally from 3,4-diaminobenzoic acid and hydrazine hydrate. The relative intensity of the two emission bands is dependent markedly on the pH of medium, making the D-CDs a robust pH sensor within the pH range of 2.0–7.6. The ratiometric emission of D-CDs is sensitive to trace amounts of Fe3+; the 490 nm emission intensity diminishes with simultaneous amplification of the 570 nm emission intensity. A discernible color change from light green to bright yellow is evident under an ultraviolet (UV) lamp with increasing Fe3+ concentrations, and the CIE color coordinate shifts from (0.33, 0.43) to (0.47, 0.50). Primarily, the complexation between D-CDs and Fe3+ and the accompanying aggregation play a dominant role in the ratiometric change of the emission intensities. For the on-spot detection of Fe3+, smartphone-based and solid-phase sensing is accomplished. The D-CDs also display decent anticancer properties against the breast cancer cell line MCF-7 and effectively detect the intracellular pH and exogenous Fe3+ inside the MCF-7 cells. [ABSTRACT FROM AUTHOR]

Biomolecule-stabilized metal nanoclusters (MNCs) exhibit enormous potential as unique luminescent materials in various applications. However, synthesis of highly stable and bio-friendly fluorescent MNCs is still a challenge. Here, we report a facile synthesis of red-emitting silver nanoclusters (LYS–AgNCs) within a dithiothreitol-reduced lysozyme (LYS) scaffold. The nanoclusters exhibit a uniform size distribution, excellent water solubility, and superior photoluminescence properties featuring a quantum yield of 6.1%, a massive (280 nm) Stokes shift, and solid-state emission, and pH stability augments their applicability in dual-mode sensing platforms for Cu2+ and vitamin B12 (VB12). Two contrasting fluorescence (FL)-quenching mechanisms are responsible for the sensing; Cu2+-induced FL quenching occurs via a multifaceted mechanism involving both static and dynamic quenching, whereas the inner filter effect and Förster resonance energy transfer are mainly responsible for VB12-induced FL quenching. An inexpensive portable paper strip is fabricated using LYS–AgNCs for on-site field application, enabling instrument-free fast and visual detection of Cu2+ and VB12. Moreover, LYS–AgNCs also possess favorable biocompatibility against human cervical cancer cells (HeLa), making them a suitable nanoprobe for cell imaging and an efficient agent for detecting Cu2+ and VB12 inside live cells as well. Finally, we demonstrate the applicability of LYS–AgNCs for real-sample analysis of VB12 with a satisfactory outcome. [ABSTRACT FROM AUTHOR]

Multiplex protein imaging technologies enable deep phenotyping and provide rich spatial information about biological samples. Existing methods have shown great success but also harbored trade-offs between various pros and cons, underscoring the persisting necessity to expand the imaging toolkits. Here we present PACIFIC: photoactive immunofluorescence with iterative cleavage, a new modality of multiplex protein imaging methods. PACIFIC achieves iterative multiplexing by implementing photocleavable fluorophores for antibody labeling with one-step spin-column purification. PACIFIC requires no specialized instrument, no DNA encoding, or chemical treatments. We demonstrate that PACIFIC can resolve cellular heterogeneity in both formalin-fixed paraffin-embedded (FFPE) samples and fixed cells. To further highlight how PACIFIC assists discovery, we integrate PACIFIC with live-cell tracking and identify phosphor-p70S6K as a critical driver that governs U87 cell mobility. Considering the cost, flexibility, and compatibility, we foresee that PACIFIC can confer deep phenotyping capabilities to anyone with access to traditional immunofluorescence platforms., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

We present a chemical approach to profile fatty acid uptake in single cells. We use azide-modified analogues to probe the fatty acid influx and surface-immobilized dendrimers with dibenzocyclooctyne (DBCO) groups for detection. A competition between the fatty acid probes and BHQ2-azide quencher molecules generates fluorescence signals in a concentration-dependent manner. By integrating this method onto a microfluidics-based multiplex protein analysis platform, we resolved the relationships between fatty acid influx, oncogenic signaling activities, and cell proliferation in single glioblastoma cells. We found that p70S6K and 4EBP1 differentially correlated with fatty acid uptake. We validated that cotargeting p70S6K and fatty acid metabolism synergistically inhibited cell proliferation. Our work provided the first example of studying fatty acid metabolism in the context of protein signaling at single-cell resolution and generated new insights into cancer biology.