Osteoprotegerin

IgG

Mouse

Osteoprotegerin

0. 1 mg/ml Storage

ELISA: Use at a concentration of 0. 5 - 1 µ g/ml. The detection limit for mouse osteoprotegerin is approximately 0. 1 ng/well. Neut: To measure the ability of the antibody to neutralize the bioactivity of mouse OPG soluble recombinant mouse OPG/Fc is incubated with various concentrations of the antibody in media containing actinomycin D for 1 hour at 37 C in 96 well plates. WB: Use at a concentration of 1 - 2 µ g/ml. This concentration detects mouse OPG at approximately 2 ng/lane and 5 ng/lane under nonreducing and reducing conditions respectively. Reduced mouse OPG/Fc has a calculated molecular mass of approximately 70. 9 kDa. As a result of glycosylation the recombinant protein migrates as a 100 kDa band in SDS-PAGE under reducing conditions. Cross-linked recombinant human TRAIL is then added and incubated for an additional 30 minutes. Following preincubation the mixture is added to confluent cultures of L929 cells in 96 well plates. The assay mixture (total volume of 150 µ l/well) over a monolayer of L929 cells containing antibody (concentrations of 0. 01-100 µ g/ml) soluble recombinant mouse OPG/Fc (0. 1 µ g/ml) actinomycin D (1 µ g/ml) and cross-linked human TRAIL (20 ng/ml) is incubated at 37 C for 24 hours in a humidified CO2 incubator. At the end of the incubation period the media is removed and the cells are fixed with 5% formaldehyde and then stained with crystal violet. The stain is subsequently dissolved in 100 µ l of 33% acetic acid and the optical density is read at 540 nm. The Neutralization Dose50 (ND50) for anti-mouse osteoprotegerin is 0. 5-2 µ g/ml in the presence of 0. 1 µ g/ml of soluble recombinant mouse OPG/Fc using the TRAIL sensitive mouse L929 cytolytic assay. The ND50 is the concentration of antibody required to yield one-half maximal inhibition of the cytokine activity on a responsive cell line when the cytokine is present at a concentration just high enough to elicit a maximum response. The exact concentration of antibody required to neutralize recombinant mouse OPG/Fc activity is dependent on the cytokine concentration cell type growth conditions and the type of activity studied. Not tested in other applications. Optimal dilutions/concentrations should be determined by the end user. Cellular

Acts as decoy receptor for RANKL and thereby neutralizes its function in osteoclastogenesis. Inhibits the activation of osteoclasts and promotes osteoclast apoptosis in vitro. Bone homeostasis seems to depend on the local RANKL/OPG ratio. May also play a role in preventing arterial calcification. May act as decoy receptor for TRAIL and protect against apoptosis. TRAIL binding blocks the inhibition of osteoclastogenesis.

Secreted.

Up-regulated by increasing calcium-concentration in the medium and estrogens. Down-regulated by glucocorticoids.

The N-terminus is blocked.

Contains 2 death domains.