固体支撑的自组装的双层类脂膜

具有通常BLM_s某些相似特性的固体支撑的双层类脂膜(S-BLM)能够通过两步自组装到新劈开的金属丝上面。如:(1)包有聚四氟乙烯的铂丝头部浸在类脂溶液里,用解剖刀把顶部切开;(2)包有类脂溶液的新铂丝末端转移到0.1mol/L KCl溶液里,电测定证实,数分钟后,在金属丝末端自动地形成了稳定的类脂双层。本文报道了这种固体支撑的BLM(S-BLM)在检测Pb²⁺离子中的应用。S-BLM为液晶结构,它可用于基础研究、生物传感器和分子电子器件等技术上的应用。     

铂支撑的碘-聚吡咯修饰双层脂膜的I-V特性

首次在铂丝新生表面上形成了碘-聚吡咯双层脂膜,用循环伏安法研究了碘掺杂浓度对双层脂膜I-V特性的影响。用此法所得到的铂支撑双层脂膜十分稳定,可进一步用于生物传感器及分子电子器件的基础与应用研究。     

一套研究黑膜的新装置和三种类脂膜电特性的测定

在板形双分子膜的研究中,目前有两种成膜技术:(1)从单层到双层,(2)从脂滴到双层。由于后者是一个自动变薄过程,在显微镜视野中,可见膜的干涉光彩色花纹逐渐消失,膜黑区面积不断扩大,最终留下一个稳定的,四周由P—G边界包围的黑色膜,早期文献中常把用这种方法产生的膜称作黑膜(Black Membrane)。本文介绍用直流法研究黑膜电特性的一套实验装置,全部采用国产器材,利用这套装置,我们初步测试了三种板形人工双层类脂膜的电特性。一、实验装置已知黑膜的实测膜电阻——Rm为10~5—     

细胞传感器与芯片的研究进展

细胞传感器（cell-based biosensor）与芯片(cell-based biochip)已成为后基因时代生物科学研究的重要工具,它们利用生活细胞作为研究对象或敏感元件,与传感器和芯片技术相结合,通过生物信号与物理、电化学等其他信号的转换,实现实时、快速、微量地检测细胞的功能信息和待测物的性质.在细胞生物学研究、环境监测和药物开发等领域有广泛的应用.综述近三年来细胞传感器与芯片技术的研究进展及应用,并提出展望.     

DNA生物传感器在环境污染监测中的应用

基于生物催化和免疫原理的生物传感器在环境领域中获得了广泛应用.近年来,随着分子生物学和生物技术的发展,人们开发了以核酸探针为识别元件,基于核酸相互作用原理的DNA生物传感器.该传感器可用于受感染微生物的核酸序列分析、优先控制污染物的检测以及污染物与DNA之间相互作用的研究,在环境污染监测中具有潜在的巨大应用前景.简要介绍了核酸杂交生物传感器的基本原理及其在环境微生物和优先控制污染物（priority pollutant）检测中的应用研究进展.     

平面双层脂膜系统用于植物离子通道研究

平面双层脂膜系统是人工用脂类涂制的脂双层(BLM)结合电测定来模拟生物膜离子通透的系统。在1963年,Mueller和Rudin首先报道将脂类的溶液涂在一个直径很小的孔上时形成了脂双层,且具有和生物膜类似的电学性质,为将模拟膜系统用于生物膜研究奠定了基础。但是到70年代后,才将一些抗生素类的物质嵌合入BLM进行离子通道的研究。将生物膜嵌入BLM则是在70年代后朗的事,但当时没有受到广泛的重视,直到80年代单离子通道记录及膜片钳技术兴起时,人们才又重新将这一技术进行改     

滚环扩增技术在电化学生物传感器中的应用

在电化学生物传感器的设计中,信号放大是实验环节中的重要步骤,特别是对靶标进行灵敏度分析时更是不可或缺。滚环扩增（rolling circle amplification,RCA）能够在短时间内得到大量产物,并在电极表面进行扩增或孵育,然后通过一定的设计使电化学信号被快速放大。RCA技术具有高度的灵敏性和特异性,电化学生物传感器则可提供实时、快速、低成本的检测。为了更好的了解RCA,介绍了RCA环化的基本原理、RCA种类,重点总结了RCA与电化学生物传感器结合的不同技术类型及应用,并对未来相关研究领域的发展趋势进行了展望,旨在为RCA技术在电化学生物传感器中的进一步发展和应用提供参考。     

