Human Leptin DuoSet ELISA

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Product Details
Citations (25)
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Human Leptin DuoSet ELISA Summary

Assay Type
Solid Phase Sandwich ELISA
96-well strip plate
Sample Volume Required
100 µL
Sufficient Materials
For five or fifteen 96-well plates*
Please see the product datasheet

* Provided that the recommended microplates, buffers, diluents, substrates and solutions are used, and the assay is run as summarized in the Assay Procedure provided.

This DuoSet ELISA Development kit contains the basic components required for the development of sandwich ELISAs to measure natural and recombinant human Leptin. The suggested diluent is suitable for the analysis of most cell culture supernate samples. Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet.

Product Features

  • Optimized capture and detection antibody pairings with recommended concentrations save lengthy development time
  • Development protocols are provided to guide further assay optimization
  • Assay can be customized to your specific needs
  • Economical alternative to complete kits

Kit Content

  • Capture Antibody
  • Detection Antibody
  • Recombinant Standard
  • Streptavidin conjugated to horseradish-peroxidase (Streptavidin-HRP)

Other Reagents Required

DuoSet Ancillary Reagent Kit 2 (5 plates): (Catalog # DY008) containing 96 well microplates, plate sealers, substrate solution, stop solution, plate coating buffer (PBS), wash buffer, and Reagent Diluent Concentrate 2.

The components listed above may be purchased separately:

PBS: (Catalog # DY006), or 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, 1.5 mM KH2PO4, pH 7.2 - 7.4, 0.2 µm filtered

Wash Buffer: (Catalog # WA126), or 0.05% Tween® 20 in PBS, pH 7.2-7.4

Reagent Diluent: (Catalog # DY995), or 1% BSA in PBS, pH 7.2-7.4, 0.2 µm filtered

Substrate Solution: 1:1 mixture of Color Reagent A (H2O2) and Color Reagent B (Tetramethylbenzidine) (Catalog # DY999)

Stop Solution: 2 N H2SO4 (Catalog # DY994)

Microplates: R&D Systems (Catalog # DY990)

Plate Sealers: ELISA Plate Sealers (Catalog # DY992)

Data Example

Human Leptin / OB ELISA Standard Curve

Product Datasheets

Preparation and Storage

Stability & Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: Leptin/OB

Leptin is a product of the mouse obese gene. Mice with mutations in the obese gene that block the synthesis of leptin are obese and diabetic, and have reduced activity, metabolism, and body temperature. Rat leptin shares approximately 96% and 82% sequence identity with the mouse and human protein, respectively. The expression of leptin mRNA is restricted to adipose tissue.

Entrez Gene IDs:
3952 (Human); 16846 (Mouse); 25608 (Rat)
Alternate Names:
FLJ94114; LEP; leptin (murine obesity homolog); leptin (obesity homolog, mouse); Leptin; OB; Obese protein; obese, mouse, homolog of; Obesity factor; OBOBS

Assay Procedure


Plate Preparation

  1. Dilute the Capture Antibody to the working concentration in PBS without carrier protein. Immediately coat a 96-well microplate with 100 μL per well of the diluted Capture Antibody. Seal the plate and incubate overnight at room temperature.
  2. Aspirate each well and wash with Wash Buffer, repeating the process two times for a total of three washes. Wash by filling each well with Wash Buffer (400 μL) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each step is essential for good performance. After the last wash, remove any remaining Wash Buffer by aspirating or by inverting the plate and blotting it against clean paper towels.
  3. Block plates by adding 300 μL Reagent Diluent to each well. Incubate at room temperature for a minimum of 1 hour.
  4. Repeat the aspiration/wash as in step 2. The plates are now ready for sample addition.

Assay Procedure

  1. Add 100 μL of sample or standards in Reagent Diluent, or an appropriate diluent, per well. Cover with an adhesive strip and incubate 2 hours at room temperature.
  2. Repeat the aspiration/wash as in step 2 of Plate Preparation.
  3. Add 100 μL of the Detection Antibody, diluted in Reagent Diluent, to each well. Cover with a new adhesive strip and incubate 2 hours at room temperature.
  4. Repeat the aspiration/wash as in step 2 of Plate Preparation.
  5. Add 100 μL of the working dilution of Streptavidin-HRP to each well. Cover the plate and incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
  6. Repeat the aspiration/wash as in step 2.
  7. Add 100 μL of Substrate Solution to each well. Incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
  8. Add 50 μL of Stop Solution to each well. Gently tap the plate to ensure thorough mixing.
  9. Determine the optical density of each well immediately, using a microplate reader set to 450 nm. If wavelength correction is available, set to 540 nm or 570 nm. If wavelength correction is not available, subtract readings at 540 nm or 570 nm from the readings at 450 nm. This subtraction will correct for optical imperfections in the plate. Readings made directly at 450 nm without correction may be higher and less accurate.

