Wu D H, Tavoni A, Garzelli C, Neri R, Vitali C, Bombardieri S
Cattedra di Immunologia Clinica, Istituto di Patologia Medica 1, Pisa, Italy.
J Immunol Methods. 1989 Jul 26;121(2):219-24. doi: 10.1016/0022-1759(89)90163-4.
In the present study, Ro/SS-A antigen has been isolated from human spleen by a two-step procedure. In the first step most of the non-antigenic material was removed by means of ammonium sulphate precipitation and ion exchange chromatography. The final purification was obtained by passing the Ro/SS-A-containing fractions twice through a Mono Q ion exchange fast protein liquid chromatography (FPLC) column. The purified antigen showed identical immunoreactivity with crude material on CIE and was composed of two polypeptides with a molecular weight of approximately 60,000 and 55,000 respectively on SDS-PAGE, both reacting on Western blotting with a panel of anti-Ro/SS-A antisera. This system permits milligrams of highly purified antigen to be obtained from grams of human spleen.
在本研究中，通过两步法从人脾脏中分离出Ro/SS-A抗原。第一步，通过硫酸铵沉淀和离子交换色谱法去除大部分非抗原性物质。通过将含Ro/SS-A的组分两次通过Mono Q离子交换快速蛋白质液相色谱（FPLC）柱进行最终纯化。纯化后的抗原在免疫双扩散试验中与粗制品显示出相同的免疫反应性，在十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）上由两条分别具有约60,000和55,000分子量的多肽组成，两者在蛋白质印迹法中均与一组抗Ro/SS-A抗血清发生反应。该系统可从克级的人脾脏中获得毫克级的高度纯化抗原。
