Dual-Channel fluorescent detection of Biothiols: A novel probe for Distinguishing Cysteine, Homocysteine, Glutathione, and N-Acetylcysteine in cellular environments.

Biothiols, including cysteine (Cys), homocysteine (Hcy), glutathione (GSH), and N-acetylcysteine (NAC), possess similar chemical structures and properties but play crucial, distinct roles in biological cells and blood serum. Imbalances in the concentrations of these biothiols are associated with various diseases, highlighting the importance of precise discrimination, especially between Cys and other biothiols. Owing to the similarity of the chemical properties of Cys, Hcy, GSH, and NAC, developing an effective methodology to differentiate these thiol compounds is challenging. In this study, we designed and synthesized a novel dual-channel fluorescent probe, hereafter referred to as CNTC, by integrating coumarin and acrylonitrile. This probe enabled the simultaneous discrimination of Cys from Hcy, GSH, and NAC, producing distinct fluorescent signals: blue for Cys and green for Hcy, GSH, and NAC. CNTC exhibited rapid response kinetics (within 30 min) and impressive detection limits of 0.31, 0.11, 0.029, and 0.032 µM for Cys, Hcy, GSH, and NAC, respectively. Furthermore, CNTC was successfully applied in the fluorescence imaging of both exogenous and endogenous Cys, Hcy, GSH, and NAC in living cells. The remarkable analytical and bioimaging capabilities of CNTCin vivo establish it as a promising tool for elucidating the pathophysiological roles of biothiols, particularly Cys, Hcy, GSH, and NAC.

Theoretical basis, state and challenges of living cell-based drug delivery systems.

The therapeutic efficacy of drugs is determined, to a certain extent, by the efficiency of drug delivery. The low efficiency of drug delivery systems (DDSs) is frequently associated with serious toxic side effects and can even prove fatal in certain cases. With the rapid development of technology, drug delivery has evolved from using traditional frameworks to using nano DDSs (NDDSs), endogenous biomaterials DDSs (EBDDSs), and living cell DDSs (LCDDSs). LCDDSs are receiving widespread attention from researchers at present owing to the unique advantages of living cells in targeted drug delivery, including their excellent biocompatibility properties, low immunogenicity, unique biological properties and functions, and role in the treatment of diseases. However, the theoretical basis and techniques involved in the application of LCDDSs have not been extensively summarized to date. Therefore, this review comprehensively summarizes the properties and applications of living cells, elaborates the various drug loading approaches and controlled drug release, and discusses the results of clinical trials. The review also discusses the current shortcomings and prospects for the future development of LCDDSs, which will serve as highly valuable insights for the development and clinical transformation of LCDDSs in the future.

A novel reaction-based fluorescent probe with a nanomolar sensitivity for detection of Hg(II) and its multiple application.

An easily synthesized reaction-based probe for the fluorescence detection of Hg2+ ions using was reported. The designed probe exhibited "turn-on" fluorescence response towards Hg2+ ions via deprotection of the thiocarbonate-protecting group to alcohol in the HEPES/EtOH=8:2 (v/v, 5 mM, pH 7.4). The investigation results of probe Bzp-1 properties for Hg2+ detection indicated that probe Bzp-1 has satisfactory high selectivity and sensitivity. The detection limit of Bzp-1 for Hg2+ was found to be 4.2 nM. The recognition mechanism of Bzp-1 for Hg2+ was confirmed by ESI-MS. Moreover, the probe Bzp-1 has been successfully used to rapidly detect trace amounts of hazardous Hg2+ ions in real samples such as tap water, seafood and soil with good recoveries and less the relative standard deviations. Moreover, the Bzp-1 can also be used for fluorescence imagining of Hg2+ in living cells.

Laser-induced stripping defect for highly selective electrochemical quantification of dopamine: Anti-interference from other catecholamine neurotransmitters.