Citations for Human Leptin DuoSet ELISA

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

25 Citations: Showing 1 - 10
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  1. Impact to short-term high intensity intermittent training on different storages of body fat, leptin and soluble leptin receptor levels in physically active non-obese men: A pilot investigation
    Authors: RS Caldeira, VLG Panissa, DS Inoue, EZ Campos, PA Monteiro, BM Giglio, GD Pimentel, P Hofmann, FS Lira
    Clin Nutr ESPEN, 2018;28(0):186-192.
    Species: Human
    Sample Types: Serum
  2. Impact of rural-urban environment on metabolic profile and response to a 5-day high-fat diet
    Authors: DL Tahapary, K de Ruiter, F Kurniawan, Y Djuardi, Y Wang, SME Nurdin, E Iskandar, D Minggu, E Yunir, B Guigas, T Supali, PCN Rensen, E Sartono, P Soewondo, DS Harbuwono, JWA Smit, M Yazdanbakh
    Sci Rep, 2018;8(1):8149.
    Species: Human
    Sample Types: Plasma
  3. Effects of the inspiratory muscle training and aerobic training on respiratory and functional parameters, inflammatory biomarkers, redox status and quality of life in hemodialysis patients: A randomized clinical trial
    Authors: PHS Figueiredo, MMO Lima, HS Costa, JB Martins, OD Flecha, PF Gonçalves, FL Alves, VGB Rodrigues, EHB Maciel, VA Mendonça, ACR Lacerda, ÉLM Vieira, AL Teixeira, F de Paula, CH Balthazar
    PLoS ONE, 2018;13(7):e0200727.
    Species: Human
    Sample Types: Plasma
  4. The effects of intensified training on resting metabolic rate (RMR), body composition and performance in trained cyclists
    Authors: AL Woods, AJ Rice, LA Garvican-L, AM Wallett, B Lundy, MA Rogers, M Welvaert, S Halson, A McKune, KG Thompson
    PLoS ONE, 2018;13(2):e0191644.
    Species: Human
    Sample Types: Serum
  5. Presence of IL-17 in synovial fluid identifies a potential inflammatory osteoarthritic phenotype
    Authors: SJ Snelling, S Bas, GJ Puskas, SG Dakin, D Suva, A Finckh, C Gabay, P Hoffmeyer, AJ Carr, A Lübbeke
    PLoS ONE, 2017;12(4):e0175109.
    Species: Human
    Sample Types: Serum
  6. Obesity-induced diet leads to weight gain, systemic metabolic alterations, adipose tissue inflammation, hepatic steatosis, and oxidative stress in gerbils (Meriones unguiculatus)
    Authors: LL Ventura, NC Fortes, HC Santiago, MV Caliari, MA Gomes, DR Oliveira
    PeerJ, 2017;5(0):e2967.
    Species: Gerbil
    Sample Types: Tissue Homogenates
  7. High Expression of STAT3 in Subcutaneous Adipose Tissue Associates with Cardiovascular Risk in Women with Rheumatoid Arthritis
    Authors: M Nadali, R Pullerits, KME Andersson, ST Silfverswä, MC Erlandsson, MI Bokarewa
    Int J Mol Sci, 2017;18(11):.
    Species: Human
    Sample Types: Serum
  8. The adipokine leptin modulates adventitial pericyte functions by autocrine and paracrine signalling
    Authors: F Riu, SC Slater, EJ Garcia, I Rodriguez-, V Alvino, E Avolio, G Mangialard, A Cordaro, S Satchell, C Zebele, A Caporali, G Angelini, P Madeddu
    Sci Rep, 2017;7(1):5443.
    Species: Human
    Sample Types: Cell Culture Supernates
  9. Associations between Maternal Body Composition and Appetite Hormones and Macronutrients in Human Milk
    Authors: S Kuganantha, Z Gridneva, CT Lai, AR Hepworth, PJ Mark, F Kakulas, DT Geddes
    Nutrients, 2017;9(3):.
    Species: Human
    Sample Types: Milk
  10. Leptin Levels Are Higher in Whole Compared to Skim Human Milk, Supporting a Cellular Contribution
    Authors: Foteini Kakulas
    Nutrients, 2016;8(11):.
    Species: Human
    Sample Types: Milk
  11. Secretory activity of subcutaneous abdominal adipose tissue in male patients with rheumatoid arthritis and osteoarthritis - association with clinical and laboratory data
    Authors: Ewa Kontny
    Reumatologia, 2016;54(5):227-235.
    Species: Human
    Sample Types: Tissue Extracts
  12. A Multifactorial Weight Reduction Programme for Children with Overweight and Asthma: A Randomized Controlled Trial
    PLoS ONE, 2016;11(6):e0157158.
    Species: Human
    Sample Types: Serum
