CROSS-REACTIVITY OF DELTA-17-6-KETO-PGF(1-ALPHA) WITH 6-KETO-PGF(1-ALPHA) ANTIBODIES

The cross-reactivity of the PGI(3) metabolite, Delta 17-6-keto-PGF(1 a lpha), with antibodies against 6-keto-PGF(1 alpha) for radioimmunoassa ys (RIA) has been investigated, Delta 7-6-keto-PGF(1 alpha) was obtain ed either from commercial sources or after its purification from endot helial cells. In the latter case, primary cultured bovine aortic endot helial cells were incubated for 20 min at 37 degrees C with 10 mu M ei cosapentaenoic acid (EPA) in the presence of 2 mu M 13-hydroperoxy-oct adecadienoic acid, an activator of the EPA cyclooxygenation, and the 6 -keto-PGF1 alpha and Delta 17-6-keto-PGF(1 alpha) produced were separa ted by RP-HPLC. Then, cross-reactivities of the commercial and purifie d Delta 17-6-keto-PGF(1 alpha) with 6-keto-PGF(1 alpha) antibodies wer e determined and found not to exceed 10%. lit addition, the amounts of prostacyclin-related compounds detected by direct measurements in med ia of cells loaded with EPA were compared with those obtained after pu rification of 6-keto-PGF(1 alpha). In accordance with the cross-reacti vity data, we found that RIA in media mainly measured 6-keto-PGF(1 alp ha), the Delta 17-6-keto-PGF(1 alpha) formed being undetected at 90%. It is concluded that 6-keto-PGF(1 alpha) antibodies generally used for RLA of 6-keto-PGF(1 alpha) are highly specific since they can discrim inate a metabolite bearing an additional double bond such as the PGI(3 ) metabolite Delta 17-6-keto-PGF(1 alpha).