Leptin ELISA Kit (LEP)

Details for Product LEP ELISA Kit No. ABIN625206, Anbieter: Anmelden zum Anzeigen
Antigen
  • ob
  • obese
  • LEPD
  • OB
  • OBS
  • leptin
  • Lep
  • LEP
  • lep
Alternativen
Cow (Bovine) Leptin ELISA Kit
Reaktivität
Ratte (Rattus)
Alternativen
Kits mit alternativen Reaktivitäten:
61
39
29
20
14
13
12
9
7
6
5
5
4
2
1
1
1
Methodentyp
Sandwich ELISA
Detektionsbereich
30-8000 pg/mL
Untere Nachweisgrenze
30 pg/mL
Applikation
ELISA
Optionen
Hersteller
Anmelden zum Anzeigen
Hersteller Produkt- Nr.
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Verwendungszweck Rat Leptin ELISA Kit for cell culture supernatants, plasma, and serum samples.
Proben Plasma, Cell Culture Supernatant, Serum
Analytische Methode Quantitative
Nachweismethode Colorimetric
Spezifität The Leptin ELISA kit shows no cross-reactivity with the following cytokines tested: rat CINC-2, CINC-3, CNTF, Fractalkine, IL-1 alpha, IL-1 beta, IL-4, IL-6, IL-10, GM-CSF, IFN-gamma, L-Selectin, Lix, MCP-1, MIP-3 alpha, beta-NGF, TIMP-1, TNF-alpha.
Kreuzreaktivität (Details) This ELISA kit shows no cross-reactivity with the following cytokines tested: rat CINC-2, CINC-3, CNTF, Fractalkine, IL-1alpha, IL-1beta, IL-4, IL-6, IL-10, GM-CSF, IFN-gamma, L-Selectin, Lix, MCP-1, MIP-3alpha, beta-NGF, TIMP-1, TNF-alpha.
Sensitivität < 30 pg/mL
Produktmerkmale
  • Strip plates and additional reagents allow for use in multiple experiments
  • Quantitative protein detection
  • Establishes normal range
  • The best products for confirmation of antibody array data
Bestandteile
  • Pre-Coated 96-well Strip Microplate
  • Wash Buffer
  • Stop Solution
  • Assay Diluent(s)
  • Lyophilized Standard
  • Biotinylated Detection Antibody
  • Streptavidin-Conjugated HRP
  • TMB One-Step Substrate
Benötigtes Material
  • Distilled or deionized water
  • Precision pipettes to deliver 2 μL to 1 μL volumes
  • Adjustable 1-25 μL pipettes for reagent preparation
  • 100 μL and 1 liter graduated cylinders
  • Tubes to prepare standard and sample dilutions
  • Absorbent paper
  • Microplate reader capable of measuring absorbance at 450nm
  • Log-log graph paper or computer and software for ELISA data analysis
Andere Bezeichnung Leptin (LEP ELISA Kit Abstract)
Hintergrund Lens epithelial cell protein LEP503
Gen-ID 113917
UniProt Q9WVB7
Forschungsgebiet Cardiovascular, Atherosclerosis, Hormones
Pathways JAK-STAT Signalweg, AMPK Signaling, Hormone Transport, Peptide Hormone Metabolism, Hormone Activity, Negative Regulation of Hormone Secretion, Regulation of Carbohydrate Metabolic Process, Feeding Behaviour, Monocarboxylic Acid Catabolic Process
Applikationshinweise Recommended Dilution for serum and plasma samples3 fold
Probenmenge 100 μL
Plattentyp Pre-coated
Protokoll
  1. Prepare all reagents, samples and standards as instructed in the manual.
  2. Add 100 μL of standard or sample to each well.
  3. Incubate 2.5 h at RT or O/N at 4 °C.
  4. Add 100 μL of prepared biotin antibody to each well.
  5. Incubate 1 h at RT.
  6. Add 100 μL of prepared Streptavidin solution to each well.
  7. Incubate 45 min at RT.
  8. Add 100 μL of TMB One-Step Substrate Reagent to each well.
  9. Incubate 30 min at RT.
  10. Add 50 μL of Stop Solution to each well.
  11. Read at 450 nm immediately.
Aufbereitung der Reagenzien
  1. Bring all reagents and samples to room temperature (18 - 25 °C) before use.
    2. Sample dilution: If your samples need to be diluted, 1x Assay Diluent D (Item K) should be used for dilution of serum/plasma/ culture supernatants. Suggested dilution for normal serum/plasma: 3 fold. Please note that levels of the target protein may vary between different specimens. Optimal dilution factors for each sample must be determined by the investigator.
    3. Assay Diluent D (Item K) and Assay Diluent B (Item E) should be diluted 5-fold with deionized or distilled water before use.
    4. Preparation of standard: Briefly spin the vial of Item C. Add 400 µL 1x Assay Diluent D (Item K) into Item C vial to prepare a 50 ng/mL standard solution. Dissolve the powder thoroughly by a gentle mix. Add 80 µL Leptin standard from the vial of Item C, into a tube with 420 µL 1x Assay Diluent D to prepare a 8,000 pg/mL standard solution. Pipette 400myl 1x Assay Diluent D into each tube. Use the stock standard solution to produce a dilution series . Mix each tube thoroughly before the next transfer. 1x Assay Diluent D serves as the zero standard (0 pg/mL). 200 µL 200 µL 200 µL 200 µL 200 µL 200myl 80 µL standard + 420 µL 8,000 2667 888.9 296.3 98.77 32.92 10.97 0 pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL
    5. If the Wash Concentrate (20x) (Item B) contains visible crystals, warm to room temperature and mix gently until dissolved. Dilute 20 ml of Wash Buffer Concentrate into deionized or distilled water to yield 400 ml of 1x Wash Buffer.
    6. Briefly spin the Detection Antibody vial (Item F) before use. Add 100 µL of 1x Assay Diluent B (Item E) into the vial to prepare a detection antibody concentrate. Pipette up and down to mix gently (the concentrate can be stored at 4 °C for 5 days). The detection antibody concentrate should be diluted 80-fold with 1x Assay Diluent B and used in step 4 of Part VI Assay Procedure.
    7. Briefly spin the HRP-Streptavidin concentrate vial (Item G) and pipette up and down to mix gently before use. HRP-Streptavidin concentrate should be diluted 200-fold with 1x Assay Diluent B (Item E). For example: Briefly spin the vial (Item G) and pipette up and down to mix gently . Add 50 µL of HRP-Streptavidin concentrate into a tube with 10 ml 1x Assay Diluent B to prepare a 200-fold diluted HRP-Streptavidin solution (don't store the diluted solution for next day use). Mix well.
Testdurchführung
  1. Bring all reagents and samples to room temperature (18 - 25 °C) before use. It is recommended that all standards and samples be run at least in duplicate.
    2. Add 100 µL of each standard (see Reagent Preparation step 2) and sample into appropriate wells. Cover well and incubate for 2.5 hours at room temperature or over night at 4 °C with gentle shaking.
    3. Discard the solution and wash 4 times with 1x Wash Solution. Wash by filling each well with Wash Buffer (300 myl) using a multi-channel Pipette or autowasher. Complete removal of liquid at each step is essential to good performance. After the last wash, remove any remaining Wash Buffer by aspirating or decanting. Invert the plate and blot it against clean paper towels.
    4. Add 100 µL of 1x prepared biotinylated antibody (Reagent Preparation step 6) to each well. Incubate for 1 hour at room temperature with gentle shaking.
    5. Discard the solution. Repeat the wash as in step
    6. Add 100 µL of prepared Streptavidin solution (see Reagent Preparation step 7) to each well. Incubate for 45 minutes at room temperature with gentle shaking.
    7. Discard the solution. Repeat the wash as in step
    8. Add 100 µL of TMB One-Step Substrate Reagent (Item H) to each well. Incubate for 30 minutes at room temperature in the dark with gentle shaking.
    9. Add 50 µL of Stop Solution (Item I) to each well. Read at 450 nm immediately.
Ergebnisberechnung

