Probes For Cell Imaging Research Articles

Excitation energy transfer based naphthalimide-boron-dipyrromethene dyads as two-photon fluorescent probes for palladium(0) detection and live cell imaging

Increasing industrial demand has accelerated the consumption and contamination of palladium (Pd) worldwide. The implementation of fluorescent probes, especially with two-photon excitation, holds great promise for Pd detection in environmental and biological analysis. In this work, two naphthalimide (NPI)-boron-dipyrromethene (BDP) dyads (named NBH and NBN) were developed as two-photon fluorescent probes for Pd(0) detection based on an excitation energy transfer (EET) strategy, employing NPI as the energy donor and BDP as the acceptor. Spectroscopic studies revealed that both probes exhibited a fast response to Pd(0) in a pronounced fluorescence "ON-OFF" manner. The Pd-catalyzed Tsuji-Trost reaction activated an intramolecular photoinduced electron transfer (PET) process between the aniline and BDP moieties, resulting in the fluorescence quenching of the BDP fluorophore. The reaction-based probes exhibited excellent sensitivity and selectivity for Pd(0) species with a detection limit as low as 2.4 nM. These probes were successfully applied to the determination of Pd residues in drug, soil, and water samples. A simple smartphone-based platform was also constructed to determine Pd(0) via an RGB reader. With negligible cytotoxicity, confocal imaging study validated their feasibility of monitoring Pd(0) in living cells through multiple excitation channels.

Water-soluble sulfur quantum dots as a potential sensitive fluorescent probe for quercetin detection and cell imaging

A simple fluorescent quenching probe based on polyethylene glycol-400 capped sulfur quantum dots (PEG-SQDs) was fabricated to determine quercetin (QT) quantitatively. As anticipated, the PEG-SQDs exhibited favourable luminescent properties, stability and low cytotoxicity. QT effectively quenched the fluorescence of the PEG-SQDs through static quenching and the inner filter effect. Moreover, the PEG-SQDs showed rapid QT detection within a linear range of 0.100–45.0 μM, with a limit of detection of 0.014 μM (3σ/k). This fluorescent probe successfully detected QT in human serum, quercetin supplement capsules and red wine, achieving a standard recovery of 92.6 %–105 %.

Amino-Functionalized Metal-Organic Framework as Fluorescence Probe for Cell Imaging and Doxorubicin Detection.

In this study, an NH2-Cu-MOF was synthesized using the one-pot method and employed as a fluorescence probe for doxorubicin (DOX) detection. The synthesized NH2-Cu-MOF exhibited remarkable fluorescence recognition capabilities for detecting DOX. Within the concentration range of 1-100 µmol/L, a linear relationship was observed between the fluorescence intensity of the NH2-Cu-MOF and the DOX concentration. Furthermore, the synthesized NH2-Cu-MOF was effectively utilized for highly selective fluorescence quenching recognition and quantitative detection of DOX in the presence of multiple metal ions and other antibiotics. Despite interference from multiple metal ions and antibiotics, DOX was identified and quantified by highly selective fluorescence quenching with a detection limit of 2.1654 µmol/L. These findings underscore the potential of NH2-Cu-MOF as a class of "on-off" fluorescent probes for the rapid detection of DOX.

BOPHY Derivatives as Phototherapeutic Agents for the Photodynamic Inactivation of Microorganisms

