Large scale, rapid purification of recombinant tissue‐type plasminogen activator

Abstract

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 × 10⁶ . Analysis by SDS-polyacrylamide gel electrophoresis followed by staining with Coomassie brilliant blue R250 showed major protein bands at M _r = 63000 and 65000; 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.