DNA Damage Pathway

DNA damage results in cell cycle arrest, apoptosis, and the stabilization and repair of the cellular genome. The Pathway includes downstream targets such as ATM, ATR, CDC25A, Chk1, Chk2, Caspase 3, p21 and signals through P53 dependent and P53 independent processes. DiscoveRx offers novel direct, cell-based, functional assays for predictive toxicology, that are reliable, easy to run, user-friendly and compatible with 384 well protocols. These assays allow you to obtain effect of your early stage compound on key cellular pathways such as DNA damage (p53, CDC25A) or ER stress. No need to wait! Start your testing early.
 

PathHunter® U2OS p53 Nuclear Translocation Assay



—Features and Benefits
  • Non-ELISA based whole cell assay
  • Avoid antibodies, fixing and cell staining
  • Simple chemiluminescent detection

—Applications 

  • Screen  targets upstream of P53 pathway
  • Study compound toxicity profiles (genotoxic markers)
  • Measure effect of compounds on P53, CDC25A or XBP1

—Assay Principle

The PathHunter®  assay monitors the interaction of two proteins in a whole cell, homogeneous assay format using enzyme fragment complementation. The cells have been engineered to express two complementing fragments of β-Gal within   different cellular compartments. In this system, a small 42 AA enzyme fragment is appended to the protein of interest (P53/ CDC25A/ XBP1). The larger enzyme fragment EA (Enzyme Acceptor) resides in the nucleus. Activation of the DNA damage pathway initiates protein translocation to the nucleus that forces complementation of the two β-Gal enzyme fragments. This action results in the formation of fully complemented β-galactosidase enzyme, the activity of which is measured using chemiluminescent substrate. The signal can be read on any standard luminometer

View Categories: Pathway Assays

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