Scanning Probe Evolution in Biology
07 Nov 2003-Science (American Association for the Advancement of Science)-Vol. 302, Iss: 5647, pp 1002-1005
TL;DR: Instruments with increased imaging rates, lower probe-specimen force interactions, and probe configurations not constrained to planar surfaces are being developed, with the goal of imaging processes at the single-molecule level—not only at surfaces but also within three-dimensional volumes—in real time.
Abstract: Twenty years ago the first scanning probe instrument, the scanning tunneling microscope, opened up new realms for our perception of the world. Atoms that had been abstract entities were now real objects, clearly seen as distinguishable individuals at particular positions in space. A whole family of scanning probe instruments has been developed, extending our sense of touching to the scale of atoms and molecules. Such instruments are especially useful for imaging of biomolecular structures because they can produce topographic images with submolecular resolution in aqueous environments. Instruments with increased imaging rates, lower probe-specimen force interactions, and probe configurations not constrained to planar surfaces are being developed, with the goal of imaging processes at the single-molecule level—not only at surfaces but also within three-dimensional volumes—in real time.
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TL;DR: The emerging ability to control the patterns of matter on the nanometer length scale can be expected to lead to entirely new types of biological sensors capable of sensing at the single-molecule level in living cells, and capable of parallel integration for detection of multiple signals.
Abstract: In the coming decade, the ability to sense and detect the state of biological systems and living organisms optically, electrically and magnetically will be radically transformed by developments in materials physics and chemistry. The emerging ability to control the patterns of matter on the nanometer length scale can be expected to lead to entirely new types of biological sensors. These new systems will be capable of sensing at the single-molecule level in living cells, and capable of parallel integration for detection of multiple signals, enabling a diversity of simultaneous experiments, as well as better crosschecks and controls.
2,960 citations
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TL;DR: This review critically analyzes what is known of phase behavior and liquid-liquid immiscibility in model systems and compares these data with what isknown of domain formation in cell membranes.
Abstract: Views of how cell membranes are organized are presently changing. The lipid bilayer that constitutes these membranes is no longer understood to be a homogeneous fluid. Instead, lipid assemblies, termed rafts, have been introduced to provide fluid platforms that segregate membrane components and dynamically compartmentalize membranes. These assemblies are thought to be composed mainly of sphingolipids and cholesterol in the outer leaflet, somehow connected to domains of unknown composition in the inner leaflet. Specific classes of proteins are associated with the rafts. This review critically analyzes what is known of phase behavior and liquid-liquid immiscibility in model systems and compares these data with what is known of domain formation in cell membranes.
1,615 citations
TL;DR: The biological functions of encapsulated drugs and cells can be dramatically enhanced by designing biomaterials with controlled organizations at the nanometer scale.
Abstract: Research in the areas of drug delivery and tissue engineering has witnessed tremendous progress in recent years due to their unlimited potential to improve human health. Meanwhile, the development of nanotechnology provides opportunities to characterize, manipulate and organize matter systematically at the nanometer scale. Biomaterials with nano-scale organizations have been used as controlled release reservoirs for drug delivery and artificial matrices for tissue engineering. Drug-delivery systems can be synthesized with controlled composition, shape, size and morphology. Their surface properties can be manipulated to increase solubility, immunocompatibility and cellular uptake. The limitations of current drug delivery systems include suboptimal bioavailability, limited effective targeting and potential cytotoxicity. Promising and versatile nano-scale drug-delivery systems include nanoparticles, nanocapsules, nanotubes, nanogels and dendrimers. They can be used to deliver both small-molecule drugs and various classes of biomacromolecules, such as peptides, proteins, plasmid DNA and synthetic oligodeoxynucleotides. Whereas traditional tissue-engineering scaffolds were based on hydrolytically degradable macroporous materials, current approaches emphasize the control over cell behaviors and tissue formation by nano-scale topography that closely mimics the natural extracellular matrix (ECM). The understanding that the natural ECM is a multifunctional nanocomposite motivated researchers to develop nanofibrous scaffolds through electrospinning or self-assembly. Nanocomposites containing nanocrystals have been shown to elicit active bone growth. Drug delivery and tissue engineering are closely related fields. In fact, tissue engineering can be viewed as a special case of drug delivery where the goal is to accomplish controlled delivery of mammalian cells. Controlled release of therapeutic factors in turn will enhance the efficacy of tissue engineering. From a materials point of view, both the drug-delivery vehicles and tissue-engineering scaffolds need to be biocompatible and biodegradable. The biological functions of encapsulated drugs and cells can be dramatically enhanced by designing biomaterials with controlled organizations at the nanometer scale. This review summarizes the most recent development in utilizing nanostructured materials for applications in drug delivery and tissue engineering.
