Recombinant human leptin

Manufactured by R&D Systems

Sourced in United States, United Kingdom

Recombinant human leptin is a protein produced using recombinant DNA technology. It is a hormone that regulates food intake and energy expenditure.

Automatically generated - may contain errors

Lab products found in correlation

Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions) 125iodine, by PerkinElmer (1 mentions) 1470 wizard, by PerkinElmer (1 mentions) Luminex immunoassay, by Thermo Fisher Scientific (1 mentions) Delfia eutda cytotoxicity kit, by PerkinElmer (1 mentions) Synergy mx, by Agilent Technologies (1 mentions) Hnepc, by PromoCell (1 mentions) D lys3 ghrp 6, by Phoenix Pharmaceuticals (1 mentions) Lipopolysaccharide lps, by Merck Group (1 mentions) Recombinant human il 6, by Thermo Fisher Scientific (1 mentions) Stat3 inhibitor, by Merck Group (1 mentions) Recombinant human adiponectin acrp30, by R&D Systems (1 mentions) Dulbecco s modified eagle s medium dmem, by Lonza (1 mentions) L glutamine, by Lonza (1 mentions) Fetal bovine serum (fbs), by Lonza (1 mentions) Ham s f 12 medium, by Lonza (1 mentions) Rpmi 1640 culture medium, by Thermo Fisher Scientific (1 mentions) Cd3 apc h7 clone sk7, by BD (1 mentions) Facsaria 2, by BD (1 mentions) Wst 1 reagent, by Roche (1 mentions)

23 protocols using recombinant human leptin

Leptin Biodistribution in Mice

Check if the same lab product or an alternative is used in the 5 most similar protocols

Recombinant human leptin (R&D Systems) was labelled with ¹²⁵Iodine (Perkin Elmer) using the Iodogen method (Thermo Scientific). Mice were injected intravenously via the lateral tail vein with 12 ng (14.4 kBq) leptin in a total volume of 100 µL made up with phosphate-buffered solution. Animals were then placed in an individual cage with access to food and water until the specified time when the animal was killed by rapid CO₂ asphyxiation with n = 4 for each time point, to observe distribution over a time course. Tissues were dissected and weighed with duplicate samples placed in polypropylene tubes and measured for total γ-radioactivity (1470 Wizard, Perkin Elmer). Background radiation was subtracted from all samples. Due to equipment failure, no data were collected for brain or digestive tract at 120 min.
Blood was collected via cardiac puncture and transferred to vial containing 20 IU heparin and measured in duplicate. Total blood volume was calculated as 84.7 mL/kg body mass as previously reported (34 (link)). The skin was removed apart from that around the snout and ‘cuffs’ around the limbs and weighed intact. Four samples were taken from the left forelimb, right hindlimb, interscapular and dorsal cervical regions. Gut tissues were measured with contents.

Hart R.A., Dobos R.C., Agnew L.L., Smart N.A, & McFarlane J.R. (2016). Leptin pharmacokinetics in male mice. Endocrine Connections, 6(1), 20-26.

+ Open protocol

+ Expand

Leptin Modulation of NK Cell Cytotoxicity

Check if the same lab product or an alternative is used in the 5 most similar protocols

For molecular investigations, NK-92 cells either remained unstimulated or were preincubated with 10 ng/mL (physiological concentration in normal weight individuals) and 100 ng/mL (pathophysiological concentration in obese individuals) recombinant human leptin (R&D Systems, Minneapolis, MN, USA) for 4 h or 24 h. Cells were collected and stored at −80°C until analysis. The cytotoxicity of NK cells was analyzed using the DELFIA EuTDA Cytotoxicity kit (PerkinElmer, Waltham, MA, USA) according to the manufacturer's manual. NK-92 cells as well as primary NK cells served as effector cells and DLD-1 cells served as target cells. NK effector cells either remained unstimulated or were preincubated with 10 ng/mL and 100 ng/mL recombinant human leptin for 4 h or 72 h. To determine the cytotoxicity, NK cells were coincubated with DLD-1 cells for 1 h in RPMI 1640 medium supplemented with 10% FBS. Fluorescence data were recorded using a time resolved fluorometer (Synergy Mx, BioTek Instruments, Winooski, VT, USA). Remaining supernatants of the cytotoxicity assay were collected for IFN-γ analyses by luminex immunoassay (eBioscience, Frankfurt am Main, Germany). In both incubation experiments with leptin as well as cytotoxicity assays including analyses of IFN-γ secretion, the incubation medium of NK-92 and primary NK cells contained 200 U/mL IL-2.