电信号分子在电化学功能核酸生物传感器中的研究进展

电信号分子是应用于功能核酸电化学生物传感器中起着信号转换作用的具有电化学活性且能够和核酸相互作用或是可以标记在核酸链上的一类分子的统称。电信号分子对于功能核酸电化学生物传感器是必不可少的一部分,它对于电化学生物传感器检测的灵敏度和应用的普及性都至关重要。简要介绍了5大类电信号分子,即染料类电信号分子、金属有机配合物类电信号分子、纳米材料类电信号分子、类过氧化氢酶类电信号分子、有机小分子类电信号分子,详细阐述了这些电信号在功能核酸电化学生物传感器中的应用,主要从产生电信号的方式、实际应用以及每种电信号的使用优缺点进行分析,并对新的电信号分子的发现或设计进行了展望,以期对后续有关电信号的研究有借鉴作用。     

Azone类透皮剂作用机理及一种新型模型膜的研究

使用电子自旋共振波谱技术,采用5—doxyl stearic acid作为自旋标记物,新设计的镶嵌JPM和胆固醇的卵壳膜作为实验模型膜,进行Azone类透皮吸收促进剂的主要作用机理研究.实验证实上述膜是一种很有前途的模型膜.由于Azone透皮剂的作用,增大膜中类脂和自旋标记物的脂肪长链的运动速率,即增强类脂的流动性,使得类脂的序参数值减小,从而证实了前人有关角质化细胞间的类脂相是药物穿透角质层的主要通道的假设.为进一步探讨透皮剂对天然皮肤的作用,采用裸鼠皮肤角质层作为实验模型膜,得到与上述实验相符的结果.     

细胞黏附压电传感响应机制分析

基于压电传感器的一维多层及传输线等效电路模型,利用声阻抗概念,将传感器响应与声阻抗直接联系,建立起压电传感器响应机制的声阻抗模型。由此模型对单、双层等基本负载分别导出相应的传感器响应方程。理论分析表明,声阻抗是生物传感的核心,可通过其阐明各种传感器响应机制的物理意义,特别是细胞黏附的压电传感响应机制分析。实验结果良好地验证了细胞黏附行为的压电传感响应声阻抗理论,据此建立了频率变化!f(Hz)与细胞浓度C(ml-1)之间良好的线性关系,相关系数R=0.98,其线性方程为"f=-246C-20.1(P<0.001)。研究对细胞黏附的压电传感及其应用具有指导意义。     

Charge-transfer processes and redox reactions in planar lipid monolayers and bilayers

Supported bilayer lipid membranes (BLMs) and lipid monolayers have been known for quite sometime and are attracting sustained interest since they open new research vista and offer practical approaches in biosensor development and molecular device applications. Central to these areas of interest are electric processes and redox reactions where the movement of ions and electrons plays a pivotal role. In this paper an overview of the major findings in this field is presented. Further, we summarize the work on planar lipid bilayers and monolayers that have been done in the past few years in a number of laboratories. Supported planar BLMs and their closely related systems provide the foundation for a variety of lipid bilayer-based molecular sensors that are sensitive, versatile, as well as potentially inexpensive (i.e., disposable), and open to all sorts of experimentation.     

Application of voltammetric techniques to membrane studies

The application of voltammetric methods to planar bilayer lipid membranes (BLM) studies is described. BLM-compound interaction experiments lead to the measurement of the membrane current underlying transport phenomena. From measurements of current/voltage of BLM in unstirred solutions as a function of scan rates, it is possible to obtain both thermodynamic and kinetic information. In past years, a variety of techniques have been used to study the electrical properties of BLMs, but in terms of versatility, the cyclic voltammetric technique is outstanding. Cyclic voltammetry is the definitive means of characterizing the redox process of electroactive membranes.     

