Mouse monoclonal antibodies were derived against a plasminogen activator inhibitor
with a mol.wt. of ∼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 ∼50-fold to homogeneity
from conditioned cell culture fluid with a yield of ∼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.
Key words
Plasminogen activator inhibitor - Monoclonal antibody - Fibrosarcoma cells - Inhibitor
neutralization - Purification