959 citations
TL;DR: The basic principles, advantages and limitations of the most common AFM bioimaging modes are reviewed, including the popular contact and dynamic modes, as well as recently developed modes such as multiparametric, molecular recognition, multifrequency and high-speed imaging.
Abstract: Atomic force microscopy (AFM) is a powerful, multifunctional imaging platform that allows biological samples, from single molecules to living cells, to be visualized and manipulated. Soon after the instrument was invented, it was recognized that in order to maximize the opportunities of AFM imaging in biology, various technological developments would be required to address certain limitations of the method. This has led to the creation of a range of new imaging modes, which continue to push the capabilities of the technique today. Here, we review the basic principles, advantages and limitations of the most common AFM bioimaging modes, including the popular contact and dynamic modes, as well as recently developed modes such as multiparametric, molecular recognition, multifrequency and high-speed imaging. For each of these modes, we discuss recent experiments that highlight their unique capabilities.
649 citations
TL;DR: This review discusses single-molecule experiments (SMEs) in biological physics from an experimental perspective, first exposing the most common experimental methodologies and later presenting various molecular systems where such techniques have been applied.
Abstract: I review single-molecule experiments (SMEs) in biological physics. Recent technological developments have provided the tools to design and build scientific instruments of high enough sensitivity and precision to manipulate and visualize individual molecules and measure microscopic forces. Using SMEs it is possible to manipulate molecules one at a time and measure distributions describing molecular properties, characterize the kinetics of biomolecular reactions and detect molecular intermediates. SMEs provide additional information about thermodynamics and kinetics of biomolecular processes. This complements information obtained in traditional bulk assays. In SMEs it is also possible to measure small energies and detect large Brownian deviations in biomolecular reactions, thereby offering new methods and systems to scrutinize the basic foundations of statistical mechanics. This review is written at a very introductory level, emphasizing the importance of SMEs to scientists interested in knowing the common playground of ideas and the interdisciplinary topics accessible by these techniques. The review discusses SMEs from an experimental perspective, first exposing the most common experimental methodologies and later presenting various molecular systems where such techniques have been applied. I briefly discuss experimental techniques such as atomic-force microscopy (AFM), laser optical tweezers (LOTs), magnetic tweezers (MTs), biomembrane force probes (BFPs) and single-molecule fluorescence (SMF). I then present several applications of SME to the study of nucleic acids (DNA, RNA and DNA condensation) and proteins (protein-protein interactions, protein folding and molecular motors). Finally, I discuss applications of SMEs to the study of the nonequilibrium thermodynamics of small systems and the experimental verification of fluctuation theorems. I conclude with a discussion of open questions and future perspectives.
386 citations
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TL;DR: The atomic force microscope as mentioned in this paper is a combination of the principles of the scanning tunneling microscope and the stylus profilometer, which was proposed as a method to measure forces as small as 10-18 N. As one application for this concept, they introduce a new type of microscope capable of investigating surfaces of insulators on an atomic scale.
Abstract: The scanning tunneling microscope is proposed as a method to measure forces as small as 10-18 N. As one application for this concept, we introduce a new type of microscope capable of investigating surfaces of insulators on an atomic scale. The atomic force microscope is a combination of the principles of the scanning tunneling microscope and the stylus profilometer. It incorporates a probe that does not damage the surface. Our preliminary results in air demonstrate a lateral resolution of 30 A and a vertical resolution less than 1 A.
