Immunocytochemical detection of androgen receptor in human temporal cortex: Characterization and application of polyclonal androgen receptor antibodies in frozen and paraffin-embedded tissues

Abstract

Immunocytochemical and biochemical studies have demonstrated the presence of androgen receptor protein in various regions of the rodent and non-human primate cortex. Localization of androgen receptor in the human brain has, however, not been studied as extensively, because of difficulties in obtaining suitable tissue samples. In the present study, we have localized androgen receptors in both frozen and paraffin-embedded temporal cortex from epileptic patients undergoing resection. Polyclonal antibodies were raised against fusion proteins containing fragments of the human androgen receptor protein. The antibodies were affinity-purified against the corresponding fusion protein. Immunoprecipitation and Western blotting using extracts from human cell lines demonstrated the specificity of the antibodies for the human androgen receptor and lack of cross-reactivity with other steroid hormone receptors. Immunocytochemistry was performed on frozen and paraffin sections of human temporal cortex and in paraffin-embedded benign hyperplastic prostates (BPH), as well as prostate and breast carcinomas, by the streptavidin-biotin-peroxidase method. Antigen-retrieval was performed in paraffin-embedded sections using microwave irradiation. Specific nuclear and cytoplasmic immunoreactivity for androgen receptor was detected in neurons, astrocytes, oligodendrocytes, and microglia cells of the temporal cortex. In contrast, only nuclear staining was observed in BPH, prostate and breast carcinomas. Immunoprecipitation of human temporal cortex lysate and subsequent Western blot analysis demonstrated the expression of a 98 kDa immunoreactive protein, slightly smaller than the reported molecular weight of the wild-type androgen receptor. These results provide further evidence for the expression of androgen receptor in the human temporal cortex. The use of these immunocytochemical techniques should enable the retrospective determination of possible changes in androgen receptor expression in a variety of archival paraffin-embedded tissues, including samples of the human central nervous system.