Analytical Imaging Techniques for Soft Matter Characterization

The answer to this question is quite obvious: in order to obtain the comprehensive impression about surrounding objects, one needs both of these sensation systems. Importantly, the information obtained visually and tactilely is not the same since different detection mechanisms are applied. This example provides a perception of diversity and similarity between detection mechanisms of three main high resolution microscopy techniques (transmission electron microscopy (TEM), scanning electron microscopy (SEM) and atomic force microscopy (AFM)) and human visual and tactile sensation systems.

The characterization of colloidal systems like pharmaceutical or hydrated chemical formulations by microscopic techniques is essential to obtain reliable data about the actual morphology of the system. Since the size range of colloidal drug delivery systems has long ago reached the lower end of the nanometer scale, classical light microscopy has been replaced by electron microscopy techniques which provide sufficient resolution for the visualisation of nano-sized structures. Indeed, the superior resolution and methodological versatility of electron microscopy has rendered this technique an indispensable tool for the analysis of nanoemulsions. Microscopic analysis of these lipid-based drug delivery systems with particle sizes in the lower submicron range provides critical information about the size, shape and internal structure of the emulsion droplets. Moreover, surfactant aggregates such as liposomes or multilamellar structures which remain unnoticed during particle size measurements can be detected in this fashion. This chapter provides a brief overview about both transmission electron microscopy (TEM) and scanning electron microscopy (SEM) techniques which have been employed to characterise colloidal solutions. Of special interest are sophisticated cryo techniques of sample preparation for both TEM and SEM which deliver high-quality images of pharmaceutical formulations in their natural state. An overview about the instrumentation and sample preparation for all presented methods is given. Important practical aspects, sources of error and common artefacts as well as recent methodological advances are discussed. Selected examples of electron microscopic studies of nanoemulsions are presented to illustrate the potential of this technique to reveal detailed and specific information.

At present, the description of biological ultrastructure is more closely related to the living state and is important as a complementary study of dynamic events in living cells by fluorescent light microscopy. However, the ultimate resolution of the optical microscopy is limited by a few hundred nanometres, which is by far not sufficient for the ultrastructural investigation at the level of individual macromolecule. Till now, TEM of ultrathin sections was the main technique used to address the problem, although the low electron microscopy contrast of biological samples, the necessity to use a two-dimensional projection of the sample volume and the issue of beam damage of sample strictly limit the number of topics which could be assigned to this method.

The ultrastructure of biological membrane is one of the hottest topics in cell biology. These organelle are involved in a variety of cellular processes such as cell adhesion, ion conductivity and cell signaling and serve as the attachment surface for several extracellular structures, including the cell wall, glycocalyx, and intracellular cytoskeleton. The aim of this chapter is to provide a brief overview about microscopical techniques which are suitable for the ultrastructural characterization of different kind of cellular membranes at the level of individual macromolecule. Important practical aspects concerning sample preparation, sources of error and common artefacts as well as recent methodological advances are discussed.

Biomineralization is the formation of nanostructured minerals by living cells and organisms [1]. The biological interest in the field of biomineralization is obvious-cells, extracellular matrices, transport, signaling, hormone control and many biomedical implications that have direct bearing on orthopedics, dentistry, urology etc. Materials scientists study mineralized tissues in order to gain inspiration for developing new synthetic composite materials that are based on natural systems. Paleontologists and archaeologists are interested in this field because mineralized tissues make up most of the fossil record and are also major constituents of the archaeological record of our planet [2]. The term “biomineralisation” implies that a mineral phase that is deposited requires or is occasioned by the intervention of a living organism. This can happen in two basic ways, either the mineral phase develops from the ambient environments as it would from a saturated solution of the requisite ions, but requires the living system to nucleate and localize mineral deposition, or the mineral phase is developed under the direct regulatory control of the organism, so that the mineral deposits are not only localized, but may be directed to form unique crystal habits not normally developed by a saturated solution of the requisite ions. Moreover, the shape, size and orientation of the crystals may be controlled by the cells involved. The first type of mineralization was called biologically induced “mineralization” and the second “organic matrix mediated mineralization”. Single-celled organisms and protoctista such as algae may deposit biologically-induced mineral either intra- or extra (inter)-cellularly. The majority of eukaryote matrix-mediated mineralization is extracellular. The variety of structures, as well as the diversity of minerals and macromolecules that make up mineralized tissues, is amazing [2–4].

A living cell can be described as a highly dynamical organism where different kinds of physiological processes may happen simultaneously. These processes take place at different length scales and are strongly influenced by each other. In order to circumvent the complexity and the dynamics of biological systems, so far many of the processes have been studied mostly in vitro through the use of two-dimensional molecular assemblies as model systems. Such an approach is very useful for monitoring physiological processes in vitro, but it hardly can be applied for an accurate description of the native process in a living cell or tissue, since it represents a multicomponent chemical context where each of cellular constituents can be involved in more than one process simultaneously. True investigations of the cellular dynamics in vivo are challenging and complicated but paramount at different levels from the cellular metabolism of each cell in particular until physiological processes on the macro scale like tissue formation, degeneration, or geo-mineral formation involving bio-mineralization.