Detecting dopamine (DA) is critical for early diagnosis of neurological and psychiatric disorders. However, the presence of other catecholamine neurotransmitters with structural similarities to DA causes significant interference in its detection. Herein, we introduce S stripping defects via laser-induced MoS2 to functionalize MoS2 electrodes and improve their selectivity for DA electrochemical detection. The sensing results show its excellent immunity to interference from other neurotransmitters, ensuring the preservation of the DA electrochemical signal even in the mixed neurotransmitters such as acetylcholine (ACh), γ-aminobutyric acid (GABA), epinephrine (EP), norepinephrine (NP), and serotonin (5-HT). DFT calculations further reveal that the negatively charged S-stripping defects enhance DA adsorption on the surface of the functionalized MoS2 electrode, contributing to its excellent performance. Moreover, this functionalized electrodes successfully monitor DA released from living PC12 cells in the presence of other interference, highlighting its potential applicability in intercellular signaling communication.

Fluorescent Nanodiamonds for High-Resolution Thermometry in Biology.

Optically active color centers in diamond and nanodiamonds can be utilized as quantum sensors for measuring various physical parameters, particularly magnetic and electric fields, as well as temperature. Due to their small size and possible surface functionalization, fluorescent nanodiamonds are extremely attractive systems for biological and medical applications since they can be used for intracellular experiments. This review focuses on fluorescent nanodiamonds for thermometry with high sensitivity and a nanoscale spatial resolution for the investigation of living systems. The current state of the art, possible further development, and potential limitations of fluorescent nanodiamonds as thermometers will be discussed here.

Multi-walled carbon nanotubes-metal-organic framework nanocomposite based sensor for the monitoring of multiple monoamine neurotransmitters in living cells.

The levels of monoamine neurotransmitters (MNTs) including dopamine (DA), adrenaline (Adr), norepinephrine (NE) and 5-hydroxytryptamine (5-HT) in cells are useful indicators to explore the pathogenesis of MNTs-related diseases such as Alzheimer's disease, Parkinson's disease and depression. Herein, we constructed a novel electrochemical sensing platform based on multi-walled carbon nanotubes (MWCNTs)-amine functionalized Zr (IV) metal-organic framework (UIO-66-NH2) nanocomposite for the detection of multiple MNTs including DA, Adr, NE and 5-HT. The synergistic effect between MWCNTs and UIO-66-NH2 endowed the nanocomposite with high specific surface area, low interface impedance and superior electrocatalytic activity, which effectively enhance the electrochemical performance of the sensor. The MWCNTs-UIO-66-NH2 nanocomposite-based sensor exhibited satisfied sensitivity for the quantitative measurement of DA, Adr, NE and 5-HT, as well as low detection limit. The outstanding biocompatibility of the constructed sensor permitted it to be successfully implemented for the real-time monitoring of DA released by PC12 and C6 cells, providing a promising strategy for clinical diagnosis of MNTs-related disorders and diseases.

Characterizing parameters and incorporating action potentials via the Hodgkin-Huxley model in a novel electric model for living cells.

To enhance our understanding of electroporation and optimize the pulses used within the frequency range of 1 kHz to 100 MHz, with the aim of minimizing side effects such as muscle contraction, we introduce a novel electrical model, structured as a 2D representation employing exclusively lumped elements. This model adeptly encapsulates the intricate dynamics of living cells' impedance variation. A distinguishing attribute of the proposed model lies in its capacity to decipher the distribution of transmembrane potential across various orientations within living cells. This aspect bears critical importance, particularly in contexts such as electroporation and cellular stimulation, where precise knowledge of potential gradients is pivotal. Furthermore, the augmentation of the proposed electrical model with the Hodgkin-Huxley (HH) model introduces an additional dimension. This integration augments the model's capabilities, specifically enabling the exploration of muscle cell stimulation and the generation of action potentials. This broader scope enhances the model's utility, facilitating comprehensive investigations into intricate cellular behaviors under the influence of external electric fields.

In our research, we've introduced an enhanced electrical model for living cells. This model simplifies cell behavior using only basic electrical components like resistors and capacitors. It's designed to mimic the real electrical properties of cells, particularly the cell membrane, which can change in response to electricity at different frequencies, ranging from 1 kHz to 100 MHz. This frequency range is essential for studying processes like electroporation, a technique used in various medical applications.Our model is represented in a two-dimensional structure, making it a handy tool for identifying transmembrane potential distributions, a critical factor in electroporation procedures. This means we can better understand how cells react to electrical impulses, which is crucial for improving electroporation techniques.Additionally, we've extended our model to include muscle cells by incorporating the Hodgkin-Huxley model, a well-established model for understanding electrical behavior in muscle cells. This allows us to study how muscles contract when exposed to different electrical pulses, a common side effect of electroporation procedures. By examining various pulse characteristics, we can determine which ones are best for minimizing muscle contractions during electroporation.In summary, our research has led to the development of a versatile electrical model for living cells. It not only helps us understand how cells respond to electricity in the context of electroporation but also provides insights into muscle contractions and how to optimize electrical pulses for medical treatments.

