cell imaging

细胞成像
  • 文章类型: Journal Article
    本文报道了一种基于MB衍生物的用于Cu2离子检测的荧光探针(MB-M)。通过1HNMR对探针进行了良好的表征,13CNMR和HR-MS谱。探针MB-M显示出从无色到靛蓝的颜色变化对Cu2+的肉眼识别。该探针表现出很有前途的特征,如高荧光和UV-vis选择性,快速反应(5薄荷),可在pH2-7和低检测限(LOD=0.33µM)下使用。探针MB-M还用于HepG-2细胞中的Cu2离子成像和日常生活中的检测(测试条和湖水)。此外,采用非共价相互作用(NCI)和分子中原子的量子理论(QTAIM)分析研究了MB-M与Cu2离子之间的相互作用。通过使用自然键轨道(NBO)检查络合物的电子特性,电子密度差(EDD),和前沿分子轨道(FMO)分析,研究了MB-M对Cu2+离子的敏感性。结果表明,MB-M和Cu2离子之间的相互作用涉及化学吸附。
    Herein we have reported a fluorescent probe (MB-M) based on MB derivative for Cu2+ ions detection. The probe was well characterized by 1H NMR, 13C NMR and HR-MS spectrum. Probe MB-M showed naked-eyes recognition to Cu2+ as color change from colorless to indigo. The probe exhibited promising features such as high fluorescence and UV-vis selectivity, fast response (5 mint), workable at pH 2-7, and low limit of detection (LOD = 0.33 µM). Probe MB-M was also used for Cu2+ ions imaging in HepG-2 cells and detection in daily life (Test Strip and lake water). Moreover, non-covalent interaction (NCI) and quantum theory of atoms in molecules (QTAIM) analysis were used to study the interaction between MB-M and Cu2+ ions. By examining the electronic characteristics of the complex using natural bond orbital (NBO), electron density difference (EDD), and frontier molecular orbital (FMO) analysis, the sensitivity of MB-M towards Cu2+ ions were investigated. The results illustrated that the interactions between MB-M and Cu2+ ions involved chemisorption.
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  • 文章类型: Journal Article
    EPO-聚吲哚-硝基-碳硼烷(E-P-INDOLCAB)的连续制备方案,L100-55-聚吲哚-硝基-碳硼烷(L-P-INDOLCAB),RS-聚吲哚-nido-碳硼烷(S-P-INDOLCAB),用RL-聚吲哚-硝基-碳硼烷(R-P-INDOLCAB)制备了四种丙烯酸树脂包覆的硝基-碳硼烷吲哚荧光聚合物。在测试了它们在各种酸性pH值(分别为3.4和5.5)下的光谱特性和荧光稳定性曲线趋势之后,L-P-INDOLCAB和S-P-INDOLCAB被确定为最佳聚合物。在原子力显微镜(AFM)测试过程中,两种聚合物的稳定状态和纳米颗粒在系统表面的分散状态由-23和-42mV的zeta电位显示。使用zeta电位和原子力显微镜(AFM)检查了纳米颗粒在系统表面的分散和两种聚合物的稳定状态。透射电子显微镜(TEM)也可以证实这些发现,表明丙烯酸树脂牢固地包裹内部以形成眼球。两种聚合物与HELA细胞的生物相容性在细胞成像中得到增强,紧密包围靶细胞。当浓度为20ug/mL时,两种复合物对PC-3和HeLa细胞均表现出较强的抑制作用,如随后的细胞增殖毒性研究所验证。
    The continuous preparation scheme EPO-Poly-indol-nido-carborane (E-P-INDOLCAB), L100-55-Poly-indol-nido-carborane (L-P-INDOLCAB), RS-Poly-indol-nido-carborane (S-P-INDOLCAB), and RL-Poly-indol-nido-carborane (R-P-INDOLCAB) were used to prepare the four types of acrylic resin-coated nido-carborane indole fluorescent polymers. After testing their spectral properties and the fluorescence stability curve trend at various acidic pH values (3.4 and 5.5, respectively), L-P-INDOLCAB and S-P-INDOLCAB were determined to be the best polymers. The stable states of the two polymers and the dispersion of the nanoparticles on the system\'s surface during Atomic Force Microscope (AFM) test are shown by the zeta potentials of -23 and -42 mV. The dispersion of nanoparticles on the system\'s surface and the stable condition of the two polymers were examined using zeta potential and atomic force microscopy (AFM). Transmission electron microscopy (TEM) can also confirm these findings, showing that the acrylic resin securely encases the interior to form an eyeball. Both polymers\' biocompatibility with HELA cells was enhanced in cell imaging, closely enclosing the target cells. The two complexes displayed strong inhibitory effects on PC-3 and HeLa cells when the concentration was 20 ug/mL, as validated by subsequent cell proliferation toxicity studies.
