Effects of various agents on staining of the maturation pattern at the surface of rat incisor enamel.

In the maturation zone, two types of ameloblasts are arranged as bands across the rat incisor; these corresponded with a staining pattern at the surface which reflected the alternating pattern of ruffle-ended and smooth-ended ameloblasts. PAS stain showed bands and stripes similar to those following glyoxal bis(2-hydroxyanil) (GBHA) staining; these stains visualize the organic (PAS) and inorganic (GBHA) components of the maturation pattern. To further elucidate the nature of these bands, dissected rat incisors were treated with various agents prior to staining with GBHA or PAS. Guanidine for 2 h at room temperature showed no maturation pattern when stained with GBHA, as did teeth treated with EDTA and a bisophosphonate (HEBP). Hydrochloric acid and nitric acid removed the layer of outer enamel and incisors did not stain with GBHA, suggesting that the staining is a surface-related phenomenon. As staining was abolished by either the removal of mineral (EDTA) or protein (guanidine), the concurrent localization of non-crystal-bound calcium by GBHA staining, and of glycoprotein by PAS staining, indicates that calcium is associated with glycoprotein at the surface of the enamel.