Ultralong-Term Super-Resolution Tracking of Lysosomes in Brain Organoids by Near-Infrared Noble Metal Nanoclusters

Abstract

To study lysosomes with an ultralong term, herein, noble-metal Au and Ag nanoclusters (MNCs) modified with biocompatible protein bovine serum albumin (BSA) called BSA–NCs were applied to image lysosomes under structured illumination microscopy (SIM). Compared with commercial lysosomal dyes, BSA–NCs exhibited remarkable anti-interference in pH fluctuation, excellent antiphotobleaching for ultralong-term lysosomal tracking, and impressive near-infrared fluorescence. The lysosomal events (i.e., fission, fusion, and kiss-and-run events) were observed with the BSA–NCs under SIM, as was the interplay between lysosomes and mitochondria leading to mitophagy. With their molecular-like properties and near-infrared fluorescence, BSA–Au NCs proved able to enter the internal cells of organoids and, for the first time, were applied to track subcellular lysosomal dynamics. The results not only demonstrate that MNCs are excellent tools for imaging organelles in living cells and organoids with super-resolution technology but also highlight new fields for using MNCs in ultralong-term bioimaging.

Enhancing Mitochondrial Functions by Optogenetic Clustering

“…The photoactivatable CRY2PHR was anchored to the surface of mitochondria, via the specific mitochondria-targeting transmembrane domain Miro1TM (transmembrane domain of Miro1),[3a] and mCherry was used as a marker for the system’s expression, generating CRY2PHR-mCherry-Miro1TM (mCherry, a member of the mFruits family of monomeric red fluorescent proteins, Figure 1A). After transfecting different concentrations of the designed plasmid into HeLa cells, structured illumination microscopy (SIM) was used to resolve mitochondria[11]. For the appropriate fluorescent intensity and low background interference, 2.0 μg (in 200 μL DMEM, 35 mm dish; DMEM, Dulbecco’s modified eagle medium) was determined as the optimal concentration for the following transfection (Figure S1).…”

Quantifying cell viability through organelle ratiometric probing

“…Resistance to photobleaching is a vital ability of probes that determines whether it can function for long-time fluorescence tracking 22,23 . According to the chemical structure and previous fluorescence stability tests, PCV-1 seems to be endowed with a high resistance to photobleaching.…”

“…10 μm) 19 , structured illumination microscopy (SIM) may be a promising alternative, especially given its super spatial resolution (i.e., approx. 100 nm), short acquisition time, and only slight photodamage [20][21][22][23] .…”

BSA-Capped Dual-Emissive Silver Nanoclusters for Detection of IO4– and Cu2+ Ions

“…Metal nanoclusters (MNCs) possessing inherent intriguing features, viz., molecule-like discrete energy levels, ultrasmall size, and remarkable fluorescence (Fl), continue gaining attention among researchers. − Their exceptional inherent properties render them an extensive platform for offering various applications like sensing toxic metal ions − and hazardous molecules, , bioimaging, photodynamic theory, catalysis, and optoelectronics . MNCs, particularly with red or near-infrared (NIR) emission, possess remarkable advantages in contrast to lower wavelength emitting MNCs due to their beneficial properties like deep cell penetrating power, minimizing the possibility of light scattering, and bypassing autofluorescence, − augmenting their utilization for various applications ranging from sensing of environmental contaminants to biologically hazardous molecules. − MNCs capped by proteins offer excellent biocompatibility and optical stability. − Following the first report of bovine serum albumin (BSA)-capped gold nanoclusters (AuNCs), several other proteins, e.g., lysozyme (LYS), ovalbumin, insulin, pea protein, and pepsin, have been utilized for capping many other MNCs. Induction of biocompatible reducing agents like dithiothreitol (DTT) in the synthesis further improves their applicability in biological systems .…”