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  • 文章类型: Journal Article
    热加工食品中的丙烯酰胺(AA)已成为全球健康问题,主要致癌,神经毒性,和生殖毒性,越来越多的研究人员深入研究了其毒理学机制。研究已经证明,在一系列浓度下通过AA暴露HepG2可以诱导miR-21和miR-221的上调。监测细胞内miRNAs的反应可以在揭示AA毒性机制中发挥重要作用。这里,通过结合AA适体的识别功能和DNAzymewalker的信号放大,由三种AIE染料构建了多色聚集诱导的发射纳米颗粒(AIENP)探针,用于细胞内AA和AA诱导的miR-21/miR-221的同时成像。这些纳米粒子的表面含有羧基,通过酰胺反应促进它们与用荧光猝灭基团修饰的底物链的连接。进行了优化实验,以确定最佳的底物与DNA酶的比例,确认其作为信号放大助行器的功效。AA的灵敏检测,miR-21和miR-221在细胞外培养基中获得,AA的检测限为0.112nM,miR-21和miR-221分别为0.007pM和0.003pM,表现出优异的选择性。细胞内,ZIF-8结构倒塌,释放Zn2+,激活DNA酶裂解活性,随着刺激时间(0-12h)和AA浓度(0-500μM)的增加,HepG2细胞内多色AIENP的荧光逐渐恢复。这种动态反应揭示了AA暴露与miR-21/miR-221表达之间的关系,进一步验证AA的致癌性。
    Acrylamide (AA) in heat-processed foods has emerged as a global health problem, mainly carcinogenic, neurotoxic, and reproductive toxicity, and an increasing number of researchers have delved into elucidating its toxicological mechanisms. Studies have demonstrated that exposure of HepG2 by AA in a range of concentrations can induce the upregulation of miR-21 and miR-221. Monitoring the response of intracellular miRNAs can play an important role in unraveling the mechanisms of AA toxicity. Here, multicolor aggregation induced emission nano particle (AIENP) probes were constructed from three AIE dyes for simultaneous imaging of intracellular AA and AA-induced miR-21/miR-221 by combining the recognition function of AA aptamers and the signal amplification of a DNAzyme walker. The surface of these nanoparticles contains carboxyl groups, facilitating their linkage to a substrate chain modified with a fluorescent quencher group via an amide reaction. Optimization experiments were conducted to determine the optimal substrate-to-DNAzyme ratio, confirming its efficacy as a walker for signal amplification. Sensitive detection of AA, miR-21 and miR-221 was achieved in extracellular medium, with detection limits of 0.112 nM for AA, 0.007 pM and 0.003 pM for miR-21 and miR-221, respectively, demonstrating excellent selectivity. Intracellularly, ZIF-8 structure collapsed, releasing Zn2+, activating DNAzyme cleavage activity, and the fluorescence of multicolor AIENPs within HepG2 cells gradually recovered with increasing stimulation time (0-12 h) and concentrations of AA (0-500 μM). This dynamic response unveiled the relationship between AA exposure and miR-21/miR-221 expression, further validating the carcinogenicity of AA.
