Akt1 (Phospho-Tyr474)Colorimetric Cell-Based ELISA Kit

Assay Genie offers a Cell-Based ELISA Kit designed for the detection of phosphorylated AKT1 at Tyrosine 474 in cell lysates. This colorimetric assay provides a reliable and sensitive tool for measuring phosphorylated AKT1, with reactivity covering Human, Mouse, and Rat samples. The kit is suitable for use with adherent cells and employs a lysate-free, high-throughput format suited to screening the effects that various treatments, inhibitors (such as siRNA or chemicals), or activators have on Akt1 phosphorylation.

Detection is achieved through an indirect ELISA format: Akt1 (Phospho-Tyr474) is captured by phospho-specific primary antibodies, while HRP-conjugated secondary antibodies bind the Fc region of the primary antibody, enabling colorimetric detection at 450 nm upon substrate addition. A monoclonal antibody specific for human GAPDH is included as an internal positive control for normalizing target absorbance values. Following colorimetric measurement of HRP activity, the Crystal Violet whole-cell staining method may be used to determine cell density, with results analysed by normalizing absorbance values to cell amounts to adjust for plating differences. The kit supports a dynamic range of > 5000 Cells.

Phosphorylation of AKT1 at Tyr474 is a critical signaling event that regulates cell growth, survival, and metabolism. Dysregulation of AKT1 signaling is implicated in various diseases, including cancer, and cardiovascular disorders, highlighting the relevance of studying AKT1 signalling for context. Easy-to-follow protocols and quick turnaround times make this kit an essential tool for researchers investigating AKT1 signalling and its implications in health and disease.