Live cell imaging

Carbon-based nano materials are developed as a cytocompatible alternative to semiconducting quantum dots for bioimaging and fluorescence-based sensing. The green alternatives for the synthesis of carbon materials are imminent. The present study demonstrates microwave based one step quick synthesis of fluorescent carbon material (FCM) having three variants: (i) un-doped fluorescent carbon material (UFCM) (ii) nitrogen doped FCM (N@FCM), and (iii) nitrogen & phosphorus co-doped FCM (N-P@FCM) using sugarcane extract as a carbon source...

A new study by Zenker et al. uses time-lapse imaging to discover how dynamic actin movements contribute to epithelialization of living mouse embryos. Together with work from other labs, this study presents exciting new ways to think about the emergence of cell fates during mammalian development.

Background & Aims: Cystic fibrosis (CF) patients and CF mouse models have increased risk for gastrointestinal tumors. CF mice show augmented intestinal proliferation of unknown etiology and an altered intestinal environment. We examined the role of the cystic fibrosis transmembrane conductance regulator (Cftr) in Wnt/β-catenin signaling, stem cell proliferation, and its functional expression in the active intestinal stem cell (ISC) population. Dysregulation of intracellular pH (pHi ) in CF ISCs was investigated for facilitation of Wnt/β-catenin signaling...

Herein, we design a smart autonomous ATP self-powered strand-displacement cascade amplification (SDCA) system for highly sensitive multiple intracellular miRNA detection. Rationally engineered Y-motif DNA structures are functionalized on mesoporous silica-coated copper sulfide nanoparticles loaded with numerous ATPs (CuS@mSiO2 -Y/ATP) through pH stimulus-responsive disulfide bonds. The SDCA system is implemented by endogenous specific miRNA as a trigger and ATP as fuel released from the nanocarrier at acidic pH and photothermal stimuli-responsive CuS...

Dopamine receptor expression is correlated with certain types of cancers, including lung, breast and colon cancers. In this study, we report luminescent iridium(iii) complexes ( 11-14 ) as intracellular dopamine receptor (D1R/D2R) cell imaging agents. Complexes 11 and 13 , which are conjugated with a dopamine receptor agonist, showed superior cell imaging characteristics, high stability and low cytotoxicity (>100 μM) in A549 lung cancer cells. siRNA knockdown and dopamine competitive assays indicated that complexes 11 and 13 could selectively bind to dopamine receptors (D1R/D2R) in A549 cells...

Carbon monoxide (CO) is a key gaseous signaling molecule in living cells and organisms. This protocol illustrates the synthesis of a highly sensitive Nile Red (NR)-Pd-based fluorescent probe, NR-PdA, and its applications for detecting endogenous CO in tissue culture cells, ex vivo organs, and zebrafish embryos. In the NR-PdA synthesis process, 3-diethylamine phenol reacts with sodium nitrite in the acidic condition to afford 5-(diethylamino)-2-nitrosophenol hydrochloride (compound 1), which is further treated with 1-naphthalenol at a high temperature to provide the NR dye via a cyclization reaction...

Minerals are versatile tools utilised to modify and control the physical-chemical and functional properties of substrates. Those properties include ones directing cell fate; thus, minerals can potentially provide a direct and inexpensive method to manipulate cell behaviour. This paper shows how different minerals influence human dermal fibroblast behaviour depending on their properties. Different calcium carbonates, calcium sulphates, silica, silicates, and titanium dioxide were characterised using TEM, ATR-FTIR, and zeta potential measurements...

A novel rhodamine based sensor RBH was developed and synthesized by rhodamine B with 3-methyl-2-benzothiazolinone hydrazone, which exhibited high selectivity and sensitivity for sensing Cu2+ ions in the presence of other important relevant metal ions in aqueous acetonitrile. As an "off-on" fluorescent sensor, RBH exhibited a UV-vis absorption increase at 570 nm and a 132-fold significant fluorescence enhancement at 580 nm with a distinct color change from colorless to purple upon the addition of Cu2+ ions...

As a critical gaseous signaling molecule, H2 S is involved in various biological processes. To deeper study the physiological and pathological roles of H2 S, convenient and efficient detection techniques for endogenous H2 S in vivo are still in urgent demand. Herein, we reported a new turn-on Near-infrared (NIR) fluorescence probe NIR-H2 S based on thiolysis reactions for detection of H2 S. The probe possessed many excellent properties including high sensitivity and selectivity, good cell-membrane permeability, and low cytotoxicity...

Giant plasma membrane vesicles (GPMVs) are isolated directly from living cells and provide an alternative to vesicles constructed of synthetic or purified lipids as an experimental model system for use in a wide range of assays. GPMVs capture much of the compositional protein and lipid complexity of intact cell plasma membranes, are filled with cytoplasm, and are free from contamination with membranes from internal organelles. GPMVs often exhibit a miscibility transition below the growth temperature of their parent cells...

BACKGROUND: Suitable fluorophores are the core of fluorescence imaging. Among the most exciting, yet controversial, labels are quantum dots (QDs) with their unique optical and chemical properties, but also considerable toxicity. This hinders QDs applicability in living systems. Surface chemistry has a profound impact on biological behavior of QDs. This study describes a two-step synthesis of QDs formed by CdTe core doped with Schiff base ligand for lanthanides [Ln (Yb3+ , Tb3+ and Gd3+ )] as novel cytocompatible fluorophores...

