Immunocytochemical detection of ornithine decarboxylase

Ornithine decarboxylase (ODC), a regulatory enzyme of polyamine biosynthesi s, is involved in cell growth and differentiation. Lack of information abou t the exact cellular and subcellular localization of ODC is one of the main obstacles to precise interpretation of the biological roles of the ODC/pol yamine system. Here we describe the development and optimization of an immu nocytochemical method to detect ODC in cells and tissues. For this purpose a monoclonal antibody (MP16-2) against a defined epitope of ODC protein was developed. Specificity of the antibody for ODC was substantiated by Wester n blotting and ELISA analysis using cell and tissue homogenates. In culture d cells, optimal staining results were obtained after fixation with crossli nking fixatives followed by permeabilization with methanol. In rat tissues, ODC immunoreactivity was best preserved in paraffin sections fixed with Bo uin's fixative. Antigen retrieval using SDS and citrate buffer substantiall y increased ODC immunostaining and decreased background staining. Localizat ion studies of ODC in different cell lines showed that strongest staining f or ODC was found in the nucleoplasm of mitotic cells, whereas confluent cel ls showed moderate perinuclear staining. Immunocytochemical studies of vari ous rat tissues showed high cytoplasmic immunostaining of ODC in epithelial cells of kidney, prostate, and adrenal medulla of testosterone-treated rat s, in glandular epithelium of small intestine, and in pancreas of neonatal and adult rats.