  13. Association study of arcuate nucleus neuropeptide Y neuron receptor gene variation and serum NPY levels in clozapine treated patients with schizophrenia
    Authors: E Leinonen
    Eur. Psychiatry, 2016;40(0):13-19.
    Species: Human
    Sample Types: Serum
  14. Effect of Human Milk Appetite Hormones, Macronutrients, and Infant Characteristics on Gastric Emptying and Breastfeeding Patterns of Term Fully Breastfed Infants
    Authors: Zoya Gridneva
    Nutrients, 2016;9(1):.
    Species: Human
    Sample Types: Human Milk
  15. Characterization of In Vitro Engineered Human Adipose Tissues: Relevant Adipokine Secretion and Impact of TNF-alpha.
    Authors: Aubin K, Safoine M, Proulx M, Audet-Casgrain M, Côté J, Tetu F, Roy A, Fradette J
    PLoS ONE, 2015;10(9):e0137612.
    Species: Human
    Sample Types: Cell Culture Supernates
  16. Human adipose tissue conditioned media from lean subjects is protective against H2O2 induced neurotoxicity in human SH-SY5Y neuronal cells.
    Authors: Wan Z, Mah D, Simtchouk S, Kluftinger A, Little J
    Int J Mol Sci, 2015;16(1):1221-31.
    Species: Human
    Sample Types: Cell Culture Supernates
  17. Endothelial cells from visceral adipose tissue disrupt adipocyte functions in a three-dimensional setting: partial rescue by angiopoietin-1.
    Authors: Pellegrinelli V, Rouault C, Veyrie N, Clement K, Lacasa D
    Diabetes, 2014;63(2):535-49.
    Species: Human
    Sample Types: Cell Culture Supernates
  18. Expression of obesity markers and Persistent Organic Pollutants levels in adipose tissue of obese patients: reinforcing the obesogen hypothesis?
    Authors: Pereira-Fernandes A, Dirinck E, Dirtu A, Malarvannan G, Covaci A, Van Gaal L, Vanparys C, Jorens P, Blust R
    PLoS ONE, 2014;9(1):e84816.
    Species: Human
    Sample Types: Serum
  19. Adipokine resistin predicts anti-inflammatory effect of glucocorticoids in asthma.
    Authors: Leivo-Korpela S, Lehtimaki L, Vuolteenaho K, Nieminen R, Kankaanranta H, Saarelainen S, Moilanen E
    J Inflamm (Lond), 2011;8(0):12.
    Species: Human
    Sample Types: Plasma
  20. LIGHT (TNFSF14) inhibits adipose differentiation without affecting adipocyte metabolism.
    Authors: Tiller G, Laumen H, Fischer-Posovszky P, Finck A, Skurk T, Keuper M, Brinkmann U, Wabitsch M, Link D, Hauner H
    Int J Obes (Lond), 2011;35(2):208-16.
    Species: Human
    Sample Types: Cell Culture Supernates
  21. Performance characteristics of combinations of host biomarkers to identify women with occult placental malaria: a case-control study from Malawi.
    Authors: Conroy AL, Liles WC, Molyneux ME
    PLoS ONE, 2011;6(12):e28540.
    Species: Human
    Sample Types: Plasma
  22. Regulation of adipokine production in human adipose tissue by propionic acid.
    Authors: Al-Lahham SH, Roelofsen H, Priebe M, Weening D, Dijkstra M, Hoek A, Rezaee F, Venema K, Vonk RJ
    Eur. J. Clin. Invest., 2010;40(5):401-7.
    Species: Human
    Sample Types: Cell Culture Supernates
  23. Homeostatic regulation of T effector to Treg ratios in an area of seasonal malaria transmission.
    Authors: Finney OC, Nwakanma D, Conway DJ, Walther M, Riley EM
    Eur. J. Immunol., 2009;39(5):1288-300.
    Species: Human
    Sample Types: Plasma
  24. Relationships between serum leptin and bone markers during stable weight, weight reduction and weight regain in male and female judoists.
    Authors: Prouteau S, Benhamou L, Courteix D
    Eur. J. Endocrinol., 2006;154(3):389-95.
    Species: Human
    Sample Types: Serum
  25. Leptin Levels Are Higher in Whole Compared to Skim Human Milk, Supporting a Cellular Contribution.
    Authors: Kugananthan S, Lai C, Gridneva Z, Mark P, Geddes D, Kakulas F
    Nutrients, 0;8(11):.
    Species: Human
    Sample Types: Milk


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Human Leptin DuoSet ELISA
By Anonymous on 12/20/2017
Sample Tested: EDTA Plasma

Human Leptin DuoSet ELISA
By Anonymous on 08/09/2017
Sample Tested: Serum

serum dilution used - 1:2.5