Visualisation of methacrylate-embedded human bone sections by infrared nanoscopy

A recently developed ultra-resolving near-field infrared nanoscope is applied to investigate methyl methacrylate embedded, un-decalcified human bone sections. Results show detail at a resolution of 30 nm. Specific contrasting of mineral components is enabled by choosing an appropriate infrared wavelength, here 9.47 μm, in the phosphate vibrational band. The method is surface-sensitive, probing to a depth of about 30 nm into the surface. The obtained infrared images are presented in direct comparison with optical and electron micrographs of the identical specimen. Lamellar bone organization, peri-cellular mineral deposition, and regional differences in mineral content are clearly detectable. Individual fibrils are resolved. - Infrared nanoscopy requires just standard hard tissue preparation techniques combined with section surface polishing. It can be integrated into existing laboratory environments without impeding subsequent routine staining and evaluation methods.