Leptin ELISA Kit (LEP)

Details for Product LEP ELISA Kit No. ABIN5665014, Supplier: Log in to see
Antigen
  • ob
  • obese
  • LEPD
  • OB
  • OBS
  • leptin
  • Lep
  • LEP
  • lep
Reactivity
Sheep (Ovine)
Alternatives
Kits with alternative reactivity to:
56
40
32
19
15
13
11
9
7
6
6
5
4
4
1
1
1
Method Type
Sandwich ELISA
Detection Range
0.15 ng/mL - 10 ng/mL
Minimum Detection Limit
0.15 ng/mL
Application
ELISA
Options
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Sample Type Cell Culture Supernatant, Cell Lysate, Plasma, Serum, Tissue Homogenate
Analytical Method Quantitative
Detection Method Colorimetric
Specificity

This assay has high sensitivity and excellent specificity for detection of Leptin (LEP).
No significant cross-reactivity or interference between Leptin (LEP) and analogues was observed.

Sensitivity 0.054 ng/mL
Components
  • Pre-coated, ready to use 96-well strip plate
  • Plate sealer for 96 wells
  • Standard Diluent
  • Assay Diluent A
  • Assay Diluent B
  • Stop Solution
  • Standard
  • Detection Reagent A
  • Detection Reagent B
  • TMB Substrate
  • Wash Buffer (30 x concentrate)
  • Instruction manual
Material not included
  • Microplate reader with 450 nm filter.
  • Precision single or multi-channel pipettes and disposable tips.
  • Eppendorf Tubes for diluting samples.
  • Deionized or distilled water.
  • Absorbent paper for blotting the microtiter plate.
  • Container for Wash Solution
Plasmids, Primers & others Plasmids, Primers & others Leptin products on genomics-online (e.g. as negative or positive controls)
Antigen Leptin (LEP)
Pathways JAK-STAT Signaling, 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
Application Notes
  • Limited by the current condition and scientific technology, we cannot completely conduct the comprehensive identification and analysis on the raw material provided by suppliers. So there might be some qualitative and technical risks to use the kit.
  • The final experimental results will be closely related to validity of the products, operation skills of the end users and the experimental environments. Please make sure that sufficient samples are available.
  • Kits from different batches may be a little different in detection range, sensitivity and color developing time.
  • Do not mix or substitute reagents from one kit lot to another. Use only the reagents supplied by manufacturer.
  • Protect all reagents from strong light during storage and incubation. All the bottle caps of reagents should be covered tightly to prevent the evaporation and contamination of microorganism.
  • There may be some foggy substance in the wells when the plate is opened at the first time. It will not have any effect on the final assay results. Do not remove microtiter plate from the storage bag until needed.
  • Wrong operations during the reagents preparation and loading, as well as incorrect parameter setting for the plate reader may lead to incorrect results. A microplate plate reader with a bandwidth of 10nm or less and an optical density range of 0-3 O.D. or greater at 450 ± 10nm wavelength is acceptable for use in absorbance measurement. Please read the instruction carefully and adjust the instrument prior to the experiment.
  • Even the same operator might get different results in two separate experiments. In order to get better reproducible results, the operation of every step in the assay should be controlled. Furthermore, a preliminary experiment before assay for each batch is recommended.
  • Each kit has been strictly passed Q.C test. However, results from end users might be inconsistent with our in-house data due to some unexpected transportation conditions or different lab equipments. Intra-assay variance among kits from different batches might arise from above factors, too.
  • Kits from different manufacturers for the same item might produce different results, since we have not compared our products with other manufacturers.
Sample Volume 100 μL
Assay Time 3 h
Plate Pre-coated
Protocol The test principle applied in this kit is Sandwich enzyme immunoassay. The microtiter plate provided in this kit has been pre-coated with an antibody specific to Leptin (LEP). Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody specific to Leptin (LEP). Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added, only those wells that contain Leptin (LEP), biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of Leptin (LEP) in the samples is then determined by comparing the O.D. of the samples to the standard curve.
Assay Procedure
  1. Prepare all reagents, samples and standards,
  2. Add 100μL standard or sample to each well. Incubate 2 hours at 37 °C,
  3. Aspirate and add 100μL prepared Detection Reagent A. Incubate 1 hour at 37 °C,
  4. Aspirate and wash 3 times,
  5. Add 100μL prepared Detection Reagent B. Incubate 30 minutes at 37 °C,
  6. Aspirate and wash 5 times,
  7. Add 90μL Substrate Solution. Incubate 10-20 minutes at 37 °C,
  8. Add 50μL Stop Solution. Read at 450nm immediately.
Assay Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level Leptin (LEP) were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level Leptin (LEP) were tested on 3 different plates, 8 replicates in each plate.
CV(%) = SD/meanX100
Intra-Assay: CV<10%
Inter-Assay: CV<12%

Restrictions For Research Use only
Precaution of Use The Stop Solution suggested for use with this kit is an acid solution. Wear eye, hand, face, and clothing protection when using this material.
Handling Advice The stability of kit is determined by the loss rate of activity. The loss rate of this kit is less than 5 % within the expiration date under appropriate storage condition.
To minimize extra influence on the performance, operation procedures and lab conditions, especially room temperature, air humidity, incubator temperature should be strictly controlled. It is also strongly suggested that the whole assay is performed by the same operator from the beginning to the end.
Storage 4 °C
Storage Comment
  • For unopened kit: All the reagents should be kept according to the labels on vials. The Standard, Detection Reagent A, Detection Reagent B and the 96-well strip plate should be stored at -20°C upon receipt while the others should be at 4°C.
  • For opened kit: When the kit is opened, the remaining reagents still need to be stored according to the above storage condition. Besides, please return the unused wells to the foil pouch containing the desiccant pack, and reseal along entire edge of zip-seal.
    Note: It is highly recommended to use the remaining reagents within 1 month provided this is within the expiration date of the kit.
  • For ELISA kit, 1 day storage at 37°C can be considered as 2 months at 4°C, which means 3 days at 37°C equaling 6 months at 4°C.
Expiry Date 6 months
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