A novel one-stepped synthesized Schiff-base fluorescence probe for specific recognition of zinc ions with highly sensitive and its application in living cells.

Fluorescent turn-on receptors are extensively employed for the detection of Zn ions contamination in the environment due to its simplicity, convenience and portability. However, developing highly sensitive and cell-imageable fluorescent turn-on probe for the recognition of Zn ions in living organisms remains a significant challenge. Herein, we have successfully synthesized a novel Schiff base probe (H2L) with a significant fluorescence turn-on response (Zn ions) by one-step synthetic method. In this work, H2L exhibited high sensitivity to Zn2+ ions upon interaction with various common metal ions in HEPES buffer solution. Its detection limit is 1.87 × 10-7 M, which is lower than the requirement of Environmental Protection Agency (EPA) and World Health Organization (WHO) guidelines. The fluorescence titration and Job's plot analysis suggested a 1:1 binding ratio between the probe and Zn ion, and the single-crystal structures obtained further confirmed this inference. In addition, the fluorescent sensor demonstrated recyclability, maintaining its fluorescence intensity for up to 6 cycles without significant decrease, which holds promise for future investigations on reversible fluorescent chemosensors. Notably, fluorescence imaging experiments demonstrated that H2L could be successfully used for the detection of Zn2+ in live cells.

High-quality AFM image acquisition of living cells by modified residual encoder-decoder network.

Atomic force microscope enables ultra-precision imaging of living cells. However, atomic force microscope imaging is a complex and time-consuming process. The obtained images of living cells usually have low resolution and are easily influenced by noise leading to unsatisfactory imaging quality, obstructing the research and analysis based on cell images. Herein, an adaptive attention image reconstruction network based on residual encoder-decoder was proposed, through the combination of deep learning technology and atomic force microscope imaging supporting high-quality cell image acquisition. Compared with other learning-based methods, the proposed network showed higher peak signal-to-noise ratio, higher structural similarity and better image reconstruction performances. In addition, the cell images reconstructed by each method were used for cell recognition, and the cell images reconstructed by the proposed network had the highest cell recognition rate. The proposed network has brought insights into the atomic force microscope-based imaging of living cells and cell image reconstruction, which is of great significance in biological and medical research.

Development of a nanozyme-based electrochemical catalyst for real-time biomarker sensing of superoxide and nitric oxide anions released from living cells and exogenous donors.

Developing quantitative biosensors of superoxide (O2•-) and nitric oxide (NO) anion is crucial for pathological research. As of today, the main challenge for electrochemical detection is to develop high-selectivity nano-mimetic materials to replace natural enzymes. In this study, the dendritic-like morphological structure of silver organic framework (Ag-MOF) was successfully synthesized via a solvothermal strategy. Owing to the introduction of polymeric composites results in improved electrical conductivity and catalytic activity, which promotes mass transfer and leads to faster electron efficiency. For monitoring the electrochemical signals of O2•- and NO, the Ag-MOF electrode substrate was produced by drop-coating, and composites were designed by cyclic voltammetric potential cycles. The designed electrode substrates demonstrate high sensitivity, wide linear concentrations of 1 nM-1000 µM and 1 nM-850 µM, and low detection limits of 0.27 nM and 0.34 nM (S/N = 3) against O2•- and NO. Aside from that, the sensor successfully monitored the cellular release of O2•-, and NO from HepG2 and RAW 264.7 living cells and has the potential to monitor exogenous NO release from donors of Diethylamine (DEA)-NONOate and sodium nitroprusside (SNP). Additionally, the developed system was applied to the analysis of O2•- and NO in real biological fluid samples, and the results were good satisfactory (94.10-99.57 ± 1.23%). The designed system provides a novel approach to obtaining a good electrochemical biosensor platform that is highly selective, stable, and flexible. Finally, the proposed method provides a quantitative way to follow the dynamic changes in O2•- and NO in biological systems.

