Biochemical Interactions through Microscopic Techniques: Structural and Molecular Characterization (original) (raw)

Invited review article: combining scanning probe microscopy with optical spectroscopy for applications in biology and materials science

The Review of scientific instruments, 2012

This is a comprehensive review of the combination of scanning probe microscopy (SPM) with various optical spectroscopies, with a particular focus on Raman spectroscopy. Efforts to combine SPM with optical spectroscopy will be described, and the technical difficulties encountered will be examined. These efforts have so far focused mainly on the development of tip-enhanced Raman spectroscopy, a powerful technique to detect and image chemical signatures with single molecule sensitivity, which will be reviewed. Beyond tip-enhanced Raman spectroscopy and/or topography measurements, combinations of SPM with optical spectroscopy have a great potential in the characterization of structure and quantitative measurements of physical properties, such as mechanical, optical, or electrical properties, in delicate biological samples and nanomaterials. The different approaches to improve the spatial resolution, the chemical sensitivity, and the accuracy of physical properties measurements will be d...

Molecular-scale studies on biopolymers using atomic force microscopy

2006

The atomic force microscope (AFM) is capable of acquiring a range of structural and physicochemical information on a wide range of biopolymers. The ability to achieve nanoscale resolution, coupled with the potential to image and manipulate realtime molecular-scale events, suggests that AFM is one of the most promising techniques available for the study of biopolymers. AFM offers the potential to obtain a wide range of both quantitative and qualitative information on biopolymers, ranging from their conformations in physiological buffers to the forces involved in bond cleavage. This review explores the most common modes of AFM operation including imaging (contact and tapping mode) and force spectroscopy. The application of these modes to biopolymer characterisation will be discussed, with an emphasis on key studies. Keywords AFM • Biopolymer • DNA • RNA • Proteins Abbreviations 3-MPA 3-Mercaptopropanoic acid 11-MUA 11-Mercaptoundecanoic acid AFM Atomic force microscope or atomic force microscopy APTES 3-Aminopropyltriethoxysilane (used to create AP-mica) AP-mica APTES-mica Bp Base pair(s) BR Bacteriarhodopsin BSA Bovine Serum Albumin BSA-MPAD BSA conjugated with a mercaptopropanoic acid derivative of atarazine CBH I An exoglucanase enzyme (a cellulase) CMA Carboxymethylamylose CM-AFM Contact mode AFM CMC Carboxymethylcellulose CNT Carbon nanotubes Cryo-AFM Cryogenic atomic force microscopy csA Glycoprotein contact site AS DMSO Dimethylsulfoxide dsDNA Double-stranded DNA dsRNA Double-stranded RNA EDC 1-Ethyl-3-(3-dimethylaminopropyl)-carbodiimide EDTA Ethylenediaminetetraacetic acid EGII Endogluconase Molecular-Scale Studies on Biopolymers Using Atomic Force Microscopy EtBr Ethidium bromide FDC Force-distance curve HPI Hexagonally packed intermediate layer Ig Immunoglobin LT Large tectosquares MoMLV Moloney murine leukaemia virus MPADT mercaptopropanoic acid derivative of atarazine NCM-AFM Noncontact mode AFM NHS N-hydroxysulfosuccinamide OT Optical tweezers PAMAM Poly(amidoamine) (a type of dendrimer) pBR322 An E.

Quantitative interrogation of micropatterned biomolecules by surface force microscopy

Ultramicroscopy, 2000

Synthetic biomaterials are widely used in medical implants with success in improving and extending quality of life. However, these materials were not originally designed to interact with cells through speci"c signaling pathways. As a result, the interaction with the body is mediated through passive adsorption of a disorganized protein monolayer. Next generation biomaterials have been proposed to be active in modifying the biological response of the host through the incorporation of speci"c biorecognition moieties. An important tool in the development of these novel active biomaterials is the scanning force microscope (SFM). The SFM allows for interrogation of bioactive biomaterials in mapping or spectroscopic modes. In this work, micropatterned protein surfaces were prepared using biomolecules implicated in wound healing. The surfaces were imaged via SFM and the speci"c binding forces between surface associated biomolecules and antibody functionalized tips were quanti"ed.