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  • 文章类型: Journal Article
    基于共轭聚合物(CPs)的近红外光学由于其高摩尔消光系数而受到越来越多的关注,宽发射波长,易于制备和优异的生物相容性。在这里,使用三苯胺(D2)和噻吩(D1)作为电子给体,苯并噻二唑(A)作为电子受体,通过一锅法偶联很容易制备出几种具有D2-D1-A结构的新型共轭聚合物。有趣,它们的光学性能和功率转换效率可以通过噻吩(D1)上的侧链来调节。将乙二氧基作为侧链引入D1显著提高了荧光成像亮度,光热转换效率和亲水性,并扩展发射波长,这对光疗是有益的。侧链修饰提供了新的机会来设计有效的光疗学,而无需构建新的荧光骨架。
    Conjugated polymers (CPs)-based near-infrared phototheranostics are receiving increasing attention due to their high molar extinction coefficient, wide emission wavelength, easy preparation and excellent biocompatibility. Herein, several new conjugated polymers with D2-D1-A structures were easily prepared through one-pot coupling using triphenylamine (D2) as well as thiophenes (D1) as electron donors and benzothiadiazole (A) as electron acceptors. Interesting, their optical performance and power conversion efficiency could be tuned by side chains on thiophenes (D1). The introduction of ethylenedioxy into D1 as side chain significantly improves fluorescence imaging brightness, photothermal conversion efficiency and hydrophilicity, and extends emission wavelength, which are beneficial for phototheranostic. The side chain modification provides new opportunity to design efficient phototheranostics without construction new fluorescent skeletons.
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  • 文章类型: Journal Article
    具有光响应性的纳米材料由于其荧光成像而引起了人们的关注,光动力,和光热治疗特性。在这项研究中,通过使用光敏剂和碳点(CD)开发了光响应性纳米组装体。由于它们的多个激发峰和多色荧光发射,特别是它们的膜渗透特性,这些纳米组件可以用多种颜色标记细胞,并实时跟踪细胞成像。此外,光敏剂和CD的掺入为纳米组装体提供了用于光动力疗法(PDT)和光热疗法(PTT)的潜力。纳米组装体通过PDT和PTT有效抑制大肠杆菌和金黄色葡萄球菌的活性。此外,纳米组装体对大肠杆菌和金黄色葡萄球菌表现出高亲和力。这些不同的特征赋予纳米组装体广谱抗菌性质。作为光响应性纳米平台,这些纳米组装体在生物成像和抗菌领域具有潜在的应用前景。
    Nanomaterials with photoresponsivity have garnered attention due to their fluorescence imaging, photodynamic, and photothermal therapeutic properties. In this study, a photoresponsivity nanoassembly was developed by using photosensitizers and carbon dots (CDs). Due to their multiple excitation peaks and multicolor fluorescence emission, especially their membrane-permeating properties, these nanoassemblies can label cells with multiple colors and track cell imaging in real time. Additionally, the incorporation of photosensitizers and CDs provides the nanoassemblies with the potential for photodynamic therapy (PDT) and photothermal therapy (PTT). The nanoassemblies effectively suppressed the activity of Escherichia coli and Staphylococcus aureus through PDT and PTT. Moreover, the nanoassemblies exhibited a high affinity for E. coli and S. aureus. These distinct features confer broad-spectrum antibacterial properties to the nanoassemblies. As a photoresponsivity nanoplatform, these nanoassemblies have demonstrated potential applications in the fields of bioimaging and antimicrobial.