A thiazole-based colorimetric and photoluminescent chemosensors for As3+ ions detection: Density functional theory, test strips, real samples, and bioimaging applications.

A Schiff-base Ethyl (E)-2-(3-((2-carbamothioylhydrazono)methyl)-4-hydroxyphenyl)-4-methylthiazole-5-carboxylate (TZTS) dual functional colorimetric and photoluminescent chemosensor which includes thiazole and thiosemicarbazide has been synthesized to detect arsenic (As3+) ions selectively in DMSO: H2O (7:3, v/v) solvent system. The molecular structure of the probe was characterized via FT-IR, 1H, and 13C NMR & HRMS analysis. Interestingly, the probe exhibits a remarkable and specific colorimetric and photoluminescence response to As3+ ions when exposed to various metal cations. The absorption spectral changes of TZTS were observed upon the addition of As3+ ions, with a naked eye detectable color change from colorless to yellow color. Additionally, the chemosensor (TZTS) exhibited a new absorption band at 412 nm and emission enhancements in photoluminescence at 528 nm after adding As3+ ions. The limit of detection (LOD) for As3+ ions was calculated to be 16.5 and 7.19 × 10-9 M by the UV-visible and photoluminescent titration methods, respectively. The underlying mechanism and experimental observations have been comprehensively elucidated through techniques such as Job's plot, Benesi-Hildebrand studies, and density functional theory (DFT) calculations. For practical application, the efficient determination of As3+ ions were accomplished using a spike and recovery approach applied to real water samples. In addition, the developed probe was successfully employed in test strip applications, allowing for the naked-eye detection of arsenic ions. Moreover, fluorescence imaging experiments of As3+ ions in the breast cancer cell line (MCF-7) demonstrated their practical applications in biological systems. Consequently, these findings highlight the significant potential of the TZTS sensor for detecting As3+ ions in environmental analysis systems.

The Emergence of Tumor-Initiating Cells in an Advanced Hypopharyngeal Tumor Model Exhibits Enhanced Angiogenesis and Nuclear Factor Erythroid 2-Related Factor 2-Associated Antioxidant Effects.

Aims: Hypopharyngeal cancer (HPC) is associated with the worst prognosis of all head and neck cancers and is typically identified in an advanced stage at the time of diagnosis. While oxidative stress might contribute to the onset of HPC in patients using tobacco or alcohol, the extent of this influence and the characteristics of HPC cells in advanced stage remain to be investigated. In this study, we explored whether HPC cells survived from necrotic xenograft tumors at late stage would display increased tumor resistance along with altered tolerance to oxidative stress. Results: The remnant living HPC cells isolated from a late-stage xenograft tumor, named FaDu ex vivo cells, showed stronger chemo- and radioresistance, tumorigenesis, and invasiveness compared with parental FaDu cells. FaDu ex vivo cells also displayed increased angiogenic ability after re-transplantation in mice visualized by in vivo near infrared-II fluorescence imaging modality. Moreover, FaDu ex vivo cells exhibited significant tumor-initiating cell (TIC)-related properties accompanied by a reduction of the level of reactive oxygen species, which was associated with the upregulation of transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2). Interestingly, inhibition of Nrf2 by the RNA interference and the chemical inhibitor could reduce the TIC-related properties of FaDu ex vivo cells. Innovation: Oxidative stress potentially initiates HPC, but elevation of Nrf2-associated antioxidant mechanisms would be essential to mitigate this effect for promoting and sustaining the stemness of HPC at the advanced stage. Conclusion: Present data suggest that the antioxidant potency of advanced HPC would be a therapeutic target for the design of adjuvant treatment. Antioxid. Redox Signal. 41, 505-521.

One-Pot Synthesis of Nanoflower-Like Zn2SnS4 as Nanozymes for Highly Sensitive Electrochemical Detection of H2O2 Released by Living Cells.

