PATTERNS OF IMMUNOREACTIVITY TO AN ANTIFIBRONECTIN POLYCLONAL ANTIBODY IN FORMALIN-FIXED, PARAFFIN-EMBEDDED ORAL-TISSUES ARE DEPENDENT ON METHODS OF ANTIGEN RETRIEVAL

The influence of antigen retrieval (AR) techniques on immunoreactivity in formalin-fixed, paraffin-embedded tissues is recognized, In focal reactive overgrowths of oral mucosa, we noted that patterns of immunoreactivity to a fibronectin polyclonal antibody were dependent on methods of AR. To establish these patterns we investigated eight pyogenic granulomas and eight Fibroepithelial polyps. In the absence of AR no immunoreactivity was observed. After a-chymotrypsin AR a band of intense immunoreactivity was associated with vascular endothelial cells, with either minimal or no staining of connective tissue, After microwave AR immunoreactivity was observed in connective tissue, especially in the subepithelial region, but there was no specific vascular staining, After autoclave AR immunoreactivity was observed in connective tissue and also in epithelial nuclei. To determine if these results represented either changes induced by tissue fixation and processing or exposure of epitopes normally masked in vivo, we investigated frozen sections from two fibroepithelial polyps and one pyogenic granuloma, In frozen sections immunoreactivity was observed around vascular endothelial cells and within connective tissue, especially in the subepithelial region, but not in epithelia. This study provides indirect evidence that alpha-chymotrypsin and heat-mediated AR protocols expose different masked epitopes, and reinforces the need to undertake appropriate pilot studies for immunohistochemical investigation of archival tissue.