Identification of collagen fibrils in cross sections of bone by electron energy loss spectroscopy (EELS)

Transmission electron microscopic (TEM) images of ion-milled bovid cortical bone cut approximately normal to the axes of fibrils show that mineral occurs in the form of plates surrounding and laying between circular or elliptical features about 50 nm in diameter. The classification of these features as either pores or collagen fibrils is highly debated. Electron energy loss spectroscopy (EELS) mapping of these features in ion milled sections shows that they are lacking significant amounts of mineral or collagen, although their appearance suggests that they are cross sections of collagen fibrils. However, analogous sections prepared using an ultramicrotome show that, while these circular features show reduced concentrations of calcium and phosphorus, some of them contain quantities of carbon and nitrogen in bonding states comparable to the composition of collagen. This work demonstrates that the observed circular features are sections of collagen fibrils, but that bombardment by argon ions during broad beam ion milling destroys the collagen and associated gap-zone mineral.