Leptin, also known as OB, is a polypeptide hormone that regulates adipose tissue mass and energy balance. In mice (ob/ob) and humans, inactivating mutations of the OB gene can cause obesity. Leptin signals through the transmembrane Leptin R in the hypothalamus but can also bind shorter and soluble forms of Leptin R which do not signal. Adipocytes increase Leptin expression as cell size increases, resulting in depressed appetite and increased energy expenditure. Low Leptin levels prompt increased appetite and decreased energy expenditure. Leptin can also block the onset of puberty or menses during starvation, extreme exercise-induced weight loss, anorexia, or cancer-induced cachexia.
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Key Product Details
Assay Type
Solid Phase Sandwich ELISA
Assay Range
31.2-2000 pg/mL
Sample Type
Cell culture supernates, serum, and plasma
Note: Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet
Note: Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet
Reactivity
Human
Human Leptin DuoSet ELISA 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
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Product Summary for Human Leptin DuoSet ELISA
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 Specifications
Assay Format
96-well strip plate (sold separately)
Sample Volume Required
100 µL
Detection Method
Colorimetric ELISA - 450nm (TMB)
Conjugate
Biotin
Specificity
Label
HRP
Scientific Data Images for Human Leptin DuoSet ELISA
Human Leptin / OB ELISA Standard Curve
Kit Contents for Human Leptin DuoSet ELISA
- 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)
Preparation and Storage
Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Background: Leptin/OB
Additional Leptin/OB Products
Product Documents for Human Leptin DuoSet ELISA
Certificate of Analysis
To download a Certificate of Analysis, please enter a lot or batch number in the search box below.
Note: Certificate of Analysis not available for kit components.
Product Specific Notices for Human Leptin DuoSet ELISA
For research use only
Citations for Human Leptin DuoSet ELISA
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Customer Reviews for Human Leptin DuoSet ELISA (7)
4.9 out of 5
7 Customer Ratings
Have you used Human Leptin DuoSet ELISA?
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Sample Tested: Stromal vascular fractionVerified Customer | Posted 03/06/2025
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Sample Tested: Human cell conditioned mediumVerified Customer | Posted 01/24/2025
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Sample Tested: SerumVerified Customer | Posted 10/17/2023
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Sample Tested: EDTA PlasmaVerified Customer | Posted 12/04/2019diluted 1:50, followed the same procedures in the protocol.
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Sample Tested: Human serumVerified Customer | Posted 08/14/2019Sample diluted 1/40, very good replicates. Very reliable across plates. Used 384-well plate format
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Sample Tested: EDTA PlasmaVerified Customer | Posted 12/20/2017
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Sample Tested: SerumVerified Customer | Posted 08/09/2017serum dilution used - 1:2.5
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Protocols
View specific protocols for Human Leptin DuoSet ELISA (DY398):
GENERAL ELISA PROTOCOL
Plate Preparation
- 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.
- 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.
- Block plates by adding 300 μL Reagent Diluent to each well. Incubate at room temperature for a minimum of 1 hour.
- Repeat the aspiration/wash as in step 2. The plates are now ready for sample addition.
Assay Procedure
- 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.
- Repeat the aspiration/wash as in step 2 of Plate Preparation.
- 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.
- Repeat the aspiration/wash as in step 2 of Plate Preparation.
- 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.
- Repeat the aspiration/wash as in step 2.
- Add 100 μL of Substrate Solution to each well. Incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
- Add 50 μL of Stop Solution to each well. Gently tap the plate to ensure thorough mixing.
- 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.
Find general support by application which include: protocols, troubleshooting, illustrated assays, videos and webinars.
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- View all Protocols, Troubleshooting, Illustrated assays and Webinars
