Human OX40/TNFRSF4 ELISA Kit
Protocol Booklet
Key features and details
- Specification:
- Sensitivity:
- Standard Curve Range:
- Standard Curve Gradient:
- Number of Incubations:
- Sample Volume:
- Assay type:
- Operation Duration:
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Brand:
CAT.NO. : AEH0120
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Product Details
Background
OX40 (CD134; TNFRSF4) is a T cell costimulatory molecule of the TNF receptor superfamily that coordinates with other membrane-bound costimulators such as CD28, CD40, CD30, CD27 and 4-1BB. OX40 is expressed on naïve CD4+ T cells only after engagement of the TCR by antigen presenting cells (APC; dendritic and B cells), and costimulation by CD40/CD40 ligand and CD28/B7. It is maximal at 2‑5 days post activation, or 4 hours post reactivation of memory T cells. Human OX40 is a 48 kDa type I transmembrane glycoprotein with a 28 amino acid (aa) signal sequence, a 185 aa extracellular domain (ECD) that has four TNFR-Cys repeats and an O-glycosylated hinge region, a 20 aa transmembrane segment, and a 41 aa cytoplasmic domain. The ECD of human OX40 shows 71%, 68%, 67%, 64% and 64% aa identity with feline, canine, rabbit, mouse and rat OX40 ECD, respectively. Engagement of OX40 on activated CD4+ T cells by OX40 ligand on activated dendritic cells promotes T cell survival and proliferation, prolongs the immune response, and enhances the number of cells making the transition from effector to memory T cells. OX40 signal transduction includes binding TNF receptor-associated factors (TRAFs), and activating NF kappa B and PI3 kinase to enhance expression of cytokines, antiapoptotic Bcl-2 family members, survivin and the chemokine receptor CXCR5. CXCR5 promotes T cell migration to germinal centers to deliver B cell help. Studies using knockout or transgenic mice, and agonistic or blocking antibodies, show that OX40/OX40L interaction is critical for establishing or reactivating memory T cells and breaking immune tolerance. Blockade of OX40 engagement is efficacious in animal models of allergic airway inflammation, graft-versus-host disease and autoimmune disease.
Typical data
| pg/ml | O.D. | Average | Corrected | |
| 0.00 | 0.0168 | 0.0150 | 0.0159 | |
| 12.29 | 0.0413 | 0.0360 | 0.0387 | 0.0228 |
| 30.72 | 0.0754 | 0.0717 | 0.0736 | 0.0577 |
| 76.80 | 0.1554 | 0.1469 | 0.1512 | 0.1353 |
| 192.00 | 0.3511 | 0.3457 | 0.3484 | 0.3325 |
| 480.00 | 0.8416 | 0.8055 | 0.8236 | 0.8077 |
| 1200.00 | 1.8010 | 1.8460 | 1.8235 | 1.8076 |
| 3000.00 | 3.1860 | 3.2240 | 3.2050 | 3.1891 |
Precision
| Intra-assay Precision | Inter-assay Precision | |||||
| Sample Number | S1 | S2 | S3 | S1 | S2 | S3 |
| 22 | 22 | 22 | 6 | 6 | 6 | |
| Average(pg/ml) | 68.3 | 374.5 | 1141.6 | 69.2 | 348.2 | 1106.9 |
| Standard Deviation | 2.1 | 23.5 | 44.4 | 3.9 | 12.7 | 52.2 |
| Coefficient of Variation(%) | 3.0 | 6.3 | 3.9 | 5.6 | 3.7 | 4.7 |
Inter-assay Precision (Precision between assays) Three samples of known concentration were tested six times on one plate to assess intra-assay precision.
Spike Recovery
The spike recovery was evaluated by spiking 3 levels of human OX40/TNFRSF4 into health human serum sample. The un-spiked serum was used as blank in this experiment.
The recovery ranged from 93% to 120% with an overall mean recovery of 103%.
The recovery ranged from 93% to 120% with an overall mean recovery of 103%.
Sample Values
| Sample Matrix | Sample Evaluated | Range (pg/ml) | Detectable (%) | Mean of Detectable (pg/ml) |
| Serum | 30 | 19.5-94.28 | 100 | 59.08 |
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