线粒体靶向的近红外HClO/ClO-荧光探针的研究进展

HClO/ClO-作为细胞质中一种重要的活性氧(ROS),源自线粒体,参与各种生理和病理过程,因此快速有效检测HClO/ClO-具有重要的生物学及生理学意义.荧光分析法因其灵敏度高、响应时间快、选择性高、成本低和操作简便等优点而备受关注.更重要的是,使用荧光探针可以在体外和体内可视化检测.近年来,为了研究HClO/Cl...     

线粒体靶向过氧化氢荧光探针的研究进展

控制线粒体过氧化氢的含量可以为细胞存活、生长、分化和维持发挥有益的作用.然而,线粒体过氧化氢的异常产生可能会破坏生物大分子以及细胞的结构,导致老化、突变甚至癌症的发生.因此,迫切需要能够在活细胞中特别是在线粒体中对过氧化氢的水平变化进行有效监测的手段.为此,开发、设计了多种监测线粒体中过氧化氢水平变化的荧光探针.以靶向...     

基于硝基还原机理的一氧化碳荧光探针的开发及细胞成像研究

一氧化碳(CO)作为一种重要的细胞气体信号传递分子,参与生物体内重要的生理和病理功能.选用经典的荧光平台——萘酰亚胺荧光平台,以硝基基团作为识别位点,通过严谨的设计和两步合成的方法,开发了一种新型用来检测一氧化碳的荧光增强型荧光探针NaLy-CO.光谱测试数据表明,该荧光探针对一氧化碳具有较好的选择性和灵敏度.利用荧光...     

基于三苯基磷的线粒体靶向荧光探针研究进展

简述了线粒体内各种相关活性物质及实时变化是探索生命体奥秘的重要组成部分,而化学荧光探针则是研究该领域的重要工具之一。近年来,具有定位和检测双功能的线粒体荧光探针被相继报道,其中,基于三苯基磷的线粒体靶向荧光探针因具有良好检测性能而成为当前研究的热点。综述了近十年以三苯基磷为定位基团的线粒体靶向体荧光探针在检测活性氧和还原性物质等方面的研究进展,并归纳了文献报道中各探针的特点,以期为进一步探索构建新的线粒体靶向荧光探针提供借鉴与参考。     

细胞内活性小分子近红外荧光成像探针

近年来,随着生命科学的不断发展,人们对细胞内活性小分子在病理、生理等方面的功能研究越来越深入。荧光成像作为一种直观、原位的可视化观测技术在小分子检测方面得到了广泛应用,其中基于近红外分子与纳米探针的荧光成像技术因具有背景干扰低、对细胞损伤小、样品穿透性强、检测灵敏度高等优点,显示了较好的应用前景。本文评述了近年来近红外荧光探针用于细胞内活性小分子成像检测的应用及进展,主要讨论该类方法在活性氧物质、金属离子、H+、阴离子及巯基化合物的分析应用,并对该方法的应用前景进行了展望。     

基于Nile Red的线粒体靶向性增强型硫化氢荧光探针及其细胞成像

硫化氢作为一类重要的信号分子, 除了在神经系统、炎症应激和心血管等生理系统方面发挥着调控作用之外, 还对线粒体ATP酶的活性和抗氧化应激起着重要的调节作用. 为了实时定量的检测线粒体中的硫化氢, 本文报道了一个可以定位于线粒体并检测外源硫化氢的荧光增强型荧光探针NRS. 探针NRS以尼尔红为母体通过引入吸电子的2,4-二硝基苯结构单元构建了基于光诱导电子转移(PET)机理的荧光探针. 探针NRS表现出快速的硫化氢响应性和较高的硫化氢选择性, 不受其他活性氧、活性氮、阴离子以及金属离子等物种的干扰. 随着硫化钠的加入, 探针NRS的最大吸收峰由565 nm蓝移至550 nm, 溶液由紫色变为红色; 同时在640 nm处的荧光光谱强度不断增强, 当硫化钠的浓度处于32～176 μmol·L^-1范围内, 荧光强度与硫化钠的浓度呈现出较好的线性关系. 细胞染色实验表明, 探针NRS能够进入到细胞内部, 具有细胞膜的透过性; 与Rh123进行共定位成像进一步证明探针NRS能够定位于细胞的线粒体中并检测硫化氢.     

近红外Cu2+比率荧光探针合成及其细胞成像

基于罗丹明染料和菁染料衍生物,设计合成了一种新型近红外比率Cu2+荧光探针(RCy7).通过核磁共振和质谱对其结构进行了表征.详细研究了该荧光探针对Cu2+的光学识别性能.在体积比为1∶1的乙腈/Tris-HCl(20 mmol/L,pH=7.2)缓冲溶液中,探针RCy7显蓝色且在722 nm波长处有较强的荧光发射峰,...     

