A rapid, quantitative functional assay for measuring leptin

Mol Cell Endocrinol. 1998 Aug 25;143(1-2):117-23. doi: 10.1016/s0303-7207(98)00129-4.

Abstract

At present, leptin is quantitated using immuno-assays that measure leptin mass. Leptin biological activity is determined using protocols that measure feed consumption and weight reduction. These in vivo protocols are semi-quantitative and require large quantities of leptin. We describe a rapid, sensitive and quantitative in vitro assay for leptin using HEK-293 cells stably co-transfected with the leptin receptor Ob-Rb isoform and a STAT-inducible promoter regulating the firefly luciferase 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-derived peptide, leptin fragment 22-56, previously shown to be capable of reducing feed intake following ICV injection does not act directly through the leptin receptor.

MeSH terms

  • Animals
  • Biological Assay*
  • Carrier Proteins
  • Cell Line
  • Leptin
  • Mice
  • Peptide Fragments
  • Proteins / analysis*
  • Receptors, Cell Surface*
  • Receptors, Leptin
  • Sensitivity and Specificity
  • Transfection

Substances

  • Carrier Proteins
  • Leptin
  • Peptide Fragments
  • Proteins
  • Receptors, Cell Surface
  • Receptors, Leptin
  • leptin receptor, mouse