Transcription Factor Activation Kits: HIF-1 alpha, Phospho-CREB and FOXO3a

These kits allow direct measurements of molecular translocation of specific proteins using a fixed end-point assay based on immunofluorescence detection in cells grown on standard high-density microplates. The primary antibodies are specific for their targets and have minimal cross-reactivity with other targets.

The Multiplex and Singleplex Transcription Factor Activation Kits have been optimized for use with the Cellomics ArrayScan® HCS Reader using the Compartmental Analysis BioApplication Software Module. These kits can be used with other Cellomics BioApplications. Thus, automated plate-handling, focusing, cell image acquisition/processing, and data analysis/management are combined in one high content screening (HCS) system to assay for test compounds regulating HIF-1 alpha, phospho-CREB, and FOXO3a expression. In addition to HCS instruments, cells labeled by the reagents in this kit can be viewed and analyzed by other fluorescence microscopes.

Figure 1. A549 cells were challenged with 250 µM desferrioxamine overnight to measure HIF-1 alpha translocation, 1 mM hydrogen peroxide for 30 minutes for phosphorylated CREB, and 10 mM hydrogen peroxide for 1 hour for FOXO3a activation. Cells were also labeled with Hoechst dye for nuclear staining. Top panels represent images from non-treated cells, and bottom panels indicate the various treatments. Stimulation by each drug results in translocation of each transcription factor from the cytosol to the nucleus. DF= desferrioxamine; H₂O₂ = hydrogen peroxide

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