Development and characterization of a radioimmunoassay to measure human tissue kallikrein in biological fluids.

A direct radioimmunoassay has been developed to measure tissue kallikrein in human biological fluids. These fluids include serum, plasma, urine, pancreatic juice and saliva. Purified kallikreins from human urine and human saliva were used to raise rabbit antibody and each was labelled with Na125I for use in the radioimmunoassay. A comparison of the different antigen-antibody systems was then made. Bound and free enzyme were separated by a double-antibody technique. The usable range of the standard curve was from 2.5 to 100 micrograms kallikrein/l. The intra-assay coefficient of variation was 4.7%, the interassay coefficient of variation 8.9% and the recoveries of purified kallikrein added to the samples were 99.3, 96.0, 110.8 and 81.2% for urine, saliva, serum and plasma respectively. Parallel dilution curves were obtained for serum and plasma, as well as for urine, saliva and pancreatic juice. However, plasma anticoagulated with EDTA or heparin gave consistently lower values than serum, when measured in the radioimmunoassay. From eight different subjects plasma (EDTA) values were on average 50% lower than those of serum. Experiments designed to determine the cause of this difference revealed that treatment of blood with some anticoagulants, in particular heparin and EDTA, resulted in a marked reduction in the amount of measurable tissue kallikrein.