Functional Leptin ELISA Assay

Additional Information

Assay Background

Leptin a hormone of 146 amino acids shows a molecular weight of 16 kDa (P41159). It consists of four anti-parallel a-helices. Leptin is primarily produced by adipocytes and thus provides a signal to the energy state of the organism. By binding to the leptin receptor (P48357) it influences the activity of the JAK -STAT pathway and thereby regulates energy metabolism, especially the food intake. Leptin levels show a circadian variation (+/- 30%) and are dependent on BMI, pubertal status, and gender. In addition to the expression itself, the bioavailability of leptin, is regulated via a binding protein, as with many hormones with type I cytokine receptors. The binding protein is the extracellular domain of the Leptin receptor formed by proteolytic cleavage of the receptor be metalloproteases (ADAM 10/17).In healthy persons food intake is reduced by increasing concentrations of leptin and by falling SOB-R quantities. In various pathological situations, this regulatory circuit is however interrupted: e.g. the leptin levels in the circulation of obese persons are increased, but resulting in no satiety. This phenomenon, known as leptin resistance, could have its cause in a reduction in the number of leptin receptors or in influencing the intracellular signal transduction by other parameters. Lately it was shown that a naturally occurring transversion in the leptin gen (c.298G → T) results in an amino acid exchange of asparagine to tyrosin in position 100 (pD100Y). An in vitro cell culture model revealed that this mutated leptin was still secreted but unable to bind to the receptor and therewith unable to exert intracellular signaling. In vivo this mutation results in disturbance of food intake regulation and in consequence in extreme obesity. Under treatment with recombinant human leptin food intake behavior normalized and concomitantly body weight was reduced significantly. Since the mutant leptin is recognized by the classical immunological leptin test systems, either a genetic test (sequencing of the ob gene), or a functional assay (binding to the receptor) is necessary. The Eagle Biosciences Functional Leptin assay allows the measurement of leptin in human serum by binding it to the soluble receptor (leptin binding protein). Based on the measurement of receptor binding leptin, the bioLEP assay provides additional information on the biological reactivity of the circulating leptin molecules. Mutations resulting in less or no binding leptin can be detected by a significant reduction of the measured bioactive Leptin. Especially in connection with measurement of total Leptin by classical immunoassays, the receptor-binding characteristics of leptin can easily be detected and quantified. This might help to evaluate mutation rates of the leptin gene by an inexpensive, fast method and accelerate obesity research by improved patient stratification in clinical studies.

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