The sensitive and reliable nanozyme-based sensor enables the detection of low concentrations of H2O2 in biological microenvironments, it has potential applications as an in-situ monitoring platform for cellular H2O2 release. The uniformly dispersed bimetallic sulfide (Zn2SnS4) nanoflowers were synthesized via a one-pot hydrothermal method and the two kinds of metal ions can serve as morphology and structure directing agents for each other in the synthetic process. The nanoparticles were utilized as nanozyme materials to fabricate a novel electrochemical sensor, and it exhibits a distinct electrochemical response towards H2O2 with excellent stability and detection capability (with a minimum detection limit of 1.79 nM (S/N=3)), the excellent characteristics facilitate the precise detection of low concentrations of H2O2 in biological microenvironments. Use the macrophages differentiated from leukemia THP-1 cells as a representative sensing model, the sensor was successfully utilized for real-time monitoring of the release of H2O2 induced by living cells, which has significant potential applications in clinical diagnosis and cancer treatment.

New approaches for low phototoxicity imaging of living cells and tissues.

Fluorescence microscopy is a powerful tool used in scientific and medical research, but it is inextricably linked to phototoxicity. Neglecting phototoxicity can lead to erroneous or inconclusive results. Recently, several reports have addressed this issue, but it is still underestimated by many researchers, even though it can lead to cell death. Phototoxicity can be reduced by appropriate microscopic techniques and carefully designed experiments. This review focuses on recent strategies to reduce phototoxicity in microscopic imaging of living cells and tissues. We describe digital image processing and new hardware solutions. We point out new modifications of microscopy methods and hope that this review will interest microscopy hardware engineers. Our aim is to underscore the challenges and potential solutions integral to the design of microscopy systems. Simultaneously, we intend to engage biologists, offering insight into the latest technological advancements in imaging that can enhance their understanding and practice.

Highly chromophoric fluorescent-labeled methionyl-initiator tRNAs applicable in living cells.

Fluorescent initiator tRNAs (tRNAi) play a crucial role in studying protein synthesis, yet generating highly fluorescent tRNAi complexes remains challenging. We present an optimized strategy to effectively generate highly fluorescent initiator-tRNA complexes in living cells. Our strategy allows the generation of Fluo-Met-tRNAiMet complexes. These complexes can have highly chromogenic N-terminal labeling. For generating such complexes, we use either purified fluorescent methionine (PFM) or non-purified fluorescently labeled methionine (NPFM). Furthermore, PFM promotes the active generation of endogenous tRNAi in cells, leading to highly efficient Fluo-Met-tRNAiMet complexes. Finally, PFM-tRNAiMet complexes also facilitate the visualization of native fluorescently labeled Tat binding to beads. This demonstrates the potential of our approach to advance precision protein engineering and biotechnology applications.

A dual-core 3D DNA nanomachine based on DNAzyme positive feedback loop for highly sensitive MicroRNA imaging in living cells.

A double 3D DNA walker nanomachine by DNAzyme self-driven positive feedback loop amplification for the detection of miRNA was constructed. This method uses two gold nanoparticles as the reaction core, and because of the spatial confinement effect the local concentration of the reactants increase the collision efficiency was greatly improved. Meanwhile, the introduction of positive feedback loop promotes the conversion efficiency. In presence of miRNA-21, a large amount of DNAzyme was released and hydrolyze the reporter probe, resulting the recovery of fluorescence signal. The linear range for miRNA-21 is 0.5-60 pmol/L, and the detection limit is 0.41 pmol/L (S/N = 3). This nanomachine has been successfully used for accurate detection of miRNA-21 expression levels in cell lysates. At the same time, it can enter cells for intracellular miRNA-21 fluorescence imaging, distinguishing tumor cells from normal cells. This combination of in vitro detection and imaging analysis of living cells can achieve the goal of jointly detecting cancer markers through multiple pathways, providing new ideas for early diagnosis and screening of diseases.

A novel ratiometric peptide-based fluorescent probe for sequential detection of Hg2+ and S2- ions and its application in living cells and zebrafish imaging.

