Human Leptin ELISA Kit
Assay range: 31.2 - 2000pg/mL
Kit Size: 96 wells/kit
INTRODUCTION
Leptin is a product of the mouse obese gene secreted by adipocytes. It controls body fat deposition and energy balance. Leptin deficiency in humans and mice can result in obesity 1. Circulating levels of Leptin are regulated by food consumption, insulin levels and pregnancy status 2. Mice with leptin synthesis-blocking mutations in the obese gene develop obesity and diabetes as well as decreased activity, metabolism, and body temperature 3.
PRINCIPLE OF THE ASSAY
This assay is a quantitative sandwich enzyme-linked immunosorbent assay (ELISA). The microtiter plate is pre-coated with a polyclonal antibody specific for human Leptin. Standards and samples are pipetted into the wells and any human Leptin present is bound by the immobilized antibody. After washing away any unbound substances, a biotin labelled polyclonal antibody specific for mouse Leptin is added to the wells. After wash step to remove any unbound reagents, streptavidin-horseradish peroxidase conjugate (STP-HRP) is added. After the last wash step, an HRP substrate solution is added and color develops in proportion to the amount of human Leptin bound initially. The assay is stopped, and the optical density of the wells is determined using a microplate reader. Since the increases in absorbance are directly proportional to the amount of captured human Leptin, the unknown sample concentration can be interpolated from a reference curve included in each assay.
ASSAY PERFORMANCE
TYPICAL STANDARD CURVE
The following standard curve is provided for demonstration only. A standard curve should be generated for each set of sample assay.
Mouse Leptin
(pg/mL)
Absorbance
(450 nm)
Blanked Absorbance
0
0.138
0
31.25
0.160
0.022
62.5
0.206
0.068
125
0.268
0.130
250
0.434
0.296
500
0.871
0.733
1000
1.667
1.529
2000
2.517
2.379
Mouse Leptin standard curve (4-parameter)
ASSAY CHARACTERISTICS
A. Sensitivity
The lowest level of mouse Leptin that can be detected by this assay is 31.25 pg/mL.
B. Precision
Intra-assay Precision (Precision within an assay) C.V. <2.9%. Inter-assay Precision (Precision between assays) C.V. <6.1%.