A cryoSEM Method for Preservation and Visualization of Calcified Shark Cartilage (And Other Stubborn Heterogeneous Skeletal Tissues)

Biological materials interest biologists and engineers for their complex interactions among constituents and unique mechanical properties. While the heterogeneous tissue structure and its material properties are responsible for intriguing biomechanics, they pose challenges for sectioning, particularly in regions with stark tissue boundaries. Microscopists level this playing field when sectioning samples by embedding them in paraffin or plastic; but for scanning electron microscopy, natural morphology must be preserved without sacrificing the sample’s surface contours. Here we outline a simple preparation method for visualizing in cryoSEM the calcified cartilage of sharks and rays (elasmobranch fishes), a layered biocomposite that has traditionally been considered difficult to prepare for microscopy.