Small Fluorogenic Amino Acids for Peptide-Guided Background-Free Imaging

The multiple applications of super-resolution microscopy have prompted the need for minimally invasive labeling strategies for peptide-guided fluorescence imaging. Many fluorescent reporters display limitations (e.g., large and charged scaffolds, non-specific binding) as building blocks for the construction of fluorogenic peptides. Herein we have built a library of benzodiazole amino acids and systematically examined them as reporters for background-free fluorescence microscopy. We have identified amine-derivatized benzoselenadiazoles as scalable and photostable amino acids for the straightforward solid-phase synthesis of fluorescent peptides. Benzodiazole amino acids retain the binding capabilities of bioactive peptides and display excellent signal-to-background ratios. Furthermore, we have demonstrated their application in peptide-PAINT imaging of postsynaptic density protein-95 nanoclusters in the synaptosomes from mouse brain tissues.     

A structural study of the retinal photoreceptor, plexiform and ganglion cell layers following exposure to UV-B and UV-C radiation in the albino rat

Over the last two decades, ultraviolet radiation levels (UV), reaching the Earth's surface, have been increasing at a rate of 1.5% per each 1% loss of the ozone layer. Moreover, artificial UV-sources have also proliferated and contributed to the rising UV-stress that many organisms have to face. To assess how the vertebrate retina responds to an exposure of short wavelength UV, we focused our attention on the rat retina, observing photoreceptor (containing outer and inner segments of rods and cones), inner plexiform, and ganglion cell layers by light and transmission electron microscopy using conventional and cytochemical techniques. We analyzed how cells of the layers in question responded to a 30 min exposure to UV-C and UV-B radiation with doses of 7200 and 590 J/cm², respectively. The results show that there are significant changes in the nuclei and cytoplasmic organelles of the exposed retinae when compared with those of the unexposed controls. The changes include an increase in heterochromatin, distension of rough endoplasmic reticulum, mitochondrial disruptions, and increases in the number of myelin bodies. The recorded morphological changes, especially those of the ganglion cells, are suggestive of apoptotic processes and show that the exposure of vertebrate retina to wavelengths ranging from 254 to 312 nm can produce alterations that are likely to impact negatively on the retina's proper functioning.     

Labeling the planar face of crystalline cellulose using quantum dots directed by type-I carbohydrate-binding modules

We report a new method for the direct labeling and visualization of crystalline cellulose using quantum dots (QDs) directed by carbohydrate-binding modules (CBMs). Two type-I (surface binding) CBMs belonging to families 2 and 3a were cloned and expressed with dual histidine tags at the N- and C-termini. Semiconductor (CdSe)ZnS QDs were used to label these CBMs following their binding to Valonia cellulose crystals. Using this approach, we demonstrated that QDs are linearly arrayed on cellulose, which implies that these CBMs specifically bind to a planar face of cellulose. Direct imaging has further shown that different sizes (colors) of QDs can be used to label CBMs bound to cellulose. Furthermore, the binding density of QDs arrayed on cellulose was modified predictably by selecting from various combinations of CBMs and QDs of known dimensions. This approach should be useful for labeling and imaging cellulose-containing materials precisely at the molecular scale, thereby supporting studies of the molecular mechanisms of lignocellulose conversion for biofuels production.     

Preparation and characterization of ordered porous carbons for increasing hydrogen storage behaviors

We prepared ordered porous carbons (PCs) by using a replication method that had well-organized mesoporous silica as a template with various carbonization temperatures in order to investigate the possibility of energy storage materials. The microstructure and morphologies of the samples are characterized by XRD, TEM, and FT-Raman spectroscopy. N₂ adsorption isotherms are analyzed by the t-plot method, as well as the BET and the H–K method in order to characterize the specific surface area, pore volume, and pore size distribution of the samples, respectively. The capacity of the hydrogen adsorption of the samples is evaluated by BEL-HP at 77 K and 1 bar. From the results, we are able to confirm that the synthesis of the samples can be accurately governed by the carbonization temperature, which is one of the effective parameters for developing the textural properties of the carbon materials, which affects the behaviors of the hydrogen storage.     

Multi-domain High-Resolution Platform for Integrated Spectroscopy and Microscopy Characterizations

In recent decades, materials science has experienced rapid development and posed increasingly high requirements for the characterizations of structures, properties, and performances. Herein, we report on our recent establishment of a multi-domain (energy, space, time) high-resolution platform for integrated spectroscopy and microscopy characterizations, offering an unprecedented way to analyze materials in terms of spectral (energy) and spatial mapping as well as temporal evolution. We present several proof-of-principle results collected on this platform, including in-situ Raman imaging (high-resolution Raman, polarization Raman, low-wavenumber Raman), time-resolved photoluminescence imaging, and photoelectrical performance imaging. It can be envisioned that our newly established platform would be very powerful and effective in the multi-domain high-resolution characterizations of various materials of photoelectrochemical importance in the near future.     

