Early studies of the mechanisms of biological calcification were hampered by confusion over the state of saturation of the body fluids with respect to calcium and inorganic phosphate. This early literature has been reviewed (1, 2) and, with newer evidence, seems more and more to support the view that the body fluids contain more than sufficient concentrations of calcium and phosphate to produce the mineral crystals of bone (3, 4). The problem of controlling the mineralization process now

more »

... to involve the induction by or prevention of the formation of effective crystal-nuclei or seeds (4, 5). There has been accumulating (4-8) both direct and indirect evidence which indicts collagen as the inducer or nucleating substance. The preventer or inhibitor of mineralization has been postulated (2, 9) but never demonstrated. In the present study, the induction of crystal formation by demineralized dentin has been investigated in vitro with inorganic solutions and simulated ultrafiltrate of serum. The reasons for employing dentin in preference to bone as a source material for collagenous inducer will be explored fully in the discussion. The results obtained lend additional support for the nucleation concept and, further, suggest that steric properties (associated with e-amino groups) are responsible for the specific ability of collagen to induce crystal formation. EXPERIMENTAL PROCEDURE Preparation of Collagenous Tissue Residues-The roots of sound human and bovine teeth, previously stored at 4", were filed free of cementum, and then split into small pieces (approximately 3 mm) by means of pliers. The pulpal material was removed and the dentin fragments were soaked overnight in 0.9% sodium chloride solution at 4". The dentin (1 g) was washed with distilled water and then demineralized by soaking in daily changes (200 ml) of freshly prepared 0.5 M potassium ethylenediaminetetraacetate at pH 7.4 for a period of 6 to 8 days at 4". The demineralized dentin was then washed with 6 changes of cold KC1 (p = 0.16) for 24 hours, rinsed with water, and after drying by pressing on filter paper, was immediately placed in the mineralizing solutions. Portions of the demineralized dentin were tested for the presence of residual mineral material by