近红外荧光探针检测金属离子、小分子和生物大分子

荧光探针技术是近年来发展迅速的一种荧光分析方法,具有灵敏度高、选择性好、操作简便和响应迅速等特点,受到环境及生命科学领域的青睐.随着荧光探针技术的发展,近红外一区荧光探针由于具有发射波长长(600～ 900 nm)、对细胞损伤小、组织穿透性强和自发荧光背景低等优点,被广泛应用于细胞、组织等复杂生物体系中生物分子的检测、...     

An Ultrasensitive Cyclization‐Based Fluorescent Probe for Imaging Native HOBr in Live Cells and Zebrafish

Bromine has been reported recently as being the 28^th essential element for human health. HOBr, which is generated in vivo from bromide, is a required factor in the formation of sulfilimine crosslinks in collagen IV. However, to date, no method for the specific detection of native HOBr in vivo has been reported. Herein, we develop a simple small molecular probe for imaging HOBr based on a specific cyclization catalyzed by HOBr. The probe can be easily synthesized in high yield through a Suzuki cross‐coupling reaction. The probe exhibits ultrahigh sensitivity at the picomole level, in addition to specificity for HOBr and real‐time response. Importantly, without Br^? stimulation, this probe reports native HOBr levels in HepG2 cells. Thus, the probe is a promising new tool for imaging endogenous HOBr and may provide a means for finding new physiological functions of HOBr in living organisms.     

细胞内铜离子的双光子荧光成像

基于无荧光的螺环结构与具有荧光的开环酰胺的平衡反应,本文合成了一个能在水基的缓冲溶液中选择性地识别Cu²⁺的罗丹明衍生物FD2.当在HEPES缓冲溶液中加入10当量的Cu²⁺时,FD2的单光子激发荧光和双光子激发荧光的强度均表现出明显的增强;更为重要的是,运用双光子荧光显微技术可以选择性地对活细胞内Cu²⁺进行成像.     

用于细胞和组织中弗林蛋白酶特异性成像的双光子荧光探针研究

弗林蛋白酶是前体蛋白转化酶家族中最具特色的酶之一,具有重要的生物学功能,其表达量水平与许多疾病有密切的关系,如癌症的发生和发展与弗林蛋白酶表达水平有着密切关联.目前文献中报道了一些单光子荧光探针用于弗林蛋白酶的检测,但这些探针不能应用于深层组织成像,且弗林蛋白酶在肿瘤发展过程的作用仍没有得到很好地研究.针对这些问题,本工作构建了一种新型双光子荧光探针Nap-F用于细胞和肿瘤组织内弗林蛋白酶的检测与双光子成像. Nap-F是由经典双光子荧光染料1,8-萘酰亚胺、弗林蛋白酶特异性多肽序列RVRR和自消除连接体整合而成.实验结果表明Nap-F对弗林蛋白酶具有很好的特异性,能够定量检测弗林蛋白酶的活性.在飞秒激光820nm激发下,Nap-F能有效降低生物背景,并提高组织穿透深度,适用于细胞和组织的双光子成像.Nap-F成功地实现了几种活细胞中弗林蛋白酶的双光子成像,揭示了癌细胞和表达缺陷细胞中弗林蛋白酶含量的差异.更重要的是,我们将该探针用于Co Cl2固定HIF-1构建的肿瘤细胞缺氧模型成像,实验结果表明弗林蛋白酶的表达与肿瘤细胞缺氧程度存在正相关性.     

一种选择性检测SO_2的荧光增强型探针的合成

SO_2作为一种重要的气体信号分子,其浓度异常与癌症、心血管疾病、神经系统疾病有关,因此有必要发展一种检测SO_2的分析方法。本文以1,1,2-三甲基-1H-苯并[e]吲哚和碘甲烷为原料合成中间体2;2与2,4-二甲基-5-醛基-1H-吡咯-3-羧酸经缩合反应合成了一个基于苯并吲哚和吡咯共轭的荧光增强型探针(3),其结构经1HNMR,13CNMR和HR-MS表征。探针3可以定量检测外源性SO_2,并且对SO_2具有良好的选择性,对其他阴离子和生物硫醇不响应或响应水平低。用350nm激发,探针3在450nm处的荧光强度随SO_2的浓度增加而增强。当SO_2浓度为0~100μmol·L~(-1)时,荧光强度与其呈良好的线性关系,相关系数R~2为0.997。     

比率型SO2荧光探针的构建及细胞成像研究

本文构建了一种基于苯并吲哚季铵盐结构的荧光探针用于检测SO₂衍生物。该荧光探针能够快速、灵敏、高选择性地检测HSO₃^-和SO₃^2-,并显示出颜色和荧光变化双重响应。其比率荧光强度（I₄₆₂/I₅₈₈）与HSO₃^-的浓度（0～16 μmol/L）之间具有良好的线性关系,检测限低至12 nmol/L。¹HNMR表明该探针的响应机制为1,4-亲核加成反应。激光共聚焦荧光成像结果表明,CZBI具有良好的细胞膜通透性,并且可以通过比率荧光成像实现对细胞内SO₂衍生物的监测。     