AbstractThe improvement of photodynamic inactivation (PDI) significantly depends on the development of new families of photosensitizers (PSs). In this sense, three BOPHY derivatives (BP, BP‐Br and BP‐I) were synthetized, studied, and compared to assess their antimicrobial photodynamic properties. BP is an interesting fluorescent probe for cell imaging, while the halogenated analogs (BP‐Br and BP‐I) are excellent oxygen photosensitizing agents. BP compound presented a fluorescence quantum yield close unity and showed no reactive oxygen species (ROS) production. In contrast, BP‐I did not show emission properties but exhibited a high production of ROS through both photodynamic mechanisms, generating singlet oxygen (type II) and superoxide radical anion (type I) under aerobic light irradiation. BP‐Br presented an adequate balance between ROS production and emission properties. The photokilling action and the binding to bacterial cells of these macrocycles were evaluated in vitro against methicillin‐resistant Staphylococcus aureus (MRSA) and Escherichia coli bacteria. Our results demonstrated that the halogenated BOPHY derivatives were effective PSs in inactivating MRSA using shorter irradiation periods. In addition, the antimicrobial action sensitized by these BOPHYs was potentiated by adding KI. The combination of halogenated BOPHY and KI led to a complete elimination of both Gram‐positive and Gram‐negative bacteria. Hence, BP‐Br and BP‐I prove to be potent broad‐spectrum antimicrobial PSs. To the best of our knowledge, this is the first time that BOPHY derivatives have been applied to photokill pathogenic microorganisms.

Cancer-Targeting and Viscosity-Activatable Near-Infrared Fluorescent Probe for Precise Cancer Cell Imaging.

Small-molecule fluorescent probes have emerged as potential tools for cancer cell imaging-based diagnostic and therapeutic applications, but their limited selectivity and poor imaging contrast hinder their broad applications. To address these problems, we present the design and construction of a novel near-infrared (NIR) biotin-conjugated and viscosity-activatable fluorescent probe, named as QL-VB, for selective recognition and imaging of cancer cells. The designed probe exhibited a NIR emission at 680 nm, with a substantial Stokes shift of 100 nm and remarkably sensitive responses toward viscosity changes in solution. Importantly, QL-VB provided an evidently enhanced signal-to-noise ratio (SNR: 6.2) for the discrimination of cancer cells/normal cells, as compared with the control probe without biotin conjugation (SNR: 1.8). Moreover, we validated the capability of QL-VB for dynamic monitoring of stimulated viscosity changes within cancer cells and employed QL-VB for distinguishing breast cancer tissues from normal tissues in live mice with improved accuracy (SNR: 2.5) in comparison with the control probe (SNR: 1.8). All these findings indicated that the cancer-targeting and viscosity-activatable NIR fluorescent probe not only enables the mechanistic investigations of mitochondrial viscosity alterations within cancer cells but also holds the potential as a robust tool for cancer cell imaging-based applications.

A sequential dual-locked luminescent copper nanocluster probe for tumor cell imaging and killing.

Asequential dual-locked luminescent copper nanoclusters (CuNCs) probe was designed and synthesizedfor the specific imaging and selective killing of tumor cells. This nanoprobe was prepared by first forming a Fe3+-coupled tannic acid (TA)-stabilized CuNCs (CuNCs-FeIII), which is in quenching state due to the electron transfer between CuNCs and Fe3+, and then coating a protectable layer of hyaluronic acid (HA) on the surface of CuNCs-FeIII to form the final dual-locked nanoprobe (CuNCs-FeIII@HA). When the nanoprobe of CuNCs-FeIII@HA target enter the tumor cells through CD44-HA receptor, HAase will first digest the HA layer of the nanoprobes, and then, GSH over-expressed in tumor cells will reduce Fe3+ to Fe2+, thus restoring the fluorescence emission of CuNCs and at the same time killing the tumor cells with the hydroxyl free radicals (∙OH) produced by the Fenton reaction between Fe2+ and H2O2. This sequential dual-locked luminescent nanoprobe of CuNCs-FeIII@HA has been successfully used for the specific imaging and selective killing of tumor cells.

