Light and electron microscopical combined staining by immunohistochemical and enzyme histochemical methods using rat...

Combined immunohistochemical staining (IHCS) and enzyme histochemical staining (EHCS) methods for light microscopy (LM) and electron microscopy (EM) are reported, using oestrogen-induced rat pituitary tumours. For LM, combined staining for alkaline phosphatase and acid phosphatase by EHCS, using the azo dye method, and for prolactin and ACTH by IHCS, using the enzyme-labelled antibody method, gave the best results on 1 microgram glycol methacrylate sections. For EM, combined staining by EHCS on 30 microgram tissue sections followed by IHCS for prolactin on ultrathin Epon sections (enzyme-labelled antibody method) provided acceptable results. By these combined staining methods, the neoplastic prolactin cells were shown to have close affinity to rich alkaline phosphatase-positive capillaries and to possess an alkaline phosphatase-positive cell membrane. Furthermore, they revealed acid phosphatase-positive lysosomal and secretory granules. These combined staining methods may be valuable in studies on the actual functional status of cells.