Stabilized lipid film based biosensor for atenolol

This work reports a technique for the stabilization after storage in air of a lipid film based biosensor for atenolol. Microporous filters composed of glass fibers (nominal pore sizes 0.7 and 1.0 microm) were used as supports for the formation and stabilization of these devices. The lipid film is formed on the filter by polymerization prior to its use. Methacrylic acid was the functional monomer, ethylene glycol dimethacrylate was the crosslinker and 2,2'-azobis-(2-methylpropionitrile) was the initiator. The method for preparation of stabilized lipid film biosensor is studied throughout this work. The response towards atenolol of these stabilized lipid membrane biosensor, for repetitive use, composed of phosphatidylcholine was compared with planar freely suspended bilayer lipid membranes (BLMs). The stabilized lipid membranes provided similar artificial ion gating events as BLMs in the form of transient signals and can function for repetitive uses after storage in air. This will allow the practical use of the techniques for chemical sensing based on lipid films and commercialization of these devices.     

Self-assembling bilayer lipid membranes on solid support

Solid-supported bilayer lipid membranes (s-BLMs) that possess some properties similar to those of conventional BLMs can be self-assembled on a freshly cleaved metal wire by a two-step procedure: (i) The tip of a Teflon-coated platinum wire, while immersed in a lipid solution, is cut off with a scalpel; (ii) the new tip of the wire, having become coated with lipid solution, is transferred into 0.1 M KCl. After a few minutes, a stable lipid bilayer forms spontaneously on the tip of the wire, as verified by electrical measurements. An application of such a supported BLM (s-BLM) is reported for the detection of Pb2+ ions. The s-BLM is liquid-crystalline in structure, which makes it amenable to modification for basic studies, as well as for technological applications such as biosensors and molecular electronic devices.     

Interactions of liposomes with planar bilayer membranes

Summary The adhesion to horizontal, planar lipid membranes of lipid vesicles containing calcein in the aqueous compartment or fluorescent phospholipids in the membranes has been examined by phase contrast, differential interference contrast and fluorescence microscopy. With water-immersion lenses, it was possible to study the interactions of vesicles with planar bilayers at magnifications up to the useful limit of light microscopy. In the presence of 15 mM calcium chloride, vesicles composed of phosphatidylserine and either phosphatidylethanolamine or soybean lipids adhere to the torus, bilayer and lenses of planar bilayers of the same composition. Lenses of solvent appear, at the site where vesicles attach to decane-based bilayers and lipid fluorophores move from the vesicles to the lenses. Because the calcein contained in such vesicles is not released, we interpret this as indicating fusion of only the outer monolayer (hemifusion) of the vesicles with the decane lenses. In the case of squalene-based black lipid membranes (BLMs), in contrast, vesicles do not nucleate lenses but they apparently do fuse with the torus at the bilayer boundary. Interactions leading to hemifusions between vesicles and planar membranes thus occur predominantly in regions where hydrocarbon solvent is present. Osmotic water flow, induced by addition of urea to the compartment containing vesicles, causes coalescence of lenses in decane-based, BLMs as well as coalescence of the aqueous spaces of the vesicles that have undergone hemifusion with the lenses. We did not observe transfer of the aqueous phase of vesicles to therans side of either decane-or squalene-based planar membranes; however, we cannot rule out the possibility particularly in the latter case, that rupture of the planar membrane may have been an immediate result of vesicle fusion and thus precluded its detection.     

Chemical and Photochemical Modification of Colicin E1 and Gramicidin A in Bilayer Lipid Membranes

Chemical modification and photodynamic treatment of the colicin E1 channel-forming domain (P178) in vesicular and planar bilayer lipid membranes (BLMs) was used to elucidate the role of tryptophan residues in colicin E1 channel activity. Modification of colicin tryptophan residues by N-bromosuccinimide (NBS), as judged by the loss of tryptophan fluorescence, resulted in complete suppression of wild-type P178 channel activity in BLMs formed from fully saturated (diphytanoyl) phospholipids, both at the macroscopic-current and single-channel levels. The similar effect on both the tryptophan fluorescence and the electric current across BLM was observed also after NBS treatment of gramicidin channels. Of the single-tryptophan P178 mutants studied, W460 showed the highest sensitivity to NBS treatment, pointing to the importance of the water-exposed Trp460 in colicin channel activity. In line with previous work, the photodynamic treatment (illumination with visible light in the presence of a photosensitizer) led to suppression of P178 channel activity in diphytanoyl-phospholipid membranes concomitant with the damage to tryptophan residues detected here by a decrease in tryptophan fluorescence. The present work revealed novel effects: activation of P178 channels as a result of both NBS and photodynamic treatments was observed with BLMs formed from unsaturated (dioleoyl) phospholipids. These phenomena are ascribed to the effect of oxidative modification of double-bond-containing lipids on P178 channel formation. The pronounced stimulation of the colicin-mediated ionic current observed after both pretreatment with NBS and sensitized photomodification of the BLMs support the idea that distortion of membrane structure can facilitate channel formation.Abbreviations: AlP_cS₃, almininum trisulfophthalocyanine; BLM, bilayer lipid membrane; DOPC, dioleoylphosphatidylcholine; DOPG, dioleoylphosphatidyl-glycerol; DPhPG, diphytanoylphos-phatidylglycerol; DPhPg, diphytanoylphosphatidylcholine; gA, gramicidin A; NBS, N-bromosuccinimideThis revised version was published online in August 2005 with a corrected cover date.     