Characterization of a Hg2+-Selective Fluorescent Probe Based on Rhodamine B and Its Imaging in Living Cells

A small organic molecule P was synthesized and characterized as a fluorometric and colorimetric dual-modal probe for Hg²⁺. The sensing characteristics of the proposed probe for Hg²⁺ were studied in detail. A fluorescent enhancing property at 583 nm (>30 fold) accompanied with a visible colorimetric change, from colorless to pink, was observed with the addition of Hg²⁺ to P in an ethanol-water solution (8:2, v/v, 20 mM HEPES, pH 7.0), which would be helpful to fabricate Hg²⁺-selective probes with “naked-eye” and fluorescent detection. Meanwhile, cellular experimental results demonstrated its low cytotoxicity and good biocompatibility, and the application of P for imaging of Hg²⁺ in living cells was satisfactory.     

A Two-Photon Fluorescent Probe for the Visual Detection of Peroxynitrite in Living Cells and Zebrafish

Peroxynitrite (ONOO⁻), as an important reactive oxygen species (ROS), holds great potential to react with a variety of biologically active substances, leading to the occurrence of various diseases such as cancer and neurodegenerative diseases. In this work, we developed a novel mitochondria-localized fluorescent probe, HDBT-ONOO⁻, which was designed as a mitochondria-targeting two-photon fluorescence probe based on 1,8-naphthylimide fluorophore and the reactive group of 4-(bromomethyl)-benzene boronic acid pinacol ester. More importantly, the probe exhibited good biocompatibility, sensitivity, and selectivity, enabling its successful application in imaging the generation of intracellular and extracellular ONOO⁻. Furthermore, exogenous and endogenous ONOO⁻ products in live zebrafish were visualized. It is greatly expected that the designed probe can serve as a useful imaging tool for clarifying the distribution and pathophysiological functions of ONOO⁻ in cells and zebrafish.     

一种快速响应的线粒体靶向荧光探针用于检测活细胞和斑马鱼中的次氯酸盐

以香豆素为荧光团,设计并合成了一种线粒体靶向荧光探针Cou-Py,用于检测ClO^-。由于肟基在激发态的C=N异构化,Cou-Py表现出非常微弱的荧光,能够实现ClO^-促发的脱氧反应,在5 s内实现荧光恢复。此外,Cou-Py对ClO^-表现出高的选择性和低的检测限（6.87 nmol·L^-1）。重要的是,Cou-Py已成功用于检测MCF-7细胞线粒体和斑马鱼幼虫中的ClO^-。     

Development of a Two-photon Ratiometric Fluorescent Probe for Glutathione and Its Applications in Living Cells

Glutathione(GSH), as the most abundant intracellular biothiol, plays an important role in the redox homeostasis of the organism. Abnormal concentrations of GSH in cells may lead to many malignant diseases, such as cancer, liver damage and neurodegenerative diseases. It is urgent to develop effective methods to detect GSH in living organisms. In this work, a new two-photon ratiometric fluorescent probe Co-GSH based on the coumarin-chalcone dye platform was judiciously developed. Based on the Michael-addition reaction, Co-GSH was able to identify GSH with high selectivity and sensitivity. Furthermore, assisted by laser-scanning confocal microscopy, Co-GSH could specifically response GSH over the other biothiols, including Cys and Hcy, in living HeLa cells by using one-and two-photon modes.     

Development of a Highly Selective and Sensitive Fluorescent Probe for Imaging RNA Dynamics in Live Cells

RNA imaging is of great importance for understanding its complex spatiotemporal dynamics and cellular functions. Considerable effort has been devoted to the development of small-molecule fluorescent probes for RNA imaging. However, most of the reported studies have mainly focused on improving the photostability, permeability, long emission wavelength, and compatibility with live-cell imaging of RNA probes. Less attention has been paid to the selectivity and detection limit of this class of probes. Highly selective and sensitive RNA probes are still rarely available. In this study, a new set of styryl probes were designed and synthesized, with the aim of upgrading the detection limit and maintaining the selectivity of a lead probe QUID−1 for RNA. Among these newly synthesized compounds, QUID−2 was the most promising candidate. The limit of detection (LOD) value of QUID−2 for the RNA was up to 1.8 ng/mL in solution. This property was significantly improved in comparison with that of QUID−1. Further spectroscopy and cell imaging studies demonstrated the advantages of QUID−2 over a commercially available RNA staining probe, SYTO RNASelect, for highly selective and sensitive RNA imaging. In addition, QUID−2 exhibited excellent photostability and low cytotoxicity. Using QUID−2, the global dynamics of RNA were revealed in live cells. More importantly, QUID−2 was found to be potentially applicable for detecting RNA granules in live cells. Collectively, our work provides an ideal probe for RNA imaging. We anticipate that this powerful tool may create new opportunities to investigate the underlying roles of RNA and RNA granules in live cells.