Mouse TGF beta1 ELISA

Additional Information

Assay Background

TGF beta (Transforming Growth Factor beta) is a multifunctional protein that controls proliferation, differentiation and other functions in many cell types. Many cells synthesize TGFB1 and have specific receptors for it. It positively and negatively regulates many other growth factors. It plays an important role in bone remodeling as it is a potent stimulator of osteoblastic bone formation, causing chemotaxis, proliferation and differentiation in committed osteoblasts (By similarity). Stimulates sustained production of collagen through the activation of CREB3L1 by regulated intramembrane proteolysis (RIP). Can promote either T-helper 17 cells (Th17) or regulatory T-cells (Treg) lineage differentiation in a concentration-dependent manner. At high concentrations, leads to FOXP3-mediated suppression of RORC and down-regulation of IL-17 expression, favoring Treg cell development. At low concentrations in concert with IL-6 and IL-21, leads to expression of the IL-17 and IL-23 receptors, favoring differentiation to Th17 cells. Mediates SMAD2/3 activation by inducing its phosphorylation and subsequent translocation to the nucleus. Can induce epithelial-to-mesenchymal transition (EMT) and cell migration in various cell types.

Assay Procedure

1. Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal it.
2. Add 100 μl of standards, samples and zero controls (standard diluent buffer) into wells. Incubate for 1.5 h at 37°C.
3. Aspirate each well and wash, repeating the process four times for a total five washes. Wash by filling each well with 1× Wash Buffer (350 μl) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each is essential to good performance. After the last wash, remove any remaining Wash Buffer by aspirating, decanting or blotting against clean paper towels.
4. Add 100 μl 1X Antibody conjugate into each well. Cover wells and incubate for 1 hour at 37°C.
5. Aspirate each well and wash as step 3.
6. Add 100 μl of 1X HRP-Streptavidin solution to each well. Cover wells and incubate for 30 minutes at 37°C.
7. Aspirate each well and wash as step 3.
8. Add 100 μl of TMB Reagent to each well. Incubate for 15 minutes at 37°C in dark.
9. Add 100 μl of Stop Solution to each well. The color of the solution should change from blue to yellow.
10. Read the OD with a microplate reader at 450nm immediately.

Typical Standard Curve