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  • 文章类型: Journal Article
    癌细胞溶酶体中较高的粘度和较低的pH值突出了它们作为癌症生物标志物的潜力。因此,酸激活粘度荧光探针的研制对癌症的早期诊断和治疗具有重要意义。基于此,设计并合成了一种基于2-(2-羟基苯基)苯并噻唑(HBT)基团的近红外荧光探针,即HBTH,监测溶酶体内的粘度变化。已经证明HBTH对粘度非常敏感,在(logη)=0-3.06(相关系数为0.98)的范围内,荧光强度与log(粘度)之间具有很强的线性关系,证明其定量粘度测量的能力。特别是,只有低pH和高粘度的联合作用才能有效地触发HBTH最明显的荧光增强。此外,HBTH可以通过低浓度(100nM)的无洗程序快速定位到溶酶体,并在20s内实现高保真成像。它还可以监测细胞中溶酶体的动态过程,药物刺激下的粘度变化,和线粒体自噬过程中的溶酶体行为。重要的是,HBTH能够通过体内成像识别荷瘤裸鼠中的肿瘤。这些功能使HBTH成为癌症早期诊断和治疗的强大工具。
    The higher viscosity and lower pH in lysosomes of cancer cells highlight their potential as biomarkers for cancer. Therefore, the development of acid-activated viscosity fluorescent probes is significant for the early diagnosis and treatment of cancer. Based on this, we have designed and synthesized a near-infrared fluorescent probe based on the 2-(2-hydroxyphenyl)benzothiazole (HBT) group, namely HBTH, to monitor the viscosity changes within lysosomes. It has been demonstrated that HBTH was extremely sensitive to viscosity, with a strong linear relationship between fluorescence intensity and log(viscosity) within the range of (logη) = 0-3.06 (a correlation coefficient of 0.98), proving its capability for quantitative viscosity measurement. In particular, the most obvious fluorescence enhancement of HBTH was only efficiently triggered by the combined effect of low pH and high viscosity. Furthermore, HBTH can rapidly localize to lysosomes by wash-free procedure at a low concentration (100 nM) and achieve high-fidelity imaging within 20 s. It can also monitor the dynamic processes of lysosomes in cells, viscosity changes under drug stimuli, and lysosomal behavior during mitophagy. Importantly, HBTH is capable of identifying tumors in tumor-bearing nude mice through in vivo imaging. These features make HBTH a powerful tool for the early diagnosis and treatment of cancer.
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  • 文章类型: Journal Article
    在生物成像领域,半菁染料具有优异的发射波长特性和生物相容性,是构建近红外荧光探针的理想结构。开发能够检测半胱氨酸(Cys)的近红外荧光探针是本研究的目的。在各种分析物存在下,新型开发的荧光探针P对Cys显示出高选择性和灵敏度。发现P的检测极限为0.329μM。MTT分析显示该探针基本上是非细胞毒性的。此外,该探针已成功用于活细胞和小鼠的半胱氨酸成像。
    Hemicyanine dyes are an ideal structure for building near-infrared fluorescent probes due to their excellent emission wavelength properties and biocompatibility in biological imaging field. Developing a near-infrared fluorescent probe capable of detecting cysteine (Cys) was the aim of this study. A novel developed fluorescent probe P showed high selectivity and sensitivity to Cys in the presence of various analytes. The detection limit of P was found to be 0.329 μM. The MTT assay showed that the probe was essentially non-cytotoxic. Furthermore, the probe was successfully used as cysteine imaging in living cells and mice.
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  • 文章类型: Journal Article
    SO2衍生物,亚硫酸盐/亚硫酸氢盐,广泛应用于食品加工和药物合成行业。尽管它们广泛应用,过量的亚硫酸盐/亚硫酸氢盐会对人体健康产生负面影响。大多数用于检测亚硫酸盐/亚硫酸氢盐的探针由于其在可见光谱范围内的荧光和在水溶液中的溶解性差而受到限制,这限制了它们在食品测试和生物成像中的使用。在这里,由氰基吡啶菁骨架组成的近红外探针,4-((Z)-2-((E)-2-氯-3-(2-氰基-2-(1-甲基吡啶-4(1H)-亚基)亚乙基)环己-1-烯-1-基)-1-氰基乙烯基)-1-甲基吡啶-1-铵(简写为CCP),已开发。该探针能够精确定量几乎纯缓冲溶液中的亚硫酸氢盐(HSO3-),在784nm处显示近红外荧光发射,检测限低至0.32μM。该探针因其出色的选择性而脱颖而出,对干扰的敏感性最小,适应性强。探针CCP利用CC键通过亲核加成引发与HSO3-的近红外荧光猝灭反应,这有效地破坏了分子内的大量离域,以进行准确的HSO3-鉴定。此外,该探针已成功应用于检测各种食品和活细胞中的HSO3-,简化了水样中HSO3-含量的测量。这种进步不仅增强了分析能力,而且有助于确保食品安全和环境保护。环境含义:SO2衍生物,包括亚硫酸盐/亚硫酸氢盐,兼有防腐剂和抗氧化剂的双重作用,广泛应用于各个领域,包括食品防腐,水卫生,和制药业。尽管它们广泛应用,过量的亚硫酸盐/亚硫酸氢盐会影响人体健康。开发精确和灵敏地检测食品和生物样品中亚硫酸盐/亚硫酸氢盐的方法对于确保食品安全和环境保护非常重要。这里,在99.9%缓冲溶液中灵敏的近红外和多功能荧光探针,随着水凝胶封装,已成功应用于食品中亚硫酸氢盐的检测,正宗水样,和生物细胞。