“AIE+ESIPT” based naphthothiazole fluorescent probes for live cell imaging

Dual-functional organic fluorophores with aggregation induced emission (AIE) behavior and excited-state intramolecular proton transfer (ESIPT) behavior are of great significance to the design of imaging probes to bioimage systems. However, the pursuit of probes exhibiting dual AIE+ESIPT effects for live cell imaging remains an ongoing research challeng. Herein, we have successfully developed naphthothiazole fluorescent probes, HNBT-H and HNBT-G, based on the "AIE+ESIPT" effect. These probes were designed by incorporating responsive units for β-galactosidase (β-gal) and H2O2 into the pre-existing "AIE+ESIPT" HNBT fluorophore framework. The HNBT-H probe demonstrated the capability to distinguish various levels of H2O2. Furthermore, the applicability of HNBT-H and HNBT-G have been examined in living systems. These probes exhibit a good biocompatibility and low toxicity, the cell imaging experiments demonstrate the capability of HNBT-H and HNBT-G for fluorescence visualization in response to β-gal and H2O2 in RAW264.7 cells and PC12 cells.

A β-catenin chromobody-based probe highlights endothelial maturation during vascular morphogenesis in vivo.

Visualization of protein dynamics is a crucial step in understanding cellular processes. Chromobodies, fluorescently labelled single-domain antibodies, have emerged as versatile probes for live cell imaging of endogenous proteins. However, how these chromobodies behave in vivo and how accurately they monitor tissue changes remain poorly explored. Here, we generated an endothelial-specific β-catenin chromobody-derived probe and analyzed its expression pattern during cardiovascular development in zebrafish. Using high-resolution confocal imaging, we show that the chromobody signal correlates with the localization of β-catenin in the nucleus and at cell-cell junctions, and thereby can be used to assess endothelial maturation. Loss of Cadherin 5 strongly affects the localization of the chromobody at the cell membrane, confirming the cadherin-based adherens junction role of β-catenin. Furthermore, using a genetic model to block blood flow, we observed that cell junctions are compromised in most endothelial cells but not in the endocardium, highlighting the heterogeneous response of the endothelium to the lack of blood flow. Overall, our data further expand the use of chromobodies for in vivo applications and illustrate their potential to monitor tissue morphogenesis at high resolution.

Enabling Universal Access to Rapid and Stable Tetrazine Bioorthogonal Probes through Triazolyl-Tetrazine Formation.

Despite the immense potential of tetrazine bioorthogonal chemistry in biomedical research, the in vivo performance of tetrazine probes is challenged by the inverse correlation between the physiological stability and reactivity of tetrazines. Additionally, the synthesis of functionalized tetrazines is often complex and requires specialized reagents. To overcome these issues, we present a novel tetrazine scaffold-triazolyl-tetrazine-that can be readily synthesized from shelf-stable ethynyl-tetrazines and azides. Triazolyl-tetrazines exhibit improved physiological stability along with high reactivity. We showcase the effectiveness of this approach by creating cell-permeable probes for protein labeling and live cell imaging, as well as efficiently producing 18F-labeled molecular probes for positron emission tomography imaging. By utilizing the readily available pool of functionalized azides, we envisage that this modular approach will provide universal accessibility to tetrazine bioorthogonal tools, facilitating applications in biomedicine and materials science.

A Fluorescent Furan-based Probe with Protected Functional Groups for Highly Selective and Non-Toxic Imaging of HT-29 Cancer Cells and 4T1 Tumors.

Most of the previously reported fluorescent organic probes for cancer cell and tumor imaging have significant limitations including chemical toxicity, structural instability, low Stokes shift value, and the inability for selective accumulations in tumors during in vivo imaging. To overcome the mentioned challenges, we synthesized the fluorescent probes with protected polar functional groups to enhance the non-toxicity nature and increase the selectivity toward tumors. In addition, the structural rigidity of the fluorescent probes was increased by embedding aromatic rings in the probe structure. This issue enables us to obtain ultrabright cell images due to enhanced fluorescence quantum yield (ΦFL) values. After synthesis and spectral characterizations, the applicability of two furan-based and imidazole-based fluorescent probes ( abbreviated as DCPEF and DBPPI, respectively) was investigated for ultrabright in vitro and in vivo imaging of cancer cells. The probe DCPEF shows the ΦFL value of 0.946 and the Stocks shift of 86 nm. In addition, probe DBPPI offers the ΦFL value of 0.400 and a Stocks shift of 150 nm. The MTT colorimetric cytotoxicity assay showed that probe DCPEF has minimal effects against HT-29 (cancer) and Vero (normal) cells. The probe DCPEF produced ultrabright fluorescence images from HT-29 cells. In addition, in vivo imaging of cancer cells showed that probe DCPEF selectively accumulates in the 4T1 tumor in mice. The spectral and chemical stability, minimal cytotoxicity, significant Stokes shift, and high degree of selectivity for tumor cells during in vivo imaging make DCPEF an appropriate candidate to be used as a standard probe for cancer cell imaging.

