IkappaB-alpha (Phospho-Tyr305)Colorimetric Cell-Based ELISA Kit

Assay Genie's cell-based ELISA kit is designed for the qualitative detection of phosphorylated IkappaB-alpha at Tyr305, with a colorimetric readout at 450 nm. It is intended for use with adherent cells, is reactive with human, mouse and rat samples and covers a dynamic range of > 5000 Cells. The assay is described as a convenient, lysate-free, high-throughput format for measuring IkappaB-alpha phosphorylation and expression in cultured cells, suited to studying NF-kB signalling and screening the effects of treatments, inhibitors such as siRNA or chemicals, or activators on IkappaB-alpha phosphorylation.

The indirect ELISA format uses phospho-specific and total primary antibodies, with HRP-conjugated secondary antibodies catalysing a colorimetric reaction on substrate addition. A monoclonal anti-GAPDH antibody is included as an internal positive control for normalising target absorbance, and a Crystal Violet whole-cell staining step is used to determine cell density so absorbance can be normalised to cell amount. The target belongs to the NF-kB inhibitor family (gene symbol NFKBIA; UniProt P25963) and falls within the immunology research area.