    SO2 derivatives, sulfite/bisulfite, are widely employed in both the food processing and drug synthesis industries. Despite their widespread application, excessive levels of sulfite/bisulfite can negatively impact human health. Most probes for detecting sulfite/bisulfite are restricted by their fluorescence within the visible spectrum range and poor solubility in aqueous solution, which limit their use in food testing and biological imaging. Herein, a near-infrared probe comprising of the cyanopyridine cyanine skeleton, 4-((Z)-2-((E)-2-chloro-3-(2-cyano-2-(1-methylpyridine-4(1H)-ylidene)ethylidene)cyclohex-1-en-1-yl)-1-cyanovinyl)-1-methylpyridin-1-ium (abbreviated as CCP), was developed. This probe enables precise quantification of bisulfite (HSO3-) in almost pure buffered solutions, showing a near-infrared fluorescence emission at 784 nm with an impressively low detection limit of 0.32 μM. The probe stands out for its exceptional selectivity, minimal susceptibility to interference, and strong adaptability. The probe CCP utilizes the CC bond to trigger a near-infrared fluorescence quenching reaction with HSO3- via nucleophilic addition, which effectively disrupts the large delocalization within the molecule for accurate HSO3- identification. Moreover, the probe has been successfully applied in detecting HSO3- in various food products and living cells, simplifying the measurement of HSO3- content in water samples. This advancement not only enhances the analytical capabilities but also contributes to ensuring food safety and environmental protection. ENVIRONMENTAL IMPLICATION: SO2 derivatives including sulfite/bisulfite, serving dual roles as preservatives and antioxidants, have widespread application across various sectors including food preservation, water sanitation, and the pharmaceutical industry. Despite their widespread application, excessive levels of sulfite/bisulfite can affect human health. Developing methods for precisely and sensitively detecting sulfite/bisulfite in food products and biological samples is important for ensuring food safety and environmental protection. Here, a sensitive near-infrared and multifunctional fluorescent probe in a 99.9 % buffered solution, along with water gel encapsulation, has been successfully applied for the detection of bisulfite in food, authentic water samples, and biological cells.
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  • 文章类型: Journal Article
    线粒体,对真核细胞功能至关重要的中央细胞器,将他们的影响力扩展到ATP生产之外,包括在细胞凋亡中的作用,钙信号,和生物合成。最近的研究聚焦了线粒体功能的两个新兴决定因素:线粒体内粘度和二氧化硫(SO2)水平。而最佳的线粒体粘度控制分子扩散和重要的过程,如氧化磷酸化,像差与神经退行性疾病有关,糖尿病,和癌症。同样,SO2是一种气体信号分子,调节能量途径和氧化应激反应;然而,失衡导致细胞毒性亚硫酸盐和亚硫酸氢盐积累,引发癌症和心血管异常等疾病。我们的研究重点是开发双通道荧光探针,在香豆素染料基质中应用吸电子受体,有助于监测活细胞中的线粒体粘度和SO2。该探针在650nm和558nm处区分荧光峰,允许按比例定量SO2,而不受其他硫物种的干扰。此外,它可以实现近红外粘度测定,特别是在线粒体内。研究采用了利用密度泛函理论(DFT)方法的理论计算来确定分子的几何形状并计算旋转能。值得注意的是,探针的吲哚部分表现出最低的旋转能量,定量为7.38kcals/mol。当包含在接受制霉菌素等药物的HeLa细胞内时,该探针具有增强的线粒体粘度动力学,莫能菌素,和细菌脂多糖(LPS)。总的来说,我们的创新方法阐明了复杂的线粒体因子,呈现对细胞能量学的变革性见解,氧化还原稳态,和线粒体相关疾病的治疗途径。
