Monoclonal antibodies to human 54,000 molecular weight plasminogen activator inhibitor from fibrosarcoma cells--inhibitor neutralization and one-step affinity purification.
Mouse monoclonal antibodies were derived against a plasminogen activator inhibitor with a mol. wt. of approximately 54,000 (54 K) from the human fibrosarcoma cell line HT-1080. Screening for hybrids producing antibodies directed against the inhibitor was performed with enzyme-linked immunosorbent assay and SDS-polyacrylamide gel electrophoresis (SDS-PAGE) followed by immunoblotting. Four clones of hybridomas producing IgG1 antibodies were further characterized. The inhibitor was purified approximately 50-fold to homogeneity from conditioned cell culture fluid with a yield of approximately 85% by a one-step procedure using Sepharose-conjugated monoclonal antibody. In the 125I-fibrin plate assay one of the antibodies neutralized the effect of the inhibitor on urokinase-type plasminogen activator. Two of the antibodies bound complexes between urokinase-type plasminogen activator and inhibitor while the remaining two antibodies did not. The antibodies could be used for immunocytochemical localization of the inhibitor in HT-1080 cells. All four antibodies cross-reacted with a plasminogen activator inhibitor derived from cultured human umbilical cord endothelial cells.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000302:isolation & purification; D002460:Cell Line; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D004727:Endothelium / Q000032:analysis / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D005354:Fibrosarcoma / Q000378:metabolism*; D006023:Glycoproteins / Q000276:immunology / Q000302:isolation & purification*; D006651:Histocytochemistry; D006801:Humans; D006825:Hybridomas / Q000032:analysis / Q000276:immunology / Q000378:metabolism; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000145:classification; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D009500:Neutralization Tests; D010960:Plasminogen Activators / Q000378:metabolism; D015849:Plasminogen Inactivators; D014470:Umbilical Cord / Q000032:analysis / Q000166:cytology; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
D000911:Antibodies, Monoclonal; D006023:Glycoproteins; D007074:Immunoglobulin G; D015849:Plasminogen Inactivators; D010960:Plasminogen Activators; D014568:Urokinase-Type Plasminogen Activator