sRANKL (total) Human ELISA (Osteoprotegerin Ligand)

Research topic

Bone and cartilage metabolism

Features

• The free assay time is about 18 hours.
• The kit measures total sRANKL

Storage/Shipping

Store the kit at 2–8°C. Under these conditions, the kit is stable until the expiration date (see label on the box).

Summary

sRANKL, receptor activator of nuclear factor (NF)-κB ligand (also: osteoprotegerin ligand, OPGL), is a part of the TNF superfamily with high similarity to other members of that protein species. (SwissProt Nr. O14788).

Three isoforms are produced by alternate splicing, two type II membrane proteins (ISOFORM 1, 317 AA, and ISOFORM 3, 270 AA), and a secreted molecule (ISOFORM 2, 244 AA). ISOFORM 1 is identical to previously reported RANKL and possesses intracellular, transmembrane, and extracellular domains; ISOFORM 2 does not have the intracellular and transmembrane domains, and ISOFORM 3 does not have the intracellular domain. A soluble form arises by proteolytic processing from membrane isoforms.

Although all forms are bioactive, the membrane-bound proteins seem to be the homeostatic forms, while the production of soluble RANKL signals pathological conditions.

RANKL, RANK, and osteoprotegerin (OPG) have been identified as the key molecular regulation system for bone remodelling. RANKL is the main stimulatory factor for the formation of mature osteoclasts and is essential for their survival. Therefore, an increase in RANKL expression leads to bone resorption and bone loss. RANKL is produced by osteoblastic lineage cells and activated T lymphocytes. It activates its specific receptor RANK, which is located on osteoclasts and dendritic cells. The effects of RANKL are counteracted by OPG, which is secreted by various tissues and acts as an endogenous soluble receptor antagonist.

Imbalances of the RANKL/OPG system have been related to the pathogenesis of Paget’s disease, benign and malignant bone tumors, postmenopausal osteoporosis, rheumatoid arthritis, bone metastases and hypercalcemia. Several studies using animal models have shown that restoring the RANKL/OPG balance (e.g. by administering OPG) reduces the severity of these disorders.

Indication

• Postmenopausal and senile osteoporosis
• Diseases with locally increased bone resorption activity
• Paget´s disease
• Periodontal disease
• Cardiovascular disease, arterial calcification
• Inflammatory diseases
• Immunological disorders
• Arthritis
• Oncology

Assay format

Sandwich ELISA, Biotin-labelled antibody

Sample requirements

5 µl/well

Applications

Plasma-Citrate, Plasma-EDTA, Plasma-Heparin, Serum

Calibration Curve

Limit of detection

Analytical Limit of Detection (LOD) is calculated from the real sRANKL values in wells and is 0.1pmol/l

Limit of quantification

Assay Sensitivity (LOQ) takes the dilution of samples into consideration and is calculated according to the formula: Assay Sensitivity = Analytical Limit of Detection x sample dilution = 0.1pmol/l x 100 = 10 pmol/l

Intra-assay (Within-Run, n=8)

CV = 6.2 %

Inter-assay (Run-to-Run, n=4)

CV = 7.2 %

Spiking Recovery

95 %

Dilution Linearity

109 %

Cross-Reactivity

Human

References to this product

• Kyrtsonis MC, Vassilakopoulos TP, Siakantaris MP, Kokoris SI, Gribabis DA, Dimopoulou MN, Angelopoulou MK, Pangalis GA . Serum syndecan-1, basic fibroblast growth factor and osteoprotegerin in myeloma patients at diagnosis and during the course of the disease. Eur J Haematol . Apr;72(4):252-8 (2004)

References to summary

• Hofbauer L.C et al. Effect of oral contraceptives on circulatingos­teoprotegerin and soluble RANK ligand serum levels in healthy young women. Clin Endocrinol (2004) 60: 214–219
• Ito S. et al. Crystal structure of the extracellular domain of mouse ligand at2.2-A resolution. J Biol Chem (2002) 22: 6631–6636.
• Lam J. et al. Crystal structure of the TRANCE/RANKL cytokine reveals determinants of receptorligand specifity. J Clin Invest (2001) 108: 971–979.
• Lacey D.L, et al., Osteoprotegerin ligand is a cytokine that regulates osteoclast differentiation and activation. Cell (1998), 93:165–176.
• Kong Y.Y. et al., OPGL is a key regulator of osteoclastogenesis, lymphocyte development and lymph-node organogenesis. Nature (1999), 397: 315–323.
• Hsu H. et al., Tumor necrosis factor receptor family member RANK mediates osteoclast differentiation and activation induced by osteoprotegerin ligand. Proc Natl Acad Sci (1999), 96:3540–3545.
• Josien R. et al, TRANCE, a tumor necrosis factor family member, enhances the longevity and adjuvant properties of dendritic cells in vivo. J Exp Med (2000), 191: 495–502.
• Fuller K. et al., TRANCE is necessary and sufficient for osteoblast-mediated activation of bone resorption in osteoclasts. J Exp Med (1998), 188: 997–1001.
• Nakashima T. et. al., Protein expression and functional difference of membrane-bound and soluble receptor activator of NF-kappaB ligand: modulation of the expression by osteotropic factors and cytokines. Biochem Biophys Res Commun (2000), 275(3):768–775.
• Kong Y.Y. et al., Activated T cells regulate bone loss and joint destruction in adjuvant arthritis through osteoprotegerin ligand. Nature (1999), 402: 304–309.
• Hofbauer L.C. & A.E. Heufelder, Role of receptor activator of nuclear factor-KB ligand and osteoprotegerin in bone cell biology. J Mol Med (2001), 79: 243–253.
• Hofbauer L.C. & A.E. Heufelder, The Role of Osteoprotegerin and Receptor Activator of Nuclear Factor κB Ligand in the Pathogenesis and Treatment of Rheumatoid Arthritis. Arthritis & Rheumatism (2001), 44:253–259.
• Hofbauer LC, et al. The role of receptor activator of nuclear factor-kappaB ligand and osteoprotegerin in the pathogenesis and treatment of metabolic bone diseases. J Clin Endocrinol Metab (2000), 85: 2355–2363.
• Teitelbaum S.L., Bone resorption by osteoclasts. Science (2000), 289: 1504–1508.