Human OX40 Ligand / TNFSF4 Protein, Mouse IgG2a Fc Tag, low endotoxin

INTRODUCTION

QC & Assay Protocol

Citation & Background

Synonym
OX40L,TNFSF4,CD252,Glycoprotein Gp34,TXGP1
Source
Human OX40 Ligand, Mouse IgG2a Fc Tag, low endotoxin (OXL-H5250) is expressed from human 293 cells (HEK293). It contains AA Gln 51 - Leu 183 (Accession # P23510-1).
Predicted N-terminus: Gln 51
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Molecular Characterization
This protein carries a mouse IgG2a Fc tag at the C-terminus.
The protein has a calculated MW of 42.3 kDa. The protein migrates as 53-60 kDa under reducing (R) condition (SDS-PAGE) due to glycosylation.
Endotoxin
Less than 0.1 EU per μg by the LAL method.
Purity
>95% as determined by SDS-PAGE.

Formulation
Lyophilized from 0.22 μm filtered solution in 50 mM Tris, 100 mM Glycine, pH7.5. Normally trehalose is added as protectant before lyophilization.
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Reconstitution
Please see Certificate of Analysis for specific instructions.
For best performance, we strongly recommend you to follow the reconstitution protocol provided in the CoA.
Storage
For long term storage, the product should be stored at lyophilized state at -20°C or lower.
Please avoid repeated freeze-thaw cycles.
No activity loss is observed after storage at:
1. 4-8^°C for 12 months in lyophilized state;
2. -70^°C for 3 months under sterile conditions after reconstitution.

SDS-PAGE

Human OX40 Ligand, Mouse IgG2a Fc Tag, low endotoxin on SDS-PAGE under reducing (R) condition. The gel was stained overnight with Coomassie Blue. The purity of the protein is greater than 95%.

Bioactivity

Immobilized Human OX40, Fc Tag (Cat. No. OX0-H5255) at 0.5 μg/mL (100 μL/well) can bind Human OX40 Ligand, Mouse IgG2a Fc Tag, low endotoxin (Cat. No. OXL-H5250) with a linear range of 0.8-13 ng/mL (QC tested).

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FACS assay shows that recombinant Biotinylated FACS

FACS assay shows that recombinant Human OX40 Ligand, Mouse IgG2a Fc Tag, low endotoxin (Cat. No. OXL-H5250) can bind to 293T cell overexpressing human OX40. The concentration of OX40 Ligand is 0.003 ug/ml (Routinely tested).

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FACS analysis shows that the binding of Human OX40 Ligand, Mouse IgG2a Fc Tag, low endotoxin (Cat. No. OXL-H5250) to 293T overexpressing OX40 was inhibited by increasing concentration of neutralizing anti-OX40 antibody. The concentration of OX40 Ligand used is 0.002 ug/ml. IC50=3.021 ug/ml (Routinely tested).

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Background
Tumor necrosis factor ligand superfamily member 4 (TNFSF4) is also known as glycoprotein Gp34, OX40 ligand (OX40L), TAX transcriptionally-activated glycoprotein 1 and CD252, which belongs to the tumor necrosis factor family. TNFSF4 is the ligand for CD134 and is expressed on such cells as DC2s (a subtype of dendritic cells) enabling amplification of Th2 cell differentiation. The interaction of TNFSF4-TNFSF4 is involved in the pathogenesis of multiple autoimmune and inflammatory diseases such as systemic lupus erythematosus (SLE), carotid artery disease and cancer. Furthermore, similar to other TNF superfamily members, membrane-bound OX40 Ligand (TNFSF4) exists as a homotrimer. Human TNFSF4 shares 46% amino acid sequence identity with its mouse counterpart.
References
- (1) Compaan D.M., et al., 2006, Structure 14:1321-1330.
- (2) Maizels RM., et al., 2003, Nat. Rev. Immunol. 3 (9): 733–44.
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