Nitrogen and Chlorine Co-Doped Carbon Dots with Yellow-Green Emission as a Ratiometric Fluorescent and Colorimetric Dual-Signal Probe for Quercetin Detection and Cell Imaging

Despite the emergence of numerous carbon dots (CDs)-based sensors, most of them are fluorescent probes. To improve the reliability of the results, using ratiometric fluorescence/UV-vis absorption dual-signal probes is an effective method. Herein, nitrogen and chlorine co-doped carbon dots (N, Cl-CDs) with yellow-green fluorescence were prepared and used as a dual-function probe for ratiometric fluorescence/UV-vis. The detection range of the fluorescence method was 0.05–78[Formula: see text][Formula: see text]M, and the limit of detection (LOD) was 1.2[Formula: see text]nM (3[Formula: see text]/s). For the colorimetric method, the linear range was 0.05–186[Formula: see text][Formula: see text]M with LOD of 10.7[Formula: see text]nM. Importantly, the N, Cl-CDs were successfully used to detect Quercetin (QT) in actual water samples. Besides, due to their good biocompatibility, the N, Cl-CDs were applied to in vivo biological imaging of oocysts and shown the potential for environmental survey and biological assay.

Construction of a human serum albumin site II-targeting supramolecular fluorescent probe for cell imaging and drug delivery

Two cationic probes (named TAB2 and TAB6) specifically for binding to site II in human serum albumin (HSA) were designed and synthesized. Both probes exhibited selective fluorescence “turn-on” responsiveness to site II in HSA, though minor differences in their cationic terminals resulted in notable variations in their albumin responsiveness. TAB2 and TAB6 also exhibited strong binding affinity toward HSA. The presence of phenylbutazone, ibuprofen, and lidocaine had minimal impact on the albumin responsiveness of these probes. Furthermore, the complex formed by TAB2/6 and water-soluble pillar[5]arene (WP5) showed enhanced HSA site II-targeting fluorescence responsiveness. More importantly, the WP5-TAB2/6 complex self-assembled into vesicular supramolecular probes in aqueous solution (pH=7.4). Those supramolecular probes were promising for HSA-mediated cell imaging and act as a pH-sensitive drug-delivery platform to enhance the anticancer efficiency of doxorubicin (DOX).

BioAIEgens based on dehydroabietic acid and tetraphenylethene: How does isopropyl group of dehydroabietic acid affect their spectral performance

Based on dehydroabietic acid (DA) derived from natural rosin, two aggregation-induced emission luminogens (BioAIEgens) labeled as 12-DTPA-2pTPE and 13-DTPA-2pTPE were synthesized by introducing tetraphenylethylene (TPE) groups to dehydroabietic acid triarylamine (DTPA). Their structures were optimized, the spectral properties and AIE characteristics were investigated, and the compounds were prepared into nanoparticles (NPs) as fluorescent probes for cell imaging. The results show that the two BioAIEgens have typical AIE characteristics and large Stokes shifts (＞ 120 nm). Due to the electronic and spatial effects of the isopropyl group, the molecular configuration of 12-DTPA-2pTPE is more distorted, and the electron cloud distribution is also different from 13-DTPA-2TPE. The absorption and the emission peaks of 12-DTPA-2TPE are blue-shifted, and the fluorescence quantum yield in the solid is higher. In addition, the 13-DTPA-2TPE nanoparticles (NPs) have low cytotoxicity and good cell imaging effects in vitro.

