Stewart D R, Henzel W J, Vandlen R
Dept. OB/GYN, U.C. Davis, California 95616.
J Protein Chem. 1992 Jun;11(3):247-53. doi: 10.1007/BF01024863.
Relaxin immunological activity has been observed in the plasma of pregnant bitches, and preliminary studies in our laboratory indicated that the highest relaxin concentrations were found in placentas. Therefore, canine placentas were collected at term and also from spay and relaxin was purified by methods developed for equine relaxin. Tissue was prepared by homogenization and purification on a C18 column. The preparation was further purified by stepwise elution ion-exchange chromatography, gel filtration, and gradient elution ion-exchange chromatography. One predominant peak in relaxin immunoactivity was collected. Canine relaxin was found to be larger than either porcine or equine relaxin as determined by SDS-PAGE. It migrated faster under reducing conditions, indicating a subunit structure. Purified canine relaxin was used for tracer and standard in a canine radioimmunoassay (RIA) using an antiporcine relaxin antibody. Concentrations of relaxin immunoactivity using the canine assay were up to 300-fold higher in placental preparations than those measured in the porcine relaxin assay. Sequence analysis of canine relaxin revealed a structure similar to other relaxins in the presence and placement of cystine residues.
在怀孕母犬的血浆中观察到松弛素免疫活性，并且我们实验室的初步研究表明，胎盘组织中松弛素的浓度最高。因此，在足月时收集犬胎盘，也从绝育母犬收集胎盘，并通过为马松弛素开发的方法纯化松弛素。通过匀浆和在C18柱上纯化来制备组织。通过分步洗脱离子交换色谱、凝胶过滤和梯度洗脱离子交换色谱对制备物进一步纯化。收集到一个松弛素免疫活性的主要峰。通过SDS-PAGE测定发现犬松弛素比猪或马松弛素更大。在还原条件下它迁移得更快，表明其具有亚基结构。使用抗猪松弛素抗体，将纯化的犬松弛素用作犬放射免疫测定（RIA）中的示踪剂和标准品。使用犬检测法测得的胎盘制剂中松弛素免疫活性浓度比用猪松弛素检测法测得的浓度高300倍。犬松弛素的序列分析显示，在胱氨酸残基的存在和位置方面，其结构与其他松弛素相似。