Calculate the mean absorbance for each set of duplicate standards, controls and samples, and subtract the average zero standard optical density. Plot the standard curve on log-log graph paper or using Sigma plot software, with standard concentration on the x-axis and absorbance on the y-axis. Draw the best-fit straight line through the standard points.
Typical Data: These standard curves are for demonstration only. A standard curve must be run with each assay. Rat Leptin concentration (pg/mL) O D =4 50 n m 0.01 0.1 1 10 Assay Diluent D 10 100 1,000 10,000
Sensitivity: The minimum detectable dose of Leptin is typically less than 30 pg/mL.
Recovery: Recovery was determined by spiking various levels of Leptin into normal rat serum, plasma and cell culture media. Mean recoveries are as follows: Sample Type Average % Recovery Range ( %) Serum 92.32 83-102 Plasma 84.25 75-94 Cell culture media 75.43 68-89
Linearity: Sample Type Serum Plasma Cell Culture Media 1:2 Average % of Expected 105.3 111.6 112.6 Range ( %) 94-113 102-122 103-124 1:4 Average % of Expected 108.1 119.2 113.5 Range ( %) 95-114 107-125 103-125
Reproducibility: Intra-Assay: CV<10 % Inter-Assay: CV<12 %

Testpräzision Intra-Assay: CV< 10 % Inter-Assay: CV< 12 %
Beschränkungen Nur für Forschungszwecke einsetzbar
Handhabung Avoid repeated freeze-thaw cycles.
Lagerung -20 °C
Informationen zur Lagerung The entire kit may be stored at -20°C for up to 1 year from the date of shipment. Avoid repeated freeze-thaw cycles. The kit may be stored at 4°C for up to 6 months. For extended storage, it is recommended to store at -80°C.
Haltbarkeit 6 months
Bilder des Herstellers
ELISA image for Leptin (LEP) ELISA Kit (ABIN625206) Leptin (LEP) ELISA Kit
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Zhang, Wang, Long, Wang, Li, Gao, Tian: "Alteration of sweet taste in high-fat diet induced obese rats after 4 weeks treatment with exenatide." in: Peptides, Vol. 47, pp. 115-23, 2013 (PubMed).

Huang, Korivi, Tsai, Yang, Tsai: "Supplementation of Lactobacillus plantarum K68 and Fruit-Vegetable Ferment along with High Fat-Fructose Diet Attenuates Metabolic Syndrome in Rats with Insulin Resistance." in: Evidence-based complementary and alternative medicine : eCAM, Vol. 2013, pp. 943020, 2013 (PubMed).

Sloboda, Fève, Thornton, Nzietchueng, Regnault, Simon, Labat, Louis, Max, Muscat, Osborne-Pellegrin, Lacolley, Benetos: "Fatty acids impair endothelium-dependent vasorelaxation: a link between obesity and arterial stiffness in very old Zucker rats." in: The journals of gerontology. Series A, Biological sciences and medical sciences, Vol. 67, Issue 9, pp. 927-38, 2012 (PubMed).

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