Strategies to prepare and characterize native membrane proteins and protein membranes by AFM

Progress in characterizing native membrane proteins and protein membranes by atomic force microscopy (AFM) opens exciting possibilities. While the structure, oligomeric state and supramolecular assembly of membrane proteins are assessed directly by AFM, single-molecule force spectroscopy (SMFS) identifies interactions that stabilize the fold, and characterize the switching between functional states of membrane proteins. But what is next? How can we approach cell biological, pharmaceutical and medical questions associated with native cellular membranes? How can we probe the functional state of cell membranes and study the dynamic formation of compartments? Such questions have been addressed by immobilizing membranes on solid supports, which ensures the integrity of the native state of membrane proteins but does not necessarily provide a native-like environment. Direct attachment of membranes to solid supports involves non-specific interactions that may change the physical state of supported lipids and proteins possibly hindering the assembly of membrane proteins into native functional compartments. Thus, to observe the dynamic assembly and working of proteins in native membranes by AFM, supports are required that mimic the native environment of the cell membrane as closely as possible. This review reports on recent progress in characterizing native membrane proteins by AFM, and surveys conventional and new approaches of supporting surfaces, which will allow the function, dynamics, and assembly of membrane proteins to be studied by AFM in native cell membranes.     

The effects of fluorescent probes on model membrane organization: photo-induced lipid sorting and soft structure formation

Liquid crystalline phase behaviour in model cell membranes prepared in-vitro has generated an enormous amount of interest in recent years and the phase diagrams of various bio-mimetic mixtures explored. Most recently photo-induced oxidation in fluorescently labelled membranes has been shown to influence these phase diagrams and can be used to generate interesting membrane-based geometries based on differences in membrane composition. In this article the different phases observed in model membranes will be discussed. Methods used to study these systems and recent developments in photo-stimulated phase changes are described along with areas for future investigation.     

原子力显微镜-荧光显微镜联用技术在活细胞单分子检测中的应用

原子力显微镜(Atomic force microscopy,AFM)及荧光显微镜(Fluorescence microscopy,FM)是目前活细胞单分子分析检测中最常用的两种工具.结合两种显微镜的优势,发展高时空分辨、多功能的AFM-FM联用技术成为近年该领域的研究热点.本文简述了AFM单分子力谱和FM单分子荧光成像的原理,总结了AFM-FM联用系统在仪器研制方面的发展概况,并结合本课题组在应用AFM-FM联用技术研究细胞膜上配受体相互作用等方面的工作,介绍了其在活细胞单分子检测中的应用进展.     

Characterization of fiber-forming peptides and proteins by means of atomic force microscopy

The atomic force microscope (AFM) is widely used in biological sciences due to its ability to perform imaging experiments at high resolution in a physiological environment, without special sample preparation such as fixation or staining. AFM is unique, in that it allows single molecule information of mechanical properties and molecular recognition to be gathered. This review sets out to identify methodological applications of AFM for characterization of fiber-forming proteins and peptides. The basics of AFM operation are detailed, with in-depth information for any life scientist to get a grasp on AFM capabilities. It also briefly describes antibody recognition imaging and mapping of nanomechanical properties on biological samples. Subsequently, examples of AFM application to fiber-forming natural proteins, and fiber-forming synthetic peptides are given. Here, AFM is used primarily for structural characterization of fibers in combination with other techniques, such as circular dichroism and fluorescence spectroscopy. More recent developments in antibody recognition imaging to identify constituents of protein fibers formed in human disease are explored. This review, as a whole, seeks to encourage the life scientists dealing with protein aggregation phenomena to consider AFM as a part of their research toolkit, by highlighting the manifold capabilities of this technique.     

qPlus型非接触原子力显微技术进展及前沿应用

原子力显微镜(AFM)通过探测针尖与样品之间的相互作用力获得样品表面的结构信息。基于qPlus传感器的非接触原子力显微镜(NC-AFM)在传统AFM的基础上进一步提升了空间分辨率,为研究表面物理和化学过程提供了一种新的成像和谱学研究技术。本文首先介绍NC-AFM的基本构造、高分辨成像机制和力谱测量等工作原理,总结了近年来NC-AFM在表面在位化学反应、低维材料表征和表面电荷分布测量等方面的应用,探讨了NC-AFM技术的发展与完善,展望了NC-AFM面临的机遇和挑战。     

Membranes and membrane processes for chiral resolution

This critical review is devoted to an active field of research on chiral separation, membrane-based enantioseparation technique, which has potential for large-scale production of single-enantiomer compounds. Adsorption-type enantioselective membranes and membrane-assisted resolution systems with non-enantioselective solid membranes have attracted much attention recently. The principles and recent developments of both enantioselective liquid and solid membranes and membrane-assisted processes for chiral resolution will be summarized comprehensively in this review, in which the contents are of interest to a wide range of readers in a variety of fields from analytical, organic and medicinal chemistry, to pharmaceutics and materials, to process engineering for fabricating pharmaceuticals, agrochemicals, fragrances and foods, and so on (148 references).     

