Shannon J M, Cunha G R, Taguchi O, Vanderslice K D, Gould S F
J Histochem Cytochem. 1982 Oct;30(10):1059-65. doi: 10.1177/30.10.7130669.
A procedure for the rapid preparation of autoradiograms from tissues incubated in vitro with 3H-estradiol is described. Slices of tissue were incubated in culture medium containing 17 nM 3H-estradiol, washed to remove unbound steroid, then processed for thaw-mount autoradiography. Exposure times were generally 3 to 4 weeks. Simultaneous in vitro competition with unlabeled progesterone, dihydrotestosterone, or hydrocortisone had no effect on the distribution or intensity of exposed silver grains, while competition with unlabeled estradiol or moxestrol abolished nuclear localization of silver grains. The exchange of labeled estradiol for bound endogenous estradiol during in vitro incubation of the tissues was demonstrated by a comparison of the pattern of incorporation of 3H-estradiol in tissues previously treated in vivo with unlabeled estradiol versus those not primed. A similar distribution and intensity of silver grains was observed in both the treated and untreated tissue groups. The rationale for the advantages of in vitro steroid autoradiography versus the in vivo technique is discussed.
本文描述了一种从体外与³H-雌二醇孵育的组织中快速制备放射自显影片的方法。将组织切片在含有17 nM³H-雌二醇的培养基中孵育，洗涤以去除未结合的类固醇，然后进行冻融放射自显影处理。曝光时间通常为3至4周。与未标记的孕酮、二氢睾酮或氢化可的松同时进行体外竞争，对曝光银颗粒的分布或强度没有影响，而与未标记的雌二醇或莫昔芬竞争则消除了银颗粒的核定位。通过比较³H-雌二醇在体内预先用未标记雌二醇处理的组织与未预先处理的组织中的掺入模式，证明了在组织体外孵育期间标记雌二醇与结合的内源性雌二醇的交换。在处理组和未处理组组织中观察到相似的银颗粒分布和强度。讨论了体外类固醇放射自显影相对于体内技术优势的基本原理。
