A radiometric microassay for ornithine decarboxylase

Abstract

A simple method for purifying [ 3H] l-ornithine and incubation conditions suitable for estimating l-ornithine decarboxylase activity are described. Routine and recycled cation exchange procedures for separating putrescine from ornithine are outlined. Blanks using the routine cation exchange method average approx. 0.04% of the radioactivity contained in the substrate; product recovery is approx. 94%. The l-ornithine decarboxylase assay is proportional to time for at least 8 h. The relationship between substrate purity and the sensitivity of the cation exchange procedures is assessed. Radiochemical purity is the critical determinant of sensitivity for recycled assays. The cation exchange method is compared with the commonly used CO 2-trapping method. The cation exchange method is more specific and approximately three orders of magnitude more sensitive than the CO 2-trapping method. l-Ornithine decarboxylase activity can be measured reliably in samples of embryonic neural tissue having wet-weights of approx. 1 μg l-ornithine decarboxylase activity in the lumbar spinal cord of the chick embryo decreases 25–30 fold from day 5 to day 18 of embryonic development. A cation exchange procedure for estimating l-lysine decarboxylase activity is also described. Failure to detect l-lysine decarboxylase activity in the chick embryo lumbar spinal cord is contrasted with the previous report of high cadaverine levels in chick embryos.