New Highly Sensitive and Specific Raman Probe for Live Cell Imaging of Mitochondrial Function.

For Raman hyperspectral detection and imaging in live cells, it is very desirable to create novel probes with strong and unique Raman vibrations in the biological silent region (1800-2800 cm-1). The use of molecular probes in Raman imaging is a relatively new technique in subcellular research; however, it is developing very rapidly. Compared with the label-free method, it allows for a more sensitive and selective visualization of organelles within a single cell. Biological systems are incredibly complex and heterogeneous. Directly visualizing biological structures and activities at the cellular and subcellular levels remains by far one of the most intuitive and powerful ways to study biological problems. Each organelle plays a specific and essential role in cellular processes, but importantly for cells to survive, mitochondrial function must be reliable. Motivated by earlier attempts and successes of biorthogonal chemical imaging, we develop a tool supporting Raman imaging of cells to track biochemical changes associated with mitochondrial function at the cellular level in an in vitro model. In this work, we present a newly synthesized highly sensitive RAR-BR Raman probe for the selective imaging of mitochondria in live endothelial cells.

The Soft Nanodots as Fluorescent Probes for Cell Imaging: Analysis of Cell and Spheroid Penetration Behavior of Single Chain Polymer Dots.

This study describes the formation, size control, and penetration behavior of polymer nanodots (Pdots) consisting of single or few chain polythiophene-based conjugated polyelectrolytes (CPEs) via nanophase separation between good solvent and poor solvent of CPE. Though the chain singularity may be associated with dilution nanophase separation suggests that molecules of a good solvent create a thermodynamically driven solvation layer surrounding the CPEs and thereby separating the single chains even in their poor solvents. This statement is therefore corroborated with emission intensity/lifetime, particle size, and scattering intensity of polyelectrolyte in good and poor solvents. Regarding the augmented features, Pdots are implemented into cell imaging studies to understand the nuclear penetration and to differentiate the invasive characteristics of breast cancer cells. The python based red, green, blue (RGB) color analysis depicts that Pdots have more nuclear penetration ability in triple negative breast cancer cells due to the different nuclear morphology in shape and composition and Pdots have penetrated cell membrane as well as extracellular matrix in spheroid models. The current Pdot protocol and its utilization in cancer cell imaging are holding great promise for gene/drug delivery to target cancer cells by explicitly achieving the very first priority of nuclear intake.

Dicyanostilbene‐Derived Highly Sensitive Two‐Photon Fluorescence Temperature Probe for Live Cell Imaging

AbstractA novel two‐photon fluorescence temperature‐sensitive probe (SP) with the push‐pull electronic architecture (Donor‐π‐Acceptor, D‐π‐A) derived from dicyanostilbene was developed. Its linearly dependent coefficients (R2) between the emission intensity of SP and temperature reached 0.998 and 0.999 in one‐ and two‐photon fluorescence (OPF and TPF), respectively, and corresponding function expressions were IF=−0.0144T+1.2751 (OPF) and IF=−0.0152T+1.2970 (TPF). Its maximum two‐photon excitation wavelength (λmax2) is 750 nm, and the corresponding maximum two‐photon absorption cross‐section (δmax) is 1550 GM. DFT (Density Functional Theory) Calculation demonstrated that the electron cloud densities of two nitrogen atoms (N29 and N31) on two cyano groups are 15.9 and 16.0 times higher than that of nitrogen atom on 9‐carbazolyl group (N13), respectively. This fully indicates that the electron cloud of nitrogen atom on 9‐carbazolyl group is highly delocalized to the fluorophore matrix. SP can be used for live cell imaging and detect cell temperature to diagnose disease within physiological temperature range with excellent photostability and without cytotoxicity.

Nitrogen-doped carbon dots as a “turn-off” fluorescence probe for mercury(ii) detection and live cell imaging

In this paper, nitrogen-doped carbon dots (NCDs) were synthesized via a facile hydrothermal method using gum arabic (GA) and tris(hydroxymethyl)methyl aminomethane (tris) as precursors.

