期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：1982
卷号：79
期号：12
页码：3720-3723
DOI：10.1073/pnas.79.12.3720
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Monoclonal antibodies against a human plasminogen activator of Mr {approx}52,000 (HPA52) were derived by immunization of mice with an impure preparation of the enzyme (urokinase), subsequent hybridization of spleen cells with NSI-Ag4/1 myeloma cells, and cloning of the hybridomas. Selection of mice for hybridization and screening of hybridomas were based solely on direct inhibition of an enzymatic assay of the plasminogen activator with the impure enzyme preparation. A cloned hybridoma produced IgG1 antibodies that bound to and inhibited the enzymatic activity of HPA52 irrespective of whether the HPA52 was derived from urokinase or from human glioblastoma cells, whereas there was no inhibition of or binding to a plasminogen activator of Mr {approx}70,000 from human melanoma cells or a plasminogen activator of Mr {approx}36,000 that is a degradation product of HPA52 and present in urokinase. Nor did the anti-HPA52 IgG1 inhibit a murine plasminogen activator of Mr {approx}48,000 derived from sarcoma virus-transformed cells. By using affinity chromatography with columns of anti-HPA52 IgG1 bound to Sepharose, HPA52 was purified from urokinase to homogeneity as evaluated by NaDodSO4/polyacrylamide gel electrophoresis. This study demonstrates that inhibitory monoclonal antibodies against enzymes can be derived with the sole use of impure enzyme preparations and shows how such antibodies subsequently can be used for enzyme purification.
