The ultrastructure of slam-frozen bone mineral
Abstract
Bone salt may be altered by preparative procedures. 'Slam' freezing can usefully be applied to bone mineral because it minimizes preparation and preserves the tissue chemistry. The structure and composition of the mineral in 'slam'-frozen neonatal mouse calvaria (which require neither previous slicing nor manipulation) was examined by transmission electron microscopy and X-ray microanalysis in unstained sections, 0.25 micron thick (i.e. unusually thick). Comparison was made with fresh intact calvaria and with 'snap'-frozen histological sections of mature rat femora. Under the optical microscope, calcified microspheres, up to 1 micron in diameter, were evident within 'young' osteocytes and within the extracellular matrix of both immature and mature, unstained and von Kossa-stained bone. In the electron microscope, microspheres of similar dimension and distribution were observed after 'slam' freezing and were divided into two groups. One group, found inside and outside cells, had a substructure of closely packed, electron-dense, rounded bodies 30-40 nm in diameter; despite their unusual stability in EDTA, X-ray microanalysis indicated high levels of both calcium and phosphorus. The other group was found at the calcification front and, although similar to the first group in size and chemical composition, these microspheres had a substructure of clusters of 5-nm-thick electron-dense filaments containing mineral that was characteristically EDTA labile. The 30- to 40-nm dense…
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