A new ratiometric peptide-based fluorescent probe DWPH was designed and synthesized, comprising dansyl fluorophore as a fluorescent dye, and tripeptide backbone (Trp-Pro-His-NH2) as a recognition group. The addition of Hg2+ caused the maximum emission peak of DWPH to blue shift from 560 nm to 510 nm. DWPH exhibited large Stokes shift (230 nm), satisfactory water solubility (100 % aqueous medium), good selectivity (only Hg2+), high sensitivity (24.6 nM), rapid response (within 50 s) and strong anti-interference ability for Hg2+ detection over a wide pH range (7-11). Additionally, the complex DWPH-Hg2+ as a relay response probe could also be applied to S2- according to displacement approach. Notably, the detection limit for S2- was calculated as 23.3 nM, exhibiting that DWPH showed great potential for environmental monitoring and bioimaging. In addition, DWPH were successfully used to determine Hg2+ and S2- in living cells and zebrafish based on excellent permeability and low cytotoxicity. What's more, the gradient concentration color changes of Hg2+ and S2- were combined with the smartphone APP to obtain red-green-blue (RGB) values, thus enabling rapid semi-quantitative detection of Hg2+ and S2- without expensive instruments.

Adhesion measurement of living cells based on electrical impedance.

BACKGROUND: Cells adherence provides specific information about physiology and pathology, the adhesion measurement between living cells and nanostructures can be measured by atomic force microscopy, but this detection technique is difficult to operate and costly. The adhesion height and effective contact area between cells and substrates are also the key factors affecting measurement value of the overall impedance. These factors change with structural parameters of the substrates, so the adhesion measurement between living cells and substrate can be indirectly reflected by the impedance value. OBJECTIVE: To establish a mapping relationship between the impedance measurement and the adhesion measurement of living cells. The possibility of dynamic measurement of adhesion is realized by this method, and the experimental process is simplified. METHODS: Laser interference technology was used to prepare nanoarray structures with different periods on the surface of silicon wafers for cells culture. Under the same experimental conditions, the impedance of living cells on the substrates of different cycle sizes were measured. The adhesion between cells and different substrates were analyzed by measuring impedance after the interaction between cells and substrate. RESULTS: The adhesion of living cells on the substrates of different sizes be analyzed, and the mapping relationship between the impedance and the adhesion measurement was established. The results showed that, the larger the impedance value between cells and substrate, the larger the effective contact area and the smaller the gap between them. CONCLUSION: The difference of adhesion height and effective adhesion area between living cells and substrates were obtained. This paper, a new method to measure the adhesion properties of living cells is presented, which provides theoretical basis for the related research.

Synthesis of a super-low detection limit fluorescent probe for Al3+ and its application in fluorescence imaging of zebrafish and cells.

A novel fluorescent probe N'-(2-hydroxybenzylidene)-indole-3-formylhydrazine (JHK) was designed and synthesized based on the condensation reaction of indole-3-formylhydrazine and salicylaldehyde. The probe JHK solution could highly selectively recognize Al3+ by the obvious fluorescence enhancement (288-fold) after adding Al3+. And the probe solution with Al3+ had a very high fluorescence quantum yield (89.29 %). The detection limit was calculated to be 1.135 nM, which was significantly lower than many reported detection limits, indicating that the probe JHK had pretty good sensitivity. The ratio of JHK to Al3+ (1:1) and the sensing mechanism were determined by Job's plot, 1H NMR spectra, FTIR spectra, ESI-MS and Gaussian calculation. The probe solution and medium-speed filter paper were successfully used to make test papers for more convenient detection of Al3+. Furthermore, the probe JHK had been successfully applied to the detection of Al3+ in real water, zebrafish and living cells.

The catalyst free synthesis of dibenzo[a,j]acridine and its applications in bioimaging of BF3 in HeLa cells.

In this paper, we discuss how tetrahydrodibenzo[a,j]acridine (4-HA) loses its hydrogen, which makes dibenzo[a,j]acridine (ARM) and also how 4-HA can be synthesized effectively using 2-tetralone in high yield. Dehydrogenative condensation and dehydrogenation are the two processes that make up the overall reaction of this synthetic approach. In addition, the presence of BF3 caused a remarkable fluorescence shift in ARM. Test paper analysis was used for examining the practical usefulness of ARM, which can be seen under UV light, resulting in this unique phenomenon. The fluorescent bio imaging experiment demonstrates that the sensor ARM has the capability to detect BF3 in living HeLa cells.