open access

Vol 6, No 2 (2003)
Published online: 2003-10-10
Submitted: 2012-01-23
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Development of radioimmunoassay for the measurement of human leptin in serum

Ana R. Lagarde, Katalin Nagy, Tibor Forgách, Gyozo A. Jánoki
Nucl. Med. Rev 2003;6(2):105-109.

open access

Vol 6, No 2 (2003)
Published online: 2003-10-10
Submitted: 2012-01-23

Abstract

BACKGROUND: Leptin is a 16 kDa polypeptide hormone encoded by the obese gene (ob) and secreted by adipose tissue. This hormone plays a major role in energy homeostasis and regulation of food intake and body weight. It also affects the metabolic, neuroendocrine and reproductive systems.
MATERIAL AND METHODS: Labelling of recombinant human leptin with 125I was best performed by the Chloramine-T method. New Zealand white rabbits were immunised with recombinant human leptin, cross-reaction of obtained antisera was analyzed with 10 different antigens. The separation of bound and free fractions was performed using the second antibody - PEG method.
RESULTS: The obtained tracer had specific activities of 2.8–3.3 kBq/µg and had a stability of 5 weeks. A highly specific polyclonal antibody was obtained without measurable cross-reaction against the analysed antigens. Concentrations of human leptin were measured by a single overnight incubation assay with a sensitivity of 0.5 ng/ml and a measuring range of 0.5–100 ng/ml. The intra-assay and inter-assay coefficient of variation was under 6% and 8%, respectively. Recovery ranged from 88% to 106%.
CONCLUSIONS: Serum human leptin concentrations can be accurately and precisely measured by this new radioimmunoassay. Preliminary results obtained from the measurement of serum leptin in lean, overweight and obese patients are presented. Serum leptin concentrations correlated with body mass index and were significantly higher in women than in men, except for obese patients.

Abstract

BACKGROUND: Leptin is a 16 kDa polypeptide hormone encoded by the obese gene (ob) and secreted by adipose tissue. This hormone plays a major role in energy homeostasis and regulation of food intake and body weight. It also affects the metabolic, neuroendocrine and reproductive systems.
MATERIAL AND METHODS: Labelling of recombinant human leptin with 125I was best performed by the Chloramine-T method. New Zealand white rabbits were immunised with recombinant human leptin, cross-reaction of obtained antisera was analyzed with 10 different antigens. The separation of bound and free fractions was performed using the second antibody - PEG method.
RESULTS: The obtained tracer had specific activities of 2.8–3.3 kBq/µg and had a stability of 5 weeks. A highly specific polyclonal antibody was obtained without measurable cross-reaction against the analysed antigens. Concentrations of human leptin were measured by a single overnight incubation assay with a sensitivity of 0.5 ng/ml and a measuring range of 0.5–100 ng/ml. The intra-assay and inter-assay coefficient of variation was under 6% and 8%, respectively. Recovery ranged from 88% to 106%.
CONCLUSIONS: Serum human leptin concentrations can be accurately and precisely measured by this new radioimmunoassay. Preliminary results obtained from the measurement of serum leptin in lean, overweight and obese patients are presented. Serum leptin concentrations correlated with body mass index and were significantly higher in women than in men, except for obese patients.
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Keywords

radioimmunoassay; leptin; 125I; obesity; BMI

About this article
Title

Development of radioimmunoassay for the measurement of human leptin in serum

Journal

Nuclear Medicine Review

Issue

Vol 6, No 2 (2003)

Pages

105-109

Published online

2003-10-10

Bibliographic record

Nucl. Med. Rev 2003;6(2):105-109.

Keywords

radioimmunoassay
leptin
125I
obesity
BMI

Authors

Ana R. Lagarde
Katalin Nagy
Tibor Forgách
Gyozo A. Jánoki

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