Soluble Leptin Receptor (sLEP-R) ELISA Kit

Sample

Serum and plasma

Species Reactivity

Human

Detection Method

sELISA

Intended Use

The sLEP-R ELISA Kit is intended to be used for quantitative measurement of soluble human leptin receptor (sLEP-R) in human serum or plasma for research purposes.

Contents of Kit

1. Microtiter plate: ready for use, coated with human anti-hLeptin-R antibody. Wells are separately breakable. (8x12) wells
2. Standards (A-H): lyophilized, (recombinant hLeptin Receptor Standards), concentrations are given on vial labels and on quality certificate. 8 x 1 mL
3. Control Serum 1 (KS1): lyophilized, (human serum), concentration is given on quality certificate. 1 x 500 μL
4. Control Serum 2 (KS2): lyophilized, (human serum), concentration is given on quality certificate. 1 x 500 μL
5. Antibody Conjugate (AK): ready for use, contains biotinylated anti-hLeptin-R-antibody. 1 x 12 mL
6. Enzyme Conjugate (EK): contains Streptavidin-Peroxidase Conjugate. 1 x 12 mL
7. Dilution Buffer (VP): ready for use. Please shake before use! 1 x 60 mL
8. Washing Buffer (WP): 20-fold concentrated solution. 1 x 50 mL
9. Substrate (S): ready for use, horseradish-peroxidase-(HRP) substrate, stabilised Tetramethylbencidine. 1 x 12 mL
10. Stopping Solution (SL): ready for use, 0.2 M sulfuric acid. 1 x 12 mL
11. Sealing Tape: for covering the microtiter plate. 3

Storage

Store the kit at 2-8°C after receipt until its expiry date. The lyophilized reagents should be stored at –20 °C after reconstitution. Avoid repeated thawing and freezing.
The shelf life of the components after initial opening is warranted for 4 weeks, store the unused strips and microtiter wells airtight together with the desiccant at 2-8°C in the clip-lock bag, use in the frame provided. The reconstituted components standards A-H and Control Sera KS1 and KS2 must be stored at –20°C (max. 4 weeks). For further use, thaw quickly but gently (avoid temperature increase above room temperature and avoid excessive vortexing). Up to 3 of the freeze-thaw cycles did not influence the assay. The 1:20 diluted Washing Buffer WP is 4 weeks stable at 2-8°C.

Precision

Intra-Assay Variance
Native human serum samples were measured (n=18) repeatedly at various positions of the microtiter plate. Intra-Assay variability was on average < 6%.

Inter-Assay-Variability
Serum samples were measured in independent tests. The coefficient of variation was 8% on average.

Sensitivity

The analytical sensitivity of the test system is defined, as the concentration sLEP-R, that is distinguishable from the blank / background. Sensitivity was assessed by measuring the blank and calculating the theoretical concentration of the blank + 2SD. The analytical sensitivity of the sLEP-R ELISA Kit is 0.0385 ng/mL on average. The limit of quantification (LOQ, as Serial dilutions with 3 different sera with max. difference of ± 20% to expected value) is 0.56 ng/mL.

General Description

The adipokine leptin realizes signal transduction via four different leptin receptor (LEP-R) isoforms. The amount of functionally active LEP-R, however, is affected by constitutive shedding of the extracellular domain. The product of the cleavage process, the so-called soluble leptin receptor (sLEP-R, soluble Leptin receptor), is the main binding protein for leptin in human blood and modulates its bioavailability. Concentrations of sLEP-R are differentially regulated in metabolic disorders, such as type 1 diabetes mellitus or obesity and can therefore enhance or reduce leptin sensitivity. Lipotoxicity and apoptosis increase LEP-R cleavage via ADAM10-dependent mechanisms. In contrast, although increased sLEP-R concentrations seem directly to inhibit leptin effects, reduced amounts of sLEP-R may reflect decreased membrane expression of LEP-R. These findings, in part, explain alterations of leptin sensitivity that are associated with changes in serum sLEP-R concentrations seen in metabolic disorders.

Standard Curve

The exemplary data and the standard curve in Figure below cannot be used for the calculation of the test results. You have to establish a standard curve for each test you conduct.