12,344 citations
TL;DR: Optical trapping of dielectric particles by a single-beam gradient force trap was demonstrated for the first reported time, confirming the concept of negative light pressure due to the gradient force.
Abstract: Optical trapping of dielectric particles by a single-beam gradient force trap was demonstrated for the first reported time. This confirms the concept of negative light pressure due to the gradient force. Trapping was observed over the entire range of particle size from 10 μm to ~25 nm in water. Use of the new trap extends the size range of macroscopic particles accessible to optical trapping and manipulation well into the Rayleigh size regime. Application of this trapping principle to atom trapping is considered.
6,434 citations
TL;DR: It is demonstrated that using protein constructs with identical ectodomains and different membrane regions and vice versa provides the viscous damping of the membrane domain in the lipid bilayer to probe the dynamics and size of lipid rafts in the membrane of living cells.
Abstract: To probe the dynamics and size of lipid rafts in the membrane of living cells, the local diffusion of single membrane proteins was measured. A laser trap was used to confine the motion of a bead bound to a raft protein to a small area (diam ≤ 100 nm) and to measure its local diffusion by high resolution single particle tracking. Using protein constructs with identical ectodomains and different membrane regions and vice versa, we demonstrate that this method provides the viscous damping of the membrane domain in the lipid bilayer. When glycosylphosphatidylinositol (GPI) -anchored and transmembrane proteins are raft-associated, their diffusion becomes independent of the type of membrane anchor and is significantly reduced compared with that of nonraft transmembrane proteins. Cholesterol depletion accelerates the diffusion of raft-associated proteins for transmembrane raft proteins to the level of transmembrane nonraft proteins and for GPI-anchored proteins even further. Raft-associated GPI-anchored proteins were never observed to dissociate from the raft within the measurement intervals of up to 10 min. The measurements agree with lipid rafts being cholesterol-stabilized complexes of 26 ± 13 nm in size diffusing as one entity for minutes.
997 citations
TL;DR: A force detection scheme that makes use of a modified cantilever beam and senses the force gradient through frequency modulation is described, achieving high resolution under ultrahigh-vacuum conditions with the force microscope for reactive surfaces.
Abstract: Achieving high resolution under ultrahigh-vacuum conditions with the force microscope can be difficult for reactive surfaces, where the interaction forces between the tip and the samples can be relatively large. A force detection scheme that makes use of a modified cantilever beam and senses the force gradient through frequency modulation is described. The reconstructed silicon (111)-(7x7) surface was imaged in a noncontact mode by force microscopy with atomic resolution (6 angstroms lateral, 0.1 angstrom vertical).
994 citations
TL;DR: In this paper, a high-speed scanner, free of resonant vibrations up to 60 kHz, small cantilevers with high resonance frequencies (450-650 kHz) and small spring constants (150-280 pN/nm), and several electronic devices of wide bandwidth are presented.
Abstract: The atomic force microscope (AFM) is a powerful tool for imaging individual biological molecules attached to a substrate and placed in aqueous solution. At present, however, it is limited by the speed at which it can successively record highly resolved images. We sought to increase markedly the scan speed of the AFM, so that in the future it can be used to study the dynamic behavior of biomolecules. For this purpose, we have developed a high-speed scanner, free of resonant vibrations up to 60 kHz, small cantilevers with high resonance frequencies (450–650 kHz) and small spring constants (150–280 pN/nm), an objective-lens type of deflection detection device, and several electronic devices of wide bandwidth. Integration of these various devices has produced an AFM that can capture a 100 × 100 pixel2 image within 80 ms and therefore can generate a movie consisting of many successive images (80-ms intervals) of a sample in aqueous solution. This is demonstrated by imaging myosin V molecules moving on mica (see http://www.s.kanazawa-u.ac.jp/phys/biophys/bmv_movie.htm).
958 citations