Characterization of various antibodies against tissue plasminogen activator using highly sensitive enzyme immunoassay

Polyclonal and three monoclonal anti-tissue plasminogen activator (a-t-PA) antibodies (1:3 C5, 1:3 G5 and 2:2 B10) were characterized by using enzyme immunoassay (EIA) in which beta-D-galactosidase was coupled to a-t-PA antibody. Monoclonal antibodies called 2:2 B10 and 1:3 G5, specific for both one-chain and two-chain t-PA, strongly bound to one-chain t-PA purified from cultured melanoma cell lines, but 1:3 C5 antibody bound weakly to such t-PA. When polyclonal t-PA antibody was used as the first reaction antibody immobilized on silicone pieces, few determinants were available for monoclonal antibody used in the second reaction due to previous interaction of these determinants in the first reaction. When t-PA levels in the plasma were determined, the presence of EDTA enhanced the sensitivity of t-PA determination by the present EIA technique. Plasma concentrations of t-PA were measured to be higher with 2:2 B10 monoclonal antibody than with polyclonal antibody as the first antibody. Tissue-PA was mainly detected in the endothelial cells, but not in the muscular layer of the inferior mesenteric artery when immunochemical technique was used with polyclonal t-PA antibody.