Calcified tissue histochemistry: from microstructures to nanoparticles

It has long been recognized that histochemistry and cytochemistry offer the only ways of gathering information about the biochemical composition of tissues and cells without disrupting their microscopic architecture. A variety of methods have been put forward for studying nuclei acids, proteins, carbohydrates, lipids, enzymes and other components of intact tissues and cells. By now, many of these have only a historical interest. Some do, however, survive in microscopic and ultramicroscopic applications, and have become incorporated in the most refined and precise techniques that are currently available. Histochemical reactions range from the classic procedures carried out on histological sections to yield final stained products recognizable under the light microscope (Figure 1), to those which are applied on ultrathin sections, using heavy metals or other electron-dense compounds to reveal specific components under the electron microscope (Figure 2A); others range from procedures based on the antigen-antibody reaction that are capable of revealing the presence of specific biological molecules (Figure 2B), to the biophysical techniques which permit the qualitative and quantitative analysis of elements (Figure 3); lastly, there are the recently proposed ultra-high resolution methods that allow nanoparticles to be recognized. This brief review, which is based on personal experience and on the data in the literature, will discuss the most important methods now being used.