Quantitation of extracts containing tumour angiogenesis factor (TAF) by radioimmunometric and radioimmunoassays.

Abstract

An antiserum which is able to inhibit TAF-induced neovascularization in vivo (TAF antiserum) was used to develop two quantitative assays for TAF-containing tumour extracts (tumor TAF). 1) Radioimmunometric assay (RIMA): the IgG of the TAF antiserum was labelled with 125I. An excess of 125I-IgG was incubated with increasing concentrations of tumour TAF and the antigen-bound fraction was precipitated by addition of Clq. 2) Radioimmunoassay (RIA): an excess of iodinated antigen (tumour TAF) was incubated with TAF antiserum diluted so that binding in the absence of unlabelled antigen represented 70-80% of the maximum binding. When tumour TAF was added, competition between labelled and unlabelled antigen for the TAF antibody binding sites resulted in displacement of the former by increasing concentrations of the latter. A second antibody was used to precipitate the bound labelled antigen. Of the two assays, the RIMA was the more sensitive and, due to the lack of a purified antigen, allowed standardization of the results in a more accurate manner. Our data show that there was a good correlation between the ability of tumour TAF to induce angiogenesis in vivo and the degree of antiserum binding detected in vitro by both assays. Preparations containing TAF, whatever the source (i.e. human or animal tumours or tissue culture) shared common antigenic determinants. It is suggested that the quantitative assays should prove valuable in determining the clinical relevance of TAF as a tumour marker.