Bähr I., Goritz V., Doberstein H., Hiller G.G., Rosenstock P., Jahn J., Pörtner O., Berreis T., Mueller T., Spielmann J, & Kielstein H. (2017). Diet-Induced Obesity Is Associated with an Impaired NK Cell Function and an Increased Colon Cancer Incidence. Journal of Nutrition and Metabolism, 2017, 4297025.

+ Open protocol

+ Expand

Ghrelin, GHSR1a, and Leptin Interactions

Check if the same lab product or an alternative is used in the 5 most similar protocols

Recombinant human ghrelin and the GHSR1a antagonist; D-Lys-3-growth hormone-releasing peptide 6 (D-Lys-3-GHRP-6) were obtained from Phoenix Pharmaceuticals. Lipopolysaccharide (LPS) was purchased from Sigma-Aldrich. Recombinant human leptin was purchased from R&D Systems. The HNEpCs were obtained from PromoCell (C-12620).

Choi Y.S., Na H.G., Bae C.H., Song S.Y, & Kim Y.D. (2022). Ghrelin Downregulates Lipopolysaccharide/ Leptin-Induced MUC5AC Expression in Human Nasal Epithelial Cells. Clinical and Experimental Otorhinolaryngology, 16(1), 49-58.

+ Open protocol

+ Expand

Antibody Detection and Neutralization Protocol

Check if the same lab product or an alternative is used in the 5 most similar protocols

The following primary monoclonal antibodies (mAbs) and polyclonal antibodies (pAbs) were used to detect proteins by immunofluorescence labeling and immunoblot analysis and/or to neutralize the effects of leptin and IL-6: mouse anti-leptin mAb (44802), anti-leptin receptor mAb (52263) (R&D Systems, Minneapolis, MN, USA), rat anti-IL-6 mAb (MQ2-13A5; Miltenyi Biotec, Bergisch Gladbach, Germany), mouse anti-phospho STAT3 (Tyr705) mAb (3E2), rabbit anti-STAT3 pAb (D47E7), anti-SOCS3 pAb (Cell Signaling Technology, Danvers, MA, USA), mouse anti-β-actin mAb (E1C605; EnoGene, New York, NY, USA), rabbit anti-claudin-5 mAb (EPR7583), anti-VE-cadherin pAb (Abcam, Cambridge, MA, USA), rabbit anti-occludin pAb, anti-ZO-1 pAb (Invitrogen, Carlsbad, CA, USA), and mouse anti-D2-40 mAb (413451; Nichirei, Tokyo, Japan). Mouse IgG (11711) isotype controls were purchased from R&D Systems. Recombinant human leptin was obtained from R&D Systems. Recombinant human IL-6 was purchased from Peprotech (Rocky Hill, NJ, USA). STAT3 inhibitor was obtained from EMD Millipore (Billerica, MA, USA).

Sato A., Kamekura R., Kawata K., Kawada M., Jitsukawa S., Yamashita K., Sato N., Himi T, & Ichimiya S. (2016). Novel Mechanisms of Compromised Lymphatic Endothelial Cell Homeostasis in Obesity: The Role of Leptin in Lymphatic Endothelial Cell Tube Formation and Proliferation. PLoS ONE, 11(7), e0158408.