The preservation of native structures of soft polymers and biological organisms during sample preparation and microscopic study is the ultimate requirement for comprehensive analyses at the level of individual macromolecules. A combination of low temperature techniques such as cryo sectioning [cryoultramicrotomy and cryo focus ion beam (FIB) milling], followed by high-resolution cryo microscopy study (cryo transmission electron microscopy, cryo TEM), has proved to be the most powerful approach available so far for ultrastructural investigation of the bulk of soft materials.

Polymeric materials are generally reinforced with inorganic fillers in order to reduce cost or to enhance mechanical, thermal, rheological and barrier properties. The fillers used have different geometrical dimensions and thus affect the polymer properties differently. For example, silica (and calcium carbonate) particles are generally spherical (aspect ratio near to 1) and enhance the strength of the polymeric materials. On the other hand, clay and graphene particles are platy in nature and enhance the mechanical, barrier and electrical properties of the polymers. Fibers or nanotubes have the highest aspect ratio and enhance the longitudinal strength as well as electrical properties of the polymer materials. Most common factor affecting the filler performance is the dispersion and distribution of the filler particles in the polymer matrix. Good dispersion and distribution of the filler particles is mandatory to achieve efficient interface between the organic inorganic components. Various microscopy techniques constitute most powerful methods to characterize the morphology of the organic–inorganic materials as described by the diverse examples presented in the following sections. The characterization of nanocomposites also poses additional challenges owing to the large number of nano-sized particles. The microscopic characterization also acts as a quality control tool as the poorly dispersed systems can be improved by changing the process parameters. Apart from characterization of the distribution and dispersion of filler particles, microscopy techniques also provide information on the alignment of the platy and fibrous particles.

In principle, this chapter is the continuation of the previous chapter where the dispersion of various types of fillers in polymer matrices was discussed. However, in this chapter, more examples pertaining to the interfacial morphology evolution in organic–inorganic composites as well as polymer blends are demonstrated. For example, in the case of nanocomposites with multiple components, specific interactions of some components with filler phase can lead to slightly different morphology at the filler matrix interface as compared to the bulk. Similarly, the interface is very dynamic in the case of multi-component blend systems and is significantly affected even when slight changes in the blend composition are made. The following sections demonstrate the characterization of these aspects associated with the interface evolution.

Both surface and volume morphology of the systems is required to be characterized as the resulting surface and bulk properties of the materials drive their applications. The control on the morphology and its tuning according to the requirement is another characteristic which is generally optimized by microscopic characterizations. These analyses can lead to vital information like surface smoothness/hardness (which in turn affects the wetting and adsorption characteristics of the surface), surface morphology (like strawberry, moon crater, hemispherical morphology etc.), particle size and its distribution, porosity of the particles, interactions between the components, defects present in the bulk of the sample, overall stability/dispersion of the filler phase in the polymer matrix, structure of the monoliths etc. The following sections demonstrate these analyses for a wide range of systems. Apart from organic and organic–inorganic systems, a brief discussion on the surface and volume characterization of inorganic particles has also been presented.

Chemical reactions are carried out on the surface of various substrates in order to generate specific functionalities or to change the surface properties of these substrates. For example, polymeric brushes are grafted on the surface of clay platelets in order to synthesize exfoliated nanocomposites. Similarly, the brushes are also grafted from the surface of polymer particles in order to generate surface properties directing their various applications. It is thus of requirement to confirm the surface reaction.

Interactions can be of different kinds, e.g. interaction of polymer chains with the surface of the filler, interactions of external species with the surface (e.g. adsorption on the surface), interactions of particles surface with stimulants like temperature, sonication, salt, solvent etc., or interactions between two inorganic species etc. (also termed as decoration of one inorganic surface with the particles of other). Such information can help to predict the behavior of the materials, their aggregation tendency etc. The characterizations of such interactions can be achieved either by analyzing the effect of such expected interactions on the morphology or by actual visualization of the morphology of the components. Though such evaluations generate information on the interaction between the components, however, the nature of interaction (chemical or physical) is not possible to be obtained. Various systems dealing with above mentioned interactions between the different components and substrates are described in the following sections.

The commercial applications of materials often involve the structuring of nanoparticles into micro or macro structures. For example, the polymer particles are generally structured to form monoliths which can then be used as chromatography columns. Similarly, inorganic nanoparticles are fused together to form macroporous networks which can be used as catalyst supports or high strength and low density metallic foams. Organic particles also form continuous films on the substrates on which they are applied or coated. Characterization of such structures for their porosity, surface roughness, uniformity as well as stability is required as these characteristics drive the applications of these networks. A number of examples describing these features are presented in the following sections.