A RAPID, QUANTITATIVE FUNCTIONAL ASSAY FOR MEASURING LEPTIN

At present, leptin is quantitated using immune-assays that measure lep tin mass. Leptin biological activity is determined using protocols tha t measure feed consumption and weight reduction. These in vivo protoco ls are semi-quantitative and require large quantities of leptin. We de scribe a rapid, sensitive and quantitative in vitro assay for leptin u sing HEK-293 cells stably co-transfected with the leptin receptor Ob-R b isoform and a STAT-inducible promoter regulating the firefly lucifer ase cDNA. The assay, performed in a 96-well format, has an EC50 of 150 pM and is linear from 3 to 700 pM of leptin. We demonstrate that the assay is capable of measuring leptin in plasma samples. We demonstrate that bacterially-expressed, recombinant leptin and in vivo expressed leptin are equipotent. Furthermore, we demonstrate that a leptin-deriv ed peptide, leptin fragment 22-56, previously shown to be capable of r educing feed intake following ICV injection does not act directly thro ugh the leptin receptor. (C) 1998 Elsevier Science Ireland Ltd. All ri ghts reserved.