Dodd I, Jalalpour S, Southwick W, Newsome P, Browne M J, Robinson J H
FEBS Lett. 1986 Dec 1;209(1):13-7. doi: 10.1016/0014-5793(86)81075-4.
Recombinant tissue-type plasminogen activator (rt-PA) from cultures of a genetically manipulated Bowes melanoma cell line (TRBM6) was purified in batches of average volume 451 using an autoclavable, reusable, continuous chromatography system comprising zinc chelate-Sepharose CL4B and lysine-Sepharose CL4B. After eight successive purifications the rt-PA was ultrafiltered to yield a preparation containing 4.9 mg protein/ml and 2.7 X 10(6) IU/ml. Analysis by SDS-polyacrylamide gel electrophoresis followed by staining with Coomassie brilliant blue R250 showed major protein bands at Mr = 63,000 and 65,000; most of the material was in the 1-chain form. The potential usefulness of a simple, rapid continuous chromatography system that can be operated under aseptic conditions is discussed.
利用包含锌螯合琼脂糖凝胶CL4B和赖氨酸琼脂糖凝胶CL4B的可高压灭菌、可重复使用的连续色谱系统，对来自基因工程改造的鲍伊斯黑色素瘤细胞系（TRBM6）培养物中的重组组织型纤溶酶原激活剂（rt-PA）进行分批纯化，平均每批体积为451。经过连续八次纯化后，对rt-PA进行超滤，得到一种制剂，其蛋白质含量为4.9 mg/ml，活性为2.7×1⁰⁶ IU/ml。通过SDS-聚丙烯酰胺凝胶电泳分析，随后用考马斯亮蓝R250染色，结果显示主要蛋白条带位于Mr = 63,000和65,000处；大部分物质为单链形式。本文讨论了一种可在无菌条件下操作的简单、快速连续色谱系统的潜在实用性。
