Lax E R, Kreuzfelder E, Schriefers H
Hoppe Seylers Z Physiol Chem. 1979 Jul;360(7):897-904. doi: 10.1515/bchm2.1979.360.2.897.
This paper described a modified method for the radiometric determination of hydroxysteroid dehydrogenase activities in rat liver. The principle advantages of this method are the improved precision and a radical reduction in the time involved in performing the assay. The procedure comprises the following steps: incubation of 14C-labelled substrate with coenzyme and cell fraction under optimized conditions; termination of the reaction by addition of organic solvent containing a defined amount of 3H-labelled reaction product; removal of precipitated protein and coenzyme by centrifugation; paper chromatographic isolation of the product; direct quantitation of 14C activity in the product zone of the paper chromatogram. The assay systems have been applied to elucidate and quantitate sex and strain differences in the activities of the above enzymes in Chbb/THOM and Sprague-Dawley rats.
本文描述了一种改良方法，用于放射性测定大鼠肝脏中羟类固醇脱氢酶的活性。该方法的主要优点是提高了精度，并大幅减少了测定所需的时间。该程序包括以下步骤：在优化条件下，将14C标记的底物与辅酶和细胞组分一起孵育；通过加入含有一定量3H标记反应产物的有机溶剂终止反应；通过离心去除沉淀的蛋白质和辅酶；用纸色谱法分离产物；直接定量纸色谱图产物区中的14C活性。该测定系统已用于阐明和定量Chbb/THOM和Sprague-Dawley大鼠中上述酶活性的性别和品系差异。