Combined AFM and two-focus SFCS study of raft-exhibiting model membranes.

Dioleoylphosphatidylcholine/sphingomyelin/cholesterol (DOPC/SM/cholesterol) model membranes exhibit liquid-liquid phase separation and therefore provide a physical model for the putative liquid-ordered domains present in cells. Here we present a combination of atomic force microscopy (AFM) imaging, force measurements, confocal fluorescence imaging and two-focus scanning fluorescence correlation spectroscopy (two-focus SFCS) to obtain structural and dynamical information about this model membrane system. Partition coefficients and diffusion coefficients in the different phases were measured with two-focus SFCS for numerous fluorescent lipid analogues and proteins, while being directly related to the lateral organization of the membrane and its mechanical properties probed by AFM. Moreover we show how the combination of these different approaches is effective in reducing artifacts resulting from the use of a single technique.     

High-resolution synchrotron radiation core-level spectroscopy of semiconductor surfaces

Thanks to recent technical developments (optics and detection systems), at several synchrotron radiation facilities in the world, it is possible to perform photoemission spectroscopy studies with very high resolution. In the present report, we will review the new insights gained from high-resolution core-level spectroscopy on the structural and electronic properties of different semiconductor surfaces, such as InAs(110), Si and Ge (100) and (111), as well as clean and adsorbate covered Si and Ge(111) surfaces.     

A concise review of mass spectrometry imaging

Mass spectrometric imaging allows the investigation of the spatial distribution of molecules at complex surfaces. The combination of molecular speciation with local analysis renders a chemical microscope that can be used for the direct biomolecular characterization of histological tissue surfaces. MS based imaging advantageously allows label-free detection and mapping of a wide-range of biological compounds whose presence or absence can be the direct result of disease pathology. Successful detection of the analytes of interest at the desired spatial resolution requires careful attention to several steps in the mass spectrometry imaging protocol. This review will describe and discuss a selected number of crucial developments in ionization, instrumentation, and application of this innovative technology. The focus of this review is on the latest developments in imaging MS. Selected biological applications are employed to illustrate some of the novel features discussed. Two commonly used MS imaging techniques, secondary ion mass spectrometric (SIMS) imaging and matrix-assisted laser desorption ionization (MALDI) mass spectrometric imaging, center this review. New instrumental developments are discussed that extend spatial resolution, mass resolving power, mass accuracy, tandem-MS capabilities, and offer new gas-phase separation capabilities for both imaging techniques. It will be shown how the success of MS imaging is crucially dependent on sample preparation protocols as they dictate the nature and mass range of detected biomolecules that can be imaged. Finally, developments in data analysis strategies for large imaging datasets will be briefly discussed.     

BODIPY-based supramolecular fluorescent metallacages

During the past few years, the construction of BODIPY-based supramolecular fluorescent metallacages through coordination-driven self-assembly has gained increasing interest due to their unique photophysical properties and applications in catalysis, sensing, and bioimaging. In consideration of the rapid development of this field, it is time to summarize recent developments involving BODIPY-based metallacages.In this review, a comprehensive summary of the construction of BODIPY-based metallacages ...     

Porous ceramic membranes for catalytic reactors — overview and new ideas

The membrane reactor (MR) concept, combining in the same unit a conversion effect (catalyst) and a separation effect (membrane), already showed various potential benefits (increased reaction rate, selectivity and yield) for a range of reactions involving the membrane as extractor, distributor or contactor. Due to the generally severe conditions of heterogeneous catalysis, most MR applications use inorganic membranes, which can be dense or porous, inert or catalytically active. After a rapid overview of the working concepts of MRs, the main types of porous ceramic membranes, which have been developed for MR applications, are reported and discussed (characteristics and limitations). Starting from these general basis, our objective is to put recent developments into focus, with a special emphasis on porous composite infiltrated membranes and related synthesis methods. Some new ideas currently explored in our group, such as the ‘chemical valve membrane’ concept and the interest of nanophase materials for oxygen transport, will be also developed. An attempt in addressing the future developments of porous membranes for MRs will be finally proposed.     

Single-molecule chemistry and physics explored by low-temperature scanning probe microscopy

It is well known that scanning probe techniques such as scanning tunnelling microscopy (STM) and atomic force microscopy (AFM) routinely offer atomic scale information on the geometric and the electronic structure of solids. Recent developments in STM and especially in non-contact AFM have allowed imaging and spectroscopy of individual molecules on surfaces with unprecedented spatial resolution, which makes it possible to study chemistry and physics at the single molecule level. In this feature article, we first review the physical concepts underlying image contrast in STM and AFM. We then focus on the key experimental considerations and use selected examples to demonstrate the capabilities of modern day low-temperature scanning probe microscopy in providing chemical insight at the single molecule level.     