+ Open protocol

+ Expand

Assessing Adiponectin and Leptin Receptor Function

Check if the same lab product or an alternative is used in the 5 most similar protocols

Functioning adiponectin and leptin receptors were assessed by changes in NPY expression in response to adiponectin/aCrp30 and leptin treatment: ~75Day neuronal cultures were held in neuronal maintenance medium free of insulin and at 5.5mM glucose 24hours and then supplemented with either 1ug/ml recombinant human adiponectin/aCrp30 or 50 nM recombinant human leptin (R&D Systems, Minneapolis MN). Cells in 24-well plates were collected at 4 hours of exposure and RNA isolated for expression analysis using qRT-PCR as described above and using 18SRNA for normalization. Four independent neuronal cultures derived from 2 iPSC were tested in triplicate. A two-tailed t-test was used to determine p-values. The sequences for the primer pairs used are given in Table S1. Functioning glutamate receptors were assessed by patch clamp as described in Supplemental Methods Figure S5.

Yao L., Liu Y., Qiu Z., Kumar S., Curran J.E., Blangero J., Chen Y, & Lehman D.M. (2017). Molecular profiling of human induced pluripotent stem cell-derived hypothalamic neurons provides developmental insights to genetic loci for body weight regulation. Journal of neuroendocrinology, 29(2), 10.1111/jne.12455.

+ Open protocol

+ Expand

Osteoarthritis Cartilage and Synovial Fluid Analysis

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Cartilage and synovial fluid (SF) samples were collected from 97 patients with OA who were undergoing knee replacement surgery. All patients fulfilled the American College of Rheumatology classification criteria for OA [30 (link)]. Cartilage samples were processed for tissue culture as previously described [15 ]. Cartilage pieces were incubated for 42 hours with or without leptin (10 μg/ml). The concentration of leptin used was chosen based on our previous studies and on existing literature [15 , 17 (link)–19 (link)]. Recombinant human leptin was purchased from R&D Systems Europe Ltd, Abindgon, UK. Synovial fluid (SF) samples from the corresponding patients were also collected at the beginning of the arthroplasty. The SF samples were centrifuged at 4000 g at 4 °C and supernatants were collected and kept at −70 °C until assayed.
The immortalized murine H4 chondrocyte cell line [31 (link)], developed in the Laboratory of Experimental Rheumatology, University Medical Center, Nijmegen, The Netherlands, was used in the siRNA experiments. The chondrocytes were cultured at 37 °C in humidified 5 % carbon dioxide atmosphere in Dulbecco’s modified Eagle’s medium (DMEM) with L-glutamine and Ham’s F-12 medium (1:1) supplemented with 5 % fetal bovine serum (all obtained from Lonza Group Ltd, Basel, Switzerland).

Koskinen-Kolasa A., Vuolteenaho K., Korhonen R., Moilanen T, & Moilanen E. (2016). Catabolic and proinflammatory effects of leptin in chondrocytes are regulated by suppressor of cytokine signaling-3. Arthritis Research & Therapy, 18, 215.

+ Open protocol

+ Expand

Isolation and Characterization of PBMC Subsets

Check if the same lab product or an alternative is used in the 5 most similar protocols

PBMCs were washed 3× with RPMI1640 culture medium (Gibco, Thermo Fisher Scientific Inc., Waltham, MA, USA) and resuspended in complete culture medium (CM: RPMI1640 supplemented with 10% fetal bovine serum, 100 U/mL penicillin and 50 µM 2-mercaptoethanol), at a concentration of 10⁶ cells/mL. PBMCs were cultured in triplicate in a 37 °C humidified chamber with 5% CO₂ for 12 h, in the presence or absence of mitogens (PI: 5 ng/mL phorbol myristate acetate and 1 μM ionomycin) (Sigma-Aldrich, Merck KGaA) or recombinant human leptin (R&D Systems) at a concentration of 200 or 500 or 800 ng/mL. At the end of culture, CD3+ T cells and CD14+ monocytes were isolated by cell sorting using a BD FACS Aria II flow cytometer (BD Biosciences, San Jose, CA, USA). The sorting strategy is shown in Figure 8. The enriched cell populations were used when purity was >95% for T cells and >90% for monocytes. The antibodies used for cell sorting and phenotyping were mouse monoclonal anti-human antibodies CD3-APC-H7 (clone SK7) and CD14-FITC (clone M5E2) (BD Biosciences). Fluorescence-minus-one (FMO) controls were used to identify any background spread of fluorochromes and to establish gating limits. Data were analyzed using BD FACS DIVA v.9 software (BD Biosciences).

