The PathScan® Hydroxy-HIF-1α (Pro564) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of hydroxy-HIF-1α (Pro564) protein. A HIF-1α antibody has been coated onto the microwells. After incubation with cell lysates, HIF-1α protein is captured by the coated antibody. Following extensive washing, a Hydroxy-HIF-1α (Pro564) Detection Antibody is added to detect the captured hydroxy-HIF-1α (Pro564) protein. Anti-rabbit IgG, HRP-linked antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of optical density for this developed color is proportional to the quantity of hydroxy-HIF-1α (Pro564) protein.
Antibodies in kit are custom formulations specific to kit.
PathScan® Hydroxy-HIF-1α (Pro564) Sandwich ELISA Kit recognizes endogenous levels of hydroxy-HIF-1α (Pro564) protein, as shown in Figure 1. The kit sensitivity is shown in Figure 2. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.
Hypoxia-inducible factor 1 (HIF1) is a heterodimeric transcription factor that plays a critical role in the cellular response to hypoxia (1). The HIF1 complex consists of two subunits, HIF-1α and HIF-1β, which are basic helix-loop-helix proteins of the PAS (Per, ARNT, Sim) family (2). HIF1 regulates the transcription of a broad range of genes that facilitate responses to the hypoxic environment, including genes regulating angiogenesis, erythropoiesis, cell cycle, metabolism, and apoptosis. The widely expressed HIF-1α is typically degraded rapidly in normoxic cells by the ubiquitin/proteasomal pathway. Under normoxic conditions, HIF-1α is proline hydroxylated leading to a conformational change that promotes binding to the von Hippel Lindau protein (VHL) E3 ligase complex; ubiquitination and proteasomal degradation follows (3,4). Both hypoxic conditions and chemical hydroxylase inhibitors (such as desferrioxamine and cobalt) inhibit HIF-1α degradation and lead to its stabilization. In addition, HIF-1α can be induced in an oxygen-independent manner by various cytokines through the PI3K-AKT-mTOR pathway (5-7).
HIF-1β is also known as AhR nuclear translocator (ARNT) due to its ability to partner with the aryl hydrocarbon receptor (AhR) to form a heterodimeric transcription factor complex (8). Together with AhR, HIF-1β plays an important role in xenobiotics metabolism (8). In addition, a chromosomal translocation leading to a TEL-ARNT fusion protein is associated with acute myeloblastic leukemia (9). Studies also found that ARNT/HIF-1β expression levels decrease significantly in pancreatic islets from patients with type 2 diabetes, suggesting that HIF-1β plays an important role in pancreatic β-cell function (10).
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