A rapid radioimmunoassay for rat plasma kininogen

The assay of plasma kininogen most often relies on bioassay techniques that measure biological effects of vasopeptides formed by proteolytic action on the substrate. These assays are tedious, time consuming, subject to the variations in tissue response and expensive. In the present report a sensitive and rapid radioimmunoassay for rat plasma kininogen is described. Polyclonal antibodies of high avidity and specificity were produced in rabbits immunized with purified rat plasma kininogen. Optimal conditions for each step of the assay were established from a series of initial experiments. Fifty percent (50%) precipitation of 125I-kininogen was obtained with a 1:100,000 dilution of antirat plasma kininogen antibody. Precipitation of the antigen-antibody complex was achieved with 10% polyethylene glycol in 0.05 M phosphate buffer, pH 7.5. Since precipitation with polyethylene glycol requires no incubation, this procedure eliminates lengthy waiting periods and the need for a second antibody. The assay can be completed within one day. A standard curve constructed by plotting the fraction of bound 125I-kininogen against the plasma kininogen in a logit-log mode yielded a straight line for kininogen concentrations between 0.1 and 30 ng. The lowest limit of detection of the rat plasma kininogen was 0.1 ng.