Near infra-red absorbing Quinolizidine fused curcuminoid-BF2 chelate and its applications in photodynamic therapy using MCF-7 and A549 cells

Metal-free near-infrared absorbing photosensitizers (PS) have been considered promising candidates for photodynamic therapy. Curcumin, curcuminoid, and its derivatives have therapeutic values due to their anti-inflammatory, antifungal, and antiproliferative properties. Curcuminoid-BF2 chelates have also been studied as cell imaging probes, however, their applications in photodynamic therapy are rare. In this article, we describe the synthesis and therapeutic evaluation of quinolizidine fused curcuminoid-BF2 chelate (Quinolizidine CUR-BF2) containing an acid-sensitive group. This donor-acceptor-donor curcuminoid-BF2 derivative exhibits absorption and emission in the deep red region with an absorption band maximum of ∼647 nm and a weak emission band at approximately 713 nm. It is interesting to note that this derivative has a high molar extinction coefficient (164,655 M−1cm−1). Quinolizidine CUR-BF2 possesses intramolecular charge transfer properties, facilitating the production of singlet oxygen (1O2), which plays a crucial role in cell death. Additionally, Quinolizidine CUR-BF2 can enable the selective release of active ingredients in an acidic medium (pH 5). Furthermore, the nanoaggregates of PS were prepared by encapsulating Quinolizidine CUR-BF2 within Pluronic F127 block co-polymer for better water-dispersibility and enhanced cellular uptake. Dark cytotoxicity of nanoaggregates was found to be negligible, whereas they exhibited significant photoinduced cytotoxicity towards cancer cells (MCF-7 and A549) under irradiation of 635 nm light. Further, the cell death pathway using Quinolizidine CUR-BF2 nanoaggregates as PS is found to occur through apoptosis. Specifically, the present study deals with the successful preparation of Quinolizidine CUR-BF2 nanoaggregates for enhanced water-dispersibility and cellular uptake as well as the efficacy evaluation of developed nanoaggregates for photodynamic therapy.

Functional chromophores synthesized via multicomponent Reactions: A review on their use as cell-imaging probes

This review aims to provide a comprehensive overview of recent advancements and applications of fluorescence imaging probes synthesized via MCRs (multicomponent reactions). These probes, also known as functional chromophores, belong to a currently investigated class of fluorophores that are presently being successfully applied in bioimaging experiments, especially in various living cell lineages. We describe some of the MCRs that have been employed in the synthesis of these probes and explore their applications in biological imaging, with an emphasis on cellular imaging. The review also discusses the challenges and future perspectives in the field, particularly considering the potential impact of MCR-based fluorescence imaging probes on advancing this field of research in the coming years. Considering that this area of research is relatively new and nearly a decade has passed since the first publication, this review also provides a historical perspective on this class of fluorophores, highlighting the pioneering works published between 2011 and 2016.

Design of a serum albumin sensitive probe for cell imaging and drug delivery by modifying a fluorescent agent

Protein-ligand interaction serves as a critical foundation for intramolecular charge transfer (ICT) probes to bind and recognize proteins. Many fluorescent agents are reported to have favorable protein binding abilities but lack protein sensitivity. Herein, a new serum albumin sensitive ICT probe (named MBD) was synthesized by modifying an existing fluorescent agent MPM. MBD not only inherited the favorable human serum albumin (HSA) binding ability of MPM but also newly obtained favorable serum albumin responsiveness. These properties were maintained after MBD formed a complex with water-soluble pillar[5]arene (WP5). Typical commercial drugs, fluorescent amino acids, fatty acids, and metal ions had negligible effects on the HSA responsiveness of the complex. The WP5-MBD complex possessed strong anti-interference ability in binding and recognizing HSA. Moreover, the WP5-MBD complex self-assembled into supramolecular fluorescent nanoparticles in solution. Such nanoparticles possessed good cell imaging and traceable drug delivery properties, which hold great potential for cancer treatment and diagnosis.