Application of bradykinin radioimmunoassay for the measurement of urinary kallikrein activity rats.

Bradykinin radioimmunoassay was applied for the measurement of urinary kallikrein activity in rats. Antisera against bradykinin were raised in rabbits by injecting synthetic bradykinin coupled to human serum albumin via a difluorodinitrobenzene reaction. For radioimmunoassay a high titre antibody was used at a final dilution of 1:65000 to bind approximately 8,000 cpm of [125I]-tyrosine8-bradykinin. Standard curves were plotted from synthetic bradykinin samples (1--1000 pg). Kinins were generated by incubating partially purified kininogen with rat urine in the presence of kininase inhibitors. Bradykinin antiserum showed crossreaction with kallidin, met-lys-bradykinin and dog-kininogen. To eliminate interference of the kininogen with kinin radioimmunoassay, alcohol precipitation of the substrate was performed after incubation. Concentration of urinary kallikrein in rats showed no significant difference between day and night 12-hours-colllecting period. Total kallikrein excretion at night, however, was about twofold higher, correlating to a twofold higher urine volume during the night.