    Mitochondria, central organelles pivotal for eukaryotic cell function, extend their influence beyond ATP production, encompassing roles in apoptosis, calcium signaling, and biosynthesis. Recent studies spotlight two emerging determinants of mitochondrial functionality: intramitochondrial viscosity and sulfur dioxide (SO2) levels. While optimal mitochondrial viscosity governs molecular diffusion and vital processes like oxidative phosphorylation, aberrations are linked with neurodegenerative conditions, diabetes, and cancer. Similarly, SO2, a gaseous signaling molecule, modulates energy pathways and oxidative stress responses; however, imbalances lead to cytotoxic sulfite and bisulfite accumulation, triggering disorders such as cancer and cardiovascular anomalies. Our research focused on development of a dual-channel fluorescent probe, applying electron-withdrawing acceptors within a coumarin dye matrix, facilitating monitoring of mitochondrial viscosity and SO2 in live cells. This probe distinguishes fluorescence peaks at 650 nm and 558 nm, allowing ratiometric quantification of SO2 without interference from other sulfur species. Moreover, it enables near-infrared viscosity determination, particularly within mitochondria. The investigation employed theoretical calculations utilizing Density Functional Theory (DFT) methods to ascertain molecular geometries and calculate rotational energies. Notably, the indolium segment of the probe exhibited the lowest rotational energy, quantified at 7.38 kcals/mol. The probe featured heightened mitochondrial viscosity dynamics when contained within HeLa cells subjected to agents like nystatin, monensin, and bacterial lipopolysaccharide (LPS). Overall, our innovative methodology elucidates intricate mitochondrial factors, presenting transformative insights into cellular energetics, redox homeostasis, and therapeutic avenues for mitochondrial-related disorders.
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  • 文章类型: Journal Article
    光学分辨率光声显微镜(OR-PAM)的威望目标选择性和成像深度已引起人们的注意,以实现先进的细胞内可视化。然而,光声信号的宽带特性容易产生噪声和由低效的光压转换引起的伪影,导致图像质量差。本研究预见了奇异值分解(SVD)的应用,以有效地从这些噪声和伪影中提取光声信号。尽管时空SVD成功地提取了超声血流信号,由于访问多个帧的负担,传统的多帧模型不适用于通过扫描OR-PAM获取的数据。要在OR-PAM上使用SVD,这项研究从探索SVD应用于光声信号的多条A线而不是帧开始。经过探索,观察到不确定存在不需要的奇异向量的障碍。为了解决这个问题,在分析时空奇异向量的基础上,设计了一个数据驱动的加权矩阵来提取相关的奇异向量。通过使用加权矩阵的SVD对提取能力的评估显示出与过去研究相比具有高效计算的出色信号质量。总之,这项研究通过探索应用于A线信号的SVD以及从噪声和伪影分量中区分和恢复光声信号的实际应用,为该领域做出了贡献。
    The prestige target selectivity and imaging depth of optical-resolution photoacoustic microscope (OR-PAM) have gained attentions to enable advanced intra-cellular visualizations. However, the broad-band nature of photoacoustic signals is prone to noise and artifacts caused by the inefficient light-to-pressure translation, resulting in poor image quality. The present study foresees application of singular value decomposition (SVD) to effectively extract the photoacoustic signals from these noise and artifacts. Although spatiotemporal SVD succeeded in ultrasound flow signal extraction, the conventional multi frame model is not suitable for data acquired with scanning OR-PAM due to the burden of accessing multiple frames. To utilize SVD on the OR-PAM, this study began with exploring SVD applied on multiple A-lines of photoacoustic signal instead of frames. Upon explorations, an obstacle of uncertain presence of unwanted singular vectors was observed. To tackle this, a data-driven weighting matrix was designed to extract relevant singular vectors based on the analyses of temporal-spatial singular vectors. Evaluation on the extraction capability by the SVD with the weighting matrix showed a superior signal quality with efficient computation against past studies. In summary, this study contributes to the field by providing exploration of SVD applied on A-line signals as well as its practical utilization to distinguish and recover photoacoustic signals from noise and artifact components.
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