Atomic force microscopy in bionanotechnology

Nano Today, 2008

Within the field of scanning probe microscopy, atomic force microscopy 1 (AFM) is extensively used in a wide range of disciplines such as life science, solid-state physics, and materials science. The AFM has evolved into an imaging method that yields structural details of biological samples such as proteins, nucleic acids, membranes, and cells in their native environment 2-5 . AFM is a unique technique for providing subnanometer resolution at a reasonable signal-to-noise ratio under physiological conditions. As a result of continuous developments in sample preparation, imaging techniques, and instrumentation, AFM is now a companion technique to X-ray crystallography and electron microscopy (EM) for the determination of protein structures 6 , for example. It complements EM by allowing visualization of biological samples in buffers that preserve their native structure over extended time periods. AFM does not rely on symmetry averaging and crystallization, therefore revealing defects and structural anomalies not observable in classical ensemble measurements 7 .

Nanoscopy of cell architecture

BioArchitecture, 2011

I t was light microscopy that first revealed the hidden world of bacteria and the unit of life the "cell." From these first observations, made in the late 1600s, it has been clear that seeing is an important tool in biology. The merging of the fields of fluorescence and microscopy created the possibility to see subcellular structures and proteins. In the 1990s the use of the confocal microscopes, where cells/tissue could be optically sectioned, further improved the resolution of object visualization. From this microworld view we now move forward to the exciting prospects of the nanoworld view of biology. In this review I propose a nanoimaging approach, nanoscopy, which could be used to reveal cell architecture at the level of proteins and protein complexes. Nanoscopy includes, the F-techniques, superresolution microscopy, correlative light and electron microscopy and atomic force microscopy. To illustrate the biology that could be investigated by nanoscopy we focus on structures formed at the actin-membrane interface. In particular, focal adhesions and stress fibres have been analyzed using nanoscopy. Many of the proteins present in focal adhesions and stress fibres are shared with structures such as filopodia, lamellipodia, endocytic vesicles, actin pedestals and invadopodia. It is likely that nanoscopy of cells will reveal mechanistic details of biology at the level of individual proteins and protein complexes and importantly in a physiological context.

Microscopic investigations of the interaction of proteins with surfaces

Biosensors and Bioelectronics, 1994

The application of light microscopy to the study of protein-surface interactions is described with several examples showing its versatility. Extensive use is made of image analysis algorithms to extract quantitative information from the digital images and it is shown how this can shed light on the processes taking place on the surface. Amongst the surfaces investigated are siloxane films on silicon wafers, glucose oxidase/poly(ethyleneglycol) films cast onto glass slides and glucose oxidase entrapped in electropolymerised poly(phenyleneoxide) films on platinum. Parameters that can be measured include : surface loading, surface heterogeneity, distribution of enzyme activity, surface mobility and film porosity. Both reflected light and fluorescence microscopy were used to characterise the surfaces and it was concluded that not only does light microscopy offer a powerful tool for the characterisation of surfaces but it may also provide an interface between bioelectronic materials and conventional computing machinery .

Scanning transmission electron microscopy of biological macromolecules

Ultramicroscopy, 1980

The practical usefulness of a STEM (Scanning Transmission Electron Microscope) for the study of the structure of biological macromolecules has been investigated using a STEM attachment connected to a TEM (Transmission Electron Microscope), which in one case was equipped with a tungsten hairpin cathode, and in the other case with a field emission gun. The point to point resolution has been determined. Results obtained in STEM dark field from light negatively stained specimens are compared with results obtained in TEM bright field from normal negatively stained specimens. In addition unstained molecules have been visualized. Some remarks are made about preparation methods suitable for STEM.

Intercellular interaction observed by atomic force microscopy

Ultramicroscopy, 2008

The cultured myoblasts, focusing on the microprocesses related to the intercellular interaction, were observed by taking topological images. For atomic force microscopy (AFM), cells were fixed and either dried as in usual scanning electron microscopy or kept in the buffer. The dried cells were used for observing intercellular interactions related to the fusion. The prefusing myoblasts aligned in a chain were mostly spindle in shape and were characterized by the presence of many microprocesses along the facing edges of adjacent aligned myoblasts. The space between fusing myoblasts and between myotubes and myoblasts were often traversed by filopodia and cellular bridges formed by the connection of microvilli. These results suggest that microprocesses may be involved in the fusion of myoblasts. The best images of the fixed cell in liquid were obtained using the contact mode of AFM. AFM observation is an efficient tool in the study on the interaction between cells, and the fixation, imaging in liquid is a good approach to understand the cellular dynamics.