Development of a specific radioimmunoassay to measure physiological changes of circulating leptin in cattle and sheep

Studies of leptin in large domestic ruminants have been limited to measurem ents of gene expression because methods to measure circulating levels are n ot available. To develop a bovine leptin radioimmunoassay, we produced reco mbinant bovine leptin and used it to immunize rabbits, and to prepare bovin e leptin tracer and standards. A single antiserum with sufficient affinity and titer was identified. Using this antiserum, logit-transformed binding o f I-125-labeled bovine leptin was linearly related (R-2 = 0.99) to the log of added bovine or ovine leptin between 0.1 to 2.0 ng. Serial dilution of b ovine and ovine plasma, chicken serum and bovine milk gave displacement cur ves that were parallel to those of bovine or ovine leptin. Recoveries of ex ternal addition of bovine leptin in ewe and cow plasma ranged between 94 an d 104%. Plasma leptin concentration measured by this assay was directly rel ated to the plans of nutrition in growing calves and lambs. At 11-14 weeks of age, ewe lambs had a higher circulating leptin concentration than ram la mbs. Finally, plasma leptin concentration was linearly related to the fat c ontent of the empty carcass in growing cattle and to body condition score i n lactating dairy cows. We conclude that circulating leptin in sheep and ca ttle is increased by fatness and plane of nutrition, consistent with result s in humans and rodents. This assay provides an important tool to investiga te mechanisms that regulate plasma leptin in cattle and sheep.