Scanning Electron Microscopy


The assessment of in vitro osteoclastic activity has, until recently, been dependent on the analysis of organ culture experiments. We have developed a single cell resorption assay so that the resorptive function of individual osteoclasts could be studied. This paper examines the biological variation in the sizes of resorption lacunae produced by bone cell cultures derived from neonate rats and rabbits, and prehatch or hatchling chicks. Cultures were run for 24h for all species; and in addition for 48h for rat, 9 or 12 hours for rabbit and 3-7 hours for chick. The numbers of the nuclei of osteoclasts seeded on to plastic were counted for all three species. SEM stereophotogrammetry was used to measure areas, volumes, and maximum and average depths of the lacunae using specially designed instruments and software. Rat osteoclasts were smallest, and more chick osteoclasts were very large. There was a species difference in the onset of resorption and the sizes of pits produced, the chick osteoclasts being more vigorous resorbers than the rabbit ones, and the rat least so. For a given plan area, chick lacunae were deeper. There was a high correlation between area and volume. The range of maximum depths for a given area was high, however. Thus the mean of a few measurements of depths should not be used to calculate volume from area. At 24 hours, 77% of the rat, 47% of the rabbit and 28% of the chick lacunae were less than 1,000 μm3 in volume; and 11% of the rat, 17% of the rabbit and 22% of the chick lacunae were between 1,000 and 2,000μm in volume. The mean values at 24 hours were 981, 2796, and 4582 μm3 for rat, rabbit and chick lacunae respectively.

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