Strategy of measuring bradykinin and kallidin and their concentration in plasma and urine.
Bradykinin (BK) and kallidin (KAL) derivatives containing a Cys residue instead of a Ser residue at positions 6 and 7, respectively [BK(Cys6), KAL(Cys7)], were synthesized. These derivatives were linked to BSA via the Cys residue by a heterobifunctional cross-linker. The coupling product containing a kinin with both free N- and C-terminal ends was used as immunogen. We obtained highly sensitive and specific antisera, simultaneously directed against both free ends. The radioimmunoassay for BK displays a sensitivity of 0.5-60 fmol BK at a dilution of 1:80,000 with 125I-BK(Tyr8) as tracer. Des-Arg9-BK, [BK(1-8)], displayed the highest cross-reactivity in the amount of 24%. Des-Arg1-BK and smaller molecular weight fragments display a cross-reactivity of less than 0.1%. The cross-reactivity of the BK antiserum with KAL is approximately 4%. In presence of 125I-KAL(Tyr9) the radioimmunoassay for KAL displays a sensitivity of 2 to 200 fmol KAL to an antiserum dilution of 1:80,000. The cross-reactivity with BK is 0.02%. KAL(Hyp4), BK(Hyp3), and des-Arg10-KAL [KAL(1-9)] show a cross-reactivity of 6.3, 4.9, and 2.4%. All other natural kinin derivatives show a cross-reactivity of less than 1%. Both assays were used to measure BK and KAL concentrations in blood and urine in humans after extraction and HPLC separation. The BK plasma level 1.97 (SD 0.54) pg/ml. The KAL plasma level is 81.0 (SD 14.3) pg/ml, indicating that KAL instead of BK is a circulating peptide. In urine, the BK level is 16.3 pg/ml.(ABSTRACT TRUNCATED AT 250 WORDS)
Hilgenfeldt
U
U
Department of Pharmacology, University of Heidelberg, Federal Republic of Germany.
Linke
R
R
Riester
U
U
König
W
W
Breipohl
G
G
D000906:Antibodies; D001920:Bradykinin / Q000097:blood* / Q000652:urine*; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D007609:Kallidin / Q000097:blood* / Q000652:urine*; D011863:Radioimmunoassay / Q000379:methods; D011869:Radioligand Assay
D000906:Antibodies; D007609:Kallidin; D001920:Bradykinin