A radioimmunoassay for 6-keto-prostaglandin F1 alpha utilizing an antiserum against 6-methoxime-prostaglandin F1 alpha. Application to study renal in vitro synthesis of prostacyclin.

PGI2 and 6-keto-PGF1 alpha were converted to 6-methoxime-PGF1 alpha (6-MeON-PGF1 alpha) by treatment with methoxyamine HCl in acetate buffer. The formed 6-MeON-PGF1 alpha was measured by radioimmunoassay. Antisera were raised in rabbits after immunization against 6-MeON-PGF1 alpha-BSA conjugate. Diluted 1:20.000 to bind 50% of the tracer (3H-6-MeON-PGF1 alpha, 100 Ci/mmol), the antiserum cross reacted 0.8% with PGE2, 1% with PGF2 alpha and less than 0.2% with PGD2, PGF1 alpha, PGF2 beta and TXB2. The radioimmunoassay was used to estimate release of PGI2 and 6-keto-PGF1 alpha from chopped rabbit renal medulla and cortex incubated in Krebs-Ringer bicarbonate buffer (37 degrees C, 30 min). The 6-keto-PGF1 alpha radioimmunoassay was validated in biological samples by mass fragmentography. The chopped medulla (n = 5) released 38-+9 ng/g/min and the cortex (n = 5) 4.7-+2-0 ng/g/min, while the release of immunoreactive PGE2 (iPGE2) and iPGF2 alpha was 171-+26 and 74-+13 ng/g/min from the medulla and 4.3-+1.3 and 2.7-+0.3 ng/g/min from the cortex, respectively. The results confirm previous findings, which indicate that in the renal medulla prostaglandin endoperoxides are mainly transformed to prostaglandins, while in the cortex transformation to PGI2 seems to be of greater relative importance.