A Monoclonal Antibody Against Leptin
| Autor: | Banafsheh Tavangar, Saeed Zarei, Reza Hadavi, Mahmood Jeddi-Tehrani, Ahmad Reza Mahmoudi, Yasaman Vojgani, Ali Ahmad Bayat, Jafar Mahmoudian |
|---|---|
| Rok vydání: | 2012 |
| Předmět: |
Leptin
medicine.medical_specialty Immunoconjugates medicine.drug_class Blotting Western Immunology Immunocytochemistry Antibody Affinity Enzyme-Linked Immunosorbent Assay Monoclonal antibody Chromatography Affinity Mice Affinity chromatography Antigen Antibody Specificity Pregnancy Cell Line Tumor Internal medicine medicine Animals Humans Immunology and Allergy Mice Inbred BALB C Hybridomas Leptin Deficiency biology digestive oral and skin physiology Antibodies Monoclonal Molecular biology Recombinant Proteins Blot Kinetics Endocrinology Hemocyanins biology.protein Female Immunization Antibody hormones hormone substitutes and hormone antagonists |
| Zdroj: | Hybridoma. 31:372-377 |
| ISSN: | 1557-8348 1554-0014 |
| DOI: | 10.1089/hyb.2012.0045 |
| Popis: | Leptin is an important protein that regulates energy storage and homeostasis in humans and animals. Leptin deficiency results in various abnormalities such as diabetes, obesity, and infertility. Producing a high affinity monoclonal antibody against human leptin provides an important tool to monitor and trace leptin function in different biological fluids. In this study, recombinant human leptin was conjugated to KLH and injected into mice. After immunization, mouse myeloma SP2/0 cells were fused with murine splenocytes followed by selection of antibody-producing hybridoma cells. After screening of different hybridoma colonies by ELISA, a high affinity antibody was selected and purified by affinity chromatography. The affinity constant of the antibody was measured by ELISA. Western blot, immunocytochemistry, and flow cytometry experiments were used to characterize the antibody. The anti-leptin antibody had a high affinity (around 1.13 × 10(-9) M) for its antigen. The saturation of the antibody with leptin (20 moles leptin per 1 mole antibody) in Western blot analysis proved that the antibody had specific binding to its antigen. Immunocytochemistry and flow cytometry on JEG-3 (human placental choriocarcinoma cell) cells revealed that the anti-leptin antibody recognized intracellular leptin. In conclusion, we report here the production and characterization of a murine anti-leptin antibody with high affinity for human leptin. |
| Databáze: | OpenAIRE |
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