Stick-Slip Mechanisms at the Nanoscale

When two surfaces slide past each other, energy is mainly dissipated by stick-slip events. Macroscopic stick-slip is usually explained by asperities that come in and out of contact. Herein, we probe stick-slip at the nanoscale at interfaces and polymer coated interfaces by pulling single polymers covalently attached to an AFM cantilever tip laterally over solid substrates in liquid environment. We find two different stick mechanisms, namely desorption stick (DS) and cooperative stick (CS). While DS-slip resembles the velocity dependence of macroscopic stick-slip, CS-slip shows an increase in friction with velocity. For various reasons we anticipate that both stick mechanisms are necessary for a molecular understanding of stick-slip at the interface and interphase.     

高分辨阿达玛变换显微荧光图像分析(Ⅰ)——系统的成像能力及其在单个肿瘤细胞分析中的应用

阿达玛变换(Hadamard transform, HT)是一种类似于傅里叶变换的光谱调制技术, 具有多通道同时检测和多通道成像能力. 实现高分辨HT成像的关键在于阿达玛模板的制作, 阿达玛模板有两种, 即移动式机械编码模板(Movable mechanical mask)和固定式光电模板(Stationary electro-optic mask). 在实际成像方面, 移动模板和固定模板各有优缺点: 前者一般用石英玻璃制作, 对光信号不会因模板吸收而导致信号损失, 因此数据很可靠, 而且模板的制作也较为容易, 但由于采用步进电机驱动而容易导致机械故障, 难以实现快速编码; 后者无移动部件, 无机械故障, 因此系统比较紧凑, 但由于它是由液晶材料制成的(可导致信号损失), 从而限制了其在某些光谱区域的使用. 此外, 它对系统的软件设计要求比前者高, 实现高分辨成像更加困难. 正是由于上述原因, 实现快速、高分辨HT成像具有一定难度, 最近有关HT成像技术的报道极少.     

Visualizing Chiral Interactions in Carbohydrates Adsorbed on Au(111) by High-Resolution STM Imaging

Carbohydrates are the most abundant organic material on Earth and the structural “material of choice” in many living systems. Nevertheless, design and engineering of synthetic carbohydrate materials presently lag behind that for protein and nucleic acids. Bottom-up engineering of carbohydrate materials demands an atomic-level understanding of their molecular structures and interactions in condensed phases. Here, high-resolution scanning tunneling microscopy (STM) is used to visualize at submolecular resolution the three-dimensional structure of cellulose oligomers assembled on Au(1111) and the interactions that drive their assembly. The STM imaging, supported by ab initio calculations, reveals the orientation of all glycosidic bonds and pyranose rings in the oligomers, as well as details of intermolecular interactions between the oligomers. By comparing the assembly of D- and L-oligomers, these interactions are shown to be enantioselective, capable of driving spontaneous enantioseparation of cellulose chains from its unnatural enantiomer and promoting the formation of engineered carbohydrate assemblies in the condensed phases.     

Trimethylsilyl Chloride Aids in Solubilization of Oxidative Addition Intermediates from Zinc Metal

Trimethylsilyl chloride (TMSCl) is commonly used to “activate” metal(0) powders toward oxidative addition of organohalides, but knowledge of its mechanism remains limited by the inability to characterize chemical intermediates under reaction conditions. Here, fluorescence lifetime imaging microscopy (FLIM) overcomes these prior limitations and shows that TMSCl aids in solubilization of the organozinc intermediate from zinc(0) metal after oxidative addition, a previously unknown mechanistic role. This mechanistic role is in contrast to previously known roles for TMSCl before the oxidative addition step. To achieve this understanding, FLIM, a tool traditionally used in biology, is developed to characterize intermediates during a chemical reaction—thus revealing mechanistic steps that are unobservable without fluorescence lifetime data. These findings impact organometallic reagent synthesis and catalysis by providing a previously uncharacterized mechanistic role for a widely used activating agent, an understanding of which is suitable for revising activation models and for developing strategies to activate currently unreactive metals.     

Controlled Grafting Expansion Microscopy

Expansion microscopy (ExM) is a recently developed technique that allows for the resolution of structures below the diffraction limit by physically enlarging a hydrogel-embedded facsimile of the biological sample. The target structure is labeled and this label must be retained in a relative position true to the original, smaller state before expansion by linking it into the gel. However, gel formation and digestion lead to a significant loss in target-delivered label, resulting in weak signal. To overcome this problem, we have here developed an agent combining targeting, fluorescent labeling and gel linkage in a single small molecule. Similar approaches in the past have still suffered from significant loss of label. Here we show that this loss is due to insufficient surface grafting of fluorophores into the hydrogel and develop a solution by increasing the amount of target-bound monomers. Overall, we obtain a significant improvement in fluorescence signal retention and our new dye allows the resolution of nuclear pores as ring-like structures, similar to STED microscopy. We furthermore provide mechanistic insight into dye retention in ExM.