HIF-1alpha Transcription Factor Assay Kit
HIF DNA binding
The HIF (hypoxia-inducible factor) transcription factor is a member of the basic-helix-loop-helix (bHLH) family of transcription factors and plays an important role in maintaining cellular oxygen homeostasis. HIF-1.alpha. has emerged as an important drug target in breast and prostate cancer, cardiovascular disease, and ischemia. Cayman's HIF-1.alpha. Transcription Factor Assay is a non-radioactive, sensitive method for detecting specific transcription factor DNA binding activity in nuclear extracts and whole cell lysate. A 96-well enzyme-linked immunosorbent assay (ELISA) replaces the cumbersome radioactive electrophoretic mobility shift assay (EMSA). A specific double stranded DNA (dsDNA) consensus sequence containing the HIF-1.alpha. response element is immobilized to the wells of a 96-well plate. HIF-1.alpha. contained in a nuclear extract or whole cell lysates, binds specifically to the HIF-1.alpha. response element. The HIF transcription factor complex is detected by addition of a specific primary antibody directed against HIF-1.alpha.. A secondary antibody conjugated to HRP is added to provide a sensitive colorimetric readout at 450 nm.