Thomas I., Panagoulias I., Aggeletopoulou I., Varvarigou A., Spiliotis B.E, & Mouzaki A. (2021). The Role of Leptin in Childhood Immune Thrombocytopenia (ITP): An Anti-Inflammatory Agent?. International Journal of Molecular Sciences, 22(14), 7636.

+ Open protocol

+ Expand

Cell Proliferation Assay with Secretomes

Check if the same lab product or an alternative is used in the 5 most similar protocols

Three thousand cells per well were seeded into 96-well plates and after the appropriate culture period, WST-1 reagent (Roche Diagnostics, Meylan, France) was added to each well and incubated for 15 h at 37 °C; absorbance was read at 490 nm. During the culture period, medium was supplemented with fat-CM or ADSC-CM or recombinant human leptin as indicated (R&D Systems) or with an antibody directed against the subunit gp130 of IL-6 family receptor (10 µg/mL; clone BK5 from Diaclone, Besançon, France) as described [41] (link).

Avril P., Le Nail L.R., Brennan M.Á., Rosset P., De Pinieux G., Layrolle P., Heymann D., Perrot P, & Trichet V. (2015). Mesenchymal stem cells increase proliferation but do not change quiescent state of osteosarcoma cells: Potential implications according to the tumor resection status. Journal of Bone Oncology, 5(1), 5-14.

+ Open protocol

+ Expand

Recombinant Human Leptin Preparation

Check if the same lab product or an alternative is used in the 5 most similar protocols

Recombinant human leptin was obtained from R&D Systems (Wiesbaden, Germany) and diluted to 50 nM.

Laue T., Wrann C.D., Hoffmann-Castendiek B., Pietsch D., Hübner L, & Kielstein H. (2015). Altered NK cell function in obese healthy humans. BMC obesity, 2, 1.

+ Open protocol

+ Expand

Leptin Modulates Cofilin-1 in NK-92 Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

In three independent experiments, human NK-92 cells (5x10⁵) were incubated in 1 mL RPMI-Medium 1640 with 20% FBS and 200 U/mL IL-2 on 13 mm cover slips coated with 20 μg/mL human Fibronectin (Culturex^® Human Fibronectin, PathClear^®, Amsbio, Abington, UK) and stimulated with 10 ng/mL (physiological concentration) and 100 ng/mL (pathophysiological concentration) recombinant human leptin (R&D Systems, Minneapolis, MN, USA) for 30 min and 24 h. Thereafter, cells were fixed and permeabilized with 4% PSA (Formalin solution, 10%, Sigma-Aldrich, St. Louis, MO, USA), incubated with the primary monoclonal anti-Cofilin-1 antibody (1:50; D3F9, Cell Signaling Technology, Danvers, MA, USA) for 1 h at room temperature and stained with Phalloidin - TRITC (1:1,000; Sigma-Aldrich, St. Louise, MO, USA) for 1 h at room temperature, donkey anti-rabbit Alexa 488 (1:310; Dianova GmbH, Hamburg, Germany) for 1 h at room temperature and DAPI (1:50,000; Sigma-Aldrich) for 5 min at room temperature. Slides were mounted with ProLong ® Gold (Thermo Fisher Scientific, Waltham, MA, USA).

Oswald J., Büttner M., Jasinski-Bergner S., Jacobs R., Rosenstock P, & Kielstein H. (2018). Leptin affects filopodia and cofilin in NK-92 cells in a dose- and time-dependent manner. European Journal of Histochemistry : EJH, 62(1), 2848.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!