ABSTRACT
These days the common ground between structural biology and molecular biology continues to grow thanks to the biomolecular insights offered by correlative microscopy, even though the vision of combining insights from different imaging tools has been around for nearly four decades. The use of correlative imaging methods to dissect the cell's internal structure is progressing faster than ever as shown by the boom in the number of methodological approaches available for correlative microscopy studies, each designed to address a specific scientific question. In this chapter, we will present a relatively straightforward approach to combining information from fluorescence microscopy and electron microscopy at the supramolecular level. The method combines live-cell and/or confocal laser microscopy with classical sample preparation for transmission electron microscopy (TEM), thereby allowing the integration of dynamic details of subcellular processes with insights about the organelles and molecular machinery involved. We illustrate the applicability of this multidimensional correlative microscopy approach on cultured Caco-2 colorectal cancer cells exposed to fluorescently labeled cisplatin, and discuss how these methods can deepen our understanding of key cellular processes, such as drug uptake and cell fate.
Subject(s)
Electron Microscope Tomography/methods , Caco-2 Cells , Cell Culture Techniques , Cisplatin/chemistry , Cisplatin/metabolism , Fixatives/chemistry , Fluorescein-5-isothiocyanate/chemistry , Fluorescein-5-isothiocyanate/metabolism , Fluorescent Dyes/chemistry , Fluorescent Dyes/metabolism , Glutaral/chemistry , Humans , Image Processing, Computer-Assisted , Microscopy, Confocal/methods , Microscopy, Fluorescence/methods , Microscopy, Interference/methods , Organelles/chemistry , Organelles/metabolism , Organelles/ultrastructure , Single-Cell Analysis/methods , Staining and Labeling , Time-Lapse Imaging , Tissue FixationABSTRACT
Correlative microscopy is the application of two or more distinct microscopy techniques to the same region of a sample, generating complementary morphological, structural and chemical information that exceeds what is possible with any single technique. As a variety of complementary microscopy approaches rather than a specific type of instrument, correlative microscopy has blossomed in recent years as researchers have recognised that it is particularly suited to address the intricate questions of the modern biological sciences. Specialised technical developments in sample preparation, imaging methods, visualisation and data analysis have also accelerated the uptake of correlative approaches. In light of these advances, this critical review takes the reader on a journey through recent developments in, and applications of, correlative microscopy, examining its impact in biomedical research and in the field of plant science. This twin emphasis gives a unique perspective into use of correlative microscopy in fields that often advance independently, and highlights the lessons that can be learned from both fields for the future of this important area of research.
Subject(s)
Cytological Techniques , Microscopy/methods , Animals , Botany/methods , Humans , PlantsABSTRACT
This work reviews recent research on the design and control of interfaces in engineering nanomaterials. Four case studies are presented that demonstrate the power of a multimodal approach to the characterization of different types of interfaces. We have used a combination of conventional, high resolution, and analytical transmission electron microscopy, microbeam electron diffraction, and three-dimensional atom probe to study polymer-clay nanocomposites, turbine rotor steels used for power generation, multicomponent aluminum alloys, and nanocrystalline magnetic materials.
Subject(s)
Alloys/chemistry , Aluminum Silicates/chemistry , Magnetics , Nanotechnology/methods , Polymers/chemistry , Aluminum , Clay , Electron Probe Microanalysis/methods , Materials Testing , Microscopy, Electron/methodsABSTRACT
The adsorption of lignosulfonate onto a commercial, modified lead zirconate titanate (PZT-PNN) powder in aqueous suspension and its effect on particle zeta potential and suspension rheology were investigated as functions of pH and lignosulfonate dosage. Langmuir analysis of the adsorption data demonstrated that a significant component of the overall driving force of adsorption at all pH values examined was specific (nonelectrostatic) bonding. Electrostatic bonding provided a significant contribution to adsorption at pH 6.0, but diminished at lower pH owing to decreased lignosulfonate ionization and at higher pH due to decreased positive surface site concentration on the PZT-PNN. The affinity of adsorption was highest at pH 6.0 because the electrostatic component was maximal at this pH. The zeta potential magnitude increased and the apparent viscosity decreased with increasing pH and increasing lignosulfonate dosage, up to approximately monolayer coverage. The lignosulfonate dosage required for monolayer coverage decreased with increasing pH owing to increasing lignosulfonate expansion and the decrease in concentration of positive surface sites on the PZT-PNN. Suspension stabilization was considered to occur by an electrosteric mechanism.
