Yamashita S, Korach K S
Laboratory of Reproductive and Developmental Toxicology, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709.
Histochemistry. 1989;90(5):325-30. doi: 10.1007/BF00508308.
A horseradish peroxidase (HRP) labeled antibody method was developed for use with a monoclonal antibody to detect estrogen receptor (ER) in mouse tissue. Combined use of HRP labeled F(ab')2 fragment absorbed with mouse liver protein to minimize background staining and imidazol-DAB reaction gave the most reliable and sensitive immunostaining. The method was applied to uterine, vaginal, pituitary and liver tissues in ovariectomized adult mice. In uterus and vagina, ER was recognized in nuclei of epithelial cells, stromal cells and smooth muscle cells of the muscle layer and blood vessels. Liver tissue showed positive nuclear immunostaining in parenchymal cells; however, no reaction was present in endothelial cells, Kupffer cells, bile ductal cells, and smooth muscle cells of blood vessels. ER was localized in the nuclei of anterior pituitary cells while weak reaction was also recognized in cells of the intermediate lobe. No staining was detected in the posterior pituitary. Results demonstrate that both occupied and unoccupied ER are localized in the cell nucleus from several target tissues. Weak immunostaining in samples could not be enhanced by multiple procedures. It is suggested that nuclear ER is partially hidden by nuclear components such as nucleic acid and chromatin proteins.
开发了一种辣根过氧化物酶（HRP）标记抗体方法，用于与单克隆抗体结合以检测小鼠组织中的雌激素受体（ER）。将HRP标记的F(ab')2片段与小鼠肝蛋白吸收相结合以尽量减少背景染色，并使用咪唑-二氨基联苯胺反应，可得到最可靠和最灵敏的免疫染色。该方法应用于去卵巢成年小鼠的子宫、阴道、垂体和肝脏组织。在子宫和阴道中，在上皮细胞、基质细胞以及肌层和血管的平滑肌细胞的细胞核中可识别出ER。肝脏组织在实质细胞中显示出阳性核免疫染色；然而，在内皮细胞、库普弗细胞、胆管细胞和血管平滑肌细胞中未出现反应。ER定位于垂体前叶细胞的细胞核中，而在垂体中叶细胞中也可识别出弱阳性反应。在垂体后叶未检测到染色。结果表明，已占据和未占据的ER均定位于几种靶组织的细胞核中。多次操作无法增强样本中的弱阳性免疫染色。提示核ER部分被核酸和染色质蛋白等核成分所掩盖。
