Rat Receptor Activator Of Nuclear Factor Kappa B Ligand (RANkL) ELISA Kit

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to RANkL. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to RANkL. Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After the TMB substrate solution is added, only those wells that contain RANkL, 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 RANkL in the samples is then determined by comparing the O.D. of the samples to the standard curve.

For use with: Serum, Plasma, Tissue homogenates, Cell lysates, Cell culture supernates, and Other biological fluids.

For Research Use Only. Not for use in diagnostic procedures.

Target Information

Cytokine that binds to TNFRSF11B/OPG and to TNFRSF11A/RANK. Osteoclast differentiation and activation factor. Augments the ability of dendritic cells to stimulate naive T-cell proliferation. May be an important regulator of interactions between T-cells and dendritic cells and may play a role in the regulation of the T-cell-dependent immune response. May also play an important role in enhanced bone-resorption in humoral hypercalcemia of malignancy. Induces osteoclastogenesis by activating multiple signaling pathways in osteoclast precursor cells, chief among which is induction of long lasting oscillations in the intracellular concentration of Ca (2+) resulting in the activation of NFATC1, which translocates to the nucleus and induces osteoclast-specific gene transcription to allow differentiation of osteoclasts. During osteoclast differentiation, in a TMEM64 and ATP2A2-dependent manner induces activation of CREB1 and mitochondrial ROS generation necessary for proper osteoclast generation.

GENE ID	117516
SWISS PROT	Q9ESE2
SYNONYMS	CD254; TNFSF11; ODF; OPGL; TRANCE; HRANKL2; SOdf; Tumor Necrosis Factor(ligand)superfamily Member 11; TNF-related activation-induced cytokine

Materials Supplied

Kit Components	96 Wells Quantity/Size
Pre-coated, ready-to-use 96-well strip plate	1 plate
Plate sealer for 96 wells	2
Standard	2 tubes
Diluent buffer	1 bottle
Detection Reagent A	1 bottle
Detection Reagent B	1 bottle
TMB Substrate	1 tube
Stop Solution	1 tube
Wash Buffer (30X concentrate)	1 tube
Product data sheet	1 copy

Storage

Storage

The TMB Substrate, Wash Buffer (30X concentrate), and the Stop Solution should be stored at 4°C upon receipt, while the other items should be stored at -20°C.

Performance Characteristics

REPEATABILITY	Intra-assay Precision (Precision within an assay): 3 samples with low, middle, and high-level RANkL were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level RANkL were tested on 3 different plates, with 8 replicates in each plate. CV(%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%
SENSITIVITY	The minimum detectable dose was 5.3pg/mL.
ASSAY RANGE	15.6–1,000pg/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of RANkL. No significant cross-reactivity or interference between RANkL and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between RANkL and all analogs, therefore, cross reactivity may still exist.