Einarsson M, Brandt J, Kaplan L
Biochim Biophys Acta. 1985 Jul 18;830(1):1-10. doi: 10.1016/0167-4838(85)90123-2.
Tissue plasminogen activator produced by a human melanoma cell line (Bowes), was purified from large volumes of supernatant fluid using immunosorbent chromatography on monoclonal antibodies, followed by chromatography on lysine-Sepharose 4B and gel filtration on Sephadex G-150. Immunosorbents based on monoclonal antibodies were shown to be preferable to those based on polyclonal antibodies for several reasons: efficient binding and desorption, high yield along with a high degree of purification. The overall recovery was about 50%. A homogeneous sample of single-chain tissue plasminogen activator with a molecular weight of approx. 65 000 was obtained as judged by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate. Amino acid sequence analysis revealed two N-terminals in equal yields, corresponding to the long and short tissue plasminogen activator structures previously reported by others. The pure preparations of tissue plasminogen activator showed specific activities of about 200 000 IU/mg protein.
从人黑色素瘤细胞系（鲍伊斯细胞系）产生的组织型纤溶酶原激活剂，是通过以下步骤从大量上清液中纯化得到的：先用单克隆抗体进行免疫吸附色谱，然后用赖氨酸-琼脂糖4B进行色谱分离，最后用葡聚糖凝胶G-150进行凝胶过滤。基于单克隆抗体的免疫吸附剂由于几个原因而被证明比基于多克隆抗体的免疫吸附剂更可取：结合和解吸效率高、产量高且纯化程度高。总体回收率约为50%。通过十二烷基硫酸钠存在下的聚丙烯酰胺凝胶电泳判断，获得了分子量约为65000的单链组织型纤溶酶原激活剂的均一样品。氨基酸序列分析显示有两个N末端，产量相等，对应于其他人先前报道的长型和短型组织型纤溶酶原激活剂结构。组织型纤溶酶原激活剂的纯制剂显示出约200000 IU/mg蛋白质的比活性。
