Cuticle matrix imaging by histochemistry, fluorescence, and electron microscopy

Biomineralized structures consist of an organic matrix and mineral constituents. Imaging of the mineralized biological tissues is demanding due to specific requirements for the preservation and visualization of chemically different constituents and due to sectioning difficulties. In this study, a characterization of the cuticular matrix of the crustacean exoskeleton was performed by a combination of microscopic methods, aiming to obtain spatial information on the matrix composition. Histochemical procedures were performed and compared in artificially decalcified and non-decalcified samples, in paraffin and resin sections. Wheat germ agglutinin (WGA) lectin-gold conjugate and a fluorescent chitin-binding probe were used to localize chitin in paraffin and resin sections of samples prepared by different fixations. Calcified regions of the matrix were determined by histochemical staining of aldehyde-fixed, methanol-fixed, and resin-embedded samples and by scanning electron microscopy with energy dispersive X-...