Peptide meets membrane: Investigating peptide-lipid interactions using small-angle scattering techniques

Peptide–lipid interactions play an important role in defining the mode of action of drugs and the molecular mechanism associated with many diseases. Model membranes consisting of simple lipid mixtures mimicking real cell membranes can provide insight into the structural and dynamic aspects associated with these interactions. Small-angle scattering techniques based on X-rays and neutrons (SAXS/SANS) allow in situ determination of peptide partition and structural changes in lipid bilayers in vesicles with relatively high resolution between 1-100 nm. With advanced instrumentation, time-resolved SANS/SAXS can be used to track equilibrium and nonequilibrium processes such as lipid transport and morphological transitions to time scales down to a millisecond. In this review, we provide an overview of recent advances in the understanding of complex peptide–lipid membrane interactions using SAXS/SANS methods and model lipid membrane unilamellar vesicles. Particular attention will be given to the data analysis, possible pitfalls, and how to extract quantitative information using these techniques.     

Recent advances in the formation of ultrathin polymeric membranes for gas separations

During the past 20 years membrane systems have been applied to a limited number of commercial gas separations. To further advance membrane-based gas separations, current research efforts focus on optimization of (i) membrane materials, (ii) membrane structures and (iii) membrane system design. In this overview, recent developments in the formation of high-performance gas separation membranes are discussed. The gas separation properties of state-of-the-art integrally skinned asymmetric membranes and thin-film composite membranes are summarized. Future directions for the preparation of advanced gas separation membranes are highlighted.     

X‐ray action on polymeric membrane surfaces: a chemical and morphological characterization

The surfaces of six polymeric membranes—two polysulphone membranes, two composite reverse‐osmosis polyamide/polysulphone membranes having polyamide as the active layer and two activated membranes containing di‐2‐ethylhexylphosphoric acid and di‐2‐ethylhexyldithiophosphoric acid as carriers, respectively—have been characterized before and after irradiation with an x‐ray source, both chemically and topographically by XPS and atomic force microscopy (AFM), respectively. Changes in atomic concentrations of the characteristic elements of the membranes and in the shape of XPS spectra as a function of irradiation time can be related to chemical modifications on the membrane surface. The most significant changes have been observed for polysulphone, which is reduced by x‐ray action; this fact also shows the inhomogeneity of the surface of the di‐2‐ethylhexyldithiophosphoric‐activated membrane. In contrast, polyamide top layers of composite membranes have been shown to be the most stable. Chemical modifications are not related directly to changes in membrane roughness because for all membranes only small changes have been observed for AFM images recorded before and after membrane irradiation. Moreover, the roughness of both polysulphone membranes decreases slightly due to x‐ray radiation but increases slightly for all polyamide‐containing membranes (composite and activated membranes). Copyright © 2003 John Wiley & Sons, Ltd.     

无机膜与无机膜催化反应

本文对无机膜的制备、应用以及膜催化反应的现状进行了较全面的综述,并对研究工作中存在的问题展开了讨论。     

Surface analysis with atomic force microscopy through measurement in air and under liquids

Atomic force microscopy offers a number of unique options that make it a valuable tool for analytical chemists. Especially the capability of imaging surfaces in gases and under liquids greatly enhances its applicability to technological problems. In this paper we demonstrate the analytical potential of AFM with respect to topographical and atomic resolution imaging, imaging under liquids and in-situ imaging of surface processes. Selected examples including metal films, glasses, alkali halides, electroluminescence displays, filtration membranes, organic textile fibers and human hair are described.     

Make them Blink: Probes for Super‐Resolution Microscopy

In recent years, a number of approaches have emerged that enable far‐field fluorescence imaging beyond the diffraction limit of light, namely super‐resolution microscopy. These techniques are beginning to profoundly alter our abilities to look at biological structures and dynamics and are bound to spread into conventional biological laboratories. Nowadays these approaches can be divided into two categories, one based on targeted switching and readout, and the other based on stochastic switching and readout of the fluorescence information. The main prerequisite for a successful implementation of both categories is the ability to prepare the fluorescent emitters in two distinct states, a bright and a dark state. Herein, we provide an overview of recent developments in super‐resolution microscopy techniques and outline the special requirements for the fluorescent probes used. In combination with the advances in understanding the photophysics and photochemistry of single fluorophores, we demonstrate how essentially any single‐molecule compatible fluorophore can be used for super‐resolution microscopy. We present examples for super‐resolution microscopy with standard organic fluorophores, discuss factors that influence resolution and present approaches for calibration samples for super‐resolution microscopes including AFM‐based single‐molecule assembly and DNA origami.     

Atomic force microscopy of microbial cells: application to nanomechanical properties, surface forces and molecular recognition forces

In recent years, the physical properties and interaction forces of microbial cell surfaces have been extensively studied using atomic force microscopy (AFM). A variety of AFM force spectroscopy approaches have been developed for investigating native cell surfaces with piconewton (nanonewton) sensitivity and nanometer lateral resolution, providing novel information on the nanomechanical properties of cell walls, on surface forces such as van der Waals and electrostatic forces, solvation and steric/bridging forces, and on the forces and localization of molecular recognition events. The intention of this article is to survey these different applications and to discuss related methodologies (how to prepare tips and samples, how to record and interpret force curves).