Comparative study of molecular dynamics,diffusion, and permeability for ligands in biomembranes of different lipid composition

A comparative study of several model lipid bilayers of different composition, which included analysis of kinetic parameters of model lipid bilayers and permeability of bilayer membranes for small molecules, has been carried out. The conformity of results of numeric experiments to experimental data (structure of membrane lipid bilayers, lateral diffusion coefficients, and relative permeability of biomembranes for ligands) is discussed in the framework of a standard molecular dynamics protocol.     

Structure of planar membrane formed from liposomes

Lipid vesicles with incorporated ion channels from polyene antibiotic amphotericin B were used to investigate structures of planar membranes formed by Shindler's techniques. A planar membrane assembled on the aperture in a lavsan film from two layers generated at the air-aqueous liposome suspension interface is not a simple bilayer but a bimolecular membrane containing numerous partly fused liposomes. A complete fusion of liposomal membranes with the planar bilayer is an unlikely event during membrane formation. A planar bimolecular lipid membrane without incorporated liposomes can be made by a method consisting of three stages: formation of a lipid layer on the air-water interface of a suspension containing liposomes, transfer of this layer along the surface of the solution into a chamber containing a solution without liposomes where a lipid monomolecular layer forms gradually (within about 20 min) at the air-water interface, assembling of the planar bilayer membrane from this monolayer. The knowledge of the planar membrane structure may be useful in experiments on incorporation of membrane proteins into a planar lipid bilayer.     

Electron density mapping of triblock copolymers associated with model biomembranes: insights into conformational states and effect on bilayer structure

One-dimensional electron-density profiles derived from synchrotron small-angle X-ray scattering (SAXS) have been constructed and used to determine the conformational state of selected poly(ethylene oxide)-b-poly(propylene oxide)-b-poly(ethylene oxide) (PEO-PPO-PEO) triblock copolymers and the region of their association with a lipid bilayer. The number of molecular repeat units in the hydrophobic PPO block has been found to determine both the nature of triblock polymer-membrane association and the conformational state of the symmetric, flanking hydrophilic PEO units. For DMPC-based biomembranes, polymers whose PPO chain length is less than that of the bilayer thickness insert weakly into the membrane with the PEO blocks on the same side of the bilayer, leading to delocalization of the PEO at the membrane-water interface. This polymer architecture has been found not to alter the membrane fluidity and roughness. Conversely, polymers whose chain length is sufficient to span the lipid bilayer are tightly integrated, projecting their PEO chains into the water channels on opposing sides of the bilayer. The coiled conformational state of the PEO chains produces steric pressure on the bilayer, causing a thinning of the membrane and leading to a rigid, less-mobile bilayer than systems where the polymer is introduced as the lipid conjugate.     

Artificial membrane-like environments for in vitro studies of purified G-protein coupled receptors

Functional reconstitution of transmembrane proteins remains a significant barrier to their biochemical, biophysical, and structural characterization. Studies of seven-transmembrane G-protein coupled receptors (GPCRs) in vitro are particularly challenging because, ideally, they require access to the receptor on both sides of the membrane as well as within the plane of the membrane. However, understanding the structure and function of these receptors at the molecular level within a native-like environment will have a large impact both on basic knowledge of cell signaling and on pharmacological research. The goal of this article is to review the main classes of membrane mimics that have been, or could be, used for functional reconstitution of GPCRs. These include the use of micelles, bicelles, lipid vesicles, nanodiscs, lipidic cubic phases, and planar lipid membranes. Each of these approaches is evaluated with respect to its fundamental advantages and limitations and its applications in the field of GPCR research. This article is part of a Special Issue entitled: Membrane protein structure and function.