Dysfunction of the human ether-a-go-go related gene (hERG)-encoded potassium channel is identified as a major cause of the long Q-T syndrome, a marker for the lethal cardiac arrhythmia. Furthermore, recent studies revealed that hERG K+ channel is a regulator of tumor cell apoptosis and proliferation. Herein, an ultrasensitive fluorescence assay combining DNA-functionalized gold nanoparticles and rolling circle amplification (RCA) is first attempted to visualize hERG channels in living cells. The spherical nucleic acids gold nanoparticles which can anchor on hERG channels in the cell membrane, not only act as the primary amplification elements, but also trigger the subsequent RCA reaction to achieve the secondary amplification...

Noninvasive imaging of reporter gene expression by two-photon excitation (2PE) laser scanning microscopy is uniquely suited to perform dynamic and multidimensional imaging down to single-cell detection sensitivity in vivo in deep tissues. Here we used 2PE microscopy to visualize green fluorescent protein (GFP) as a reporter gene in human melanoma cells implanted into the dermis of the mouse ear skin. We first provide a step-by-step methodology to set up a 2PE imaging model of the mouse ear's skin and then apply it for the observation of the primary tumor and its associated vasculature in vivo...

A series of benzothiazolium-based hemicyanines (3a-3f) have been synthesized. Evaluation of their photophysical properties shows that they exhibit improved photophysical characteristics. In comparison with the available commercial MitoTrackers, the new probes revealed an enhanced Stokes shift (Δλ ∼ 80 nm) and minimized aggregation for increased sensitivity. The synthesized probes are found to exhibit excellent selectivity for mitochondrial staining in an oligodendrocyte cell line. Probes show almost no fluorescence in aqueous environments, while the fluorescence is increased by ∼10-fold in organic solvents, making it possible for mitochondrial imaging without the need for post-staining washing...

Viruses threaten humans, livestock, and plants, and are difficult to combat. Imaging of viruses by light microscopy is key to uncover the nature of known and emerging viruses in the quest for finding new ways to treat viral disease and deepening the understanding of virus–host interactions. Here, we provide an overview of recent technology for imaging cells and viruses by light microscopy, in particular fluorescence microscopy in static and live-cell modes. The review lays out guidelines for how novel fluorescent chemical probes and proteins can be used in light microscopy to illuminate cells, and how they can be used to study virus infections...

The anisotropic organization of plasma membrane constituents is indicative of mechanisms that drive the membrane away from equilibrium. However, defining these mechanisms is challenging due to the short spatio-temporal scales at which diffusion operates. Here, we use high-density single protein tracking combined with photoactivation localization microscopy (sptPALM) to monitor Cdc42 in budding yeast, a system in which Cdc42 exhibits anisotropic organization. Cdc42 exhibited reduced mobility at the cell pole, where it was organized in nanoclusters...

CRISPR-Cas RNA-guided endonucleases hold great promise for disrupting or correcting genomic sequences through site-specific DNA cleavage and repair. However, the lack of methods for cell- and tissue-selective delivery currently limits both research and clinical uses of these enzymes. We report the design and in vitro evaluation of S. pyogenes Cas9 proteins harboring asialoglycoprotein receptor ligands (ASGPrL). In particular, we demonstrate that the resulting ribonucleoproteins (Cas9-ASGPrL RNP) can be engineered to be preferentially internalized into cells expressing the corresponding receptor on their surface...

Recently, although several unconventional luminescent polymers have been synthesized, it still remains a significant challenge to prepare various new fluorescent polymers by functionalization of nonflurescent polymers. In the present study, a nonfluorescent 1H,1H,2H,2H-perfluoro-1-decanol grafted to nonflurescent polystyrene-b-poly(acrylic acid) block copolymers through simply esterification reaction can exhibit strong blue emission. Based on control experiments and theoretical simulation, we have proposed that the luminescence stems from interchain n→π* interaction between the lone pair (n) of hydroxyl O atoms of carboxyl units and empty π* orbital of ester carbonyl unit...

Multi-stimuli-responsive L-tyrosine based amphiphilic poly(ester-urethane)s nano-carriers were designed and developed, for the first time, to administrate anticancer drugs at cancer tissue environment via thermo-responsiveness and lysosomal enzymatic-biodegradation from single polymer platform. For this purpose, multifunctional L-tyrosine monomer was tailor-made with PEG-lated side chain at the phenolic position along with urethane and carboxylic ester functionalities. Under melt dual ester-urethane polycondensation, the tyrosine-monomer reacted with diols to produce high molecular weight amphiphilic poly(ester-urethane)s...

Nitroreductase and acidity are common features of a tumor, which is associated with serious disease with a high mortality rate. However, solo detection of nitroreductase or acidic environments may cause false-positive results. Thus, it is very meaningful to develop dual functional probes for detecting nitroreductase and acidity, as these could be used for accurate tumor imaging. Here we report on the first dual functional ratiometric fluorescent probe, NAFP, for acidity and nitroreductase detection. This probe enables the measurement of the acidic microenvironment and the concentration of nitroreductase at the same time and the same site...