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대한생식의학회지 제16권 제2호 2010년
냉동절편을 이용한 돼지 자궁내 에스트로겐 수용체의 측정
한양대학교 자연과학대학 생물학과;한양대학교 자연과학대학 생물학과;한국에너지연구소 방사션응용연구실;
윤용달;박철홍;이영근;,
A Method for the Determination of Estrogen Receptor Level in Frozen Sections of Porcine Uterus
Yoon, Yong-Dal;Park, Chor-Hong;Lee, Young-Keun;
Department of Biology, College of Natural Sciences , Hanyang University;Department of Biology, College of Natural Sciences , Hanyang University;Radiation Application Department, Korea Advanced Energy Research institute;
The present study was designed to develop a new method for the determination of estrogen receptor in porcine uterus using frozen sections. Cryostat sections were incubated with $^3H$-estradiol($^3H$-$E_2$) in the presence or absence of diethylstilbestrol(DES) and the radioactivity of 3H-E2 bound to estrogen receptor(ER) was detected. The level of specific estrogen receptor was determined by Scatchard analysis. The highest ratio of specific binding against total binding was achieved in 3 sec. tions(5mm x 5mm) which was corresponded to lOO${\mu}$/ml protein concentration. Optimal binding was obtained during incubation with $^3H$-$E_2$ for 30 minutes at 23$^{\circ}C$ after treatment of sections with acetone for 20 seconds. Three time-washing of sections was proved to be appropriate for the removal of unbound 3H-E2. 200-fold molar excess of DES was substituted for the binding of $^3H$-$E_2$ to ER sufficiently(binding efficiency of 54.8%). ER was saturated with 4nM of $^3H$-$E_2$ and its dissociation constant was 0.1nM. ER assay using frozen sections(Histological radioreceptor assay, HRRA) was significantly correlated with radioreceptor assay for estradiol(RRA, 0.976 , p
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