Immunocytochemical identification of Type I collagen in acid-etched dentin

The hypothesis tested in this study is that the application of phosphoric acid prevents collagen fibrils (CF) from maintaining their structural morphology, as assessed by their immunochemical antigenicity. Dentin was conditioned with EDTA and with 35% H₃PO₄ for 15, 30, and 60 s. For a control there was no treatment. A pre-embedding immunohistochemical procedure was evaluated under high-resolution SEM and a postembedding immunolocalization technique was performed on ultrathin sections at TEM level. Conditioning with EDTA resulted in a weaker labeling signal than for dentin conditioned with H₃PO₄. The most intense labeling was observed after dentin had been etched with H₃PO₄ for 15 s. The least intensive labeling was evident when the acid was applied for 30 s or for 60 s. A very weak signal was detected on the untreated dentin surface. These results provide evidence that a 15-s application of 35% H ₃PO₄ causes mineral dissolution of the crystals enveloping the superficial CF without damaging the ultrastructure of the CF while longer applications of 35% H₃PO₄ cause alterations in the CF that decrease their antigenicity.